Viral particles targeting hematopoietic stem cells

Abstract

Provided herein are targeted lipid particles, such as targeted lentiviral particles, having a targeting agent that binds to a target molecule selected from the group consisting of ASCT1, ASCT2, CD105, CD110, CD117, CD133, CD146, CD164, CD34, CD46, CD49f, CD90, EPCR, and ITGA3, and related polynucleotides, cells, and methods for preparation and use of the targeted lipid particles, such as lenti viral particles. Also provided herein are methods involving mobilizing bone marrow cells in combination with administering targeted or non-targeted lipid particles, such as lentiviral particles, for treatment of a disease or condition in a subject.

Description

VIRAL PARTICLES TARGETING HEMATOPOIETIC STEM CELLSCross-Reference to Related Applications

[0001] This application claims priority from U.S. provisional application No. 63 / 415,975, filed October, 13, 2022, entitled “VIRAL PARTICLES TARGETING HEMATOPOIETIC STEM CELLS”, U.S. provisional application No. 63 / 460,296, filed April 18, 2023, entitled “VIRAL PARTICLES TARGETING HEMATOPOIETIC STEM CELLS”, to U.S. provisional application No. 63 / 466,683, filed May 15, 2023, entitled “VIRAL PARTICLES TARGETING HEMATOPOIETIC STEM CELLS”, and U.S. provisional application No. 63 / 468,526, filed May 23, 2023, entitled “VIRAL PARTICLES TARGETING HEMATOPOIETIC STEM CELLS”, the contents of which are incorporated by reference in their entirety.Incorperation by Reference of Sequence Listing

[0002] The present application is being filed along with a Sequence Listing in electronic format. The Sequence Listing is provided as a file entitled 18615-20080.40.xml created October 12, 2023, which is 751,853 bytes in size. The information in the electronic format of the Sequence Listing is incorporated by reference in its entirety.Field

[0003] The present disclosure relates to targeted lipid particles, such as targeted lentiviral particles, having a targeting agent that binds to a target molecule selected from the group consisting of ASCT1, ASCT2, CD105, CD110, CD117, CD133, CD146, CD164, CD34, CD46, CD49f, CD90, EPCR, and ITGA3, and related polynucleotides, cells, and methods for preparation and use of the targeted lipid particles. The present disclosure also relates to methods involving mobilizing bone marrow cells in combination with administering targeted or non-targeted lipid particles, such as lentiviral particles, for treatment of a disease or condition in a subject.Background

[0004] Lipid particles, including viral and virus-like particles, such as lentiviral particles, are commonly used for delivery of exogenous agents to cells. However, delivery to certain target cells can be challenging. Improved lipid particles, such as lentiviral particles, and methods for delivery are needed. The provided disclosure addresses this need.Summary

[0005] The embodiments herein provide for lipid particles that allow for efficient targeting and delivery of an exogenous agent contained therein to hematopoietic stem / progenitor cells (HSPC),including to naive hematopoietic stem cells (HSCs). In some embodiments, the targeting can be to such stem or progenitor cells in a human subject, such as human HSPC. In provided embodiments, the lipid particles can include viral vectors such as lentiviral vectors that are pseudotyped with a fusogen with a preferential ability to target HSPC. In some embodiments, the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof, such as the exemplarly fusogens BaEVTR or BaEVRLess. In some embodiments, the lipid particle includes a targeting agent to target the fusogen to HSPCs, such as to CD34+ cells. In some embodiments, the targeting agent is fused to a fusogen, such as a viral envelope attachment glycoprotein, exposed on the lipid bilayer to provide a retargeted fusogen.

[0006] Provided herein in some embodiments is a targeted lipid particle, comprising a lipid bilayer, a fusogen, and a targeting agent that binds to a target molecule selected from the group consisting of ASCT1, ASCT2, CD105, CD110, CD117, CD133, CD146, CD164, CD34, CD46, CD49f, CD90, EPCR, and ITGA3, wherein the fusogen and the targeting agent are exposed on the surface of the lipid bilayer.

[0007] In some embodiments, the targeting agent is fused to a transmembrane domain incorporated into the lipid bilayer. In some embodiments, the targeting agent is fused to the fusogen.

[0008] In some of any embodiments, the particle is a viral particle or virus-like particle. In some of any embodiments, the lipid bilayer is derived from a membrane of a producer cell used for producing a viral particle or virus-like particle. In some of any embodiments, the viral particle or virus-like particle is a retroviral particle or retrovirus-like particle. In some of any embodiments, the viral particle or virus-like particle is a lentivirus-like particle. In some of any embodiments, the viral particle or virus-like particle is a lentiviral particle.

[0009] Provided herein in some embodiments is a targeted lentiviral particle, comprising a fusogen and a targeting agent that binds to a target molecule selected from the group consisting of ASCT1, ASCT2, CD105, CD110, CD117, CD133, CD146, CD164, CD34, CD46, CD49f, CD90, EPCR, and ITGA3, wherein the fusogen and the targeting agent are exposed on the surface of the lentiviral particle.

[0010] In some of any embodiments, the particle is pseudotyped with the fusogen.

[0011] In some of any embodiments, the targeting agent is fused to the fusogen. In some of any embodiments, the targeting agent is fused to the fusogen directly. In some of any embodiments, the targeting agent is fused to the fusogen via a linker.

[0012] In some of any embodiments, the targeting agent is fused to a transmembrane domain incorporated into the viral envelope.

[0013] In some of any embodiments, the particle is prepared by a method including transducing a producer cell with plasmids encoding the fusogen and targeting agent and a Gag-pol and Rev. In some of any embodiments, the producer cell is selected from the group consisting of CHO cells, BHK cells, MDCK cells, C3H 10T1 / 2 cells, FLY cells, Psi-2 cells, BOSC 23 cells, PA317 cells, WEHI cells, COS cells, BSC 1 cells, BSC 40 cells, BMT 10 cells, VERO cells, W138 cells, MRC5 cells, A549 cells,HT1080 cells, 293 cells, 293T cells, B-50 cells, 3T3 cells, NIH3T3 cells, HepG2 cells, Saos-2 cells, Huh7 cells, HeLa cells, W163 cells, 211 cells, and 211 A cells. In some of any embodiments, the producer cell is a 293T cell.

[0014] In some of any embodiments, the particle is replication defective.

[0015] In some of any embodiments, the particle comprises a viral nucleic acid. In some embodiments, the viral nucleic acid is a retroviral nucleic acid. In some of any embodiments, the viral nucleic acid is a lentiviral nucleic acid. In some of any embodiments, the viral nucleic acid comprises one or more, optionally all, of the following nucleic acid sequences: 5’ LTR (e.g., containing U5 and lacking a functional U3 domain), Psi packaging element (Psi), Central polypurine tract (cPPT) / central termination sequence (CTS) (e.g., DNA flap), Poly A tail sequence, a posttranscriptional regulatory element (e.g., WPRE), a Rev response element (RRE), and 3’ LTR (e.g., containing U5 and lacking a functional U3).

[0016] In some of any embodiments, the particle is devoid of viral genomic nucleic acids.

[0017] In some of any embodiments, wherein the target molecule is ASCT2. In some of any embodiments, the target molecule is CD 117.

[0018] In some of any embodiments, the fusogen is a viral fusion protein. In some of any embodiments, the fusogen is a viral envelope protein.

[0019] In some of any embodiments, the fusogen is a VSV-G protein or a functional variant thereof.

[0020] In some of any embodiments, the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof. In some of any embodiments, the fusogen is a truncated BaEV envelope glycoprotein that comprises a cytoplasmic tail with a partial inhibitory R peptide that is less than the full length wild-type BaEV inhibitory R peptide. In some of any embodiments, the fusogen is set forth in any of SEQ ID NO:254-260. In some of any embodiments, the fusogen is set forth in SEQ ID NO:261 (BaEVRLess). In some of any embodiments, the fusogen is set forth in SEQ ID NO:262 (BaEVTR). Provided herein is a targeted lipid particle, comprising a lipid bilayer, a fusogen, wherein the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof and is exposed on the surface of the lipid bilayer.

[0021] In some of any embodiments, the fusogen is a Cocal virus G protein or a functional variant thereof.

[0022] In some of any embodiments, the fusogen is an Alphavirus fusion protein (e.g., Sindbis virus) or a functional variant thereof.

[0023] In some of any embodiments, the fusogen is a Paramyxovirus fusion protein (e.g., a Morbilli virus or Henipavirus) or a functional variant thereof. In some of any embodiments, the fusogen is a Morbillivirus fusion protein (e.g., measles virus (MeV), canine distemper virus, Cetacean morbilli virus, Peste-des-petits-ruminants virus, Phocine distemper virus, or Rinderpest virus) or afunctional variant thereof. In some of any embodiments, the fusogen is a Henipavirus fusion protein (e.g., Nipah virus, Hendra virus, Cedar virus, Kumasi virus, Mojiang virus, or Langya virus) or a functional variant thereof. In some of any embodiments, the fusogen is a Nipah virus fusion protein or a functional variant thereof. In some of any embodiments, the fusogen comprises a Nipah virus F glycoprotein (NiV-F) or a biologically active portion thereof and a Nipah virus G glycoprotein (NiV-G) or a biologically active portion thereof.

[0024] In some embodiments, the targeting agent is fused to the NiV-G protein or biologically active portion thereof. In some of any embodiments, the targeting agent is fused to the C-terminus of the NiV-G protein or biologically active portion thereof.

[0025] In some of any embodiments, the NiV-G or biologically active portion thereof is a wildtype NiV-G protein or a functionally active variant or biologically active portion thereof. In some of any embodiments, the NiV-G protein or biologically active portion thereof is truncated and lacks up to 40 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein set forth in SEQ ID NO:4. In some of any embodiments, the NiV-G protein or biologically active portion thereof is a biologically active portion that is a truncated NiV-G that has a deletion of amino acids 2-34 at or near the N-terminus of wild-type NiV-G set forth in SEQ ID NO:4. In some of any embodiments, the NiV-G protein or biologically active portion thereof does not include an initial methionine. In some of any embodiments, NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO:42 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:42. In some of any embodiments, the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO:42.

[0026] In some of any embodiments, the fusogen comprises one or modifications to reduce binding to its native receptor.

[0027] In some of any embodiments, the NiV-G protein or biologically active portion thereof is a mutant NiV-G protein or biologically active portion thereof that exhibits reduced binding to Ephrin B2 or Ephrin B3. In some of any embodiments, the NiV-G protein or biologically active portion thereof comprises one or more amino acid substitutions corresponding to amino acid substitutions selected from the group consisting of E501A, W504A, Q530A, and E533A with reference to numbering set forth in SEQ ID NO:4. In some of any embodiments, the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO: 17 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 17. In some of any embodiments, the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 17. In some of any embodiments, the NiV-G protein or the biologically active portion has the amino acid sequence set forth in SEQ ID NO: 18 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95%sequence identity to the sequence set forth in SEQ ID NO: 18. In some of any embodiments, the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 18.

[0028] In some of any embodiments, the NiV-F protein or the biologically active portion thereof is a wild-type NiV-F protein or a functionally active variant or biologically active portion thereof. In some of any embodiments, the NiV-F protein or biologically active portion thereof is a truncated NiV-F that is truncated by or by at least 22 amino acids or by or by at least 20 amino acids at or near the C-terminus of wild-type NiV-F set forth in SEQ ID NO:30. In some embodiments, the NiV-F protein or biologically active portion thereof has a 22 amino acid truncation at or near the C-terminus of the wild-type NiV-F protein set forth in SEQ ID NO:30. In some of any embodiments, the NiV-F protein or biologically active portion thereof comprises a deletion in its cytoplasmic tail and lacks amino acid residues 525-546 of SEQ ID NO:30. In some of any embodiments, the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 16 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 16. In some of any embodiments, the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO: 16. In some of any embodiments, the NiV-F protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO:21 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:21. In some of any embodiments, the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO:21.

[0029] In some of any embodiments, the NiV-G protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 18, and the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO:21.

[0030] In some of any embodiments, the targeting agent is an antibody or antibody fragment, a Design ankyrin repeat protein (DARPin), or an antigen-binding fibronectin type III (Fn3) scaffold that binds to the target molecule. In some of any embodiments, the targeting agent is a single domain antibody that binds to the target molecule. In some of any embodiments, the targeting agent is a VHH that binds to the target molecule. In some of any embodiments, the targeting agent is a single chain variable fragment (scFv) that binds to the target molecule.

[0031] In some of any embodiments, the target molecule is expressed on a target cell.

[0032] In some of any embodiments, the particle comprises an exogenous agent for delivery to a target cell. In some embodiments, the target molecule is expressed on the target cell. In some of any of the provided embodiments, the particle comprises a fusion protein between a viral structural protein and an exogenous agent. In some of any of the provided embodiments, the fusion protein is a cleavable fusion protein comprising a cleavable linker positioned between the viral structural protein and the exogenous agent. In some of any of the provided embodiments, the fusion protein comprises one or more nuclearlocalization sequences. In some of any of the provided embodiments, the fusion protein comprises one or more nuclear export sequences.

[0033] In some of any embodiments, the exogenous agent is present in the lumen. In some of any embodiments, the exogenous agent is a protein or a nucleic acid. In some of any embodiments, the exogenous agent is or encodes a therapeutic agent or a diagnostic agent.

[0034] In some of any embodiments, the exogenous agent is or encodes a factor associated with gene editing. In some of any embodiments, the exogenous agent is or encodes a genome-modifying protein for gene editing a target gene encoding an endogenous protein. In some embodiments, the genome-modifying protein is associated with gene editing by a sequence-specific nuclease, a CRISPR- associated transposase (CAST), prime editing, or Programmable Addition via Site-specific Targeting Elements (PASTE).

[0035] In some of any embodiments, the genome -modifying protein is a sequence-specific nuclease. In some of any embodiments, the sequence-specific nuclease is selected from the group consisting of an RNA-guided nuclease, a meganuclease, a transcription activator-like effector nuclease (TALEN), and a zinc-finger nuclease (ZFN). In some of any embodiments, the sequence-specific nuclease is an RNA-guided nuclease. In some of any embodiments, the RNA-guided nuclease is TnpB. In some of any embodiments, the RNA-guided nuclease comprises a Cas nuclease and a guide RNA (CRISPR-Cas combination). In some embodiments, the CRISPR-Cas combination is a ribonucleoprotein (RNP) complex containing the gRNA and the Cas nuclease. In some of any embodiments, the Cas nuclease is a Type II or Type V Cas protein. In some of any embodiments, the genome-modifying protein is selected from the group consisting of Cas3, Cas4, Cas5, Cas8a, Cas8b, Cas8c, Cas9, CaslO, Casl2, Casl2a (Cpfl), Casl2b (C2cl), Casl2c (C2c3), Casl2d (CasY), Casl2e (CasX), Casl2f (C2cl0), Casl2g, Casl2h, Casl2i, Casl2k (C2c5), Casl3, Casl3a (C2c2), Casl3b, Casl3c, Casl3d, C2c4, C2c8, C2c9, Cmr5, Csel, Cse2, Csfl, Csm2, Csn2, CsxlO, Csxll, Csyl, Csy2, Csy3, Mad7. In some of any embodiments, the Cas is a Cas9 or a Cas 12.

[0036] In some of any embodiments, the exogenous agent is or encodes a factor associated with base editing or prime editing (e.g., target-primed reverse transcription (TPRT)). In some of any embodiments, the exogenous agent is or encodes a transposase, integrase, or recombinase. In some of any embodiments, the exogenous agent is or encodes a DNA polymerase, RNA polymerase, or reversetranscriptase.

[0037] In some of any embodiments, the exogenous agent is for use in gene therapy to correct a genetic deficiency in the target cell. In some embodiments, the exogenous agent is a nucleic acid containing a payload gene for correcting the genetic deficiency.

[0038] In some of any embodiments, the exogenous agent is or encodes a membrane protein. In some of any embodiments, the membrane protein is an antigen receptor. In some embodiments, the antigen receptor binds to an antigen expressed on cells associated with a disease or condition. In someembodiments, the disease or condition is a cancer. In some of any embodiments, the antigen receptor binds to an antigen expressed on tumor cells. In some of any embodiments, the antigen receptor is a chimeric antigen receptor (CAR). In some of any embodiments, the antigen receptor is an engineered T cell receptor (TCR).

[0039] In some of any embodiments, the target cell is a hematopoietic cell. In some of any embodiments, the target cell is CD34+. In some of any embodiments, the target cell is a CD34+ progenitor cell. In some of any embodiments, the target cell is a hematopoietic stem cell.

[0040] In some of any embodiments, the delivery to target cells expressing the target molecule is increased by or by greater than 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 1.6-fold, 1.7-fold, 1.8-fold, 1.9-fold, 2-fold, 2.5-fold, 3-fold, 3.5-fold, 4-fold, 4.5-fold, 5-fold, 5.5-fold, 6-fold, 6.5-fold, 7-fold, 7.5- fold, 8-fold, 8.5-fold, 9-fold, 9.5-fold, 10-fold, or more compared to delivery to non-target cells not expressing the target molecule.

[0041] Provided herein in some embodiments is a polynucleotide comprising a nucleic acid sequence encoding a fusogen and a targeting agent that binds to a target molecule selected from the group consisting of ASCT1, ASCT2, CD105, CD110, CD117, CD133, CD146, CD164, CD34, CD46, CD49f, CD90, EPCR, and ITGA3.

[0042] In some embodiments the polynucleotide encodes a transmembrane domain that is fused to the targeting agent.

[0043] In some embodiments, the targeting agent is fused to the fusogen. In some of any embodiments, the targeting agent is fused to the fusogen directly. In some of any embodiments, the targeting agent is fused to the fusogen via a linker.

[0044] In some of any embodiments, the target molecule is ASCT2. In some of any embodiments, the target molecule is CD 117.

[0045] In some of any embodiments, the fusogen is a viral fusion protein. In some of any embodiments, the fusogen is a viral envelope protein.

[0046] In some of any embodiments, the fusogen is a VSV-G protein or a functional variant thereof.

[0047] In some of any embodiments, the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof. In some of any embodiments, the fusogen is a truncated BaEV envelope glycoprotein that comprises a cytoplasmic tail with a partial inhibitory R peptide that is less than the full length wild-type BaEV inhibitory R peptide. In some of any embodiments, the fusogen is set forth in any of SEQ ID NO:254-260. In some of any embodiments, the fusogen is set forth in SEQ ID NO:261 (BaEVRLess). In some of any embodiments, the fusogen is set forth in SEQ ID NO:262 (BaEVTR). Provided herein is a nucleic acid sequence encoding a fusogen, wherein the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof.

[0048] In some of any embodiments, the fusogen is a Cocal virus G protein or a functional variant thereof.

[0049] In some of any embodiments, the fusogen is an Alphavirus fusion protein (e.g., Sindbis virus) or a functional variant thereof.

[0050] In some of any embodiments, the fusogen is a Paramyxovirus fusion protein (e.g., a Morbilli virus or Henipavirus) or a functional variant thereof. In some of any embodiments, the fusogen is a Morbillivirus fusion protein (e.g., measles virus (MeV), canine distemper virus, Cetacean morbilli virus, Peste-des-petits-ruminants virus, Phocine distemper virus, or Rinderpest virus) or a functional variant thereof. In some of any embodiments, the fusogen is a Henipavirus fusion protein (e.g., Nipah virus, Hendra virus, Cedar virus, Kumasi virus, Mojiang virus, or Langya virus) or a functional variant thereof. In some of any embodiments, the fusogen is a Nipah virus fusion protein or a functional variant thereof. In some of any embodiments, the fusogen comprises a Nipah virus F glycoprotein (NiV-F) or a biologically active portion thereof and a Nipah virus G glycoprotein (NiV-G) or a biologically active portion thereof.

[0051] In some embodiments, the targeting agent is fused to the NiV-G protein or biologically active portion thereof. In some of any embodiments, the targeting agent is fused to the C-terminus of the NiV-G protein or biologically active portion thereof.

[0052] In some of any embodiments, the NiV-G or biologically active portion thereof is a wildtype NiV-G protein or a functionally active variant or biologically active portion thereof. In some of any embodiments, the NiV-G protein or biologically active portion thereof is truncated and lacks up to 40 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein set forth in SEQ ID NO:4. In some of any embodiments, the NiV-G protein or biologically active portion thereof is a biologically active portion that is a truncated NiV-G that has a deletion of amino acids 2-34 at or near the N-terminus of wild-type NiV-G set forth in SEQ ID NO:4. In some of any embodiments, the NiV-G protein or biologically active portion thereof does not include an initial methionine. In some of any embodiments, the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO:42 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:42. In some of any embodiments, the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO:42.

[0053] In some of any embodiments, the fusogen comprises one or modifications to reduce binding to its native receptor.

[0054] In some of any embodiments, the NiV-G protein or biologically active portion thereof is a mutant NiV-G protein or biologically active portion thereof that exhibits reduced binding to Ephrin B2 or Ephrin B3. In some of any embodiments, the NiV-G protein or biologically activeportion thereof comprises one or more amino acid substitutions corresponding to amino acid substitutions selected from the group consisting of E501A, W504A, Q530A, and E533A with reference to numbering set forth in SEQ ID NO:4. In some of any embodiments, the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO: 17 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 17. In some of any embodiments, the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 17. In some of any embodiments, the NiV-G protein or the biologically active portion has the amino acid sequence set forth in SEQ ID NO: 18 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 18. In some of any embodiments, the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 18.

[0055] In some of any embodiments, the NiV-F protein or the biologically active portion thereof is a wild-type NiV-F protein or a functionally active variant or biologically active portion thereof. In some of any embodiments, the NiV-F protein or biologically active portion thereof is a truncated NiV-F that is truncated by or by at least 22 amino acids or by or by at least 20 amino acids at or near the C-terminus of wild-type NiV-F set forth in SEQ ID NO:30. In some of any embodiments, the NiV-F protein or biologically active portion thereof has a 22 amino acid truncation at or near the C- terminus of the wild-type NiV-F protein set forth in SEQ ID NO:30. In some of any embodiments, the NiV-F protein or biologically active portion thereof comprises a deletion in its cytoplasmic tail and lacks amino acid residues 525-546 of SEQ ID NO:30. In some of any embodiments, the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 16 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 16. In some of any embodiments, the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO: 16. In some of any embodiments, the NiV-F protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO:21 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:21. In some of any embodiments, the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO:21.

[0056] In some of any embodiments, the NiV-G protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 18, and the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO:21.

[0057] In some of any embodiments, the targeting agent is an antibody or antibody fragment, a Design ankyrin repeat protein (DARPin), or an antigen-binding fibronectin type III (Fn3) scaffold that binds to the target molecule. In some of any embodiments, the targeting agent is a single domain antibody that binds to the target molecule. In some of any embodiments, the targeting agent is a VHHthat binds to the target molecule. In some of any embodiments, the targeting agent is a single chain variable fragment (scFv) that binds to the target molecule.

[0058] In some of any embodiments, the polynucleotide is codon optimized.

[0059] In some of any embodiments, the polynucleotide comprises at least one promoter that is operatively linked to control expression of the targeting agent and fusogen. In some embodiments, the promoter is a constitutive promoter. In some embodiments, the promoter is an inducible promoter.

[0060] Provided herein in some embodiments is a plasmid, comprising the polynucleotide of some of any embodiments. In some embodiments, the plasmid comprises one or more nucleic acids encoding proteins for lentivirus production.

[0061] Provided herein in some embodiments is a vector, comprising the polynucleotide of any of some of any embodiments.

[0062] Provided herein in some embodiments is a cell comprising the polynucleotide of some of any embodiments, the plasmid of some of any embodiments, or the vector of some of any embodiments. Also provided here is a cell comprising (i) a first polynucleotide encoding a nucleic acid sequence encoding a fusogen, wherein the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof.

[0063] Provided herein in some embodiments is a cell comprising (i) a first polynucleotide containing a nucleic acid sequence encoding a fusogen and (ii) a second polynucleotide containing a nucleic acid sequence encoding a targeting agent that binds to a target molecule selected from the group consisting of ASCT1, ASCT2, CD105, CD110, CD117, CD133, CD146, CD164, CD34, CD46, CD49f, CD90, EPCR, and ITGA3.

[0064] In some embodiments, the second polynucleotide encodes a transmembrane domain that is fused to the targeting agent.

[0065] In some of any embodiments, the first and / or second polynucleotide is a plasmid. In some embodiments, the plasmid of the first and / or second polynucleotide comprises one or more nucleic acids encoding proteins for lentivirus production.

[0066] In some of any embodiments, the first and / or second polynucleotide is a vector.

[0067] In some of any embodiments, the first and / or second polynucleotide is codon optimized.

[0068] In some of any embodiments, the first polynucleotide comprises at least one promoter that is operatively linked to control expression of the fusogen; and / or the second polynucleotide comprises at least one promoter that is operatively linked to control expression of the targeting agent. In some embodiments, the promoter of the first and / or second polynucleotide is a constitutive promoter. In some embodiments, the promoter of the first and / or second polynucleotide is an inducible promoter.

[0069] In some of any embodiments, the cell is a producer cell used for producing a viral particle or virus-like particle. In some embodiments, the viral particle or virus-like particle is a retroviral particleor retrovirus-like particle. In some of any embodiments, the viral particle or virus-like particle is a lentiviral particle or lentivirus-like particle. In some of any embodiments, the cell is selected from the group consisting of CHO cells, BHK cells, MDCK cells, C3H 10T1 / 2 cells, FLY cells, Psi-2 cells, BOSC 23 cells, PA317 cells, WEHI cells, COS cells, BSC 1 cells, BSC 40 cells, BMT 10 cells, VERO cells, W138 cells, MRC5 cells, A549 cells, HT1080 cells, 293 cells, 293T cells, B-50 cells, 3T3 cells, NIH3T3 cells, HepG2 cells, Saos-2 cells, Huh7 cells, HeLa cells, W163 cells, 211 cells, and 211 A cells. In some of any embodiments, the cell is a 293T cell.

[0070] In some of any embodiments, the cell comprises a viral nucleic acid. In some embodiments, the viral nucleic acid is a retroviral nucleic acid. In some of any embodiments, the viral nucleic acid is a lentiviral nucleic acid. In some of any embodiments, the viral nucleic acid lacks one or more genes involved in viral replication. In some of any embodiments, the viral nucleic acid comprises a nucleic acid sequence encoding a viral packaging protein selected from one or more of Gag, Pol, and Rev. In some of any embodiments, the viral nucleic acid comprises one or more, optionally all, of the following nucleic acid sequences: 5’ LTR (e.g., containing U5 and lacking a functional U3 domain), Psi packaging element (Psi), Central polypurine tract (cPPT) / central termination sequence (CTS) (e.g., DNA flap), Poly A tail sequence, a posttranscriptional regulatory element (e.g., WPRE), a Rev response element (RRE), and 3’ LTR (e.g., containing U5 and lacking a functional U3).

[0071] In some of any embodiments, the target molecule is ASCT2. In some of any embodiments, the target molecule is CD 117.

[0072] In some of any embodiments, the fusogen is a viral fusion protein. In some of any embodiments, the fusogen is a viral envelope protein.

[0073] In some of any embodiments, the fusogen is a VSV-G protein or a functional variant thereof.

[0074] In some of any embodiments, the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof. In some of any embodiments, the fusogen is a truncated BaEV envelope glycoprotein that comprises a cytoplasmic tail with a partial inhibitory R peptide that is less than the full length wild-type BaEV inhibitory R peptide. In some of any embodiments the fusogen is set forth in any of SEQ ID NO:254-260. In some of any embodiments, the fusogen is set forth in SEQ ID NO:261 (BaEVRLess). In some of any embodiments, the fusogen is set forth in SEQ ID NO:262 (BaEVTR).

[0075] In some of any embodiments, the fusogen is a Cocal virus G protein or a functional variant thereof.

[0076] In some embodiments, the fusogen is an Alphavirus fusion protein (e.g., Sindbis virus) or a functional variant thereof.

[0077] In some of any embodimtents, the fusogen is a Paramyxovirus fusion protein (e.g., a Morbilli virus or Henipavirus) or a functional variant thereof. In some of any embodiments, the fusogenis a Morbillivirus fusion protein (e.g., measles virus (MeV), canine distemper virus, Cetacean morbilli virus, Peste-des-petits-ruminants virus, Phocine distemper virus, or Rinderpest virus) or a functional variant thereof. In some of any embodiments, the fusogen is a Henipavirus fusion protein (e.g., Nipah virus, Hendra virus, Cedar virus, Kumasi virus, Mojiang virus, or Langya virus) or a functional variant thereof. In some of any embodiments, the fusogen is a Nipah virus fusion protein or a functional variant thereof. In some of any embodiments, the fusogen comprises a Nipah virus F glycoprotein (NiV-F) or a biologically active portion thereof and a Nipah virus G glycoprotein (NiV-G) or a biologically active portion thereof.

[0078] In some embodiments, the NiV-G or biologically active portion thereof is a wild-type NiV- G protein or a functionally active variant or biologically active portion thereof. In some of any embodiments, the NiV-G protein or biologically active portion thereof is truncated and lacks up to 40 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein set forth in SEQ ID NO:4. In some of any embodiments, the NiV-G protein or biologically active portion thereof is a biologically active portion that is a truncated NiV-G that has a deletion of amino acids 2-34 at or near the N-terminus of wild-type NiV-G set forth in SEQ ID NO:4. In some of any embodiments, the NiV-G protein or biologically active portion thereof does not include an initial methionine. In some of any embodiments, the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO:42 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:42. In some of any embodiments, the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO:42.

[0079] In some of any embodiments, the fusogen comprises one or more modifications to reduce binding to its native receptor.

[0080] In some of any embodiments, the NiV-G protein or biologically active portion thereof is a mutant NiV-G protein or biologically active portion thereof that exhibits reduced binding to Ephrin B2 or Ephrin B3. In some of any embodiments, the NiV-G protein or biologically active portion thereof comprises one or more amino acid substitutions corresponding to amino acid substitutions selected from the group consisting of E501A, W504A, Q530A, and E533A with reference to numbering set forth in SEQ ID NO:4. In some of any embodiments, the NiV-G protein or biologically active portion thereof comprises the amino acid sequence set forth in SEQ ID NO: 17 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 17. In some of any embodiments, the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 17. In some of any embodiments, the NiV-G protein or the biologically active portion has the amino acid sequence set forth in SEQ ID NO: 18 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 18. In some of anyembodiments, the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO:18.

[0081] In some of any embodiments, the NiV-F protein or the biologically active portion thereof is a wild-type NiV-F protein or a functionally active variant or biologically active portion thereof. In some of any embodiments, the NiV-F protein or biologically active portion thereof is a truncated NiV-F that is truncated by or by at least 22 amino acids or by or by at least 20 amino acids at or near the C-terminus of wild-type NiV-F set forth in SEQ ID NO:30. In some of any embodiments, the NiV-F protein or biologically active portion thereof has a 22 amino acid truncation at or near the C- terminus of the wild-type NiV-F protein set forth in SEQ ID NO:30. In some of any embodiments, the NiV-F protein or biologically active portion thereof comprises a deletion in its cytoplasmic tail and lacks amino acid residues 525-546 of SEQ ID NO:30. In some of any embodiments, the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 16 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 16. In some of any embodiments, the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO: 16. In some of any embodiments, the NiV-F protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO:21 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:21. In some of any embodiments, the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO:21.

[0082] In some of any embodiments, the NiV-G protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 18, and the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO:21.

[0083] In some of any embodiments, the targeting agent is an antibody or antibody fragment, a Design ankyrin repeat protein (DARPin), or an antigen-binding fibronectin type III (Fn3) scaffold that binds to the target molecule. In some of any embodiments, the targeting agent is a single domain antibody that binds to the target molecule. In some of any embodiments, the targeting agent is a VHH that binds to the target molecule. In some of any embodiments, the targeting agent is a single chain variable fragment (scFv) that binds to the target molecule.

[0084] Provided herein in some embodiments is a method of making a targeted lipid particle, comprising: (a) introducing into a source cell the polynucleotide of any of some of any embodiments, the plasmid of some of any embodiments or the vector of some of any embodiments; (b) culturing the source cell under conditions for producing a lipid particle containing a lipid bilayer and the targeting agent and fusogen exposed on the surface of the lipid bilayer; and (c) separating, enriching, or purifying the lipid particle from the source cell, thereby making the lipid particle.

[0085] Provided herein in some embodiments is a method of making a targeted lipid particle, comprising: (a) introducing into a source cell (i) a first polynucleotide containing a nucleic acid sequenceencoding a fusogen and (ii) a second polynucleotide containing a nucleic acid sequence encoding a targeting agent that binds to a target molecule selected from the group consisting of ASCT1, ASCT2, CD105, CD110, CD117, CD133, CD146, CD164, CD34, CD46, CD49f, CD90, EPCR, and ITGA3; (b) culturing the source cell under conditions for producing a lipid particle containing a lipid bilayer and the targeting agent and fusogen exposed on the surface of the lipid bilayer; and (c) separating, enriching, or purifying the lipid particle from the source cell, thereby making the lipid particle.

[0086] In some embodiments, the second polynucleotide encodes a transmembrane domain that is fused to the targeting agent.

[0087] In some of any embodiments, the first and / or second polynucleotide is a plasmid. In some embodiments, the plasmid of the first and / or second polynucleotide comprises one or more nucleic acids encoding proteins for lentivirus production.

[0088] In some of any embodiments, the first and / or second polynucleotide is a vector.

[0089] In some of any embodiments, the first and / or second polynucleotide is codon optimized.

[0090] In some of any embodiments, the first polynucleotide comprises at least one promoter that is operatively linked to control expression of the fusogen; and / or the second polynucleotide comprises at least one promoter that is operatively linked to control expression of the targeting agent. In some embodiments, the promoter of the first and / or second polynucleotide is a constitutive promoter. In some embodiments, the promoter of the first and / or second polynucleotide is an inducible promoter.

[0091] In some of any embodiments, the source cell is a mammalian cell. In some of any embodiments, the source cell is a producer cell used for producing a viral particle or virus-like particle. In some embodiments, the particle is a viral particle or virus-like particle. In some of any embodiments, the viral particle or virus-like particle is a retroviral particle or retrovirus-like particle. In some of any embodiments, the viral particle or virus-like particle is a lentivirus-like particle. In some of any embodiments, the viral particle or virus-like particle is a lentiviral particle. In some of any embodiments, the particle is pseudotyped with the fusogen. In some of any embodiments, the producer cell is selected from the group consisting of CHO cells, BHK cells, MDCK cells, C3H 10T1 / 2 cells, FLY cells, Psi-2 cells, BOSC 23 cells, PA317 cells, WEHI cells, COS cells, BSC 1 cells, BSC 40 cells, BMT 10 cells, VERO cells, W138 cells, MRC5 cells, A549 cells, HT1080 cells, 293 cells, 293T cells, B-50 cells, 3T3 cells, NIH3T3 cells, HepG2 cells, Saos-2 cells, Huh7 cells, HeLa cells, W163 cells, 211 cells, and 211 A cells. In some of any embodiments, the producer cell is a 293T cell.

[0092] In some of any embodiments, the method comprises transducing the source cell with packaging plasmids encoding a Gag-pol and Rev.

[0093] In some of any embodiments, the target molecule is ASCT2. In some of any embodiments, the target molecule is CD 117.

[0094] In some of any embodiments, the fusogen is a viral fusion protein. In some of any embodiments, the fusogen is a viral envelope protein.

[0095] In some of any embodiments, the fusogen is a VSV-G protein or a functional variant thereof.

[0096] In some of any embodiments, the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof. In some of any embodiments, the fusogen is a truncated BaEV envelope glycoprotein that comprises a cytoplasmic tail with a partial inhibitory R peptide that is less than the full length wild-type BaEV inhibitory R peptide. In some of any embodiments, the fusogen is set forth in any of SEQ ID NO:254-260. In some of any embodiments, the fusogen is set forth in SEQ ID NO:261 (BaEVRLess). In some of any embodiments, the fusogen is set forth in SEQ ID NO:262 (BaEVTR).

[0097] In some of any embodiments, the fusogen is a Cocal virus G protein or a functional variant thereof.

[0098] In some of any embodiments, the fusogen is an Alphavirus fusion protein (e.g., Sindbis virus) or a functional variant thereof.

[0099] In some of any embodiments, the fusogen is a Paramyxovirus fusion protein (e.g., a Morbilli virus or Henipavirus) or a functional variant thereof. In some of any embodiments, the fusogen is a Morbillivirus fusion protein (e.g., measles virus (MeV), canine distemper virus, Cetacean morbilli virus, Peste-des-petits-ruminants virus, Phocine distemper virus, or Rinderpest virus) or a functional variant thereof. In some of any embodiments, the fusogen is a Henipavirus fusion protein (e.g., Nipah virus, Hendra virus, Cedar virus, Kumasi virus, Mojiang virus, or Langya virus) or a functional variant thereof. In some of any embodiments, the fusogen is a Nipah virus fusion protein or a functional variant thereof. In some of any embodiments, the fusogen comprises a Nipah virus F glycoprotein (NiV-F) or a biologically active portion thereof and a Nipah virus G glycoprotein (NiV-G) or a biologically active portion thereof.

[0100] In some embodiments, the NiV-G or biologically active portion thereof is a wild-type NiV- G protein or a functionally active variant or biologically active portion thereof. In some of any embodiments, the NiV-G protein or biologically active portion thereof is truncated and lacks up to 40 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein set forth in SEQ ID NO:4. In some of any embodiments, the NiV-G protein or biologically active portion thereof is a biologically active portion that is a truncated NiV-G that has a deletion of amino acids 2-34 at or near the N-terminus of wild-type NiV-G set forth in SEQ ID NO:4. In some of any embodiments, the NiV-G protein or biologically active portion thereof does not include an initial methionine. In some of any embodiments, the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO:42 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:42. Insome of any embodiments, the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO:42.

[0101] In some of any embodiments, the fusogen comprises one or modifications to reduce binding to its native receptor.

[0102] In some of any embodiments, the NiV-G protein or biologically active portion thereof is a mutant NiV-G protein or biologically active portion thereof that exhibits reduced binding to Ephrin B2 or Ephrin B3. In some of any embodiments, the NiV-G protein or biologically active portion thereof comprises one or more amino acid substitutions corresponding to amino acid substitutions selected from the group consisting of E501A, W504A, Q530A, and E533A with reference to numbering set forth in SEQ ID NO:4. In some of any embodiments, the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO: 17 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 17. In some of any embodiments, the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 17. In some of any embodiments, the NiV-G protein or the biologically active portion has the amino acid sequence set forth in SEQ ID NO: 18 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 18. In some of any embodiments, the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 18.

[0103] In some of any embodiments, the NiV-F protein or the biologically active portion thereof is a wild-type NiV-F protein or a functionally active variant or biologically active portion thereof. In some of any embodiments, the NiV-F protein or biologically active portion thereof is a truncated NiV-F that is truncated by or by at least 22 amino acids or by or by at least 20 amino acids at or near the C-terminus of wild-type NiV-F set forth in SEQ ID NO:30. In some of any embodiments, the NiV-F protein or biologically active portion thereof has a 22 amino acid truncation at or near the C- terminus of the wild-type NiV-F protein set forth in SEQ ID NO:30. In some of any embodiments, the NiV-F protein or biologically active portion thereof comprises a deletion in its cytoplasmic tail and lacks amino acid residues 525-546 of SEQ ID NO:30. In some of any embodiments, the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 16 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 16. In some of any embodiments, the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO: 16. In some of any embodiments, the NiV-F protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO:21 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:21. In some of any embodiments, the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO:21.

[0104] In some of any embodiments, the NiV-G protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 18, and the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO:21.

[0105] In some of any embodiments, the targeting agent is an antibody or antibody fragment, a Design ankyrin repeat protein (DARPin), or an antigen-binding fibronectin type III (Fn3) scaffold that binds to the target molecule. In some of any embodiments, the targeting agent is a single domain antibody that binds to the target molecule. In some of any embodiments, the targeting agent is a VHH that binds to the target molecule. In some of any embodiments, the targeting agent is a single chain variable fragment (scFv) that binds to the target molecule.

[0106] Provided herein in some embodiments is a targeted lipid particle produced by the method of some of any embodiments.

[0107] Provided herein in some embodiments is a targeted lentiviral particle produced by the method of some of any embodiments.

[0108] Provided herein in some embodiments is a composition comprising a plurality of the lipid particle of some of any embodiments.

[0109] Provided herein in some embodiments is a composition comprising a plurality of the lentiviral particle of some of any embodiments.

[0110] In some of any embodiments, the composition further comprises a pharmaceutically acceptable excipient.

[0111] Provided herein in some embodiments is a method of transducing a target cell, the method containing contacting a target cell with the lentiviral particle of some of any embodiments or the composition of some of any embodiments.

[0112] In some embodiments, the particle comprises an exogenous agent, and the transduction introduces the exogenous agent into the target cell.

[0113] Provided herein in some embodiments is a method of delivering an exogenous agent to a target cell, the method containing contacting a target cell with the particle of some of any embodiments or the composition of some of any embodiments, the particle or plurality of particles containing an exogenous agent for delivery of the target cell.

[0114] In some of any embodiments, the contacting is in vitro or ex vivo.

[0115] In some of any embodiments, wherein the contacting is in vivo in a subject.

[0116] Provided herein in some embodiments is a method of delivering an exogenous agent to a target cell in a subject, which includes administering to a subject the particle of some of any embodiments or the composition of some of any embodiments, the particle or plurality of particles containing an exogenous agent for delivery to a target cell in the subject.

[0117] In some of any embodiments, the exogenous agent is or encodes a therapeutic agent or a diagnostic agent. In some of any embodiments, the exogenous agent is for treating a disease or condition in the subject.

[0118] Provided herein in some embodiments is a method of treating a disease or condition in a subject, which includes administering to a subject the particle of some of any embodiments or the composition of some of any embodiments, the particle or plurality of particles containing an exogenous agent for treating a disease or condition in the subject.

[0119] In some of any embodiments, the method comprises administering to the subject one or more agents that stimulate mobilization of bone marrow cells from the bone marrow to the peripheral blood.

[0120] In some of any embodiments, the subject has previously been administered one or more agents that stimulate mobilization of bone marrow cells from the bone marrow to the peripheral blood.

[0121] Provided herein in some of any embodiments is a method of treating a disease or condition in a subject, which includes administering to a subject: (a) one or more agents that stimulate mobilization of bone marrow cells from the bone marrow to the peripheral blood; and (b) a lipid particle containing a lipid bilayer, a fusogen exposed on the surface of the lipid bilayer, and an exogenous agent for treating a disease or condition in the subject.

[0122] In some of any embodiments, the administration of at least one of the one or more agents that stimulate mobilization is initiated prior to the administration of the particle.

[0123] Provided herein in some of any embodiments is a method of treating a disease or condition in a subject, containing administering to a subject a lipid particle containing a lipid bilayer, a fusogen exposed on the surface of the lipid bilayer, and an exogenous agent for treating a disease or condition in the subject, wherein the subject has previously been administered one or more agents that stimulate mobilization of bone marrow cells from the bone marrow to the peripheral blood.

[0124] In some of any embodiments, the exogenous agent is for delivery to a target cell for treating the disease or condition in the subject.

[0125] In some of any embodiments, the particle is a viral particle or virus-like particle. In some of any embodiments, the lipid bilayer is derived from a membrane of a producer cell used for producing a viral particle or virus-like particle. In some of any embodiments, the particle is prepared by a method containing transducing a producer cell with plasmids encoding the fusogen and a Gag-pol and Rev. In some of any embodiments, the viral particle or virus-like particle is a retroviral particle or retrovirus-like particle. In some of any embodiments, the viral particle or virus-like particle is a lentivirus-like particle. In some of any embodiments, the viral particle or virus-like particle is a lentiviral particle.

[0126] In some of any embodiments, the particle is pseudotyped with the fusogen.

[0127] In some of any embodiments, the producer cell is selected from the group consisting of CHO cells, BHK cells, MDCK cells, C3H 10T1 / 2 cells, FLY cells, Psi-2 cells, BOSC 23 cells, PA317 cells, WEHI cells, COS cells, BSC 1 cells, BSC 40 cells, BMT 10 cells, VERO cells, W138 cells, MRC5 cells, A549 cells, HT1080 cells, 293 cells, 293T cells, B-50 cells, 3T3 cells, NIH3T3 cells, HepG2 cells, Saos-2 cells, Huh7 cells, HeLa cells, W163 cells, 211 cells, and 211A cells. In some of any embodiments, the producer cell is a 293T cell.

[0128] In some of any embodiments, the particle is replication defective.

[0129] In some of any embodiments, the particle comprises a viral nucleic acid. In some embodiments, the viral nucleic acid is a retroviral nucleic acid. In some of any embodiments, the viral nucleic acid is a lentiviral nucleic acid. In some of any embodiments, the viral nucleic acid comprises one or more, optionally all, of the following nucleic acid sequences: 5’ LTR (e.g., containing U5 and lacking a functional U3 domain), Psi packaging element (Psi), Central polypurine tract (cPPT) / central termination sequence (CTS) (e.g., DNA flap), Poly A tail sequence, a posttranscriptional regulatory element (e.g., WPRE), a Rev response element (RRE), and 3’ LTR (e.g., containing U5 and lacking a functional U3).

[0130] In some of any embodiments, the particle is devoid of viral genomic nucleic acids.

[0131] In some of any embodiments, the fusogen is a viral fusion protein. In some of any embodiments, the fusogen is a viral envelope protein.

[0132] In some of any embodiments, the fusogen is a VSV-G protein or a functional variant thereof.

[0133] In some of any embodiments, the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof. In some of any embodiments, the fusogen is a truncated BaEV envelope glycoprotein that comprises a cytoplasmic tail with a partial inhibitory R peptide that is less than the full length wild-type BaEV inhibitory R peptide. In some of any embodiments, the fusogen is set forth in any of SEQ ID NO:254-260. In some of any embodiments, the fusogen is set forth in SEQ ID NO:261 (BaEVRLess). In some of any embodiments, the fusogen is set forth in SEQ ID NO:262 (BaEVTR).

[0134] In some of any embodiments, the fusogen is a Cocal virus G protein or a functional variant thereof.

[0135] In some of any embodiments, the fusogen is an Alphavirus fusion protein (e.g., Sindbis virus) or a functional variant thereof.

[0136] In some of any embodiments, the fusogen is a Paramyxovirus fusion protein (e.g., a Morbilli virus or Henipavirus) or a functional variant thereof. In some of any embodiments, the fusogen is a Morbillivirus fusion protein (e.g., measles virus (MeV), canine distemper virus, Cetacean morbilli virus, Peste-des-petits-ruminants virus, Phocine distemper virus, or Rinderpest virus) or afunctional variant thereof. In some of any embodiments, the fusogen is a Henipavirus fusion protein (e.g., Nipah virus, Hendra virus, Cedar virus, Kumasi virus, Mojiang virus, or Langya virus) or a functional variant thereof. In some of any embodiments, the fusogen is a Nipah virus fusion protein or a functional variant thereof. In some of any embodiments, the fusogen comprises a Nipah virus F glycoprotein (NiV-F) or a biologically active portion thereof and a Nipah virus G glycoprotein (NiV-G) or a biologically active portion thereof.

[0137] In some embodiments, the NiV-G or biologically active portion thereof is a wild-type NiV- G protein or a functionally active variant or biologically active portion thereof. In some of any embodiments, the NiV-G protein or biologically active portion thereof is truncated and lacks up to 40 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein set forth in SEQ ID NO:4. In some of any embodiments, the NiV-G protein or biologically active portion thereof is a biologically active portion that is a truncated NiV-G that has a deletion of amino acids 2-34 at or near the N-terminus of wild-type NiV-G set forth in SEQ ID NO:4. In some of any embodiments, the NiV-G protein or biologically active portion thereof does not include an initial methionine. In some of any embodiments, the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO:42 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:42. In some of any embodiments, the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO:42.

[0138] In some of any embodiments, the fusogen comprises one or modifications to reduce binding to its native receptor.

[0139] In some of any embodiments, the NiV-G protein or biologically active portion thereof is a mutant NiV-G protein or biologically active portion thereof that exhibits reduced binding to Ephrin B2 or Ephrin B3. In some of any embodiments, the NiV-G protein or biologically active portion thereof comprises one or more amino acid substitutions corresponding to amino acid substitutions selected from the group consisting of E501A, W504A, Q530A, and E533A with reference to numbering set forth in SEQ ID NO:4. In some of any embodiments, the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO: 17 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 17. In some of any embodiments, the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 17. In some of any embodiments, the NiV-G protein or the biologically active portion has the amino acid sequence set forth in SEQ ID NO: 18 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 18. In some of any embodiments, the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 18.

[0140] In some of any embodiments, the NiV-F protein or the biologically active portion thereof is a wild-type NiV-F protein or a functionally active variant or biologically active portion thereof. In some of any embodiments, the NiV-F protein or biologically active portion thereof is a truncated NiV-F that is truncated by or by at least 22 amino acids or by or by at least 20 amino acids at or near the C-terminus of wild-type NiV-F set forth in SEQ ID NO:30. In some of any embodiments, the NiV-F protein or biologically active portion thereof has a 22 amino acid truncation at or near the C- terminus of the wild-type NiV-F protein set forth in SEQ ID NO:30. In some of any embodiments, the NiV-F protein or biologically active portion thereof comprises a deletion in its cytoplasmic tail and lacks amino acid residues 525-546 of SEQ ID NO:30. In some of any embodiments, the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 16 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 16. In some of any embodiments, the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO: 16. In some of any embodiments, the NiV-F protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO:21 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:21. In some of any embodiments, the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO:21.

[0141] In some of any embodiments, the NiV-G protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 18, and the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO:21.

[0142] In some of any embodiments, the bone marrow cells are CD34+. In some of any embodiments, the bone marrow cells are CD34+ progenitor cells. In some of any embodiments, the bone marrow cells are hematopoietic stem cells.

[0143] In some of any embodiments, the one or more agents that stimulate mobilization are selected from the group consisting of stem cell factor (SCF), small molecule VLA-4 inhibitor BI05192, BOP (N-(benzenesnlfonyl)-L-prolyl-L-0-(l-pyrrolidinylcarbonyl)tyrosine), heparin, granulocyte colonystimulating factor (G-CSF), MGTA-145, and plerixafor (AMD3100).

[0144] In some of any embodiments, the one or more agents that stimulate mobilization include G- CSF. In some embodiments, the G-CSF is administered to the subject daily on the two days, three days, four days, or five days prior to the administration of the first dose of the particle. In some of any embodiments, the G-CSF is administered to the subject on the day of the administration of the first dose of the particle. In some of any embodiments, the G-CSF is administered to the subject on the day of administration of a second or later dose of the particle.

[0145] In some of any embodiments, the one or more mobilization agents comprise plerixafor. In some of any embodiments, the plerixafor is administered to the subject on the day of the administrationof the first dose of the particle. In some of any embodiments, the plerixafor is administered to the subject on the day of the administration of a second or later dose of the particle.

[0146] In some of any embodiments, the one or more agents that stimulate mobilization are G-CSF and plerixafor. In some embodiments, the G-CSF is administered to the subject daily on the four days prior to the administration of the first dose of the particle; and the plerixafor is administered to the subject on the day of the administration of the first dose of the particle. In some embodiments, the G-CSF is administered to the subject (i) daily on the two days prior to the administration of the particle; (ii) on the day of the administration of the first dose of the particle; and (iii) on the day of administration of a second or later dose of the particle; and the plerixafor is administered to the subject on the day of administration of a second or later dose of the particle.

[0147] In some of any embodiments, the method comprises administering to the subject a transduction adjuvant. In some of any embodiments, the transduction adjuvant is administered to the subject on the day of the administration of at least one dose of the particle. In some of any embodiments, the transduction adjuvant is administered to the subject on the days of the administration of at least two doses of the particle.

[0148] In some of any embodiments, the transduction adjuvant is a cationic peptide. In some embodiments, the transduction adjuvant is a histidine-rich cationic peptide. In some of any embodiments, the transduction adjuvant is a cationic amphipathic peptide. In some of any embodiments, the transduction adjuvant is derived from the LAH4 peptide family. In some of any embodiments, the transduction adjuvant is Vectofusin-1.

[0149] In some embodiments, the target molecule is expressed on the target cell. In some of any embodiments, the target cell is a hematopoietic cell. In some of any embodiments, the target cell is CD34+. In some of any embodiments, the target cell is a CD34+ progenitor cell. In some of any embodiments, the hematopoietic cell is a hematopoietic stem cell.

[0150] In some of any embodiments, the exogenous agent is present in the lumen. In some of any embodiments, the exogenous agent is a protein or a nucleic acid.

[0151] In some of any embodiments, the exogenous agent is or encodes a factor associated with gene editing. In some of any embodiments, the exogenous agent is or encodes a genome-modifying protein for gene editing a target gene encoding an endogenous protein. In some embodiments, the exogenous agent is a fusion protein between a viral structural protein and an exogenous agent. In some of any of the provided embodiments, the fusion protein is a cleavable fusion protein comprising a cleavable linker positioned between the viral structural protein and the exogenous agent. In some of any of the provided embodiments, the fusion protein comprises one or more nuclear localization sequences. In some of any of the provided embodiments, the fusion protein comprises one or more nuclear export sequences.

[0152] In some embodiments, the genome-modifying protein is associated with gene editing by a sequence-specific nuclease, a CRISPR-associated transposase (CAST), prime editing, or Programmable Addition via Site-specific Targeting Elements (PASTE).

[0153] In some of any embodiments, the genome -modifying protein is a sequence-specific nuclease. In some of any embodiments, the sequence-specific nuclease is selected from the group consisting of an RNA-guided nuclease, a meganuclease, a transcription activator-like effector nuclease (TALEN), and a zinc-finger nuclease (ZFN). In some of any embodiments, the sequence-specific nuclease is an RNA-guided nuclease. In some of any embodiments, the RNA-guided nuclease is TnpB. In some of any embodiments, the RNA-guided nuclease comprises a Cas nuclease and a guide RNA (CRISPR-Cas combination). In some embodiments, the CRISPR-Cas combination is a ribonucleoprotein (RNP) complex containing the gRNA and the Cas nuclease. In some of any embodiments, the Cas nuclease is a Type II or Type V Cas protein. In some of any embodiments, the genome-modifying protein is selected from the group consisting of Cas3, Cas4, Cas5, Cas8a, Cas8b, Cas8c, Cas9, CaslO, Casl2, Casl2a (Cpfl), Casl2b (C2cl), Casl2c (C2c3), Casl2d (CasY), Casl2e (CasX), Casl2f (C2cl0), Casl2g, Casl2h, Casl2i, Casl2k (C2c5), Casl3, Casl3a (C2c2), Casl3b, Casl3c, Casl3d, C2c4, C2c8, C2c9, Cmr5, Csel, Cse2, Csfl, Csm2, Csn2, CsxlO, Csxll, Csyl, Csy2, Csy3, Mad7. In some of any embodiments, the Cas is a Cas9 or a Cas 12.

[0154] In some of any embodiments, the exogenous agent is or encodes a factor associated with base editing or prime editing (e.g., target-primed reverse transcription (TPRT)). In some of any embodiments, the exogenous agent is or encodes a transposase, integrase, or recombinase. In some of any embodiments, the exogenous agent is or encodes a DNA polymerase, RNA polymerase, or reversetranscriptase.

[0155] In some of any embodiments, the exogenous agent is for use in gene therapy to correct a genetic deficiency in the target cell. In some embodiments, the exogenous agent is a nucleic acid containing a payload gene for correcting the genetic deficiency.

[0156] In some of any embodiments, the exogenous agent is or encodes a membrane protein. In some embodiments, the membrane protein is an antigen receptor. In some embodiments, the antigen receptor binds to an antigen expressed on cells associated with a disease or condition in the subject. In some embodiments, the disease or condition is a cancer. In some of any embodiments, the antigen receptor binds to an antigen expressed on tumor cells in the subject. In some of any embodiments, the antigen receptor is a chimeric antigen receptor (CAR). In some of any embodiments, the antigen receptor is an engineered T cell receptor (TCR).

[0157] In some of any embodiments, the delivery to target cells expressing the target molecule is increased by or by greater than 1.1-fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 1.6-fold, 1.7-fold, 1.8-fold, 1.9-fold, 2-fold, 2.5-fold, 3-fold, 3.5-fold, 4-fold, 4.5-fold, 5-fold, 5.5-fold, 6-fold, 6.5-fold, 7-fold, 7.5-fold, 8-fold, 8.5-fold, 9-fold, 9.5-fold, 10-fold, or more compared to delivery to non-target cells not expressing the target molecule. In some of any embodiments, the subject is a human subject.

[0158] In some of any of the provided embodiments, the target molecule is CD 133. In some of any of the provided embodiments, the fusogen is fused to a targeting agent that binds to a target molecule selected from the group consisting of ASCT1, ASCT2, CD105, CD110, CD117, CD133, CD146, CD164, CD34, CD46, CD49f, CD90, EPCR, and ITGA3. In some of any of the provided embodiments, the target molecule is ASCT2. In some of any of the provided embodiments, is CD117.

[0159] In some of any of the provided embodiments, the target molecule is CD 133. In some of any of the provided embodiments, the targeting agent is a CD 133 binding agent that is an antibody or antigen-binding fragment comprising a variable heavy (VH) chain and a variable light (VL) chain selected from a) a VH chain comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 271, 272, 273, respectively, and a VL chain comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 275, 276, 277, respectively; (b) a VH chain comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 280, 281, 282, respectively, and a VL chain comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 284, 285, and 286, respectively; (c) a VH chain comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 298, 299, and 300, respectively, and a VL chain comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 302, 303, and 304, respectively; (d) a VH chain comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 307, 308, and 309, respectively, and a VL chain comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 311, 312, and 313, respectively; or (e) a VH chain comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 298, 290, and 291, respectively, and a VL chain comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 293, 294, and 295, respectively. In some of any of the provided embodiments, the targeting agent is a CD 133 binding agent that is an antibody or antigen-binding fragment comprising a variable heavy (VH) chain and a variable light (VL) chain selected from a) a VH comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 314, 315, and 273, respectively, and a VL comprising a CDR- Ll, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 275, 276, 277, respectively; (b) a VH comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 316, 317, and 282, respectively, and a VL comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 284, 285, and 286, respectively; (c) a VH comprising a a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 318, 319, and 300, respectively, and a VL comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 302, 303, and 304, respectively; (d) a VH comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 320, 321, and 309, respectively, and a VL comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 311, 312, and 313, respectively; and (e) a VH comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 322, 323, and 291, respectively, and a VL comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 293, 294, and 295, respectively.

[0160] In some of any of the provided embodiments, the CD 133 binding agent is selected from a) a CD 133 binding agent comprising a heavy chain variable (VH) region comprising the amino acid sequence of SEQ ID NO: 270, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a light chain variable (VL) region comprising the amino acid sequence of SEQ ID NO: 274, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; b) a CD133 binding agent comprising a VH region comprising the amino acid sequence of SEQ ID NO: 279, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a VL region comprising the amino acid sequence of SEQ ID NO: 283, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; c) a CD 133 binding agent comprising a VH region comprising the amino acid sequence of SEQ ID NO: 297, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a VL region comprising the amino acid sequence of SEQ ID NO: 301, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; d) a CD133 binding agent comprising a VH region comprising the amino acid sequence of SEQ ID NO: 306, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a VL region comprising the amino acid sequence of SEQ ID NO: 310, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; e) a CD133 binding agent comprising a VH region comprising the amino acid sequence of SEQ ID NO: 288, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a VL region comprising the amino acid sequence of SEQ ID NO: 292, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto.

[0161] In some of any of the provided embodiments, the CD 133 binding agent is a scFv. In some of any of the provided embodiments, the CD 133 binding agent comprises the amino acid sequence set forth in any one of SEQ ID NOs: 269, 278, 287, 296 or 305, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto.

[0162] In some of any of the provided embodiments, the targeting agent is a CD117 binding agent that is a VHH comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 324-374, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto. In some of any of the provided embodiments, the CD117 binding agent comprises the amino acid sequence set froth in any one of SEQ ID NOS: 324-374.

[0163] Provided herein is an anti-CDl 17 binding agent comprising a VHH single domain antibody comprising a CDR-H1, a CDR-H2, and a CDR-H3 contained within an amino acid sequence selected from the group consisting of SEQ ID NOs: 324-374.

[0164] Provided herein is an anti-CDl 17 binding agent comprising a VHH single domain antibody comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 324-374, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto.

[0165] In some of any of the provided embodiments, the VHH comprises the amino acid sequence set forth in any one of SEQ ID NOS: 324-374.

[0166] Provided herein is a viral fusion protein comprising a viral envelope protein and the antiCD 117 binding agent.

[0167] Provided herein is a viral fusion protein comprising a viral envelope protein and an antiCD 133 binding agent, wherein the anti-CDl 33 binding agent is an antibody or antigen-binding fragment comprising a variable heavy (VH) chain and a variable light (VL) chain selected from a) a VH comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 314, 315, and 273, respectively, and a VL comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 275, 276, 277, respectively; (b) a VH comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 316, 317, and 282, respectively, and a VL comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 284, 285, and 286, respectively; (c) a VH comprising a a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 318, 319, and 300, respectively, and a VL comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 302, 303, and 304, respectively; (d) a VH comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 320, 321, and 309, respectively, and a VL comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 311, 312, and 313, respectively; and (e) a VH comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 322, 323, and 291, respectively, and a VL comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 293, 294, and 295, respectively.

[0168] In some of any of the provided embodiments, the CD 133 binding agent is selected from a) a CD 133 binding agent comprising a heavy chain variable (VH) region comprising the amino acid sequence of SEQ ID NO: 270, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a light chain variable (VL) region comprising the amino acid sequence of SEQ ID NO: 274, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; b) a CD133 binding agent comprising a VH region comprising the amino acid sequence of SEQ ID NO: 279, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a VL region comprising the amino acid sequence of SEQ ID NO: 283, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; c) a CD 133 binding agent comprising a VH region comprising the amino acid sequence of SEQ ID NO: 297, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a VL region comprising the amino acid sequence of SEQ ID NO: 301, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; d) a CD133 binding agent comprising a VH region comprising the amino acid sequence of SEQ ID NO: 306, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a VL region comprising the amino acid sequence of SEQ ID NO: 310, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; e) a CD133 binding agent comprising a VH region comprising the amino acid sequence of SEQ ID NO: 288, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a VL region comprising the amino acid sequence of SEQ ID NO: 292, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto.

[0169] In some of any of the provided embodiments, the CD 133 binding agent is a scFv. In some of any of the provided embodiments, the CD 133 binding agent comprises the amino acid sequence set forth in any one of SEQ ID NOs: 269, 278, 287, 296 or 305, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto. In some of any of the provided embodiments, the viral envelope protein is a VSV-G protein or a functional variant thereof. In some of any of the provided embodiments, the viral envelope protein is a Cocal virus G protein or a functional variant thereof. In some of any of the provided embodiments, the viral envelope protein is an Alphavirus fusion protein (e.g., Sindbis virus) or a functional variant thereof. In some of any of the provided embodiments, the viral envelope protein is a Paramyxovirus fusion protein (e.g., a Morbillivirus or Henipavirus) or a functional variant thereof. In some of any of the provided embodiments, the viral envelope protein is a Morbillivirus fusion protein (e.g., measles virus (MeV), canine distemper virus, Cetacean morbillivirus, Peste-des-petits-ruminants virus, Phocine distemper virus, or Rinderpest virus) ora functional variant thereof. In some of any of the provided embodiments, the viral envelope protein is a Henipavirus fusion protein (e.g., Nipah virus, Hendra virus, Cedar virus, Kumasi virus, Mojiang virus, or Langya virus) or a functional variant thereof.

[0170] In some of any of the provided embodiments, the viral envelope protein is a Nipah virus fusion protein or a functional variant thereof. In some of any of the provided embodiments, the viral envelope protein comprises a Nipah virus G glycoprotein (NiV-G) or a biologically active portion thereof. In some of any of the provided embodiments, the binding agent is fused to the C-terminus of the NiV-G protein or biologically active portion thereof. In some of any of the provided embodiments, the NiV-G or biologically active portion thereof is a wild-type NiV-G protein or a functionally active variant or biologically active portion thereof. In some of any of the provided embodiments, the NiV-G protein or biologically active portion thereof is truncated and lacks up to 40 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein set forth in SEQ ID NO:4. In some of any of the provided embodiments, the NiV-G protein or biologically active portion thereof is a biologically active portion that is a truncated NiV-G that has a deletion of amino acids 2-34 at or near the N-terminus of wild-type NiV-G set forth in SEQ ID NO:4. In some of any of the provided embodiments, the NiV-G protein or biologically active portion thereof does not include an initial methionine. In some of any of the provided embodiments, the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO:42 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:42. In some of any of the provided embodiments, the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 42. In some of any of the provided embodiments, the viral envelope protein comprises one or more modifications to reduce binding to its native receptor. In some of any of the provided embodiments, the NiV-G protein or biologically active portion thereof is a mutant NiV-G protein or biologically active portion thereof that exhibits reduced binding to Ephrin B2 or Ephrin B3. In some of any of the provided embodiments, the NiV-G protein or biologically active portion thereof comprises one or more amino acid substitutions corresponding to amino acid substitutions selected from the group consisting of E501A, W504A, Q530A, and E533A with reference to numbering set forth in SEQ ID NO:4. In some of any of the provided embodiments, the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO: 17 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 17. In some of any of the provided embodiments, the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 17. In some of any of the provided embodiments, the NiV-G protein or the biologically active portion has the amino acid sequence set forth in SEQ ID NO: 18 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 18. In some of any of the provided embodiments, the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 18.Brief Description of The Drawings

[0171] FIG. 1A depicts a schematic of experiments performed to assess the capacity of VSV-G and BaEVTR vectors to transduce human hematopoietic stem cells (HSCs), blood lineage cells, and hepatocytes in vitro.

[0172] FIG. IB shows results for transduction of hematopoietic cells with VSV-G and BaEVTR lentiviral vectors.

[0173] FIG. 1C shows results of transduction of hematopoietic stem cells with increasing dilutions with a BaEVTR or VSV-G lentiviral vector.

[0174] FIG. ID shows additional results of transduction of hematopoietic stem cells with increasing dilutions with a BaEVTR or VSV-G lentiviral vector.

[0175] FIG. IE shows results of transduction of hematopoietic stem cells with two different Multiplicity of infection values of BaEVTR vector.

[0176] FIG. 2 shows results for the ex vivo transduction of hematopoietic cells and subsequent transgene engraftment in a mouse model.

[0177] FIG. 3 shows the phenotypic distribution of hematopoietic cells following humanization of two mouse models.

[0178] FIG.4A shows an exemplary humanization protocol for mouse experiments described herein in Example 4. FIG. 4B shows distribution of hematopoietic cells following early-stage humanization of a mouse models.

[0179] FIG. 5A shows the effect of dose schedules on transduction of hematopoietic cells with a BaEVTR vector or a VSV-G vector.

[0180] FIG.5B shows the effect of mobilization on transduction of hematopoietic cells with a BaEVTR vector.

[0181] FIG. 6A shows the effect of mobilization on in vivo distribution of hematopoietic cells following early-stage humanization.

[0182] FIG.6B shows additional results of the effect of mobilization on in vivo distribution of hematopoietic cells following early-stage humanization.

[0183] FIG. 6C shows the effect of mobilization on transduction ofhematopoietic cells with a BaEVTR vector in early-stage humanized mice.

[0184] FIG.6D shows additional results of the effect of mobilization on transduction of hematopoietic cells with a BaEVTR vector in early-stage humanized mice.

[0185] FIG. 7A shows the effect of co-infusion of BaEVTR vectors and cord-blood CD34+ cells on bone marrow transduction. FIG. 7B depicts the frequency of various cell types in a huCD34+ cell population.

[0186] FIG.8A shows the effect of co-infusion of BaEVTR vectors and cord-blood CD34+ cells on myeloid cell expression.

[0187] FIG.8B shows the effect of co-infusion of BaEVTR vectors blood CD34+ cells on stable transduction of hematopoietic cells after 16 weeks.

[0188] FIG.8C shows additional results of the the effect of co-infusion of BaEVTR vectors and cord-blood CD34+ cells on hematopoietic cell and progenitor cells sub-populations.

[0189] FIG. 9 shows the effect of different administration schedules on transduction of hematopoietic cells with VSV-G and BaEVTR vectors.

[0190] FIG. 10A shows results for transduction of hematopoietic cells with VSV-G and BaEVTR vectors and retargeted Nipah vectors.

[0191] FIG. 10B shows additional results for transduction of hematopoietic cells with VSV-G and BaEVTR vectors and retargeted Nipah vectors.

[0192] FIG. 11 shows the phenotypic distribution of hematopoietic cells transduced with BaEVTR vectors and retargeted Nipah vectors.

[0193] FIG. 12 shows an additional phenotypic distribution of hematopoietic cells transduced with BaEVTR vectors and retargeted Nipah vectors.

[0194] FIG. 13 shows the phenotypic distribution of peripheral blood hematopoietic cells following humanization.

[0195] FIG. 14 shows the results for transduction of hematopoietic cells with BaEVTR virus-like particles.

[0196] FIG. 15A shows a representative flow plot of B2M gene editing in human hematopoietic cells with the BaEVTR virus-like particles.

[0197] FIG. 15B shows the results for transduction of hematopoietic cells with BaEVTR virus-like particles in three humanized mouse models.

[0198] FIG. 16A depicts percentage of cells which were subjected to gene editing in vitro as a function of increasing dose of BaEVTR virus-like particles carrying a gene editing cargo. Percent edited cells are similarly shown for in vivo gene editing of cells of the bone marrow and Lin- / CD34+ progenitor cells in FIG. 16B.

[0199] FIG. 16 C shows , the in vivo knockout of B2M gene (e.g., B2M editing) in different HSPC subpopulations. FIG. 16D shows an exemplary long-term humanization protocol for mice experiments herein in Example 12. FIG. 16E shows the level of editing in long-term humanized mice. FIG. 16F shows true in vivo B2M editing in huCD45+ cells.

[0200] FIG. 17A shows an exemplary study design for an experiment described in Example 13. FIG. 17B shows in vivo cell transduction of a GFP transgene.

[0201] A model study design for non-human primates is shown in FIG. 18.Detailed Description

[0202] Provided herein in some embodiments are lipid particles and methods of delivering the same to a target cell. In some embodiments, the target cell is a hematopoietic stem / progenitor cell (HSPC), such as a CD34+ cell. In some embodiments, the lipid particle is a targeted lipid particle that preferentially targets a target molecule expressed on the target cell. Provided herein in some embodiments are targeted lipid particles having a targeting agent that binds to a target molecule. In some embodiments, the target molecule is expressed on a target cell. In some embodiments, the provided particle preferentially targets a target cell compared to a non-target cell. Among provided embodiments, the lipid particles allow for targeted delivery to hematopoietic stem / progenitor cells (HSPC), including to naive hematopoietic cells (HSCs).

[0203] Exogenous agent delivery that is preferentially targeted to certain target cells, such as for the preferential genetic engineering of the target cells, can result in long-term and / or systemic genetic engineering of the target cells and / or cells derived therefrom, for instance when engineered target cells are able to differentiate into a variety of cell types. As an example, CD34+ progenitor cells, which are a heterogeneous cell population that include a subpopulation of hematopoietic stem cells (HSCs), pluripotent stem cells, and cells in the early stages of lineage commitment, can differentiate to produce all hematopoietic cell types found in circulation. While CD34 is expressed in a diversity of hematopoietic progenitor cells at various stages of differentiation across the hematopoietic tree, CD34+ progenitor cells that are HSCs or multipotent progenitors (MPPs) at the top of the hematopoietic tree may be the most appealing cell targets for achieving long-term and / or systemic genetic engineering across a broad range of hematopoietic cell types. Yet other appealing cell targets for genetic engineering among CD34+ progenitor cells include myeloid progenitors and lymphoid progenitors, such as myelo-lymphoid progenitors (MLPs), early T progenitors (ETPs), megakaryo-erythroid progenitors (MEPs), common myeloid progenitors (CMPs), and granulo-monocyte progenitors (GMPs). There is a need for reagents and methods for targeted delivery of exogenous agents to highly pluripotent subpopulations of hematopoietic cells, such as to CD34+ progenitor cells, including HSCs and MPPs. Achieving in vivo genetic engineering of hematopoietic stem / progenitor cells (HSPC) would dramatically expand patients’ access to gene therapy.

[0204] The provided disclosure addresses this need. The embodiments herein provide for lipid particles that allow for efficient targeting and delivery of an exogenous agent contained therein to human HSPC. In provided embodiments, the lipid particles can include viral vectors such as lentiviral vectors that are pseudotyped with a fusogen with a preferential ability to target HSPC. In some embodiments, the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof, such as the exemplarly fusogens BaEVTR or BaEVRLess. In some embodiments, the lipid particle includes a targeting agent to target the fusogen to HSPCs, such as to CD34+ cells. In some embodiments,the targeting agent is fused to a fusogen, such as a viral envelope attachment glycoprotein, exposed on the lipid bilayer to provide a retargeted fusogen. Results herein demonstrate that such lipid particles can efficiently target HSPC, including naive HSC. Notably, results show that the lipid particles can be delivered in vivo not only to cells in the peripheral blood but that they also can establish basal accesss to human HPSC in the bone marrow (BM) and peripheral blood without relying on high vector doses or selective enrichment. The provided delivery methods also can be combined with mobilization agents that stimulate mobilization of bone marrow cells from the bone marrow to the peripheral blood in the subject. Results herein show that improved transduction of cells in peripheral blood can be achieved when combined with mobilization. Moreover, combined mobilization strategies also allow transduction to naive HSC in the bone marrow even where there is not substantial egress of human HSPC into the peripheral blood. For instance, examples herein show that embodiments using a provided lipid particle able to target HSPC (e.g. BaEVTR fusogen) allow about 2% targeting of phenotypic HSC compared to undetectable transduction without mobilization.

[0205] In some embodiments, the fusogen of a provided lipid particle is not fused to a targeting agent. For instance, observations herein indicate that a lipid particle (e.g. lentiviral vector) pseudotyped with a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof, such as the exemplarly fusogens BaEVTR, exhibits highly efficient targeting of HSPC. In some embodiments, such lipid particles can additionally include a targeting agent.

[0206] In some embodiments, the fusogen of a provided lipid particle is a retargeted fusogen in which a viral envelope attachment glycoprotein is linked or fused to a targeted agent. In such embodiments, a retargeted fusogen is engineered to recognize receptors on chosen target cell types resulting in highly cell-specific in vivo gene transfer. For example, provided embodiments relate to lipid particles (e.g. lentiviral vector) pseudotyped with a retargeted fusogenof a parmamyxovirus envelope attachment glycoprotein, such as a Nipah G protein or functional variant thereof, linked or fused with a targeting agent. In some aspects, the use of a retargeted fusogen can reduce off-target delivery. In some embodiments, a retargeted fusogen as shown herein can exhibit striking (e.g. lOOx) specificity increase over other broadly tropic pseudotyped lipid particles in targeting a receptor-positive population, including in target cells exclusively localized in the bone marrow even where such cells make up as little as 0.15% of total human cells.

[0207] In some embodiments, the target molecule bound by the targeting agent of the provided particle is expressed on CD34+ progenitor cells. In some embodiments, the target cell is a CD34+ progenitor cell. In some embodiments, the target molecule is expressed on all CD34+ progenitor cells. In some embodiments, the target molecule is expressed on at least a subset of CD34+ progenitor cells. In some embodiments, the target molecule is expressed on HSCs. In some embodiments, the target molecule is expressed on MPPs.

[0208] In some embodiments, the target molecule is selected from the group consisting of ASCT1, ASCT2, CD105, CD110, CD117, CD133, CD146, CD164, CD34, CD46, CD49f, CD90, EPCR, and ITGA3. As demonstrated herein, preferential genetic engineering, e.g., transduction, of CD34+ progenitor cells, e.g., subpopulations thereof that included highly pluripotent HSCs and MPPs, as well as myeloid progenitors that included MEPs, CMPs, and GMPs, was achieved with exemplary targeted lipid particles, e.g., targeted lentiviral particles, having a targeting agent that binds to a target molecule selected from the foregoing, e.g., CD117. Also demonstrated herein are such effects for lymphoid progenitors, such as MLPs and ETPs, using exemplary targeted lipid particles, e.g., targeted lentiviral particles, having a targeting agent that binds to a target molecule selected from the foregoing, e.g., ASCT2.

[0209] In some embodiments, the provided particle is any described in Section II. In some embodiments, the provided particle is a lipid particle, such as any described in Section II-A. In some embodiments, the provided particle is a lipid particle that is a viral particle, such as any described in Section II-A-1. In some embodiments, the provided particle is a targeted lentiviral particle. In some embodiments, the provided particle is a lipid particle that is a virus-like particle, such as any described in Section II-A-2. In some embodiments, the provided particle is a lipid particle that is a cell-based particle, such as any described in Section II-A-3.

[0210] In some embodiments, the provided particle has a fusogen, such as any described in Section II-B. In some embodiments, the fusogen contains a mammalian protein, such as any described in Section II-B-1. In some embodiments, the fusogen contains a viral protein, such as any described in Section II-B- 2. In particular embodiments, the fusogen is a baboon endogenous retrovirus, such as BaEVTR.In some embodiments, the targeting agent of the provided particle is any described in Section II-C. In some embodiments, the target molecule or target cell for any of the provided particles is any described in Section II-C.

[0211] In some embodiments, the provided particle contains an exogenous agent. In some embodiments, the exogenous agent is for delivery to the target cell. In some embodiments, the provided particle exhibits preferential delivery of the exogenous agent to a target cell compared to a non-target cell. In some embodiments, the delivery to target cells is increased by or by greater than 1.1 -fold, 1.2- fold, 1.3-fold, 1.4-fold, 1.5-fold, 1.6-fold, 1.7-fold, 1.8-fold, 1.9-fold, 2-fold, 2.5-fold, 3-fold, 3.5-fold, 4- fold, 4.5-fold, 5-fold, 5.5-fold, 6-fold, 6.5-fold, 7-fold, 7.5-fold, 8-fold, 8.5-fold, 9-fold, 9.5-fold, 10-fold, or more compared to delivery to non-target cells.

[0212] In some embodiments, the exogenous agent of the provided particle is any described in Section II-D. In some embodiments, the exogenous agent is a nucleic acid, such as any described in Section II-D-1. In some embodiments, the nucleic acid contains a payload gene encoding a payload agent, for instance a protein, such as any described in Section II-D-2. In some embodiments, the exogenous agent is a protein, such as any described in Section II-D-2. In some embodiments, the proteinis a payload agent. In some embodiments, the exogenous agent is a small molecule, such as any described in Section II-D-3.

[0213] In some embodiments, the pay load agent encoded or contained by the exogenous agent is any described in Section III. In some embodiments, the exogenous agent is or encodes an engineered receptor, such as any described in Section III-A. In some embodiments, the exogenous agent is or encodes a gene-editing agent, such as any described in Section III-B.

[0214] Also provided herein in some embodiments are polynucleotides, such as any described in Section IV-A, encoding the fusogen and targeting agent of any of the provided particles. Also provided herein in some embodiments are cells, such as any described in Section IV-B, containing polynucleotides encoding one or both of the fusogen and targeting agent of any of the provided particles, for instance any of the provided polynucleotides, such as any described in Section IV-A. Also provided herein in some embodiments are methods, such as any described in Section IV-B, of making a targeted lipid particle, such as any described in Section II, using polynucleotides encoding one or both of the fusogen and targeting agent of any of the provided particles, for instance any of the provided polynucleotides, such as any described in Section IV-A. In some embodiments, the provided methods are for making any of the provided particles containing an exogenous agent, such as any described in Section II-D.

[0215] Also provided herein in some embodiments are compositions, such as any described in Section V, containing a plurality of any of the provided particles. In some embodiments, the provided composition contains a plurality of any of the provided lenti viral particles.

[0216] Also provided herein in some embodiments are methods, such as any described in Section VI, of using any of the provided particles. In some embodiments, the provided method involves administering any of the provided particles or compositions to a subject. In some embodiments, the provided method involves administering to the subject an agent that stimulates mobilization of bone marrow cells from the bone marrow to the peripheral blood in the subject. In some embodiments, the bone marrow cells are CD34+ progenitor cells. As demonstrated herein using the provided targeted particles, e.g., ASCT2- and CD117-targeted lentiviral particles, the mobilization of bone marrow cells to the peripheral blood prior to the in vivo genetic engineering, e.g., transduction, of cells improved the transduction of CD34+ progenitor cells, including HSCs and MPPs, following intravenous administration of the particles. Such effects are also demonstrated herein using non-targeted particles, e.g., not containing a targeting agent, such as those with fusogens having natural tropism for bone marrow cells like CD34+ progenitor cells, e.g., BaEV envelope glycoproteins. During steady state, CD34+ progenitors including HSCs reside in specialized niches within the bone marrow (Hoggatt et al. Stem Cell Res Ther. 2011 Mar 14;2(2): 13). The mobilization of bone marrow cells may increase the number of CD34+ progenitor cells in the peripheral blood that are accessible to the intravenously administered particles, thereby improving transduction.

[0217] Also provided herein in some embodiments are methods involving administering to a subject (i) an agent that stimulates mobilization of bone marrow cells from the bone marrow to the peripheral blood in the subject and (ii) a particle, such as any described in Section II, either with or without any of the targeting agents described herein. In some embodiments, the method is any described in Section VI.

[0218] All publications, including patent documents, scientific articles and databases, referred to in this application are incorporated by reference in their entirety for all purposes to the same extent as if each individual publication were individually incorporated by reference. If a definition set forth herein is contrary to or otherwise inconsistent with a definition set forth in the patents, applications, published applications and other publications that are herein incorporated by reference, the definition set forth herein prevails over the definition that is incorporated herein by reference.

[0219] The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described.I. DEFINITIONS

[0220] Unless defined otherwise, all terms of art, notations and other technical and scientific terms or terminology used herein are intended to have the same meaning as is commonly understood by one of ordinary skill in the art to which the claimed subject matter pertains. In some cases, terms with commonly understood meanings are defined herein for clarity and / or for ready reference, and the inclusion of such definitions herein should not necessarily be construed to represent a substantial difference over what is generally understood in the art.

[0221] As used herein, the articles “a” and “an” refer to one or to more than one (i.e. to at least one) of the grammatical object of the article. By way of example, “an element” means one element or more than one element.

[0222] As used herein, the term “about” will be understood by persons of ordinary skill in the art and will vary to some extent on the context in which it is used. As used herein, “about” when referring to a measurable value such as an amount, a temporal duration, and the like, is meant to encompass variations of ±20% or ±10%, more preferably ±5%, even more preferably ±1%, and still more preferably ±0.1% from the specified value, as such variations are appropriate to perform the disclosed methods.

[0223] By “wild type” or “WT” or “native” herein is meant an amino acid sequence that is found in nature, including allelic variations. A wild type protein or polypeptide has an amino acid sequence that has not been intentionally modified.

[0224] As used herein, “lipid particle” refers to any biological or synthetic particle that contains a bilayer of amphipathic lipids enclosing a lumen or cavity. Typically, a lipid particle does not contain a nucleus. Examples of lipid particles include solid particles such as nanoparticles, viral-derived particles, and cell-derived particles. Exemplary lipid particles also include viral-based particles, such as virus-like particles or viral particles (e.g., lentiviral particles), exosomes, enucleated cells, various vesicles, such asa microvesicle, a membrane vesicle, an extracellular membrane vesicle, a plasma membrane vesicle, a giant plasma membrane vesicle, an apoptotic body, a mitoparticle, a pyrenocyte, or a lysosome. In some embodiments, a lipid particle can be a fusosome. In some embodiments, the lipid particle is not a platelet. In some embodiments, the lipid particle is derived from a source cell. A lipid particle also may include an exogenous agent, which may be present in the lumen of the lipid particle.

[0225] The term “viral-based particle” can be any type of lipid particle that is derived from a virus or from viral protein, for example viral particles and virus-like particles.

[0226] The terms “viral vector”, “viral particle”, and “viral vector particle” are used interchangeably herein. A viral particle can be any type of lipid particle which comprises one or more viral structural proteins in addition to at least one non-structural viral genomic component or functional fragment thereof (e.g., a polymerase, an integrase, a protease or other non-structural component).

[0227] The term “virus-like particle” or VLP can be any type of particle that features at least one viral structural protein and is devoid of viral genetic material.

[0228] The term “retroviral particle” refers to a viral particle that contains retroviral nucleic acid or is derived from a retrovirus. A retroviral particle includes the following components: a vector genome (retrovirus nucleic acid), a nucleocapsid encapsidating the nucleic acid, and a membrane envelope surrounding the nucleocapsid. Typically, a retroviral particle contains sufficient retroviral genetic information to allow packaging of an RNA genome, in the presence of packaging components, into a viral particle capable of infecting a target cell. Infection of the target cell may include reverse transcription and integration into the target cell genome. A retroviral particle may be a recombinant retroviral particle that is replication defective and lacks genes essential for replication, such as a functional gag-pol and / or env gene and / or other genes essential for replication. A retroviral particle also may be a self-inactivating (SIN) vector.

[0229] As used herein, a “lentiviral particle” or LV refers to a viral particle that contains lentiviral nucleic acid or is derived from a lentivirus. A lentiviral particle includes the following components: a vector genome (lentivirus nucleic acid), a nucleocapsid encapsidating the nucleic acid, and a membrane stirrounding the nucleocapsid. Typically, a lentiviral particle contains sufficient lentiviral genetic information to allow packaging of an RN genome, in the presence of packaging components, Into a viral particle capable of infecting a target cell. Infection of the target cell may include reverse transcription and integration into the target cell genome. A lentiviral particle may be a recombinant lentiviral particle that is replication defective and lacks genes essential for replication, such as a functional gag-pol and / or env gene and / or other genes essential for replication. A lentiviral particle also may be a self-inactivating (SIN) vector.

[0230] As used herein, a “retroviral nucleic acid,” refers to a nucleic acid containing at least the minimal sequence requirements for packaging into a retroviral particle, alone or in combination with a helper cell, helper virus, or helper plasmid. In the case of “lentiviral nucleic acid” the nucleic acid refersto at least the minimal sequence requirements for packaging into a lentiviral particle, alone or in combination with a helper cell, helper virus, or helper plasmid. In some embodiments, the viral nucleic acid comprises one or more of (e.g., all of) a 5’ LTR (e.g., to promote integration), U3 (e.g., to activate viral genomic RNA transcription), R (e.g., a Tat-binding region), U5, a 3’ LTR (e.g., to promote integration), a packaging site (e.g., psi ( )), RRE (e.g., to bind to Rev and promote nuclear export). The viral nucleic acid can comprise RNA (e.g., when part of a virion) or DNA (e.g., when being introduced into a source cell or after reverse transcription in a recipient cell). In some embodiments, the viral nucleic acid is packaged using a helper cell, helper virus, or helper plasmid which comprises one or more of (e.g., all of) gag, pol, and env.

[0231] As used herein, “fusosome” refers to a particle containing a bilayer of amphipathic lipids enclosing a lumen or cavity and a fusogen that interacts with the amphipathic lipid bilayer. In some embodiments, the fusosome comprises an exogenous agent. In some embodiments, the exogenous agent is a nucleic acid (e.g., DNA or RNA), a peptide, or a protein. In some embodiments, the fusosome is a membrane enclosed preparation. In some embodiments, the fusosome is derived from a source cell.

[0232] As used herein, “fusosome composition” refers to a composition comprising one or more fusosomes.

[0233] As used herein, “fusogen” refers to an agent or molecule that creates an interaction between two membranes, including membrane enclosed lumens. In embodiments, the fusogen facilitates fusion of the membranes. In other embodiments, the fusogen creates a connection, e.g., a pore, between two membranes or lumens (e.g., a lumen of a retroviral particle and a cytoplasm of a target cell). In some embodiments, the fusogen comprises a complex of two or more proteins, e.g., wherein neither protein has fusogenic activity alone. In some embodiments, the fusogen comprises a targeting domain.

[0234] As used herein, a “re-targeted fusogen” refers to a fusogen that comprises a targeting moiety having a sequence that is not part of the naturally-occurring form of the fusogen. In embodiments, the fusogen comprises a different targeting agent relative to the targeting agent in the naturally-occurring form of the fusogen. In embodiments, the naturally-occurring form of the fusogen lacks a targeting domain, and the re-targeted fusogen comprises a targeting agent that is absent from the naturally- occurring form of the fusogen. In embodiments, the fusogen is modified to comprise a targeting agent. In embodiments, the fusogen comprises one or more sequence alterations outside of the targeting agent relative to the naturally-occurring form of the fusogen, e.g., in a transmembrane domain, fusogenically active domain, or cytoplasmic domain.

[0235] As used herein, a “target cell” refers to a cell of a type to which it is desired that a targeted lipid particle or viral vector delivers an exogenous agent. In embodiments, a target cell is a cell of a specific tissue type or class. In some embodiments, the targeting agent or fusogen, e.g., re-targeted fusogen, leads to preferential delivery of the exogenous agent to a target cell compared to a non-target cell.

[0236] As used herein a “non-target cell” refers to a cell of a type to which it is not desired that a targeted lipid particle or viral particle delivers an exogenous agent. In some embodiments, a non-target cell is a cell of a specific tissue type or class. In some embodiments, the targeting agent or fusogen, e.g., re-targeted fusogen leads to lower delivery of the exogenous agent to a non-target cell compared to a target cell.

[0237] As used herein a “biologically active portion,” such as with reference to a protein such as a G protein or an F protein, refers to a portion of the protein that exhibits or retains an activity or property of the full-length of the protein. For example, a biologically active portion of an F protein retains fusogenic activity in conjunction with the G protein when each are embedded in a lipid bilayer. A biologically active portion of the G protein retains fusogenic activity in conjunction with an F protein when each is embedded in a lipid bilayer. The retained activity can include 10%-150% or more of the activity of a full-length or wild-type F protein or G protein. Examples of biologically active portions of F and G proteins include proteins with truncations of the cytoplasmic domain, such as any of the described NiV-F with a truncated cytoplasmic tail.

[0238] As used herein, “percent (%) amino acid sequence identity” and “homology” with respect to a peptide, polypeptide or antibody sequence are defined as the percentage of amino acid residues in a candidate sequence that are identical with the amino acid residues in the specific peptide or polypeptide sequence, after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity, and not considering any conservative substitutions as part of the sequence identity. Alignment for purposes of determining percent amino acid sequence identity can be achieved in various ways that are within the skill in the art, for instance, using publicly available computer software such as BLAST, BLAST-2, ALIGN or MEGALIGN (DNASTAR) software. Those skilled in the art can determine appropriate parameters for measuring alignment, including any algorithms needed to achieve maximal alignment over the full length of the sequences being compared.

[0239] An amino acid substitution may include but are not limited to the replacement of one amino acid in a polypeptide with another amino acid. Exemplary substitutions are shown in Table 19. Amino acid substitutions may be introduced into an antibody of interest and the products screened for a desired activity, for example, retained / improved binding.Table 19

[0240] Amino acids may be grouped according to common side-chain properties:(1) hydrophobic: Norleucine, Met, Ala, Vai, Leu, He;(2) neutral hydrophilic: Cys, Ser, Thr, Asn, Gin;(3) acidic: Asp, Glu;(4) basic: His, Lys, Arg;(5) residues that influence chain orientation: Gly, Pro;(6) aromatic: Trp, Tyr, Phe.

[0241] Non-conservative substitutions will entail exchanging a member of one of these classes for another class.

[0242] The term, “corresponding to” with reference to positions of a protein, such as recitation that nucleotides or amino acid positions “correspond to” nucleotides or amino acid positions in a disclosed sequence, such as set forth in the Sequence listing, refers to nucleotides or amino acid positions identified upon alignment with the disclosed sequence based on structural sequence alignment or using a standard alignment algorithm, such as the GAP algorithm. For example, corresponding residues of a similar sequence (e.g. fragment or species variant) can be determined by alignment to a reference sequence by structural alignment methods. By aligning the sequences, one skilled in the art can identify corresponding residues, for example, using conserved and identical amino acid residues as guides.

[0243] The term “isolated” as used herein refers to a molecule that has been separated from at least some of the components with which it is typically found in nature or produced. For example, a polypeptide is referred to as “isolated” when it is separated from at least some of the components of the cell in which it was produced. Where a polypeptide is secreted by a cell after expression, physically separating the supernatant containing the polypeptide from the cell that produced it is considered to be “isolating” the polypeptide. Similarly, a polynucleotide is referred to as “isolated” when it is not part of the larger polynucleotide (such as, for example, genomic DNA or mitochondrial DNA, in the case of a DNA polynucleotide) in which it is typically found in nature, or is separated from at least some of the components of the cell in which it was produced, for example, in the case of an RNA polynucleotide.Thus, a DNA polynucleotide that is contained in a vector inside a host cell may be referred to as “isolated”.

[0244] The term “effective amount” as used herein means an amount of a pharmaceutical composition which is sufficient enough to significantly and positively modify the symptoms and / or conditions to be treated (e.g., provide a positive clinical response). The effective amount of an active ingredient for use in a pharmaceutical composition will vary with the particular condition being treated, the severity of the condition, the duration of treatment, the nature of concurrent therapy, the particular active ingredient(s) being employed, the particular pharmaceutically-acceptable excipient(s) and / or carrier(s) utilized, and like factors with the knowledge and expertise of the attending physician.

[0245] An “exogenous agent” as used herein with reference to a viral particle refers to an agent that is neither comprised by nor encoded in the corresponding wild-type virus or fusogen made from a corresponding wild-type source cell. In some embodiments, the exogenous agent does not naturally exist, such as a protein or nucleic acid that has a sequence that is altered (e.g., by insertion, deletion, or substitution) relative to a naturally occurring protein. In some embodiments, the exogenous agent does not naturally exist in the source cell. In some embodiments, the exogenous agent exists naturally in the source cell but is exogenous to the virus. In some embodiments, the exogenous agent does not naturally exist in the recipient cell. In some embodiments, the exogenous agent exists naturally in the recipient cell, but is not present at a desired level or at a desired time. In some embodiments, the exogenous agent comprises nucleic acids or protein.

[0246] As used herein, a “promoter” refers to a cis- regulatory DNA sequence that, when operably linked to a gene coding sequence, drives transcription of the gene. The promoter may comprise a transcription factor binding sites. In some embodiments, a promoter works in concert with one or more enhancers which are distal to the gene.

[0247] As used herein, “operably linked” or “operably associated” includes reference to a functional linkage of at least two sequences. For example, operably linked includes linkage between a promoter and a second sequence, wherein the promoter sequence initiates and mediates transcription of the DNA sequence corresponding to the second sequence. Operably associated includes linkage between an inducing or repressing element and a promoter, wherein the inducing or repressing element acts as a transcriptional activator of the promoter.

[0248] As used herein, a composition refers to any mixture of two or more products, substances, or compounds, including cells. It may be a solution, a suspension, liquid, powder, a paste, aqueous, nonaqueous or any combination thereof.

[0249] As used herein, the term “pharmaceutically acceptable” refers to a material, such as a carrier or diluent, which does not abrogate the biological activity or properties of the compound, and is relatively nontoxic, i.e., the material may be administered to an individual without causing undesirable biologicaleffects or interacting in a deleterious manner with any of the components of the composition in which it is contained.

[0250] As used herein, the term “pharmaceutical composition” refers to a mixture of at least one particle with other chemical components, such as carriers, stabilizers, diluents, dispersing agents, suspending agents, thickening agents, and / or excipients. The pharmaceutical composition facilitates administration of the compound to an organism. Multiple techniques of administering a compound exist in the art including, but not limited to, intravenous, oral, aerosol, parenteral, ophthalmic, pulmonary and topical administration.

[0251] A “disease” or “disorder” as used herein refers to a condition where treatment is needed and / or desired.

[0252] As used herein, the terms “treat,” “treating,” or “treatment” refer to ameliorating a disease or disorder, e.g., slowing or arresting or reducing the development of the disease or disorder or reducing at least one of the clinical symptoms thereof. For purposes of this disclosure, ameliorating a disease or disorder can include obtaining a beneficial or desired clinical result that includes, but is not limited to, any one or more of: alleviation of one or more symptoms, diminishment of extent of disease, preventing or delaying spread (for example, metastasis, for example metastasis to the lung or to the lymph node) of disease, preventing or delaying recurrence of disease, delay or slowing of disease progression, amelioration of the disease state, inhibiting the disease or progression of the disease, inhibiting or slowing the disease or its progression, arresting its development, and remission (whether partial or total).

[0253] The terms “individual” and “subject” are used interchangeably herein to refer to an animal; for example a mammal. The term patient includes human and veterinary subjects. In some embodiments, methods of treating mammals, including, but not limited to, humans, rodents, simians, felines, canines, equines, bovines, porcines, ovines, caprines, mammalian laboratory animals, mammalian farm animals, mammalian sport animals, and mammalian pets, are provided. The subject can be male or female and can be any suitable age, including infant, juvenile, adolescent, adult, and geriatric subjects. In some examples, an “individual” or “subject” refers to an individual or subject in need of treatment for a disease or disorder. In some embodiments, the subject to receive the treatment can be a patient, designating the fact that the subject has been identified as having a disorder of relevance to the treatment, or being at adequate risk of contracting the disorder. In particular embodiments, the subject is a human, such as a human patient.

[0254] As used herein, the terms “effective amount” and “pharmaceutically effective amount” refer to a nontoxic but sufficient amount of an agent or drug to provide the desired biological result. That result can be reduction and / or alleviation of the signs, symptoms, or causes of a disease or disorder, imaging or monitoring of an in vitro or in vivo system (including a living organism), or any other desired alteration of a biological system. An appropriate effective amount in any individual case may be determined by one of ordinary skill in the art using routine experimentation.

[0255] As used herein, the terms “nuclear export sequence” (NES) or “nuclear export signal” (NES) refer to a nuclear export signal or other sequence or domain that is present in a protein and capable of targeting the protein for export from the cell nucleus to the cytoplasm through the nuclear pore complex using nuclear transport. A nuclear export domain can be fused (e.g., fused in-frame) with a polypeptide.

[0256] As used herein, the terms “nuclear localization siequence” (NLS) or “nuclear localization sequence” (NLS) refer to a nuclear localization signal or other seuqence or domain that is present in a protien and capable of targeting the protein for import from the cytoplams to the cell nucleus through the nuclear pore complex using nuclear transport. A nucelar lozalization can be fused (e.g., fused in-frame) with a polypeptide.II. TARGETED PARTICLES

[0257] Provided herein are lipid particles that allow for efficient targeting and delivery of an exogenous agent contained therein to hematopoietic stem / progenitor cells (HSPC), including to naive hematopoietic stem cells (HSCs). In some embodiments, the targeting can be to such stem or progenitor cells in a human subject, such as human HSPC. In provided embodiments, the lipid particles can include viral vectors such as lentiviral vectors that are pseudotyped with a fusogen with a preferential ability to target HSPC. In some embodiments, the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof, such as the exemplarly fusogens BaEVTR or BaEVRLess. In some embodiments, the lipid particle includes a targeting agent to target the fusogen to HSPCs, such as to CD34+ cells. In some embodiments, the targeting agent is fused to a fusogen, such as a viral envelope attachment glycoprotein, exposed on the lipid bilayer to provide a retargeted fusogen.

[0258] Provided herein in some embodiments are targeted particles having a targeting agent that binds to a target molecule. In some embodiments, the target molecule is expressed on a target cell. In some embodiments, the provided particle preferentially targets a target cell compared to a non-target cell.

[0259] In some embodiments, the provided particle is a lipid particle, such as any described in Section II-A. In some embodiments, the provided particle is a lipid particle that is a viral particle, such as any described in Section II-A-1. In some embodiments, the provided particle is a targeted lentiviral particle. In some embodiments, the provided particle is a lipid particle that is a virus-like particle, such as any described in Section II-A-2. In some embodiments, the provided particle is a lipid particle that is a cell-based particle, such as any described in Section II-A-3.

[0260] In some embodiments, the provided particle has a fusogen, such as any described in Section II-B. In some embodiments, the fusogen promotes mixing between lipids in the particle and lipids in the target cell. In some embodiments, the fusogen facilitates the fusion of the particle to a membrane of the target cell. In some embodiments, the particle integrates into the membrane of the target cell. In some embodiments, the membrane is the plasma membrane of the target cell. In some embodiments, thefusogen promotes formation of one or more pores between the interior of the particle and the cytosol of the target cell.

[0261] In some embodiments, the fusogen is endogenous to the particle. In some embodiments, the particle is pseudotyped with the fusogen, such as when the particle is a viral particle or virus-like particle.

[0262] In some embodiments, the fusogen is exposed on the surface of the particle. In some embodiments, the fusogen is exposed on the surface of the lipid bilayer of the particle. In some embodiments, a portion of the fusogen is embedded in the lipid bilayer of the particle.

[0263] In some embodiments, the particle is a viral particle or a virus-like particle. In some embodiments, the fusogen is exposed on the surface of the viral envelope of the particle. In some embodiments, a portion of the fusogen is embedded in the viral envelope of the particle.

[0264] In some embodiments, the fusogen contains a mammalian protein, such as any described in Section II-B-1. In some embodiments, the fusogen contains a viral protein, such as any described in Section II-B-2.

[0265] In some embodiments, the lipid particle includes a targeting agent to target the fusogen to HSPCs. In some embodiments, the targeting agent of the provided particle is any described in Section II- C. Exemplary target molecules and target cells for any of the provided particles are also described in Section II-C. In some embodiments, the fusogen is fused to the targeting agent of the particle. In some aspects, the fusogen is retargeted by the targeting agent to display altered tropism. In other embodiments, the targeting agent is fused to a separate transmembrane domain incorporated into the lipid bilayer, not the fusogen.

[0266] In some embodiments, the provided particle contains an exogenous agent. In some embodiments, the exogenous agent is in the lumen of the provided particle. In some embodiments, the sequence of the exogenous agent is exogenous to the particle. In some embodiments, the sequence of the exogenous agent is non-viral. In some embodiments, the exogenous agent is for delivery to the target cell. In some embodiments, the exogenous agent of the provided particle is any described in Section II-D. In some embodiments, the exogenous agent is a nucleic acid, such as any described in Section II-D-1. In some embodiments, the nucleic acid contains a payload gene encoding a payload agent. In some embodiments, the exogenous agent is a protein, such as any described in Section II-D-2. In some embodiments, the protein is a payload agent. In some embodiments, the exogenous agent is a small molecule, such as any described in Section II-D-3.

[0267] In some embodiments, the pay load agent encoded or contained by the exogenous agent is any described in Section III. In some embodiments, the exogenous agent is or encodes an engineered receptor, such as any described in Section III-A. In some embodiments, the exogenous agent is or encodes a gene-editing agent, such as any described in Section III-B.A. Lipid Particle

[0268] In some embodiments, the particle is a lipid particle. In some embodiments, the particle includes a bilayer of amphipathic lipids that encloses a lumen or cavity. In some embodiments, the particle has a lipid bilayer as the outermost surface. In some embodiments, the particle is spherical. In some embodiments, the particle is of elongated or irregular shape.

[0269] In some embodiments, the lipid bilayer encloses a lumen. In some embodiments, the lumen is aqueous. In some embodiments, the lumen is in contact with the hydrophilic head groups on the interior of the lipid bilayer. In some embodiments, the lumen is a cytosol. In some embodiments, the cytosol contains cellular components present in a source cell. In some embodiments, the cytosol does not contain components present in a source cell. In some embodiments, the lumen is a cavity. In some embodiments, the cavity contains an aqueous environment. In some embodiments, the cavity does not contain an aqueous environment.

[0270] In some embodiments, the particle comprises several different types of lipids. In some embodiments, the lipids are amphipathic lipids. In some embodiments, the amphipathic lipids are phospholipids. In some embodiments, the phospholipids comprise phosphatidylcholine, phosphatidylethanolamine, phosphatidylinositol, and phosphatidylserine. In some embodiments, the lipids comprise phospholipids such as phosphocholines and phosphoinositols. In some embodiments, the lipids comprise DMPC, DOPC, and DSPC.

[0271] In some embodiments, a particle or composition of particles can be assessed for one or more features related to their size, including diameter, average (e.g., mean or median) diameter, range of variation thereof above and below an average (e.g., mean or median) value of the diameter, coefficient of variation, polydispersity index, or other measure of size of particles. Various methods for particle characterization can be used, including laser diffraction, dynamic light scattering (DLS; also known as photon correlation spectroscopy) or image analysis, such as microscopy or automated image analysis.

[0272] In some embodiments, the particle has a diameter of less than about 3 pm, less than about 2 pm, less than about 1 pm, less than about 900 nm, less than about 800 nm, less than about 700 nm, less than about 600 nm, less than about 500 m, less than about 400 nm, less than about 300, less than about 200 nm, less than about 150 nm, less than about 100 nm, less than about 50 nm, or less than about 20 nm. In some embodiments, the particle has a diameter of less than about 400 nm. In some embodiments, the particle has a diameter of less than about 150 nm. In some embodiments, the particle has a diameter of between at or about 2 pm and at or about 1 pm, between at or about 1 pm and at or about 900 nm, between at or about 900 nm and at or about 800 nm, between at or about 800 and at or about 700 nm, between at or about 700 nm and at or about 600 nm, between at or about 600 nm and at or about 500 nm, between at or about 500 nm and at or about 400 nm, between at or about 400 nm and at or about 300 nm, between at or about 300 nm and at or about 200 nm, between at or about 200 and at or about 100 nm, between at or about 100 and at or about 50 nm, or between at or about 20 nm and at or about 50 nm.

[0273] In some embodiments, the particle has a diameter between at or about 10 nm and at or about 1000 nM, between at or about 25 nm and at or about 500 nm, between at or about 40 nm and at or about 300 nm, between at or about 50 nm and at or about 250 nm, between at or about 60 nm and at or about 225 nm, between at or about 70 nm and at or about 200 nm, between at or about 80 nm and at or about 175 nm, or between at or about 90 nm and at or about 150 nm.

[0274] In some embodiments, the particle has a hydrodynamic radius, e.g., as determined by DLS, of about 100 nm to about two microns. In some embodiments, the particle has a hydrodynamic radius between at or about 2 pm and at or about 1 pm, between at or about 1 pm and at or about 900 nm, between at or about 900 nm and at or about 800 nm, between at or about 800 and at or about 700 nm, between at or about 700 nm and at or about 600 nm, between at or about 600 nm and at or about 500 nm, between at or about 500 nm and at or about 400 nm, between at or about 400 nm and at or about 300 nm, between at or about 300 nm and at or about 200 nm, between at or about 200 and at or about 100 nm, between at or about 100 and at or about 50 nm, or between at or about 20 nm and at or about 50 nm.

[0275] In some embodiments, the particle has a geometric radius, e.g., as determined by a multiangle light scattering, of about 100 nm to about two microns. In some embodiments, the particle has a geometric radius between at or about 2 pm and at or about 1 pm, between at or about 1 pm and at or about 900 nm, between at or about 900 nm and at or about 800 nm, between at or about 800 and at or about 700 nm, between at or about 700 nm and at or about 600 nm, between at or about 600 nm and at or about 500 nm, between at or about 500 nm and at or about 400 nm, between at or about 400 nm and at or about 300 nm, between at or about 300 nm and at or about 200 nm, between at or about 200 and at or about 100 nm, between at or about 100 and at or about 50 nm, or between at or about 20 nm and at or about 50 nm.

[0276] Various particles are known, any of which can be generated in accord with the provided embodiments. Non-limiting examples of particles include any as described in, or containing any features as described in, International Published PCT Application No. WO 2017 / 095946; WO 2017 / 095944; WO 2017 / 095940; WO 2019 / 157319; WO 2018 / 208728; WO 2019 / 113512; WO 2019 / 161281; WO 2020 / 102578; WO 2019 / 222403; WO 2020 / 014209; WO 2020 / 102485; WO 2020 / 102499; WO 2020 / 102503; WO 2013 / 148327; WO 2017 / 182585; WO 2011 / 058052; or WO 2017 / 068077, each of which are incorporated by reference in their entirety.

[0277] In some embodiments, the particle is a viral-based particle, a viral particle, a virus-like particle (VLP), a nanoparticle, a vesicle, an exosome, a dendrimer, a lentiviral particle, an enucleated cell, a microvesicle, a membrane vesicle, an extracellular membrane vesicle, a plasma membrane vesicle, a giant plasma membrane vesicle, an apoptotic body, a mitoparticle, a pyrenocyte, a lysosome, another membrane enclosed vesicle, or a cell based particle

[0278] In some embodiments, the lipid bilayer includes a synthetic lipid complex. In some embodiments, the synthetic lipid complex is a liposome. In some embodiments, the lipid bilayer is avesicular structure characterized by a phospholipid bilayer membrane and an inner aqueous medium. In some embodiments, the lipid bilayer has multiple lipid layers separated by aqueous medium. In some embodiments, the lipid bilayer forms spontaneously when phospholipids are suspended in an excess of aqueous solution. In some embodiments, the lipid components undergo self-rearrangement before the formation of closed structures and entrap water and dissolved solutes between the lipid bilayers

[0279] In some aspects, the lipid bilayer is derived from a source cell during a process to produce a lipid-containing particle. Exemplary methods for producing lipid-containing particles are described in Section IV. In some embodiments, the lipid bilayer includes membrane components of the host cell from which the lipid bilayer is derived, e.g., phospholipids, membrane proteins, etc. In some embodiments, the lipid bilayer includes a cytosol that includes components found in the cell from which the vehicle is derived, e.g., solutes, proteins, nucleic acids, etc., but not all of the components of a cell, e.g., lacking a nucleus. In some embodiments, the lipid bilayer is considered to be exosome-like.

[0280] In particular embodiments, the particle is virally derived. In some embodiments, the particle is a viral or virus-like particle, such as any described in Section II-A-1. In some embodiments, the particle can be a viral-based particle, such as a viral particle (e.g., lentiviral particle) or a virus-like particle (e.g., a lentiviral-like particle). In some embodiments, the provided particle is a targeted lentiviral particle. In some embodiments, the lipid bilayer is a viral envelope. In some embodiments, the viral envelope is obtained from a host cell. In some embodiments, the viral particle includes one or more viral structural proteins (e.g., capsid proteins). In some embodiments, the lipid bilayer is obtained from a membrane other than the plasma membrane of a host cell. In some embodiments, the viral envelope lipid bilayer is embedded with viral proteins, including viral glycoproteins.

[0281] In particular embodiments, the lipid particle is not virally derived. In some embodiments, the lipid particle can be a nanoparticle, a vesicle, an exosome, a dendrimer, an enucleated cell, a microvesicle, a membrane vesicle, an extracellular membrane vesicle, a plasma membrane vesicle, a giant plasma membrane vesicle, an apoptotic body, a mitoparticle, a pyrenocyte, a lysosome, another membrane enclosed vesicle, or a cell derived particle. In some embodiments, the provided particle is a cell-based particle, such as any described in Section II-A-3.1. Viral Particle

[0282] In some embodiments, the particle is a viral particle, e.g., a retroviral particle, e.g., a lentiviral particle. In some embodiments, the viral particle is recombinant. In some embodiments, the particle is a recombinant lentiviral particle.

[0283] In some embodiments, the lipid bilayer of the particle is or comprises the viral envelope. The viral envelope may comprise a fusogen that is endogenous to the virus or is a pseudotyped fusogen. In some embodiments, the particle’s lumen or cavity comprises a viral nucleic acid, e.g., a retroviral nucleic acid, e.g., a lentiviral nucleic acid. The viral nucleic acid may be a viral genome. In some embodiments,the particle may further comprise one or more viral non-structural proteins, e.g., in its cavity or lumen. In some embodiments, the particles are lenti viral particles. In some embodiments, the lenti viral particle is Human Immunodeficiency Virus-1 (HIV-1).

[0284] In some aspects, the particle is limited in the number of polynucleotides that can be packaged. In some embodiments, nucleotides encoding polypeptides to be packaged can be modified such that they retain functional activity with fewer nucleotides in the coding region than that which encodes for the wild-type peptide. Such modifications can include truncations, or other deletions. In some embodiments, more than one polypeptide can be expressed from the same promoter, such that they are fusion polypeptides. In some embodiments, the insert size to be packaged (i.e., viral genome, or portions thereof; or heterologous polynucleotides as described) can be between 500-1000, 1000-2000, 2000-3000, 3000-4000, 4000-5000, 5000-6000, 6000-7000, or 7000-8000 nucleotides in length. In some embodiments, the insert can be over 8000 nucleotides, such as 9000, 10,000, or 11,000 nucleotides in length.

[0285] In some embodiments, the particle, such as retroviral particle, comprises one or more of gag polyprotein, polymerase (e.g., pol), integrase (e.g., a functional or non-functional variant), protease, and a fusogen. In some embodiments, the particle further comprises rev. In some embodiments, one or more of the aforesaid proteins are encoded in the retroviral genome (e.g., the insert as described above), and in some embodiments, one or more of the aforesaid proteins are provided in trans, e.g., by a helper cell, helper virus, or helper plasmid. In some embodiments, the nucleic acid (e.g., retroviral nucleic acid) comprises one or more of the following nucleic acid sequences: 5’ LTR (e.g., comprising U5 and lacking a functional U3 domain), Psi packaging element (Psi), Central polypurine tract (cPPT) Promoter operatively linked to a payload gene encoding a payload agent, payload gene (optionally comprising an intron before the open reading frame), Poly A tail sequence, WPRE, and 3’ LTR (e.g., comprising U5 and lacking a functional U3). In some embodiments, the nucleic acid further comprises a retroviral cisacting RNA packaging element and a cPPT / CTS element. In some embodiments the nucleic acid further comprises one or more insulator element. In some embodiments, the recognition sites are situated between the poly A tail sequence and the WPRE.

[0286] In some embodiments, the particle comprises supramolecular complexes formed by viral proteins that self-assemble into capsids. In some embodiments, the particle is a viral particle derived from viral capsids. In some embodiments, the particle is a viral particle derived from viral nucleocapsids. In some embodiments, the particle comprises nucleocapsid-derived proteins that retain the property of packaging nucleic acids.

[0287] In some embodiments, the particle packages one or more viral nucleic acids (e.g., retroviral nucleic acids) during the expression process. In some embodiments, the nucleic acids do not encode any genes involved in virus replication. In particular embodiments, the particle is replication defective.

[0288] In some embodiments, the retroviral nucleic acid comprises one or more of (e.g., all of): a 5’ promoter (e.g., to control expression of the entire packaged RNA), a 5’ LTR (e.g., that includes R (poly adenylation tail signal) and / or U5 which includes a primer activation signal), a primer binding site, a psi packaging signal, a RRE element for nuclear export, a promoter directly upstream of the payload gene to control payload gene expression, a payload gene, a polypurine tract, and a 3’ LTR (e.g., that includes a mutated U3, a R, and U5). In some embodiments, the retroviral nucleic acid further comprises one or more of a cPPT, a WPRE, and / or an insulator element.

[0289] A retrovirus typically replicates by reverse transcription of its genomic RNA into a linear double-stranded DNA copy and subsequently covalently integrates its genomic DNA into a host genome. Illustrative retroviruses suitable for use in particular embodiments include Moloney murine leukemia virus (M-MuLV), Moloney murine sarcoma virus (MoMSV), Harvey murine sarcoma virus (HaMuSV), murine mammary tumor virus (MuMTV), gibbon ape leukemia virus (GaLV), feline leukemia virus (FLV), spuma virus, Friend murine leukemia virus, Murine Stem Cell Virus (MSCV) and Rous Sarcoma Virus (RSV)) and lentivirus.

[0290] In some embodiments the retrovirus is a Gammretro virus. In some embodiments the retrovirus is an Epsilonretrovirus. In some embodiments the retrovirus is an Alpharetrovirus. In some embodiments the retrovirus is a Betaretro virus. In some embodiments the retrovirus is a Deltaretro virus. In some embodiments the retrovirus is a Lentivirus. In some embodiments the retrovirus is a Spumaretrovirus. In some embodiments the retrovirus is an endogenous retrovirus.

[0291] Illustrative lenti viruses include HIV (human immunodeficiency virus; including HIV type 1, and HIV type 2); visna-maedi virus (VMV) virus; the caprine arthritis-encephalitis virus (CAEV); equine infectious anemia virus (EIAV); feline immunodeficiency virus (FIV); bovine immune deficiency virus (BIV); and simian immunodeficiency virus (SIV). In some embodiments, HIV based particle backbones (e.g., HIV cis-acting sequence elements) are used.

[0292] A viral particle can comprise a nucleic acid molecule (e.g., a transfer plasmid) that includes virus-derived nucleic acid elements that typically facilitate transfer of a nucleic acid molecule or integration into the genome of a cell or to a viral particle that mediates nucleic acid transfer. Viral particles will typically include various viral components and sometimes also host cell components in addition to nucleic acids. Viral particles and transfer plasmids can comprise structural and / or functional genetic elements that are primarily derived from a virus. A retroviral particle can comprise a viral particle or plasmid containing structural and functional genetic elements, or portions thereof, that are primarily derived from a retrovirus. A lenti viral particle can comprise a viral particle or plasmid containing structural and functional genetic elements, or portions thereof, including LTRs that are primarily derived from a lentivirus.

[0293] In embodiments, a lentiviral particle may comprise a lentiviral transfer plasmid (e.g., as naked DNA) or an infectious lentiviral particle. With respect to elements such as cloning sites,promoters, regulatory elements, heterologous nucleic acids, etc., it is to be understood that the sequences of these elements can be present in RNA form in lentiviral particles and can be present in DNA form in DNA plasmids.

[0294] In some particles described herein, at least part of one or more protein coding regions that contribute to or are essential for replication may be absent compared to the corresponding wild- type virus. In some embodiments, this makes the viral particle replication-defective. In some embodiments, the particle is capable of transducing a target non-dividing host cell and / or integrating its genome into a host genome.

[0295] The structure of a wild-type retrovirus genome often comprises a 5' long terminal repeat (LTR) and a 3' LTR, between or within which are located a packaging signal to enable the genome to be packaged, a primer binding site, integration sites to enable integration into a host cell genome and gag, pol and env genes encoding the packaging components which promote the assembly of viral particles. More complex retroviruses have additional features, such as rev and RRE sequences in HIV, which enable the efficient export of RNA transcripts of the integrated provirus from the nucleus to the cytoplasm of an infected target cell. In the provirus, the viral genes are flanked at both ends by regions called long terminal repeats (LTRs). The LTRs are involved in pro viral integration and transcription. LTRs also serve as enhancer-promoter sequences and can control the expression of the viral genes. Encapsidation of the retroviral RNAs occurs by virtue of a psi sequence located at the 5' end of the viral genome.

[0296] The LTRs themselves are typically similar (e.g., identical) sequences that can be divided into three elements, which are called U3, R and U5. U3 is derived from the sequence unique to the 3' end of the RNA. R is derived from a sequence repeated at both ends of the RNA and U5 is derived from the sequence unique to the 5' end of the RNA. The sizes of the three elements can vary considerably among different retroviruses.

[0297] For the viral genome, the site of transcription initiation is typically at the boundary between U3 and R in one LTR and the site of poly (A) addition (termination) is at the boundary between R and U5 in the other LTR. U3 contains most of the transcriptional control elements of the provirus, which include the promoter and multiple enhancer sequences responsive to cellular and in some cases, viral transcriptional activator proteins. Some retroviruses comprise any one or more of the following genes that code for proteins that are involved in the regulation of gene expression: tot, rev, tax and rex. With regard to the structural genes gag, pol and env themselves, gag encodes the internal structural protein of the virus. Gag protein is proteolytically processed into the mature proteins MA (matrix), CA (capsid) and NC (nucleocapsid). The pol gene encodes the reverse transcriptase (RT), which contains DNA polymerase, associated RNase H and integrase (IN), which mediate replication of the genome. The env gene encodes the surface (SU) glycoprotein and the transmembrane (TM) protein of the virion, whichform a complex that interacts specifically with cellular receptor proteins. This interaction promotes infection, e.g., by fusion of the viral membrane with the cell membrane.

[0298] In a replication-defective retroviral genome, gag, pol and env may be absent or not functional. The R regions at both ends of the RNA are typically repeated sequences. U5 and U3 represent unique sequences at the 5' and 3' ends of the RNA genome respectively.

[0299] Retroviruses may also contain additional genes which code for proteins other than gag, pol and env. Examples of additional genes include (in HIV), one or more of vif, vpr, vpx, vpu, tat, rev and nef. El AV has (amongst others) the additional gene S2. Proteins encoded by additional genes serve various functions, some of which may be duplicative of a function provided by a cellular protein. In EIAV, for example, tat acts as a transcriptional activator of the viral LTR (Derse and Newbold 1993 Virology 194:530-6; Maury et al. 1994 Virology 200:632- 42). It binds to a stable, stem-loop RNA secondary structure referred to as TAR. Rev regulates and co-ordinates the expression of viral genes through rev-response elements (RRE) (Martarano et al. 1994 J. Virol. 68:3102-11). The mechanisms of action of these two proteins are thought to be broadly similar to the analogous mechanisms in the primate viruses. In addition, an EIAV protein, Ttm, has been identified that is encoded by the first exon of tat spliced to the env coding sequence at the start of the transmembrane protein.

[0300] In addition to protease, reverse transcriptase and integrase, non-primate lentiviruses contain a fourth pol gene product which codes for a dUTPase. This may play a role in the ability of these lentiviruses to infect certain non-dividing or slowly dividing cell types.

[0301] In embodiments, a recombinant lenti viral particle (RLV) is a particle with sufficient retroviral genetic information to allow packaging of an RNA genome, in the presence of packaging components, into a viral particle capable of infecting a target cell. Infection of the target cell can comprise reverse transcription and integration into the target cell genome. The RLV typically carries non-viral coding sequences which are to be delivered by the particle to the target cell, such as a nucleic acid encoding a payload agent as described herein. In embodiments, an RLV is incapable of independent replication to produce infectious retroviral particles within the target cell. Usually the RLV lacks a functional gag-pol and / or env gene and / or other genes involved in replication. The particle may be configured as a split-intron particle, e.g., as described in PCT patent application WO 99 / 15683, which is herein incorporated by reference in its entirety.

[0302] In some embodiments, the lentiviral particle comprises a minimal viral genome, e.g., the viral particle has been manipulated so as to remove the non-essential elements and to retain the essential elements in order to provide the required functionality to infect, transduce and deliver a nucleotide sequence of interest to a target host cell, e.g., as described in WO 98 / 17815, which is herein incorporated by reference in its entirety.

[0303] A minimal lentiviral genome may comprise, e.g., (5')R-U5-one or more first nucleotide sequences-U3-R(3'). However, the plasmid used to produce the lentiviral genome within a source cell canalso include transcriptional regulatory control sequences operably linked to the lentiviral genome to direct transcription of the genome in a source cell. These regulatory sequences may comprise the natural sequences associated with the transcribed retroviral sequence, e.g., the 5' U3 region, or they may comprise a heterologous promoter such as another viral promoter, for example the CMV promoter. Some lentiviral genomes comprise additional sequences to promote efficient virus production. For example, in the case of HIV, rev and RRE sequences may be included. Alternatively or combination, codon optimization may be used, e.g., the payload gene encoding the payload agent may be codon optimized, e.g., as described in WO 01 / 79518, which is herein incorporated by reference in its entirety. Alternative sequences which perform a similar or the same function as the rev / RRE system may also be used. For example, a functional analogue of the rev / RRE system is found in the Mason Pfizer monkey virus. This is known as CTE and comprises an RRE-type sequence in the genome which is believed to interact with a factor in the infected cell. The cellular factor can be thought of as a rev analogue. Thus, CTE may be used as an alternative to the rev / RRE system. In addition, the Rex protein of HTLV-I can functionally replace the Rev protein of HIV-I. Rev and Rex have similar effects to IRE-BP.

[0304] In some embodiments, a retroviral nucleic acid (e.g., a lentiviral nucleic acid, e.g., a primate or non-primate lentiviral nucleic acid) (1) comprises a deleted gag gene wherein the deletion in gag removes one or more nucleotides downstream of about nucleotide 350 or 354 of the gag coding sequence; (2) has one or more accessory genes absent from the retroviral nucleic acid; (3) lacks the tat gene but includes the leader sequence between the end of the 5' LTR and the ATG of gag; and (4) combinations of (1), (2) and (3). In an embodiment, the lentiviral particle comprises all of features (1) and (2) and (3). This strategy is described in more detail in WO 99 / 32646, which is herein incorporated by reference in its entirety.

[0305] In some embodiments, a primate lentivirus minimal system requires none of the HIV / SIV additional genes vif, vpr, vpx, vpu, tat, rev and nef for either particle production or for transduction of dividing and non-dividing cells. In some embodiments, an EIAV minimal particle system does not require S2 for either particle production or for transduction of dividing and non-dividing cells.

[0306] The deletion of additional genes may permit particles to be produced without the genes associated with disease in lentiviral (e.g. HIV) infections. In particular, tat is associated with disease. Secondly, the deletion of additional genes permits the particle to package more heterologous DNA. Thirdly, genes whose function is unknown, such as S2, may be omitted, thus reducing the risk of causing undesired effects. Examples of minimal lentiviral particles are disclosed in WO 99 / 32646 and in WO 98 / 17815.

[0307] In some embodiments, the retroviral nucleic acid is devoid of at least tat and S2 (if it is an EIAV particle system), and possibly also vif, vpr, vpx, vpu and nef. In some embodiments, the retroviral nucleic acid is also devoid of rev, RRE, or both.

[0308] In some embodiments the retroviral nucleic acid comprises vpx. The Vpx polypeptide binds to and induces the degradation of the SAMHD1 restriction factor, which degrades free dNTPs in the cytoplasm. Thus, the concentration of free dNTPs in the cytoplasm increases as Vpx degrades SAMHD1 and reverse transcription activity is increased, thus facilitating reverse transcription of the retroviral genome and integration into the target cell genome.

[0309] Different cells differ in their usage of particular codons. This codon bias corresponds to a bias in the relative abundance of particular tRNAs in the cell type. By altering the codons in the sequence so that they are tailored to match with the relative abundance of corresponding tRNAs, it is possible to increase expression. By the same token, it is possible to decrease expression by deliberately choosing codons for which the corresponding tRNAs are known to be rare in the particular cell type. Thus, an additional degree of translational control is available. An additional description of codon optimization is found, e.g., in WO 99 / 41397, which is herein incorporated by reference in its entirety.

[0310] Many viruses, including HIV and other lentiviruses, use a large number of rare codons and by changing these to correspond to commonly used mammalian codons, increased expression of the packaging components in mammalian producer cells can be achieved.

[0311] In some embodiments, codon optimization has a number of other advantages. In some embodiments, by virtue of alterations in their sequences, the nucleotide sequences encoding the packaging components may have RNA instability sequences (INS) reduced or eliminated from them. At the same time, the amino acid sequence coding sequence for the packaging components is retained so that the viral components encoded by the sequences remain the same, or at least sufficiently similar that the function of the packaging components is not compromised. In some embodiments, codon optimization also overcomes the Rev / RRE requirement for export, rendering optimized sequences Rev independent. In some embodiments, codon optimization also reduces homologous recombination between different constructs within the particle system (for example between the regions of overlap in the gag-pol and env open reading frames). In some embodiments, codon optimization leads to an increase in viral titer and / or improved safety.

[0312] In some embodiments, only codons relating to INS are codon optimized. In other embodiments, the sequences are codon optimized in their entirety, with the exception of the sequence encompassing the frameshift site of gag-pol.

[0313] The gag-pol gene comprises two overlapping reading frames encoding the gag-pol proteins. The expression of both proteins depends on a frameshift during translation. This frameshift occurs as a result of ribosome "slippage" during translation. This slippage is thought to be caused at least in part by ribosome-stalling RNA secondary structures. Such secondary structures exist downstream of the frameshift site in the gag-pol gene. For HIV, the region of overlap extends from nucleotide 1222 downstream of the beginning of gag (wherein nucleotide 1 is the A of the gag ATG) to the end of gag (nt 1503). Consequently, a 281 bp fragment spanning the frameshift site and the overlapping region of thetwo reading frames is preferably not codon optimized. In some embodiments, retaining this fragment will enable more efficient expression of the gag-pol proteins. For EIAV, the beginning of the overlap is at nt 1262 (where nucleotide 1 is the A of the gag ATG). The end of the overlap is at nt 1461. In order to ensure that the frameshift site and the gag-pol overlap are preserved, the wild type sequence may be retained from nt 1156 to 1465.

[0314] In some embodiments, derivations from optimal codon usage may be made, for example, in order to accommodate convenient restriction sites, and conservative amino acid changes may be introduced into the gag-pol proteins.

[0315] In some embodiments, codon optimization is based on codons with poor codon usage in mammalian systems. The third and sometimes the second and third base may be changed.

[0316] In some embodiments, due to the degenerate nature of the genetic code, it will be appreciated that numerous gag-pol sequences can be achieved by a skilled worker. Also, there are many retroviral variants described which can be used as a starting point for generating a codon optimized gag-pol sequence. Lentiviral genomes can be quite variable. For example there are many quasi-species of HIV-I which are still functional. This is also the case for EIAV. These variants may be used to enhance particular parts of the transduction process. Examples of HIV-I variants may be found in the HIV databases maintained by Los Alamos National Laboratory. Details of EIAV clones may be found at the NCBI database maintained by the National Institutes of Health.

[0317] It is within the level of a skilled artisan to empirically determine appropriate codon optimization of viral sequences. The strategy for codon optimized sequences, including gag-pol sequences, can be used in relation to any retrovirus, e.g., EIAV, FIV, BIV, CAEV, VMR, SIV, HIV-I and HIV -2. In addition this method could be used to increase expression of genes from HTLV-I, HTLV-2, HFV, HSRV and human endogenous retroviruses (HERV), MLV and other retroviruses.

[0318] In embodiments, the retroviral particle comprises a packaging signal that comprises from 255 to 360 nucleotides of gag in particles that still retain env sequences, or about 40 nucleotides of gag in a particular combination of splice donor mutation, gag and env deletions. In some embodiments, the retroviral particle includes a gag sequence which comprises one or more deletions, e.g., the gag sequence comprises about 360 nucleotides derivable from the N-terminus.

[0319] In some embodiments, the retroviral particle, helper cell, helper virus, or helper plasmid may comprise retroviral structural and accessory proteins, for example gag, pol, env, tat, rev, vif, vpr, vpu, vpx, or nef proteins or other retroviral proteins. In some embodiments, the retroviral proteins are derived from the same retrovirus. In some embodiments, the retroviral proteins are derived from more than one retrovirus, e.g. 2, 3, 4, or more retroviruses.

[0320] In some embodiments, the gag and pol coding sequences are generally organized as the Gag- Pol Precursor in native lentivirus. The gag sequence codes for a 55-kD Gag precursor protein, also calledp55. The p55 is cleaved by the virally encoded protease (a product of the pol gene) during the process of maturation into four smaller proteins designated MA (matrix [pl7]), CA (capsid [p24]), NC (nucleocapsid [p9]) , and p6. The pol precursor protein is cleaved away from Gag by a virally encoded protease, and further digested to separate the protease (plO), RT (p50), RNase H (pl5), and integrase (p31) activities.

[0321] In some embodiments, the lentiviral particle is integration-deficient. In some embodiments, the pol is integrase deficient, such as by encoding due to mutations in the integrase gene. For example, the pol coding sequence can contain an inactivating mutation in the integrase, such as by mutation of one or more of amino acids involved in catalytic activity, i.e. mutation of one or more of aspartic 64, aspartic acid 116 and / or glutamic acid 152. In some embodiments, the integrase mutation is a D64V mutation. In some embodiments, the mutation in the integrase allows for packaging of viral RNA into a lentivirus. In some embodiments, the mutation in the integrase allows for packaging of viral proteins into a lentivirus. In some embodiments, the mutation in the integrase reduces the possibility of insertional mutagenesis. In some embodiments, the mutation in the integrase decreases the possibility of generating replication- competent recombinants (RCRs) (Wanisch et al. 2009. Mol Ther. 1798): 1316-1332). In some embodiments, native Gag-Pol sequences can be utilized in a helper particle (e.g., helper plasmid or helper virus), or modifications can be made. These modifications include chimeric Gag-Pol, where the Gag and Pol sequences are obtained from different viruses (e.g., different species, subspecies, strains, clades, etc.), and / or where the sequences have been modified to improve transcription and / or translation, and / or reduce recombination.

[0322] In some embodiments, the retroviral nucleic acid includes a polynucleotide encoding a 150- 250 (e.g., 168) nucleotide portion of a gag protein that (i) includes a mutated INS1 inhibitory sequence that reduces restriction of nuclear export of RNA relative to wild-type INS1, (ii) contains two nucleotide insertion that results in frame shift and premature termination, and / or (iii) does not include INS2, INS3, and INS4 inhibitory sequences of gag.

[0323] In some embodiments, a particle described herein is a hybrid particle that comprises both retroviral (e.g., lentiviral) sequences and non-lentiviral viral sequences. In some embodiments, a hybrid particle comprises retroviral, e.g., lentiviral, sequences for reverse transcription, replication, integration and / or packaging.

[0324] According to certain specific embodiments, most or all of the viral particle backbone sequences are derived from a lentivirus, e.g., HIV-1. However, it is to be understood that many different sources of retroviral and / or lentiviral sequences can be used, or combined and numerous substitutions and alterations in certain of the lentiviral sequences may be accommodated without impairing the ability of a transfer particle to perform the functions described herein. A variety of lentiviral particles are described in Naldini et ah, (1996a, 1996b, and 1998); Zufferey et al., (1997); Dull et al., 1998, U.S. Pat. Nos. 6,013,516; and 5,994,136, many of which may be adapted to produce a retroviral nucleic acid.

[0325] At each end of the provirus, long terminal repeats (LTRs) are typically found. An LTR typically comprises a domain located at the ends of retroviral nucleic acid which, in their natural sequence context, are direct repeats and contain U3, R and U5 regions. LTRs generally promote the expression of retroviral genes (e.g., promotion, initiation and poly adenylation of gene transcripts) and viral replication. The LTR can comprise numerous regulatory signals including transcriptional control elements, polyadenylation signals and sequences for replication and integration of the viral genome. The viral LTR is typically divided into three regions called U3, R and U5. The U3 region typically contains the enhancer and promoter elements. The U5 region is typically the sequence between the primer binding site and the R region and can contain the polyadenylation sequence. The R (repeat) region can be flanked by the U3 and U5 regions. The LTR is typically composed of U3, R and U5 regions and can appear at both the 5' and 3' ends of the viral genome. In some embodiments, adjacent to the 5' LTR are sequences for reverse transcription of the genome (the tRNA primer binding site) and for efficient packaging of viral RNA into particles (the Psi site).

[0326] In some embodiments, a packaging signal can comprise a sequence located within the retroviral genome which mediate insertion of the viral RNA into the viral capsid or particle, see e.g., Clever et al., 1995. J. of Virology, Vol. 69, No. 4; pp. 2101-2109. Several retroviral particles use a minimal packaging signal (a psi [Y] sequence) for encapsidation of the viral genome.

[0327] In various embodiments, retroviral nucleic acids comprise modified 5' LTR and / or 3' LTRs. Either or both of the LTR may comprise one or more modifications including, but not limited to, one or more deletions, insertions, or substitutions. Modifications of the 3' LTR are often made to improve the safety of lentiviral or retroviral systems by rendering viruses replication-defective, e.g., virus that is not capable of complete, effective replication such that infective virions are not produced (e.g., replicationdefective lentiviral progeny).

[0328] In some embodiments, a particle is a self-inactivating (SIN) particle, e.g., replicationdefective particle, e.g., retroviral or lentiviral particle, in which the right (3') LTR enhancer- promoter region, known as the U3 region, has been modified (e.g., by deletion or substitution) to prevent viral transcription beyond the first round of viral replication. In some aspects, provided herein is a replication incompetent (also referred to herein as replication defective) particle, that cannot participate in replication in the absence of the packaging cell (i.e., viral particles are not produced from the transduced cell). In some aspects, this is because the right (3') LTR U3 region can be used as a template for the left (5') LTR U3 region during viral replication and, thus, absence of the U3 enhancer-promoter inhibits viral replication. In embodiments, the 3' LTR is modified such that the U5 region is removed, altered, or replaced, for example, with an exogenous poly(A) sequence The 3' LTR, the 5' LTR, or both 3' and 5' LTRs, may be modified LTRs. Other modifications to the viral particle, i.e., retroviral or lentiviral particle, to render said particle replication incompetent are known in the art.

[0329] In some embodiments, the U3 region of the 5' LTR is replaced with a heterologous promoter to drive transcription of the viral genome during production of viral particles. Examples of heterologous promoters which can be used include, for example, viral simian virus 40 (SV40) (e.g., early or late), cytomegalovirus (CMV) (e.g., immediate early), Moloney murine leukemia virus (MoMLV), Rous sarcoma virus (RSV), and herpes simplex virus (HSV) (thymidine kinase) promoters. In some embodiments, promoters are able to drive high levels of transcription in a Tat- independent manner. In certain embodiments, the heterologous promoter has additional advantages in controlling the manner in which the viral genome is transcribed. For example, the heterologous promoter can be inducible, such that transcription of all or part of the viral genome will occur only when the induction factors are present. Induction factors include, but are not limited to, one or more chemical compounds or the physiological conditions such as temperature or pH, in which the host cells are cultured.

[0330] In some embodiments, viral particles comprise a TAR (trans-activation response) element, e.g., located in the R region of lentiviral (e.g., HIV) LTRs. This element interacts with the lentiviral trans-activator (tat) genetic element to enhance viral replication. However, this element is not required, e.g., in embodiments wherein the U3 region of the 5' LTR is replaced by a heterologous promoter.

[0331] The R region, e.g., the region within retroviral LTRs beginning at the start of the capping group (i.e., the start of transcription) and ending immediately prior to the start of the poly A tract can be flanked by the U3 and U5 regions. The R region plays a role during reverse transcription in the transfer of nascent DNA from one end of the genome to the other.

[0332] The retroviral nucleic acid can also comprise a FLAP element, e.g., a nucleic acid whose sequence includes the central polypurine tract and central termination sequences (cPPT and CTS) of a retrovirus, e.g., HIV-1 or HIV-2. Suitable FLAP elements are described in U.S. Pat. No. 6,682,907 and in Zennou, et ah, 2000, Cell, 101:173, which are herein incorporated by reference in their entireties. During HIV-1 reverse transcription, central initiation of the plus-strand DNA at the central polypurine tract (cPPT) and central termination at the central termination sequence (CTS) can lead to the formation of a three- stranded DNA structure: the HIV-1 central DNA flap. In some embodiments, the retroviral or lentiviral particle backbones comprise one or more FLAP elements upstream or downstream of the payload gene encoding the payload agent. For example, in some embodiments a transfer plasmid includes a FLAP element, e.g., a FLAP element derived or isolated from HIV-L

[0333] In embodiments, a retroviral or lentiviral nucleic acid comprises one or more export elements, e.g., a cis-acting post-transcriptional regulatory element which regulates the transport of an RNA transcript from the nucleus to the cytoplasm of a cell. Examples of RNA export elements include the human immunodeficiency virus (HIV) rev response element (RRE) (see e.g., Cullen et al., 1991. J. Virol. 65: 1053; and Cullen et al., 1991. Cell 58: 423), and the hepatitis B virus post-transcriptional regulatory element (HPRE), which are herein incorporated by reference in their entireties. Generally, the RNA export element is placed within the 3' UTR of a gene, and can be inserted as one or multiple copies.

[0334] In some embodiments, expression of heterologous sequences (e.g., nucleic acid encoding a payload agent) in viral particles is increased by incorporating one or more of, e.g., all of, posttranscriptional regulatory elements, polyadenylation sites, and transcription termination signals into the particles. A variety of posttranscriptional regulatory elements can increase expression of a heterologous nucleic acid at the protein, e.g., woodchuck hepatitis virus posttranscriptional regulatory element (WPRE; Zufferey et al., 1999, J. Virol., 73:2886); the posttranscriptional regulatory element present in hepatitis B virus (HPRE) (Huang et al., Mol. Cell. Biol., 5:3864); and the like (Liu et al., 1995, Genes Dev., 9:1766), each of which is herein incorporated by reference in its entirety. In some embodiments, a retroviral nucleic acid described herein comprises a posttranscriptional regulatory element such as a WPRE or HPRE

[0335] In some embodiments, a retroviral nucleic acid described herein lacks or does not comprise a posttranscriptional regulatory element such as a WPRE or HPRE.

[0336] Elements directing the termination and polyadenylation of the heterologous nucleic acid transcripts may be included, e.g., to increase expression of the payload agent. Transcription termination signals may be found downstream of the polyadenylation signal. In some embodiments, particles comprise a polyadenylation sequence 3' of a polynucleotide encoding the payload agent. A polyA site may comprise a DNA sequence which directs both the termination and polyadenylation of the nascent RNA transcript by RNA polymerase II. Polyadenylation sequences can promote mRNA stability by addition of a polyA tail to the 3' end of the coding sequence and thus, contribute to increased translational efficiency. Illustrative examples of polyA signals that can be used in a retroviral nucleic acid, include AATAAA, ATT AAA, AGTAAA, a bovine growth hormone polyA sequence (BGHpA), a rabbit b- globin polyA sequence (rPgpA), or another suitable heterologous or endogenous polyA sequence.

[0337] In some embodiments, a retroviral or lenti viral particle further comprises one or more insulator elements, e.g., an insulator element described herein.

[0338] In various embodiments, the particles comprise a promoter operably linked to a polynucleotide encoding a payload agent. The particles may have one or more LTRs, wherein either LTR comprises one or more modifications, such as one or more nucleotide substitutions, additions, or deletions. The particles may further comprise one of more accessory elements to increase transduction efficiency (e.g., a cPPT / FLAP), viral packaging (e.g., a Psi packaging signal, RRE), and / or other elements that increase payload gene expression (e.g., poly (A) sequences), and may optionally comprise a WPRE or HPRE.

[0339] In some embodiments, a lentiviral nucleic acid comprises one or more of, e.g., all of, e.g., from 5’ to 3’, a promoter (e.g., CMV), an R sequence (e.g., comprising TAR), a U5 sequence (e.g., for integration), a PBS sequence (e.g., for reverse transcription), a DIS sequence (e.g., for genome dimerization), a psi packaging signal, a partial gag sequence, an RRE sequence (e.g., for nuclear export), a cPPT sequence (e.g., for nuclear import), a promoter to drive expression of the payload agent, apayload gene encoding the payload agent, a WPRE sequence (e.g., for efficient transgene expression), a PPT sequence (e.g., for reverse transcription), an R sequence (e.g., for polyadenylation and termination), and a U5 signal (e.g., for integration).2. Virus-Like Particle

[0340] In some embodiments, the particle is a virus-like particle (VLP) that is derived from virus. In some embodiments, the viral envelope may comprise a fusogen, e.g., a fusogen that is endogenous to the virus or a pseudotyped fusogen. The VLPS include those derived from retroviruses or lentiviruses. While VLPs mimic native virion structure, they lack the viral genomic information necessary for independent replication within a host cell. Therefore, in some aspects, VLPs are non-infectious. In particular embodiments, a VLP does not contain a viral genome. In some embodiments, the VLP’s bilayer of amphipathic lipids is or comprises the viral envelope. In some embodiments, the particle’s bilayer of amphipathic lipids is or comprises lipids derived from a cell. In some embodiments, a VLP contains at least one type of structural protein from a virus. In most cases this protein will form a proteinaceous capsid. In some cases the capsid will also be enveloped in a lipid bilayer originating from the cell from which the assembled VLP has been released (e.g., VLPs comprising a human immunodeficiency virus structural protein such as GAG). In some embodiments, the VLP further comprises a targeting moiety as an envelope protein within the lipid bilayer.

[0341] In some embodiments, the particle comprises supramolecular complexes formed by viral proteins that self-assemble into capsids. In some embodiments, the particle is a virus-like particle derived from viral capsid proteins. In some embodiments, the particle is a virus-like particle derived from viral nucleocapsid proteins. In some embodiments, the particle comprises nucleocapsid-derived proteins that retain the property of packaging nucleic acids. In some embodiments, the particle comprises only viral structural glycoproteins among proteins from the viral genome. In some embodiments, the particle does not contain a viral genome.

[0342] In some embodiments, the particle packages nucleic acids during the expression process, such as a nucleic acid encoding a payload agent. In some embodiments, the nucleic acids do not encode any genes involved in virus replication. In particular embodiments, the particle is a virus-like particle, e.g., retrovirus-like particle, such as a lentivirus-like particle, that is replication defective.

[0343] In some embodiments, the particle is a virus-like particle which comprises a sequence that is devoid of or lacking viral RNA, which may be the result of removing or eliminating the viral RNA from the sequence. In some embodiments, this may be achieved by using an endogenous packaging signal binding site on gag. In some embodiments, the endogenous packaging signal binding site is on pol. In some embodiments, the RNA which is to be delivered will contain a cognate packaging signal. In some embodiments, a heterologous binding domain (which is heterologous to gag) located on the RNA to be delivered, and a cognate binding site located on gag or pol, can be used to ensure packaging of the RNAto be delivered. In some embodiments, the heterologous sequence could be non-viral or it could be viral, in which case it may be derived from a different virus. In some embodiments, the particles could be used to deliver therapeutic RNA, in which case functional integrase and / or reverse transcriptase is not required. In some embodiments, the particles could also be used to deliver a therapeutic gene of interest, in which case pol is typically included.3. Cell-Based Particle

[0344] In some embodiments, the particle is a cell based particle that comprises a naturally derived membrane. In some embodiments, the naturally derived membrane comprises membrane vesicles prepared from cells or tissues. In some embodiments, the cell based particle comprises a vesicle that is obtainable from a cell. In some embodiments, the cell based particle comprises a microvesicle, an exosome, a membrane enclosed body, an apoptotic body (from apoptotic cells), a particle (which may be derived from e.g. platelets), an ectosome (derivable from, e.g., neutrophiles and monocytes in serum), a prostatosome (obtainable from prostate cancer cells), or a cardiosome (derivable from cardiac cells).

[0345] In some embodiments, the source cell is an endothelial cell, a fibroblast, a blood cell (e.g., a macrophage, a neutrophil, a granulocyte, a leukocyte), a stem cell (e.g., a mesenchymal stem cell, an umbilical cord stem cell, bone marrow stem cell, a hematopoietic stem cell, an induced pluripotent stem cell e.g., an induced pluripotent stem cell derived from a subject’s cells), an embryonic stem cell (e.g., a stem cell from embryonic yolk sac, placenta, umbilical cord, fetal skin, adolescent skin, blood, bone marrow, adipose tissue, erythropoietic tissue, hematopoietic tissue), a myoblast, a parenchymal cell (e.g., hepatocyte), an alveolar cell, a neuron (e.g., a retinal neuronal cell) a precursor cell (e.g., a retinal precursor cell, a myeloblast, myeloid precursor cells, a thymocyte, a meiocyte, a megakaryoblast, a promegakaryoblast, a melanoblast, a lymphoblast, a bone marrow precursor cell, a normoblast, or an angioblast), a progenitor cell (e.g., a cardiac progenitor cell, a satellite cell, a radial gial cell, a bone marrow stromal cell, a pancreatic progenitor cell, an endothelial progenitor cell, a blast cell), or an immortalized cell (e.g., HeEa, HEK293, HFF-1, MRC-5, WI-38, IMR 90, IMR 91, PER.C6, HT-1080, or BJ cell). In some embodiments, the source cell is other than a 293 cell, HEK cell, human endothelial cell, or a human epithelial cell, monocyte, macrophage, dendritic cell, or stem cell.

[0346] In some embodiments, the cell based particle has a density of <1, 1-1.1, 1.05-1.15, 1.1-1.2, 1.15-1.25, 1.2-1.3, 1.25-1.35, or >1.35 g / ml. In embodiments, the cell based particle has a size that is less than about 0.01%, 0.05%, 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, of that of the source cell. In some embodiments, a composition containing a plurality of the cell based particle comprises less than 0.01%, 0.05%, 0.1%, 0.5%, 1%, 1.5%, 2%, 2.5%, 3%, 4%, 5%, or 10% source cells by protein mass or less than 0.01%, 0.05%, 0.1%, 0.5%, 1%, 1.5%, 2%, 2.5%, 3%, 4%, 5%, or 10% of cells having a functional nucleus. In some embodiments, the source cell used to make the cell based particle will not be available for testing after the particle is made.

[0347] In some embodiments the cell based particle is an extracellular vesicle, e.g., a cell based vesicle comprising a membrane that encloses an internal space and has a smaller diameter than the cell from which it is derived. In embodiments the extracellular vesicle has a diameter from 20 nm to 1000 nm. In some embodiments, the cell based particle is an apoptotic body, a fragment of a cell, a vesicle derived from a cell by direct or indirect manipulation, a vesiculated organelle, and a vesicle produced by a living cell (e.g., by direct plasma membrane budding or fusion of the late endosome with the plasma membrane). In embodiments the extracellular vesicle is derived from a living or dead organism, explanted tissues or organs, or cultured cells.

[0348] In embodiments, the cell based particle is a nanovesicle, e.g., a cell-derived small (e.g., between 20-250 nm in diameter, or 30-150 nm in diameter) vesicle comprising a membrane that encloses an internal space, and which is generated from said cell by direct or indirect manipulation. The production of nanovesicles can, in some instances, result in the destruction of the source cell. The nanovesicle may comprise a lipid or fatty acid and polypeptide.

[0349] In embodiments, the cell based particle is an exosome. In embodiments, the exosome is a cell-derived small (e.g., between 20-300 nm in diameter, or 40-200 nm in diameter) vesicle comprising a membrane that encloses an internal space, and which is generated from said cell by direct plasma membrane budding or by fusion of the late endosome with the plasma membrane. In embodiments, production of exosomes does not result in the destruction of the source cell. In embodiments, the exosome comprises lipid or fatty acid and polypeptide. Exemplary exosomes and other membrane- enclosed bodies are also described in WO / 2017 / 161010, WO / 2016 / 077639, US20160168572, US20150290343, and US20070298118, each of which is incorporated by reference herein in its entirety.

[0350] In some embodiments, the cell based particle is a microvesicle. In some embodiments the micro vesicle has a diameter of about 100 nm to about 2000 nm.

[0351] In some embodiments, the cell based particle is a cell ghost. In some embodiments, a vesicle is a plasma membrane vesicle, e.g. a giant plasma membrane vesicle.

[0352] In some embodiments, the cell based particle is derived from a source cell with a genetic modification which results in increased expression of an immunomodulatory agent, such as an immunosuppressive agent. In some embodiments, the immunosuppressive agent is on an exterior surface of the cell. In some embodiments, the immunosuppressive agent is incorporated into the exterior surface of the particle. In some embodiments, the particle comprises an immunomodulatory agent attached to the surface of the solid particle by a covalent or non-covalent bond

[0353] In some embodiments, cell based particles are generated by inducing budding of an exosome, microvesicle, membrane vesicle, extracellular membrane vesicle, plasma membrane vesicle, giant plasma membrane vesicle, apoptotic body, mitoparticle, pyrenocyte, lysosome, or other membrane enclosed vesicle.

[0354] In some embodiments, cell based particles are generated by inducing cell enucleation.Enucleation may be performed using assays such as genetic, chemical (e.g., using Actinomycin D, see Bayona-Bafaluyet al., “A chemical enucleation method for the transfer of mitochondrial DNA to p° cells” Nucleic Acids Res. 2003 Aug 15; 31(16): e98), mechanical methods (e.g., squeezing or aspiration, see Lee et al., “A comparative study on the efficiency of two enucleation methods in pig somatic cell nuclear transfer: effects of the squeezing and the aspiration methods.” Anim Biotechnol. 2008;19(2):71- 9), or combinations thereof.

[0355] In some embodiments, the cell based particles are generated by inducing cell fragmentation. In some embodiments, cell fragmentation can be performed using the following methods, including, but not limited to: chemical methods, mechanical methods (e.g., centrifugation (e.g., ultracentrifugation, or density centrifugation), freeze-thaw, or sonication), or combinations thereof.

[0356] In some embodiments, a characteristic of a cell based particle is described by comparison to a reference cell. In embodiments, the reference cell is the source cell. In embodiments, the reference cell is a HeLa, HEK293, HFF-1, MRC-5, WI-38, IMR 90, IMR 91, PER.C6, HT-1080, or BJ cell. In some embodiments, a characteristic of a composition of cell based particles is described by comparison to a population of reference cells, e.g., a population of source cells, or a population of HeLa, HEK293, HFF- 1, MRC-5, WI-38, IMR 90, IMR 91, PER.C6, HT-1080, or BJ cells.B. Fusogen

[0357] In some embodiments, the particle has a fusogen. In some embodiments, the particle has more than one fusogen. In some embodiments, the fusogen contains a mammalian protein, such as any described in Section II-B-1. In some embodiments, the fusogen contains a viral protein, such as any described in Section II-B-2.

[0358] In some embodiments, the fusogen promotes mixing between lipids in the particle and lipids in the target cell. In some embodiments, the fusogen facilitates the fusion of the particle to a membrane of the target cell. In some embodiments, the particle integrates into the membrane of the target cell. In some embodiments, the membrane is the plasma membrane of the target cell. In some embodiments, the fusogen promotes formation of one or more pores between the interior of the particle and the cytosol of the target cell.

[0359] In some embodiments, the fusogen is endogenous to the particle. In some embodiments, the particle is pseudotyped with the fusogen, such as when the particle is a viral particle or virus-like particle.

[0360] In some embodiments, the fusogen is exposed on the surface of the particle. In some embodiments, the fusogen is exposed on the surface of the lipid bilayer of the particle. In some embodiments, a portion of the fusogen is embedded in the lipid bilayer of the particle.

[0361] In some embodiments, the particle is a viral particle or a virus-like particle. In some embodiments, the fusogen is exposed on the surface of the viral envelope of the particle. In some embodiments, a portion of the fusogen is embedded in the viral envelope of the particle.

[0362] In some embodiments, the fusogen is present at a copy number of at least, or no more than, 10, 50, 100, 500, 1,000, 2,000, 5,000, 10,000, 20,000, 50,000, 100,000, 200,000, 500,000, 1,000,000, 5,000,000, 10,000,000, 50,000,000, 100,000,000, 500,000,000, or 1,000,000,000 copies. In some embodiments, at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, or 99% of the fusogen comprised by the particle is disposed in the cell membrane. In embodiments, the particle also comprises fusogen internally, e.g., in the cytoplasm or an organelle. In some embodiments, the fusogen comprises (or is identified as comprising) about 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1%, 5%, 10%, 11%, 12%, 13%, 14%, 15%, 20%, or more, or about 1-30%, 5- 20%, 10- 15%, 12-15%, 13-14%, or 13.6% of the total protein in a particle, e.g., as determined by a mass spectrometry assay. In embodiments, the fusogen comprises (or is identified as comprising) about 13.6% of the total protein in the particle. In some embodiments, the fusogen is (or is identified as being) more or less abundant than one or more additional proteins of interest. In an embodiment, the fusogen has (or is identified as having) a ratio to EGFP of about 140, 145, 150, 151, 152, 153, 154, 155, 156, 157 (e.g., 156.9), 158, 159, 160, 165, or 170. In another embodiment, the fusogen has (or is identified as having) a ratio to CD63 of about 2700, 2800, 2900, 2910 (e.g., 2912), 2920, 2930, 2940, 2950, 2960, 2970, 2980, 2990, or 3000, or about 1000-5000, 2000-4000, 2500-3500, 2900-2930, 2910-2915, or 2912.0, e.g., by a mass spectrometry assay. In an embodiment, the fusogen has (or is identified as having) a ratio to ARRDC1 of about 600, 610, 620, 630, 640, 650, 660 (e.g., 664.9), 670, 680, 690, or 700. In another embodiment, the fusogen has (or is identified as having) a ratio to GAPDH of about 50, 55, 60, 65, 70 (e.g., 69), 75, 80, or 85, or about 1-30%, 5-20%, 10-15%, 12-15%, 13-14%, or 13.6%. In another embodiment, the fusogen has (or is identified as having) a ratio to CNX of about 500, 510, 520, 530, 540, 550, 560 (e.g., 558.4), 570, 580, 590, or 600, or about 300-800, 400-700, 500-600, 520-590, 530-580, 540-570, 550-560, or 558.4, e.g., by a mass spectrometry assay.

[0363] In some embodiments, the fusogen is a protein fusogen. In some embodiments, the fusogen is a native protein or a derivative of a native protein. In some embodiments, the fusogen is a synthetic protein. In some embodiments, the fusogen is a mammalian protein or a homologue of a mammalian protein (e.g., having 50%, 60%, 70%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or greater sequence identity to the mammalian protein), such as any described in Section II-B-1. In some embodiments, the fusogen is a non-mammalian protein. In some embodiments, the fusogen is a viral protein or a homologue of a viral protein (e.g., having 50%, 60%, 70%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or greater sequence identity to the viral protein), such as any described in Section II-B-2. In some embodiments, the fusogen is a fragment of any of the foregoing. In some embodiments, the fusogen is a variant of any of the foregoing. In some embodiments, the fusogen is a protein fusion containing one ormore proteins or fragments thereof. In some embodiments, the fusogen is a protein fusion containing one or more of any of the foregoing.

[0364] In some embodiments, the fusogen is mutated to reduce binding for the native binding partner of the fusogen. In some embodiments the fusogen is randomly mutated. In some embodiments the fusogen is rationally mutated. In some embodiments the fusogen is subjected to directed evolution. In some embodiments the fusogen is truncated and only a subset of the peptide is used in the particle.1. Mammalian Proteins

[0365] In some embodiments, the fusogen is or contains a mammalian protein. Exemplary mammalian fusogens include a SNARE family protein such as vSNAREs or tSNAREs, a syncytin protein such as Syncytin-1 (DOI: 10.1128 / JVI.76.13.6442-6452.2002) and Syncytin-2, myomaker (biorxiv.org / content / early / 2017 / 04 / 02 / 123158, doi.org / 10.1101 / 123158, doi: 10.1096 / fj.201600945R, doi: 10.1038 / nature 12343), myomixer (www.nature.com / nature / journal / v499 / n7458 / full / naturel2343.html, doi: 10.1038 / nature 12343), myomerger (science.sciencemag.org / content / early / 2017 / 04 / 05 / science.aam9361, DOI: 10.1126 / science.aam9361), FGFRL1 (fibroblast growth factor receptor-like 1), Minion (doi.org / 10.1101 / 122697), an isoform of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (e.g., as disclosed in US 6,099,857A), a gap junction protein such as connexin 43, connexin 40, connexin 45, connexin 32, or connexin 37 (e.g., as disclosed in US 2007 / 0224176), Hap2, any protein capable of inducing syncytium formation between heterologous cells (see Table 2), any protein with fusogenic properties, a homologue thereof, a fragment thereof, a variant thereof, and a protein fusion containing one or more proteins or fragments thereof, such as one or more of any of the foregoing. In some embodiments, the fusogen is encoded by a human endogenous retroviral element (hERV) found in the human genome. Additional exemplary fusogens are disclosed in US 6,099,857A and US 2007 / 0224176, the entire contents of each of which are hereby incorporated by reference.2. Viral Proteins

[0366] In some embodiments, the fusogen is or contains a non-mammalian protein. In some embodiments, the fusogen is or contains a viral protein. In some embodiments, the fusogen is a viral fusion protein. In some embodiments, the fusogen is a viral envelope protein.

[0367] In some embodiments, a viral fusogen is a Class I viral membrane fusion protein, a Class II viral membrane protein, a Class III viral membrane fusion protein, a viral membrane glycoprotein, or other viral fusion proteins, or a homologue thereof, a fragment thereof, a variant thereof, or a protein fusion containing one or more proteins or fragments thereof, such as one or more of any of the foregoing.

[0368] In some embodiments, Class I viral membrane fusion proteins include Baculovirus F protein, e.g., F proteins of the nucleopolyhedrovirus (NPV) genera, e.g., Spodoptera exigua MNPV (SeMNPV) F protein and Lymantria dispar MNPV (LdMNPV), and paramyxovirus F proteins.

[0369] In some embodiments, Class II viral membrane proteins include tick bone encephalitis E (TBEV E) and Semliki Forest Virus E1 / E2.

[0370] In some embodiments, Class III viral membrane fusion proteins include rhabdovirus G (e.g., fusogenic protein G of the Vesicular Stomatatis Virus (VSV-G), Cocal virus G protein), herpesvirus glycoprotein B (e.g., Herpes Simplex virus 1 (HSV-1) gB), Epstein Barr Virus glycoprotein B (EBV gB), thogotovirus G, baculovirus gp64 (e.g., Autographa California multiple NPV (AcMNPV) gp64), and Borna disease virus (BDV) glycoprotein (BDV G).

[0371] Examples of other viral fusogens, e.g., membrane glycoproteins and viral fusion proteins, include viral syncytia proteins such as influenza hemagglutinin (HA) or mutants, or fusion proteins thereof; human immunodeficiency virus type 1 envelope protein (HIV-1 ENV), gpl20 from HIV binding LFA-1 to form lymphocyte syncytium, HIV gp41, HIV gpl60, or HIV Trans- Activator of Transcription (TAT); viral glycoprotein VSV-G, viral glycoprotein from vesicular stomatitis virus of the Rhabdoviridae family; glycoproteins gB and gH-gL of the varicella-zoster virus (VZV); murine leukaemia virus (MLV)-lOAl; Gibbon Ape Leukemia Virus glycoprotein (GaLV); type G glycoproteins in Rabies, Mokola, vesicular stomatitis virus and Togaviruses; murine hepatitis virus JHM surface projection protein; porcine respiratory coronavirus spike- and membrane glycoproteins; avian infectious bronchitis spike glycoprotein and its precursor; bovine enteric coronavirus spike protein; the F and H, HN or G genes of a Morbillivirus (e.g., measles virus (MeV), canine distemper virus, Cetacean morbilli virus, Peste-des-petits-ruminants virus, Phocine distemper virus, Rinderpest virus), Newcastle disease virus, human parainfluenza virus 3, simian virus 41, Sendai virus and human respiratory syncytial virus; gH of human herpesvirus 1 and simian varicella virus, with the chaperone protein gL; human, bovine and cercopithicine herpesvirus gB; envelope glycoproteins of Friend murine leukaemia virus and Mason Pfizer monkey virus; mumps virus hemagglutinin neuraminidase, and glycoproteins Fl and F2; membrane glycoproteins from Venezuelan equine encephalomyelitis; paramyxovirus F protein; SIV gpl60 protein; Ebola virus G protein; or Sendai virus fusion protein, or a homologue thereof, a fragment thereof, a variant thereof, and a protein fusion containing one or more proteins or fragments thereof, such as one or more of any of the foregoing.

[0372] Non-mammalian fusogens include viral fusogens, homologues thereof, fragments thereof, and fusion proteins comprising one or more proteins or fragments thereof, such as one or more of any of the foregoing. Viral fusogens include class I fusogens, class II fusogens, class III fusogens, and class IV fusogens. In some embodiments, class I fusogens such as human immunodeficiency virus (HIV) gp41 have a characteristic postfusion conformation with a signature trimer of a-helical hairpins with a central coiled-coil structure. Class I viral fusion proteins include proteins having a central postfusion six-helixbundle. Class I viral fusion proteins include influenza HA, parainfluenza F, HIV Env, Ebola GP, hemagglutinins from orthomyxoviruses, F proteins from paramyxoviruses (e.g. Measles, (Katoh et al. BMC Biotechnology 2010, 10:37)), ENV proteins from retroviruses, and fusogens of filoviruses and coronaviruses. In some embodiments, class II viral fusogens such as dengue E glycoprotein, have a structural signature of P- sheets forming an elongated ectodomain that refolds to result in a trimer of hairpins. In some embodiments, the class II viral fusogen lacks the central coiled coil. Class II viral fusogen can be found in alphaviruses (e.g., El protein) and flaviviruses (e.g., E glycoproteins). Class II viral fusogens include fusogens from Semliki Forest virus, Sinbis, rubella virus, and dengue virus. In some embodiments, class III viral fusogens such as the vesicular stomatitis virus G glycoprotein, combine structural signatures found in classes I and II. In some embodiments, a class III viral fusogen comprises a helices (e.g., forming a six-helix bundle to fold back the protein as with class I viral fusogens), and sheets with an amphiphilic fusion peptide at its end, reminiscent of class II viral fusogens. Class III viral fusogens can be found in rhabdo viruses and herpesviruses. In some embodiments, class IV viral fusogens are fusion-associated small transmembrane (FAST) proteins (doi:10.1038 / sj.emboj.7600767, Nesbitt, Rae L., "Targeted Intracellular Therapeutic Delivery Using Liposomes Formulated with Multifunctional FAST proteins" (2012). Electronic Thesis and Dissertation Repository. Paper 388), which are encoded by nonenveloped reoviruses. In some embodiments, the class IV viral fusogens are sufficiently small that they do not form hairpins (doi: 10.1146 / annurev-cellbio- 101512-122422, doi:10.1016 / j.devcel.2007.12.008).

[0373] In some embodiments, the fusogen is any of the fusogenic moieties described in WO2017 / 182585; WO2022 / 164935; WO2021 / 076788; Hamilton et al. bioRxiv 2022.08.24.505004; Nikolic et al. Nat Commun 9, 1029 (2018); Dobson et al. Nat. Methods. 19, 449-460 (2022); and Yu et al. bioRxiv 2021.12.13.472464, for instance any of the VSV or variant VSV glycoproteins described therein, such as VSV glycoproteins that have reduced binding to native receptors. a. Baboon Endogenous Retrovirus

[0374] In some embodiments, the fusogen is a Baboon Endogenous Retrovirus (BaEV) envelope glycoprotein. Exemplary BaEV envelope glycoproteins and variants thereof are described in PCT / US2022 / 031459; US9249426; Aguila et al. Journal of Virology 2003 77(2):1281-1291; Bernadin et al. Blood Advances 2019 3(3):461-475; Colamartino et al. Frontiers in Immunology 2019 10:2873; Girard-Gagnepain et al. Blood 2014 124(8): 1221-1231; and Levy et al. Journal of Thrombosis and Haemostasis 2016 14:2478-2492.

[0375] Wild-type BaEV envelope glycoproteins are retroviral envelope proteins containing a C- terminal cytoplasmic tail (e.g., corresponding to amino acids 512-545 of SEQ ID NO:252), a transmembrane domain (e.g., corresponding to amino acids 489-511 of SEQ ID NO:252), and an extracellular domain (e.g., corresponding to amino acids 1-488 of SEQ ID NO:252). Maturation of theprecursor protein in the Golgi, which requires the minimal sequence [KR]-X-[KR]-R (wherein X is any amino acid), results in two subunits, the surface unit protein or gp70, and the transmembrane protein p20E. The surface unit protein or gp70 (e.g., corresponding to amino acids 1-358 of SEQ ID NO:252) and the transmembrane protein p20E (e.g., corresponding to amino acids 359-545 of SEQ ID NO:252) remain associated in a labile interaction that may include a disulfide bond. In wild-type BaEV envelope glycoproteins, fusogenicity is controlled by a short, 17 amino acid sequence termed a fusion inhibitory R peptide (e.g., set forth in SEQ ID NO:253), which is localized on the C-terminal of the cytoplasmic tail domain. The fusion inhibitory R peptide harbors the tyrosine endocytosis signal YXXL, and its cleavage by the viral protease is thought to potentiate fusogenic activation through molecular rearrangements in the membrane-spanning domain and the extracellular region of the envelope glycoprotein (Salamango et al (2015) Journal of virology 89(24): 12492-12500). In wild-type BaEV envelope glycoproteins, the gp70 mediates receptor binding to the ASCT-2 and ASCT-1 receptors on host cells. In some embodiments, the glycoprotein 70 (g70) subunit or a biologically active portion thereof binds the ASCT-2 and ASCT-1 receptors. In wild-type BaEV envelope glycoproteins, the p20E acts as a class I viral fusion protein. The interaction of the gp70 subunit with a host cell membrane triggers refolding of the p20E and is believed to activate the fusogenic potential by unmasking the fusion peptide.

[0376] In some embodiments, the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof.

[0377] In some embodiments, the fusogen is a truncated BaEV envelope glycoprotein. Exemplary BaEV envelope glycoproteins and truncates thereof are described in PCT / US2022 / 031459. In some embodiments, the truncated BaEV envelope glycoprotein comprises a cytoplasmic tail with a partial fusion inhibitory R peptide relative to a wild-type BaEV envelope glycoprotein, wherein the R peptide contains a contiguous portion of the inhibitory R peptide but lacks the full length R peptide of wild-type BaEV envelope glycoprotein. In some embodiments, the truncated BaEV envelope glycoprotein has a cytoplasmic tail that is composed of a partial inhibitory R peptide with at least one, at least two, or at least three contiguous amino-terminal amino acids of the inhibitory R peptide but less than the full-length R peptide relative to wild-type BaEV envelope glycoprotein. In some embodiments, the truncated BaEV envelope glycoprotein has a cytoplasmic tail that has a partial inhibitory R peptide composed of 1 to 16 contiguous amino-terminal amino acids of the inhibitory R peptide of the wild- type BaEV envelope glycoprotein, such as is composed of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 12, 14, 15 or 16 amino-terminal amino acids of the inhibitory R peptide of the wild-type BaEV envelope glycoprotein. In some embodiments, the truncated BaEV envelope glycoprotein is set forth in any of SEQ ID NO:254-260. In som embodiments, the truncated BaEV envelope glycoprotein s BaEV R+8 set fort hin SEQ ID NO:259.

[0378] In some embodiments, the fusogen is a modified BaEV envelope glycoprotein. In some embodiments, the cytoplasmic tail domain of the BaEV envelope glycoprotein is devoid of the fusion inhibitory R peptide. The expression “fusion inhibitory R peptide” refers to the C-terminal portion of thecytoplasmic tail domain of the envelope glycoprotein which harbours a tyrosine endocytosis signal — YXXL — and which is cleaved by viral protease during virion maturation, thus enhancing membrane fusion of the envelope glycoprotein. The fusion inhibitory R peptide of the BaEV envelope glycoprotein is typically located between amino acids 547 and 564 of the wild-type BaEV envelope glycoprotein. In some embodiments, the modified BaEV envelope glycoprotein is set forth in SEQ ID NO: 261 (BaEVRLess).

[0379] In some embodiments, the cytoplasmic tail domain of the BaEV envelope glycoprotein is replaced by the cytoplasmic tail domain of a murine leukemia virus (MLV) envelope glycoprotein. The Murine Leukemia Virus envelope glycoprotein is notably described in Ott et al. (1990) J. Virol. 64:757- 766. In some embodiments, the Murine Leukemia Virus envelope glycoprotein is that of strain 4070A. The term “MLV envelope glycoprotein” refers to the wild-type form of the MLV envelope glycoprotein or to a mutant of said wild-type MLV envelope glycoprotein which is at least 80%, preferably at least 85%, still preferably at least 90%, more preferably at least 95%, still more preferably at least 99% identical to said wild-type MLV envelope glycoprotein, provided that said mutant glycoprotein retains the capacity of the wild-type envelope glycoprotein of interacting with viral core proteins, in particular with lentiviral core proteins. Typically, the cytoplasmic tail domain of the MLV envelope glycoprotein is located between amino acids 622 and 654 of the wild-type MLV envelope glycoprotein. In some embodiments, the fusogen is BaEVTR. In some embodiments, the modified BaEV envelope glycoprotein is set forth in SEQ ID NO: 262 (BaEVTR).

[0380] In some embodiments, the fusogen is able to target the CD34+ progenitor subpopulations indicated in FIG. 7 for BaEVTR. In some embodiments, the fusogen is able to target HSCs, MPPs, CMPs, MLPs, or ETPs. In some embodiments, the fusogen is able to target HSCs, MPPs, CMPs, MLPs, and ETPs. In some embodiments, the target cells are HSCs, MPPs, CMPs, MLPs, or ETPs. In some embodiments, the target cells are MPPs, CMPs, MLPs, and ETPs.

[0381] In some embodiments where a VLP includes a BaEVTR glycoprotein fusogen (e.g., a fusogen comprising a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof or a truncated BaEV envelope glycoprotein or a functional variant thereof), the VLP also comprises a gag protein (e.g. an MLV-gag protein). In some embodiments, the MLV-gag protein is part of a fusion protein, where the the fusion protein is a cleavable fusion protein between (i) a viral structural protein (e.g. GAG (e.g. MLV-gag or HIV-gag)) and (ii) a nuclease protein (e.g. Cas protein (e.g. any of the Cas protein described herein). In some embodiments where the VLP includes a BaEVTR glycoprotein fusogen and a gag protein (e.g. an MLV-gag protein), production of such a VLP includes varying the ratio of polynucleotide encoding the BaEVTR glycoprotein fusogen and the polynucleotide encoding the gag protein. In some embodiments, the ratio of polynucleotide encoding the BaEVTR glycoprotein fusogen to the polynucleotide encoding the gag protein includes a 1:1, 1:2, 1:3, 1:4, or 1:5 ratio.b. G / H Proteins

[0382] In some embodiments, the fusogen is or contains a G or H protein. In some embodiments, the G or H protein is a Paramyxovirus (e.g., Morbillivirus or Henipavirus) G or H protein or a biologically active portion thereof. In some embodiments, the Henipavirus G protein is a Hendr a (HeV) virus G protein, a Nipah (NiV) virus G-protein (NiV-G), a Cedar (CedPV) virus G-protein, a Mojiang virus G- protein, a bat Paramyxovirus G-protein, a Kumasi virus G-protein, a Langya virus G-protein, or a biologically active portion thereof. A non-limited list of exemplary G proteins is shown in Table 1.

[0383] The Henipavirus attachment G proteins are type II transmembrane glycoproteins containing an N-terminal cytoplasmic tail (e.g., corresponding to amino acids 1-49 of SEQ ID NO:1), a transmembrane domain (e.g., corresponding to amino acids 50-70 of SEQ ID NO:1, and an extracellular domain containing an extracellular stalk (e.g., corresponding to amino acids 71-187 of SEQ ID NO:1), and a globular head (corresponding to amino acids 188-602 of SEQ ID NO:1). The N-terminal cytoplasmic domain is within the inner lumen of the lipid bilayer and the C-terminal portion is the extracellular domain that is exposed on the outside of the lipid bilayer. Regions of the stalk in the C- terminal region (e.g. corresponding to amino acids 159-167 of NiV-G) have been shown to be involved in interactions with F protein and triggering of F protein fusion (Liu et al. 2015 J of Virology 89:1838). In wild-type NiV-G protein, the globular head mediates receptor binding to henipavirus entry receptors ephrin B2 and ephrin B3, but is dispensable for membrane fusion (Brandel-Tretheway et al. Journal of Virology. 2019. 93(13)e00577-19).

[0384] In particular embodiments herein, tropism of the G protein is modified. Binding of the G protein to a binding partner can trigger fusion mediated by a compatible F protein or biologically active portion thereof. G protein sequences disclosed herein are predominantly disclosed as expressed sequences including an N-terminal methionine required for start of translation. As such N-terminal methionines are commonly cleaved co- or post-translationally, the mature protein sequences for all G protein sequences disclosed herein are also contemplated as lacking the N-terminal methionine.

[0385] G glycoproteins are highly conserved between henipavirus species. For example, the G protein of NiV and HeV viruses share 79% amino acids identity. Studies have shown a high degree of compatibility among G proteins with F proteins of different species as demonstrated by heterotypic fusion activation (Brandel-Tretheway et al. Journal of Virology. 2019). As described below, a retargeted lipid particle can contain heterologous proteins from different species.

[0386] In some embodiments, the G protein has a sequence set forth in any of SEQ ID NOs: 1-11 or is a functionally active variant or biologically active portion thereof that has a sequence that is at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at least at or about 84%, at least at or about 85%, at least at or about 86%, at least at or about 87%, at least at or about 88%, at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at least at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% identical to any one of SEQ ID NOs:l, 2, 3, 4, 5, 6, 7, 8, 9, 10 or 11. In some embodiments, the G protein has a sequence set forth in SEQ ID NO:1 or is a functionally active variant or biologically active portion thereof that has a sequence that is at least at or about 80%, at least at or about 90%, at least at or about 95%, or at least at or about 99% identical to SEQ ID NO:1. In some embodiments, the G protein has a sequence set forth inSEQ ID N0:4 or is a functionally active variant or biologically active portion thereof that has a sequence that is at least at or about 80%, at least at or about 90%, at least at or about 95%, or at least at or about 99% identical to SEQ ID NO:4. In some embodiments, the G protein has a sequence set forth in SEQ ID NO:5 or is a functionally active variant or biologically active portion thereof that has a sequence that is at least at or about 80%, at least at or about 90%, at least at or about 95%, or at least at or about 99% identical to SEQ ID NO:5.

[0387] In particular embodiments, the G protein or functionally active variant or biologically active portion is a protein that retains fusogenic activity in conjunction with a Henipa virus F protein, e.g. NiV-F or HeV-F. Fusogenic activity includes the activity of the G protein in conjunction with a Henipavirus F protein to promote or facilitate fusion of two membrane lumens, such as the lumen of the targeted lipid particle having embedded in its lipid bilayer a henipavirus F and G protein, and a cytoplasm of a target cell, e.g. a cell that contains a surface receptor or molecule that is recognized or bound by the targeted envelope protein. In some embodiments, the F protein and G protein are from the same Henipavirus species (e.g. NiV-G and NiV-F). In some embodiments, the F protein and G protein are from different Henipavirus species (e.g. NiV-G and HeV-F).

[0388] In particular embodiments, the G protein has the sequence of amino acids set forth in SEQ ID NO:1, SEQ ID NO:2, SEQ ID NOG, SEQ ID NO:4, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NO:9, SEQ ID NO: 10 or SEQ ID NO: 11 or is a functionally active variant thereof or a biologically active portion thereof that retains fusogenic activity. In some embodiments, the functionally active variant comprises an amino acid sequence having at least at or about 80%, at least at or about 85%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO: 1, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NO:9, SEQ ID NO: 10 or SEQ ID NO: 11 and retains fusogenic activity in conjunction with a Henipavirus F protein (e.g., NiV-F or HeV-F). In some embodiments, the biologically active portion has an amino acid sequence having at least at or about 80%, at least at or about 85%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO:1, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NO:9, SEQ ID NO: 10 or SEQ ID NO: 11 and retains fusogenic activity in conjunction with a Henipavirus F protein (e.g., NiV-F or HeV-F).

[0389] Reference to retaining fusogenic activity includes activity (in conjunction with a Henipavirus F protein) that is between at or about 10% and at or about 150% or more of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO: 1, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NO:9, SEQID NO: 10 or SEQ ID NO: 11 such as at least or at least about 10% of the level or degree of fusogenic activity of the corresponding wild-type G protein, such as at least or at least about 15% of the level or degree of fusogenic activity of the corresponding wild-type G protein, such as at least or at least about 20% of the level or degree of fusogenic activity of the corresponding wild-type G protein, such as at least or at least about 25% of the level or degree of fusogenic activity of the corresponding wild-type G protein, such as at least or at least about 30% of the level or degree of fusogenic activity of the corresponding wild-type G protein, such as at least or at least about 35% of the level or degree of fusogenic activity of the corresponding wild-type G protein, such as at least or at least about 40% of the level or degree of fusogenic activity of the corresponding wild-type G protein, such as at least or at least about 45% of the level or degree of fusogenic activity of the corresponding wild-type G protein, such as at least or at least about 50% of the level or degree of fusogenic activity of the corresponding wild-type G protein, such as at least or at least about 55% of the level or degree of fusogenic activity of the corresponding wild-type G protein, such as at least or at least about 60% of the level or degree of fusogenic activity of the corresponding wild-type G protein, such as at least or at least about 65% of the level or degree of fusogenic activity of the corresponding wild-type G protein, such as at least or at least about 70% of the level or degree of fusogenic activity of the corresponding wild-type G protein, such as at least or at least about 75% of the level or degree of fusogenic activity of the corresponding wild-type G protein, such as at least or at least about 80% of the level or degree of fusogenic activity of the corresponding wild-type G protein, such as at least or at least about 85% of the level or degree of fusogenic activity of the corresponding wild-type G protein, such as at least or at least about 90% of the level or degree of fusogenic activity of the corresponding wild-type G protein, such as at least or at least about 95% of the level or degree of fusogenic activity of the corresponding wild-type G protein, such as at least or at least about 100% of the level or degree of fusogenic activity of the corresponding wild-type G protein, or such as at least or at least about 120% of the level or degree of fusogenic activity of the corresponding wild-type G protein.1) Truncated Paramyxovirus G / H proteins

[0390] In some embodiments the G protein is a mutant G protein that is a functionally active variant or biologically active portion containing one or more amino acid mutations, such as one or more amino acid insertions, deletions, substitutions or truncations. In some embodiments, the mutations described herein relate to amino acid insertions, deletions, substitutions or truncations of amino acids compared to a reference G protein sequence. In some embodiments, the reference G protein sequence is the wild- type sequence of a G protein or a biologically active portion thereof. In some embodiments, the functionally active variant or the biologically active portion thereof is a mutant of a wild-type Hendra (HeV) virus G protein, a wild-type Nipah (NiV) virus G-protein (NiV-G), a wild-type Cedar (CedPV) virus G-protein, a wild-type Mojiang virus G-protein, a wild-type bat Paramyxovirus G-protein, a Kumasi virus G-protein,a Langya virus G-protein, or biologically active portion thereof. In some embodiments, the wild-type G protein has the sequence set forth in any one of SEQ ID NO:1, SEQ ID NO:2, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NO:7, SEQ ID NOG, SEQ ID NO:9, SEQ ID NO: 10 or SEQ ID NO: 11.

[0391] In some embodiments, the G protein is a mutant G protein that is a biologically active portion that is an N-terminally and / or C-terminally truncated fragment of a wild- type Hendra (HeV) virus G protein, a wild-type Nipah (NiV) virus G-protein (NiV-G), a wild-type Cedar (CedPV) virus G- protein, a wild-type Mojiang virus G-protein, a wild-type bat Paramyxovirus G-protein, a Kumasi virus G-protein, or a Langya virus G-protein. In particular embodiments, the truncation is an N-terminal truncation of all or a portion of the cytoplasmic domain. In some embodiments, the mutant G protein is a biologically active portion that is truncated and lacks up to 49 contiguous amino acid residues at or near the N-terminus of the wild-type G protein, such as a wild-type G protein set forth in any one of SEQ ID NO:1, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NOG, SEQ ID NO:7, SEQ ID NOG, SEQ ID NO:9, SEQ ID NO:10 or SEQ ID NO:11. In some embodiments, the mutant F protein is truncated and lacks up to 49 contiguous amino acids, such as up to 49, 48, 47, 46, 45, 44, 43, 42, 41, 40, 30, 38, 37, 36, 35, 34, 33, 32, 31, 30, 29, 28, 27, 26, 25, 24, 23, 22, 21, 20, 19, 18, 17, 16, 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2 or 1 contiguous amino acids at the N-terminus of the wild-type G protein.

[0392] In some embodiments, the G protein is a wild-type Nipah virus G (NiV-G) protein or a Hendra virus G protein, or is a functionally active variant or biologically active portion thereof. In some embodiments, the G protein is a NiV-G protein that has the sequence set forth in SEQ ID NO:1, SEQ ID NOG or SEQ ID NOG, or is a functional variant or a biologically active portion thereof that has an amino acid sequence having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at least at or about 84%, at least at or about 85%, at least at or about 86%, at least at or about 87%, at least at or about 88%, at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, at least at or about 99% sequence identity to SEQ ID NO:1, SEQ ID NOG or SEQ ID NOG. In some embodiments, the G protein is a NiV-G protein that has the sequence set forth in SEQ ID NO:1, or is a functional variant or a biologically active portion thereof that has an amino acid sequence having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at least at or about 84%, at least at or about 85%, at least at or about 86%, at least at or about 87%, at least at or about 88%, at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, at least at or about 99% sequence identity to SEQ ID NO:1. In some embodiments, the G protein is a NiV-G protein that has the sequence set forth in SEQ ID NO:1. In some embodiments, the G protein is a NiV-G protein that has the sequence set forth in SEQID N0:4, or is a functional variant or a biologically active portion thereof that has an amino acid sequence having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at least at or about 84%, at least at or about 85%, at least at or about 86%, at least at or about 87%, at least at or about 88%, at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, at least at or about 99% sequence identity to SEQ ID NO:4. In some embodiments, the G protein is a NiV-G protein that has the sequence set forth in SEQ ID NO:4. In some embodiments, the G protein is a NiV-G protein that has the sequence set forth in SEQ ID NO:5, or is a functional variant or a biologically active portion thereof that has an amino acid sequence having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at least at or about 84%, at least at or about 85%, at least at or about 86%, at least at or about 87%, at least at or about 88%, at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, at least at or about 99% sequence identity to SEQ ID NO:5. In some embodiments, the G protein is a NiV-G protein that has the sequence set forth in SEQ ID NO:5.

[0393] In some embodiments, the G protein is a mutant NiV-G protein that is a biologically active portion of a wild-type NiV-G. In some embodiments, the biologically active portion is an N- terminally truncated fragment. In some embodiments, the mutant NiV-G protein is truncated and lacks up to 5 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 6 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 7 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 8 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 9 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5) up to 10 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 11 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 12 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein SEQ ID NO:1, SEQ ID NO:4, or SEQ ID NO:5), up to 13 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 14 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 15 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5) up to 16 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 17 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 18 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 19 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 20 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 21 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 22 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 23 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 24 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 25 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 26 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 27 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 28 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 29 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 30 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 31 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 32 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 33 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 34 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 35 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 36 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 37 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5) up to 38 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 39 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 40 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 41 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 42 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 43 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), up to 44 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), or up to 45 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5).

[0394] In some embodiments, the mutant NiV-G protein is truncated and lacks 5 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5). In some embodiments, the mutant NiV-G protein comprises the amino acid sequence set forth in SEQ ID NO: 12. In some embodiments, the mutant NiV-G protein is truncated and lacks 10 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5). In some embodiments, the mutant NiV-G protein comprises the amino acid sequence set forth in SEQ ID NO:44. In some embodiments, the mutant NiV-G protein is truncated and lacks 15 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5). In some embodiments, the mutant NiV-G protein comprises the amino acid sequence set forth in SEQ ID NO:45. In some embodiments, the mutant NiV-G protein is truncated and lacks 20 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5). In some embodiments, the mutant NiV-G protein comprises the amino acid sequence set forth in SEQ ID NO: 13. In some embodiments, the mutant NiV-G protein is truncated and lacks 25 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5). In some embodiments, the mutant NiV-G protein comprises the amino acid sequence set forth in SEQ ID NO: 14. In some embodiments, the mutant NiV-G protein is truncated and lacks 30 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5). In some embodiments, the mutant NiV-G protein comprises the amino acid sequence set forth in SEQ ID NO:43. In some embodiments, the mutant NiV-G protein is truncated and lacks 34 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5). In some embodiments, the mutant NiV-G protein comprises the amino acid sequence set forth in SEQ ID NO:42.

[0395] In some embodiments, the NiV-G protein is a biologically active portion that does not contain a cytoplasmic domain. In some embodiments, the NiV-G protein without the cytoplasmic domain is encoded by SEQ ID NO:22.

[0396] In some embodiments, the mutant NiV-G protein comprises a sequence set forth in any of SEQ ID NOS: 12-14, 17, 18 and 22, or 42-45 or is a functional variant thereof that has an amino acid sequence having at least at or 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at or about 84%, at least at or about 85%, at least at or about 86%, or at least at or about 87%, at least at or about 88%, or at least at or about 89%, about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NOS: 12-14, 17, 18 and 22 or 42-45.

[0397] In some embodiments, the mutant NiV-G protein has a 5 amino acid truncation at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), such as set forth in SEQ ID NO: 12 or a functional variant thereof having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at or about 84%, at least at or about 85%, at least at or about 86%, or at least at or about 87%, at least at or about 88%, or at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO: 12 or such as set forth in SEQ ID NO: 17 or a functional variant thereof having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at or about 84%, at least at or about 85%, at least at or about 86%, or at least at or about 87%, at least at or about 88%, or at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO: 17. In some embodiments, the mutant NiV-G protein has a 10 amino acid truncation at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), such as set forth in SEQ ID NO:44 or a functional variant thereof having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at or about 84%, at least at or about 85%, at least at or about 86%, or at least at or about 87%, at least at or about 88%, or at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO:44. In some embodiments, the mutant NiV-G protein has a 20 amino acid truncation at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), such as set forth in SEQ ID NO: 13 or a functional variant thereof having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at or about 84%, at least at or about 85%, at least at or about 86%, or at least at or about 87%, at least at or about 88%, or at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO: 13. In some embodiments, the mutant NiV-G protein has a 25 amino acid truncation at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), such as set forth in SEQ ID NO: 14 or a functional variant thereof having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at or about 84%, at least at or about 85%, at least at or about 86%, or at least at or about 87%, at least at or about 88%, or at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO: 14. In some embodiments, the mutant NiV-G protein has a 33 amino acid truncation at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), such as set forth in SEQ ID NO: 17 or a functional variant thereof having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at or about 84%, at least at or about 85%, at least at or about 86%, or at least at or about 87%, at least at or about 88%, or at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO: 17. In some embodiments, the mutant NiV-G protein has a 34 amino acid truncation at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), such as set forth in SEQ ID NO: 18 or a functional variant thereof having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at or about 84%, at least at or about 85%, at least at or about 86%, or at least at or about 87%, at least at or about 88%, or at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO:18. In some embodiments, the mutant NiV-G protein has a 48 amino acid truncation at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4 or SEQ ID NO:5), such as set forth in SEQ ID NO:22 or a functional variant thereof having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at or about 84%, at least at or about 85%, at least at or about 86%, or at least at or about 87%, at least at or about 88%, or at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO:22.

[0398] In some embodiments, the mutant NiV-G protein has a 15 amino acid truncation at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4, or SEQ ID NO:5), such as set forth in SEQ ID NO:45 or a functional variant thereof that has an amino acid sequence having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about83%, at or about 84%, at least at or about 85%, at least at or about 86%, or at least at or about 87%, at least at or about 88%, or at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO:45.

[0399] In some embodiments, the mutant NiV-G protein has a 20 amino acid truncation at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4, or SEQ ID NO:5), such as set forth in SEQ ID NO: 13 or a functional variant thereof that has an amino acid sequence having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at or about 84%, at least at or about 85%, at least at or about 86%, or at least at or about 87%, at least at or about 88%, or at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO: 13.

[0400] In some embodiments, the mutant NiV-G protein has a 25 amino acid truncation at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4, or SEQ ID NO:5), such as set forth in SEQ ID NO: 14 or a functional variant thereof that has an amino acid sequence having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at or about 84%, at least at or about 85%, at least at or about 86%, or at least at or about 87%, at least at or about 88%, or at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO: 14.

[0401] In some embodiments, the mutant NiV-G protein has a 30 amino acid truncation at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4, or SEQ ID NO:5), such as set forth in SEQ ID NO:43 or a functional variant thereof that has an amino acid sequence having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at or about 84%, at least at or about 85%, at least at or about 86%, or at least at or about 87%, at least at or about 88%, or at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO:43.

[0402] In some embodiments, the mutant NiV-G protein has a 34 amino acid truncation at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:, SEQ ID NO:4, or SEQ ID NO:5), such as set forth in SEQ ID NO:42 or a functional variant thereof that has an amino acid sequence having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about83%, at or about 84%, at least at or about 85%, at least at or about 86%, or at least at or about 87%, at least at or about 88%, or at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO:42.

[0403] In some embodiments, the mutant NiV-G protein has a 48 amino acid truncation at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:1, SEQ ID NO:4, or SEQ ID NO:5), such as set forth in SEQ ID NO:22 or a functional variant thereof that has an amino acid sequence having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at or about 84%, at least at or about 85%, at least at or about 86%, or at least at or about 87%, at least at or about 88%, or at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO:22.

[0404] In some embodiments, the G protein is a mutant HeV-G protein that is a biologically active portion of a wild- type HeV-G. In some embodiments, the biologically active portion is an N-terminally truncated fragment.

[0405] In some embodiments, the G protein is a wild-type HeV-G protein that has the sequence set forth in SEQ ID NO:23 or 24, or is a functional variant or biologically active portion thereof that has an amino acid sequence having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at least at or about 84%, at or about 85%, at least at or about 86%, at least at or about 87%, at or about 88%, at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO:23or 24.

[0406] In some embodiments, the G protein is a mutant HeV-G protein that is a biologically active portion of a wild-type HeV-G (SEQ ID NO:23 or SEQ ID NO:24). In some embodiments, the biologically active portion is an N-terminally truncated fragment. In some embodiments, the mutant HeV-G protein is truncated and lacks up to 5 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 6 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 7 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24) or up to 8 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 9 contiguous amino acid residues at or near the N- terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 10 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 11contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 12 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 13 contiguous amino acid residues at or near the N- terminus of the wild-type HeV-G protein ( SEQ ID NO:23 or 24), up to 14 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 15 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 16 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 17 contiguous amino acid residues at or near the N- terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 18 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 19 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 20 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 21 contiguous amino acid residues at or near the N- terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 22 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 23 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein SEQ ID NO:23 or 24), up to 24 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 25 contiguous amino acid residues at or near the N- terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 26 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 27 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 28 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 29 contiguous amino acid residues at or near the N- terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 30 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 31 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 32 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 33 contiguous amino acid residues at or near the N- terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 34 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 35 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 36 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 37 contiguous amino acid residues at or near the N- terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 38 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 39 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ IDNO:23 or 24), up to 40 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 41 contiguous amino acid residues at or near the N- terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 42 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 43 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), up to 44 contiguous amino acid residues at or near the N-terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24), or up to 45 contiguous amino acid residues at or near the N- terminus of the wild-type HeV-G protein (SEQ ID NO:23 or 24).

[0407] In some embodiments, the HeV-G protein is a biologically active portion that does not contain a cytoplasmic domain. In some embodiments, the mutant HeV-G protein lacks the N-terminal cytoplasmic domain of the wild-type HeV-G protein (SEQ ID NO:23 or 24), such as set forth in SEQ ID NO:25 or a functional variant thereof having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at or about 84%, at least at or about 85%, at least at or about 86%, or at least at or about 87%, at least at or about 88%, or at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO:25. In some embodiments, the mutant HeV-G protein lacks the N-terminal cytoplasmic domain of the wild-type HeV-G protein (SEQ ID NO:23 or 24), such as set forth in SEQ ID NO:26 or a functional variant thereof having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at or about 84%, at least at or about 85%, at least at or about 86%, or at least at or about 87%, at least at or about 88%, or at least at or about 89%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO:26.

[0408] In some embodiments, the G protein or the functionally active variant or biologically active portion thereof binds to Ephrin B2 or Ephrin B3. In some aspects, the G protein has the sequence of amino acids set forth in any one of SEQ ID NO:24, SEQ ID NO:23, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10, or is a functionally active variant thereof or a biologically active portion thereof that is able to bind to Ephrin B2 or Ephrin B3. In some embodiments, the functionally active variant or biologically active portion has an amino acid sequence having at least at or about 80%, at least at or about 81%, at least at or about 82%, at least at or about 83%, at least at or about 84%, at least at or about 85%, at or about 86%, at least at or about 87%, at least at or about 88%, or at least at or about 89% , at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to any of SEQ ID NO:24,SEQ ID NO:23, SEQ ID N0:4, SEQ ID N0:6, SEQ ID N0:5, SEQ ID N0:8 or SEQ ID NO: 10, or a functionally active variant or biologically active portion thereof, and retains binding to Ephrin B2 or B3.

[0409] In some embodiments, the functionally active variant or biologically active portion has an amino acid sequence having at least about 80%, at least about 85%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10, or a functionally active variant or biologically active portion thereof, and retains binding to Ephrin B2 or B3. Reference to retaining binding to Ephrin B2 or B3 includes binding that is at least or at least about 5% of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10, or a functionally active variant or biologically active portion thereof, 10% of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10, or a functionally active variant or biologically active portion thereof, 15% of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10, or a functionally active variant or biologically active portion thereof, 20% of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10, or a functionally active variant or biologically active portion thereof, 25% of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10, or a functionally active variant or biologically active portion, 30% of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10, or a functionally active variant or biologically active portion thereof, 35% of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10, or a functionally active variant or biologically active portion thereof, 40% of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10, or a functionally active variant or biologically active portion thereof, 45% of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10, or a functionally active variant or biologically active portion thereof, 50% of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10, or afunctionally active variant or biologically active portion thereof, 55% of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10, or a functionally active variant or biologically active portion thereof, 60% of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10, or a functionally active variant or biologically active portion thereof, 65% of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10,, or a functionally active variant or biologically active portion thereof, 70% of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10 or a functionally active variant or biologically active portion thereof, such as at least or at least about 75% of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10, or a functionally active variant or biologically active portion thereof, such as at least or at least about 80% of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4, NO:4,SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO:10„ or a functionally active variant or biologically active portion thereof, such as at least or at least about 85% of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4,SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10, or a functionally active variant or biologically active portion thereof, such as at least or at least about 90% of the level or degree of binding of the corresponding wild-type G protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4,SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO: 10, or a functionally active variant or biologically active portion thereof, or such as at least or at least about 95% of the level or degree of binding of the corresponding wild-type protein, such as set forth in SEQ ID NO:27, SEQ ID NO:23, SEQ ID NO:4,SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:8 or SEQ ID NO:10„ or a functionally active variant or biologically active portion thereof. In some embodiments, the G protein is NiV-G or a functionally active variant or biologically active portion thereof and binds to Ephrin B2 or Ephrin B3. In some aspects, the NiV-G has the sequence of amino acids set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27, or is a functionally active variant thereof or a biologically active portion thereof that is able to bind to Ephrin B2 or Ephrin B3. In some embodiments, the functionally active variant or biologically active portion has an amino acid sequence having at least about 80%, at least about 85%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27 and retains binding to Ephrin B2 or B3. Exemplary biologically activeportions include N-terminally truncated variants lacking all or a portion of the cytoplasmic domain, e.g. 1 or more, such as 1 to 49 contiguous N-terminal amino acid residues. Reference to retaining binding to Ephrin B2 or B3 includes binding that is at least or at least about 5% of the level or degree of binding of the corresponding wild-type NiV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27, 10% of the level or degree of binding of the corresponding wild-type NiV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27, 15% of the level or degree of binding of the corresponding wildtype NiV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27, 20% of the level or degree of binding of the corresponding wild-type NiV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27, 25% of the level or degree of binding of the corresponding wild-type NiV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27, 30% of the level or degree of binding of the corresponding wild-type NiV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27, 35% of the level or degree of binding of the corresponding wild-type NiV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27, 40% of the level or degree of binding of the corresponding wild-type NiV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27, 45% of the level or degree of binding of the corresponding wild-type NiV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27 50% of the level or degree of binding of the corresponding wildtype NiV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27, 55% of the level or degree of binding of the corresponding wild-type NiV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27, 60% of the level or degree of binding of the corresponding wild-type NiV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27, 65% of the level or degree of binding of the corresponding wild-type NiV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27, 70% of the level or degree of binding of the corresponding wild-type NiV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27, such as at least or at least about 75% of the level or degree of binding of the corresponding wild-type NiV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27, such as at least or at least about 80% of the level or degree of binding of the corresponding wild-type NIV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27, such as at least or at least about 85% of the level or degree of binding of the corresponding wild-type NiV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27, such as at least or at least about 90% of the level or degree of binding of the corresponding wild-type NiV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27, or such as at least or at least about 95% of the level or degree of binding of the corresponding wild-type NiV-G, such as set forth in SEQ ID NO:4, SEQ ID NO:5 or SEQ ID NO:27.

[0410] In some embodiments, the G protein or the biologically thereof is a mutant G protein that exhibits reduced binding for the native binding partner of a wild-type G protein. In some embodiments, the mutant G protein or the biologically active portion thereof is a mutant of wild-type Niv-G and exhibits reduced binding to one or both of the native binding partners Ephrin B2 or Ephrin B3. In someembodiments, the mutant G-protein or the biologically active portion, such as a mutant NiV-G protein, exhibits reduced binding to the native binding partner. In some embodiments, the reduced binding to Ephrin B2 or Ephrin B3 is reduced by greater than at or about 5%, at or about 10%, at or about 15%, at or about 20%, at or about 25%, at or about 30%, at or about 40%, at or about 50%, at or about 60%, at or about 70%, at or about 80%, at or about 90%, or at or about 100%.

[0411] In some embodiments, the mutations described herein can improve transduction efficiency. In some embodiments, the mutations described herein allow for specific targeting of other desired cell types that are not Ephrin B2 or Ephrin B3. In some embodiments, the mutations described herein result in at least the partial inability to bind at least one natural receptor, such has reduce the binding to at least one of Ephrin B2 or Ephrin B3. In some embodiments, the mutations described herein interfere with natural receptor recognition.

[0412] In some embodiments, the G protein is HeV-G or a functionally active variant or biologically active portion thereof and binds to Ephrin B2 or Ephrin B3. In some aspects, the HeV- G has the sequence of amino acids set forth in SEQ ID NO:23 or 24, or is a functionally active variant thereof or a biologically active portion thereof that is able to bind to Ephrin B2 or Ephrin B3. In some embodiments, the functionally active variant or biologically active portion has an amino acid sequence having at least about 80%, at least about 85%, at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO:23 or 24 and retains binding to Ephrin B2 or B3. Exemplary biologically active portions include N- terminally truncated variants lacking all or a portion of the cytoplasmic domain, e.g. 1 or more, such as 1 to 49 contiguous N-terminal amino acid residues. Reference to retaining binding to Ephrin B2 or B3 includes binding that is at least or at least about 5% of the level or degree of binding of the corresponding wild-type HeV-G, such as set forth in SEQ ID NO:23 or 24, 10% of the level or degree of binding of the corresponding wild-type HeV-G, such as set forth in SEQ ID NO:23 or 24, 15% of the level or degree of binding of the corresponding wild-type HeV-G, such as set forth in SEQ ID NO:23 or 24, 20% of the level or degree of binding of the corresponding wild-type HeV-G, such as set forth in SEQ ID NO:23 or 24, 25% of the level or degree of binding of the corresponding wild-type HeV-G, such as set forth in SEQ ID NO:23 or 24, 30% of the level or degree of binding of the corresponding wild-type HeV-G, such as set forth in SEQ ID NO:23 or 24, 35% of the level or degree of binding of the corresponding wild-type HeV-G, such as set forth in SEQ ID NO:23 or 24, 40% of the level or degree of binding of the corresponding wild-type HeV-G, such as set forth in SEQ ID NO:23 or 24, 45% of the level or degree of binding of the corresponding wild-type HeV-G, such as set forth in SEQ ID NO:23 or 24, 50% of the level or degree of binding of the corresponding wild-type HeV-G, such as set forth in SEQ ID NO:23 or 24, 55% of the level or degree of binding of the corresponding wild-type HeV-G, such as set forth in SEQ ID NO:23 or 24, 60% of the level or degree of binding of the corresponding wild-type HeV-G, suchas set forth in SEQ ID NO:23 or 24, 65% of the level or degree of binding of the corresponding wild-type HeV-G, such as set forth in SEQ ID NO:23 or 24, 70% of the level or degree of binding of the corresponding wild-type HeV-G, such as set forth in SEQ ID NO:23 or 24, such as at least or at least about 75% of the level or degree of binding of the corresponding wild-type HeV-G, such as set forth in SEQ ID NO:23 or 24, such as at least or at least about 80% of the level or degree of binding of the corresponding wild-type NIV-G, such as set forth in SEQ ID NO:23 or 24, such as at least or at least about 85% of the level or degree of binding of the corresponding wild-type HeV-G, such as set forth in SEQ ID NO:23 or 24, such as at least or at least about 90% of the level or degree of binding of the corresponding wild-type HeV-G, such as set forth in SEQ ID NO:23 or 24, or such as at least or at least about 95% of the level or degree of binding of the corresponding wild-type HeV-G, such as set forth in SEQ ID NO:23 or 24.

[0413] In some embodiments, the G protein or the biologically thereof is a mutant G protein that exhibits reduced binding for the native binding partner of a wild-type G protein. In some embodiments, the mutant G protein or the biologically active portion thereof is a mutant of wildtype Niv-G and exhibits reduced binding to one or both of the native binding partners Ephrin B2 or Ephrin B3. In some embodiments, the mutant G-protein or the biologically active portion, such as a mutant NiV-G protein, exhibits reduced binding to the native binding partner. In some embodiments, the reduced binding to Ephrin B2 or Ephrin B3 is reduced by greater than at or about 5%, at or about 10%, at or about 15%, at or about 20%, at or about 25%, at or about 30%, at or about 40%, at or about 50%, at or about 60%, at or about 70%, at or about 80%, at or about 90%, or at or about 100%.

[0414] In some embodiments, the G protein contains one or more amino acid substitutions in a residue that is involved in the interaction with one or both of Ephrin B2 and Ephrin B3. In some embodiments, the amino acid substitutions correspond to mutations E501 A, W504A, Q530A and E533A with reference to numbering set forth in SEQ ID NO:4.

[0415] In some embodiments, the G protein is a mutant G protein. In some embodiments, the G protein is a mutant G protein containing one or more amino acid substitutions selected from the group consisting of E501A, W504A, Q530A and E533A with reference to numbering set forth in SEQ ID NO:4. In some embodiments, the G protein is a mutant G protein that contains one or more amino acid substitutions elected from the group consisting of E501A, W504A, Q530A and E533A with reference to SEQ ID NO:4 and is a biologically active portion thereof containing an N-terminal truncation. In some embodiments, the mutant NiV-G protein or the biologically active portion thereof is truncated and lacks up to 5 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), 6 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:4), 7 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), 8 contiguous amino acidresidues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), 9 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), up to 10 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), 11 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), 12 contiguous amino acid residues at or near the N-terminus of the wildtype NiV-G protein (SEQ ID NO:4), 13 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), 14 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:4), up to 15 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), 16 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), 17 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), 18 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), 19 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), up to 20 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), 21 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4) 22 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:4), 23 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), 24 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), up to 25 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), 26 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), 27 contiguous amino acid residues at or near the N-terminus of the wild-type NiV- G protein (SEQ ID NO:4), 28 contiguous amino acid residues at or near the N-terminus of the wildtype NiV-G protein (SEQ ID NO:4), 29 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), up to 30 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO: 4), up to 31 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), 32 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), 33 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4) 34 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), 35 contiguous amino acid residues at or near the N-terminus of the wild-type NiV- G protein (SEQ ID NO:4) up to 36 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), up to 37 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein (SEQ ID NO:4), up to 38 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO: 4), up to 39 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO:4), or up to 40contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein (SEQ ID NO: 4).

[0416] In some embodiments, the mutant NiV-G protein has the amino acid sequence set forth in SEQ ID NO: 17 or 18 or an amino acid sequence having at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO: 17 or 18. In particular embodiments, the G protein has the sequence of amino acids set forth in SEQ ID NO: 17 or 18. In some embodiments, the mutant NiV-G protein has the amino acid sequence set forth in SEQ ID NO: 17 or an amino acid sequence having at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO: 17. In particular embodiments, the G protein has the sequence of amino acids set forth in SEQ ID NO 17. In some embodiments, the mutant NiV-G protein has the amino acid sequence set forth in SEQ ID NO: 18 or an amino acid sequence having at least at or about 90%, at least at or about 91%, at least at or about 92%, at least at or about 93%, at least at or about 94%, at least at or about 95%, at or about 96%, at least at or about 97%, at least at or about 98%, or at least at or about 99% sequence identity to SEQ ID NO: 18. In particular embodiments, the G protein has the sequence of amino acids set forth in SEQ ID NO 18.

[0417] In some embodiments, the G protein is a mutant G protein containing one or more amino acid substitutions selected from the group consisting of E501A, W504A, Q530A and E533A with reference to numbering set forth in SEQ ID NO:4. In some embodiments, the G protein is a mutant G protein that contains one or more amino acid substitutions elected from the group consisting of E501 A, W504A, Q530A and E533A with reference to SEQ ID NO:4 and is a biologically active portion thereof containing an N-terminal truncation.2) Mutated Paramyxovirus G / H Proteins

[0418] Provided herein are mutant Paramyxovirus G / H glycoproteins (e.g., variant Paramyxovirus G / H glycoproteins) comprising one or more amino acid mutations that result in decreased glycosylation of the protein. The one or more amino acid mutations, also called deglycosylation mutations, can be one or more amino acid substitutions (also referred to as mutations).

[0419] In some embodiments, the mutant Paramyxovirus G / H glycoprotein comprises an amino acid substitution at one or more amino acid positions that reduce glycosylation of the G / H glycoprotein. In some embodiments, the one or more amino acid substitutions disrupts an N-linked glycosylation site. In some embodiments, the one or more amino acid substitutions disrupts an O-linked glycosylation site.

[0420] In some embodiments, the mutant Paramyxovirus G / H glycoprotein is derived from Morbillivirus (e.g., measles virus (MeV), canine distemper virus, Cetacean morbilli virus, Peste-des-petits-ruminants virus, Phocine distemper virus, Rinderpest virus), Henipavirus (e.g., Hendra (HeV) virus, Nipah (NiV) virus, a Cedar (CedPV) virus, Mojiang virus, a Langya virus or bat Paramyxovirus). In some embodiments, the mutant Paramyxovirus G / H glycoprotein is a mutant of a Paramyxovirus G / H glycoprotein derived from Nipah virus or Measles virus. In some embodiments, the mutant Paramyxovirus G / H protein is selected from the group consisting of SEQ ID NOs: 2-11, 375, and 376, or a modified Paramyxovirus G / H glycoprotein derived from any one of 2-11, 375, and 376 containing an altered cytoplasmic tail . In some embodiments, the mutant Paramyxovirus G / H protein has a sequence of amino acids that has at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94% or at least 95% to any one of SEQ ID NOs: 2-11, 375, and 376 and contains the acid substitution at one or more amino acid positions that reduce glycosylation of the G / H glycoprotein as provided herein.

[0421] The location of precited glycosylation sites can be determined using the sequence of a protein. For example, N-glycosylation often occurs at sites with the sequence N-X-S / T in which “X” is any amino acid except P. Various algorithms and tools are available for prediction of both N- and O- linked glycosylation, including SprintGly (http: / / sparks-lab.org / server / sprint-gly / ), NetNGlyc (https: / / services.healtlttech.dtu.dk / service.php7NetNGlyc-l.0), NetOGlyc (https: / / services.healtlttech.dtu.dk / service.php?NetOGlyc-4.0), and GlycoMinestruct(bttp: / / glycomine. erc.monash.edu / Lab / GlycoMi e.. Struct / ), and methods described in Pitti et al., Sci. Reports, 9:15975 (2019) and Pakhrin et al., Molecules 26:7314 (2021). Any predicted glycosylation site may be substituted as described herein.

[0422] In some embodiments, the Paramyxovirus G / H glycoprotein to which the deglycosylation mutation is made is a NiV-G set forth in SEQ ID NO: 4 or a modified Nipah G glycoprotein (NiV-G) that has an altered cytoplasmic tail compared to native NiV-G (e.g., SEQ ID NO: 4). In some embodiments, the variant Paramyxovirus G / H protein has a sequence of amino acids that has at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94% or at least 95% to SEQ ID NO: 4 and contains the acid substitution at one or more amino acid positions that reduce glycosylation of the G / H glycoprotein as provided herein Exemplary modified NiV- G proteins with altered cytoplasmic tails to which the one or more amino acid substitutions for reducing glycosylation can be incorporated are described in Section II.B.

[0423] Amino acid positions for substitutions are described herein with positions “corresponding to” positions of a reference sequence. It is understood that the amino acid substitutions are not limited to being made in only the reference sequence but also can be made in similar sequences by identification of residues that align or correspond with the reference positions. For instance, positions “corresponding to” to positions of a protein in a reference sequence can be identified upon alignment of a similar sequence with the referenced sequence based on structural sequence alignment or using a standard alignment algorithm, such as the GAP algorithm. By aligning the sequences, one skilled in the art can identifycorresponding residues, for example, using conserved and identical amino acid residues as guides. For instance, amino acid positions for mutations are described herein with reference to the exemplary truncated NiV-G sequence set forth in SEQ ID NO:42; however, similar amino acid positions for mutations as described can be made in other modified NiV-G sequences, such as any as described in Section II.B, by sequence alignment and identification of the corresponding residues.

[0424] In some embodiments, the one or more amino acid mutations are at positions corresponding to positions 39, 126, 128, 273, 345, 384, 448, and 496 of SEQ ID NO:42. In some embodiments, the variant Paramyxovirus G / H glycoprotein comprises an amino acid mutation at any one of positions 39, 126, 128, 273, 345, 384, 448, and 496 of SEQ ID NO:42. In some embodiments, the variant Paramyxovirus G / H glycoprotein comprises two or more amino acid mutations at any of positions corresponding to positions 39, 126, 128, 273, 345, 384, 448, and 496 of SEQ ID NO:42., such as mutations at 2, 3, 4, 5, 7, or 8 of the positions.

[0425] In some embodiments, the one or more amino acid mutations is at a position corresponding to position 39 of SEQ ID NO:42. In some embodiments, the one or more amino acid mutations is at a position corresponding to position 126 of SEQ ID NO:42. In some embodiments, the one or more amino acid mutations is at a position corresponding to position 128 of SEQ ID NO:42. In some embodiments, the one or more amino acid mutations is at a position corresponding to position 273 of SEQ ID NO: 42. In some embodiments, the one or more amino acid mutations is at a position corresponding to position 345 of SEQ ID NO:42. In some embodiments, the one or more amino acid mutations is at a position corresponding to position 384 of SEQ ID NO:42. In some embodiments, the one or more amino acid mutations is at a position corresponding to position 448 of SEQ ID NO:42. In some embodiments, the one or more amino acid mutations is at a position corresponding to position 496 of SEQ ID NO:42.

[0426] In some embodiments, the native amino acid at the position comprising the amino acid mutation is asparagine or serine. In some embodiments, the amino acid mutation is an amino acid substitution. In some embodiments, the mutation is an asparagine to glutamine substitution. In some embodiments, the mutation is a serine to alanine substitution.

[0427] In some embodiments, the mutation is an asparagine to glutamine substitution at a position corresponding to position 39 (N39Q) of SEQ ID NO:42. In some embodiments, the mutation is an asparagine to glutamine substitution at a position corresponding to position 126 (N126Q) of SEQ ID NO: 42. In some embodiments, the mutation is an asparagine to glutamine substitution at a position corresponding to position 273 (N273Q) of SEQ ID NO: 42. In some embodiments, the mutation is an asparagine to glutamine substitution at a position corresponding to position 345 (N345Q) of SEQ ID NO: 42. In some embodiments, the mutation is an asparagine to glutamine substitution at a position corresponding to position 384 (N384Q) of SEQ ID NO: 42. In some embodiments, the mutation is an asparagine to glutamine substitution at a position corresponding to position 448 (N448Q) of SEQ ID NO:42. In some embodiments, the mutation is an asparagine to glutamine substitution at a position corresponding to position 496 (N496Q) of SEQ ID NO: 42.

[0428] In some embodiments, the mutation is a serine to alanine substitution at a position corresponding to position 128 (S128A) of SEQ ID NO: 42.

[0429] In some embodiments, the G / H glycoprotein is derived from Nipah virus G protein and the one or more amino acid substitutions are at positions corresponding to positions selected from the group consisting of 39, 126, 128, 273, 345, 384, 448, and 496 of SEQ ID NO: 42. In some embodiments, the one or more amino acid substitutions are selected from N39Q, N126Q, S128A, N273Q, N345Q, N384Q, N448Q, N496Q or any combination thereof. In some embodiments, the G / H glycoprotein is a mutant NiV-G containing one amino acid substitution from any one of N39Q, N126Q, S128A, N273Q, N345Q, N384Q, N448Q, N496Q. In some embodiments, the G / H glycoprotein is a mutant NiV-G containing two amino acid substitutions from any two of N39Q, N126Q, S128A, N273Q, N345Q, N384Q, N448Q, N496Q. In some embodiments, the G / H glycoprotein is a mutant NiV-G containing three amino acid substitutions from any three of N39Q, N126Q, S128A, N273Q, N345Q, N384Q, N448Q, N496Q. In some embodiments, the G / H glycoprotein is a mutant NiV-G containing four amino acid substitutions from any one of N39Q, N126Q, S128A, N273Q, N345Q, N384Q, N448Q, N496Q. In some embodiments, the G / H glycoprotein is a mutant NiV-G containing five amino acid substitutions from any one of N39Q, N126Q, S128A, N273Q, N345Q, N384Q, N448Q, N496Q. In some embodiments, the G / H glycoprotein is a mutant NiV-G containing six amino acid substitutions from any one of N39Q, N126Q, S128A, N273Q, N345Q, N384Q, N448Q, N496Q. In some embodiments, the G / H glycoprotein is a mutant NiV-G containing seven amino acid substitutions from any one of N39Q, N126Q, S128A, N273Q, N345Q, N384Q, N448Q, N496Q. In some embodiments, the G / H glycoprotein is a mutant NiV-G containing eight amino acid substitutions from any one of N39Q, N126Q, S128A, N273Q, N345Q, N384Q, N448Q, N496Q. In some embodiments, the one or more amino acid substitutions are in the SEQ ID NO: 147 or a or a modified Nipah G glycoprotein (NiV-G) that has an altered cytoplasmic tail compared to native NiV-G (e.g., SEQ ID NO:42). In some embodiments, the amino acid substitutions are in a modified NiV-G protein described in Section II.B. In some embodiments, the amino acid substitutions are in the NiV-G set forth in SEQ ID NO:42.

[0430] In some embodiments, the variant Nipah-G protein comprises at least three amino acid substitutions. In some embodiments, the amino acid substitutions are at positions 273, 384, and 496 of SEQ ID NO:42. In some embodiments, the amino acid substitutions are at positions 273, 345, and 496 of SEQ ID NO:42. In some embodiments, the amino acid substitutions are at positions 39, 126, and 128 of SEQ ID NO:42. In some embodiments, the amino acid substitutions are at positions 39, 273, and 345 of SEQ ID NO:42. In some embodiments, the amino acid substitutions are at positions 39, 384, and 448 of SEQ ID NO:42. In some embodiments, the amino acid substitutions are at positions 39, 448, and 496 of SEQ ID NO:42. In some embodiments, the amino acid substitutions are at positions 39, 128, and 273 ofSEQ ID NO:42. In some embodiments, the amino acid substitutions are at positions 39, 345, and 384 of SEQ ID NO:42. In some embodiments, the amino acid substitutions are at positions 39, 384, and 448 of SEQ ID NO:42.

[0431] In some embodiments, the variant Nipah-G protein comprises at least two amino acid substitutions. In some embodiments, the amino acid substitutions are at positions 273, and 496 of SEQ ID NO:42. In some embodiments, the amino acid substitutions are at positions 345, and 496 of SEQ ID NO:42. In some embodiments, the amino acid substitutions are at positions 39 and 128 of SEQ IDNO:42. In some embodiments, the amino acid substitutions are at positions 39, and 345 of SEQ IDNO:42. In some embodiments, the amino acid substitutions are at positions 39, and 448 of SEQ IDNO:42. In some embodiments, the amino acid substitutions are at positions 39 and 496 of SEQ IDNO:42. In some embodiments, the amino acid substitutions are at positions 39 and 273 of SEQ IDNO:42. In some embodiments, the amino acid substitutions are at positions 39 and 384 of SEQ IDNO:42. In some embodiments, the amino acid substitutions are at positions 384 and 448 of SEQ ID NO:42.

[0432] In some embodiments, the amino acid substitution is at position 39 of SEQ ID NO:42. In some embodiments, the amino acid substitution is at position 126 of SEQ ID NO:42. In some embodiments, the amino acid substitution is at position 128 of SEQ ID NO:42. In some embodiments, the amino acid substitution is at position 273 of SEQ ID NO:42. In some embodiments, the amino acid substitution is at position 345 of SEQ ID NO:42. In some embodiments, the amino acid substitution is at position 384 of SEQ ID NO:42. In some embodiments, the amino acid substitution is at position 448 of SEQ ID NO:42. In some embodiments, the amino acid substitution is at position 496 of SEQ ID NO:42.

[0433] In some embodiments, the mutant Nipah-G protein comprises an asparagine to glutamine substitution at position 39 of SEQ ID NO:42. In some embodiments, the mutant Nipah-G protein comprises an asparagine to glutamine substitution at position 126 of SEQ ID NO:42. In some embodiments, the mutant Nipah-G protein comprises an asparagine to glutamine substitution at position 273 of SEQ ID NO:42. In some embodiments, the mutant Nipah-G protein comprises an asparagine to glutamine substitution at position 345 of SEQ ID NO:42. In some embodiments, the mutant Nipah-G protein comprises an asparagine to glutamine substitution at position 384 of SEQ ID NO:42. In some embodiments, the mutant Nipah-G protein comprises an asparagine to glutamine substitution at position 448 of SEQ ID NO:42. In some embodiments, the mutant Nipah-G protein comprises an asparagine to glutamine substitution at position 496 of SEQ ID NO:42. In some embodiments, the mutant Nipah-G protein comprises a serine to alanine substitution at position 128 of SEQ ID NO:42.

[0434] In some embodiments, the mutant Nipah-G protein comprises the sequence selected from the group consisting of any one of SEQ ID NOs: 377-503, such as any exemplary mutant Nipah-G proteins set forth in Table 2 A below. In some embodiments, the mutant Nipah-G protein comprises the sequence of SEQ ID NO: 379. In some embodiments, the variant Nipah-G protein comprises the sequence of SEQID NO: 400. In some embodiments, the variant Nipah-G protein comprises the sequence of SEQ ID NO: 404.

[0435] In some embodiments, the Paramyxovirus G / H glycoprotein to which the deglycosylation mutations is made is a Measles virus H (Mev-H) protein or a modified MeV-H protein that has an altered cytoplasmic tail compared to native MeV-H (e.g., SEQ ID NO:375). In some embodiments, the mutant Paramyxovirus G / H protein has a sequence of amino acids that has at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94% or at least 95% to SEQ ID NO: 375 and contains the acid substitution at one or more amino acid positions that reduce glycosylation of the G / H glycoprotein as provided herein.

[0436] In some embodiments, the G / H glycoprotein is derived from Measles virus H (Mev-H) protein and the one or more amino acid substitutions are at positions corresponding to positions selected from the group consisting of 168, 187, 200, 215, 238 of SEQ ID NO: 375. In some embodiments, the mutant Mev-H protein comprises at least two amino acid substitutions, such as 2, 3, 4, or 5 substitutions at positions 168, 187, 200, 215, 238 of SEQ ID NO: 375.

[0437] In some embodiments, the Paramyxovirus G / H glycoprotein to which the deglycosylation mutations is made is a Canine distemper virus H (CDV-H) protein or a modified CDV-H protein that has an altered cytoplasmic tail compared to native CDV-H (e.g., SEQ ID NO:376). In some embodiments, the mutant Paramyxovirus G / H protein has a sequence of amino acids that has at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94% or at least 95% to SEQ ID NO: 376 and contains the acid substitution at one or more amino acid positions that reduce glycosylation of the G / H glycoprotein as provided herein.

[0438] In some embodiments, the G / H glycoprotein is derived from Canine distemper virus H (CDV-H) protein and the one or more amino acid substitutions are at positions corresponding to positions selected from the group consisting of 19, 149, 422 of SEQ ID NO: 376. In some embodiments, the variant CDV-H protein comprises at least two amino acid substitutions, such as 2 or 3 substitutions at positions 19, 149, 422 of SEQ ID NO: 376. c. F Proteins

[0439] In some embodiments, the fusogen contains a protein with a hydrophobic fusion peptide domain. In some embodiments, the fusogen is or contains a F protein. In some embodiments, the fusogen contains a henipavirus F protein molecule or biologically active portion thereof. In some embodiments, the Henipavirus F protein is a Hendra (Hev) virus F protein, a Nipah (NiV) virus F-protein, a Cedar (CedPV) virus F protein, a Mojiang virus F protein, a bat Paramyxovirus F protein, a Kumasi virus F protein, a Eangya virus F protein, or a biologically active portion thereof.

[0440] Table 2 provides non-limiting examples of F proteins. In some embodiments, the N- terminal hydrophobic fusion peptide domain of the F protein molecule or biologically active portion thereof is exposed on the outside of lipid bilayer.

[0441] F proteins of henipaviruses are encoded as Fo precursors containing a signal peptide (e.g. corresponding to amino acid residues 1-26 of SEQ ID NO:28). Following cleavage of the signal peptide, the mature Fo (e.g. SEQ ID NO:29) is transported to the cell surface, then endocytosed and cleaved by cathepsin L into the mature fusogenic subunits Fl and F2. In some embodiments, the signal peptide comprises the amino acid sequence set forth in SEQ ID NO: 38. In some embodiments, the Fo comprises the amino acid sequence of SEQ ID NO:41. In some embodiments, the Fl subunit comprises the sequence amino acid sequence set forth in SEQ ID NO:46. In some embodiments, the F2 subunit comprises the sequence amino acid sequence set forth in SEQ ID NO:39. The Fl and F2 subunits are associated by a disulfide bond and recycled back to the cell surface. The Fl subunit contains the fusion peptide domain located at the N terminus of the Fl subunit, where it is able to insert into a cell membrane to drive fusion. In some aspects, fusion is blocked by association of the F protein with G protein, until the G protein engages with a target molecule resulting in its disassociation from F and exposure of the fusion peptide to mediate membrane fusion.

[0442] Among different henipavirus species, the sequence and activity of the F protein is highly conserved. For examples, the F protein of NiV and HeV viruses share 89% amino acid sequence identity. Further, in some cases, the henipavirus F proteins exhibit compatibility with G proteins from other species to trigger fusion (Brandel-Tretheway et al. Journal of Virology. 2019. 93(13):e00577-19). In some aspects or the provided re-targeted lipid particles, the F protein is heterologous to the G protein, i.e. the F and G pro...

Claims

ClaimsWhat is claimed is:

1. A targeted lipid particle, comprising a lipid bilayer, a fusogen, and a targeting agent that binds to a target molecule selected from the group consisting of ASCT1, ASCT2, CD105, CD110, CD117, CD133, CD146, CD164, CD34, CD46, CD49f, CD90, EPCR, and ITGA3, wherein the fusogen and the targeting agent are exposed on the surface of the lipid bilayer.

2. The particle of claim 1, wherein the targeting agent is fused to a transmembrane domain incorporated into the lipid bilayer.

3. The particle of claim 1, wherein the targeting agent is fused to the fusogen.

4. The particle of any of claims 1-3, wherein the particle is a viral particle or virus-like particle.

5. The particle of any of claims 1-4, wherein the lipid bilayer is derived from a membrane of a producer cell used for producing a viral particle or virus-like particle.

6. The particle of claim 4 or claim 5, wherein the viral particle or virus-like particle is a retroviral particle or retrovirus-like particle.

7. The particle of any of claims 4-6, wherein the viral particle or virus-like particle is a lentivirus-like particle.

8. The particle of any of claims 4-6, wherein the viral particle or virus-like particle is a lentiviral particle.

9. A targeted lentiviral particle, comprising a fusogen and a targeting agent that binds to a target molecule selected from the group consisting of ASCT1, ASCT2, CD105, CD110, CD117, CD133, CD146, CD164, CD34, CD46, CD49f, CD90, EPCR, and ITGA3, wherein the fusogen and the targeting agent are exposed on the surface of the lentiviral particle.

10. The particle of any of claims 4-9, wherein the particle is pseudotyped with the fusogen.

11. The particle of claim 9 or claim 10, wherein the targeting agent is fused to the fusogen.

12. The particle of any of claims 1 and 3-11, wherein the targeting agent is fused to the fusogen directly.

13. The particle of any of claims 1 and 3-11, wherein the targeting agent is fused to the fusogen via a linker.

14. The particle of any of claims 4-10, wherein the targeting agent is fused to a transmembrane domain incorporated into the viral envelope.

15. The particle of any of claims 1-14, wherein the particle is prepared by a method comprising transducing a producer cell with plasmids encoding the fusogen and targeting agent and a Gag-pol and Rev.

16. The particle of any of claims 5-8 and 10-15, wherein the producer cell is selected from the group consisting of CHO cells, BHK cells, MDCK cells, C3H 10T1 / 2 cells, FLY cells, Psi-2 cells, BOSC 23 cells, PA317 cells, WEHI cells, COS cells, BSC 1 cells, BSC 40 cells, BMT 10 cells, VERO cells, W138 cells, MRC5 cells, A549 cells, HT1080 cells, 293 cells, 293T cells, B-50 cells, 3T3 cells, NIH3T3 cells, HepG2 cells, Saos-2 cells, Huh7 cells, HeLa cells, W163 cells, 211 cells, and 211 A cells.

17. The particle of any of claims 5-8 and 10-16, wherein the producer cell is a 293T cell.

18. The particle of any of claims 4-17, wherein the particle is replication defective.

19. The particle of any of claims 1-6 and 8-18, wherein the particle comprises a viral nucleic acid.

20. The particle of claim 19, wherein the viral nucleic acid is a retroviral nucleic acid, optionally wherein the viral nucleic acid is a lentiviral nucleic acid.

21. The particle of any of claims 19-20, wherein the viral nucleic acid comprises one or more, optionally all, of the following nucleic acid sequences: 5’ LTR (e.g., comprising U5 and lacking a functional U3 domain), Psi packaging element (Psi), Central polypurine tract (cPPT) / central termination sequence (CTS) (e.g., DNA flap), Poly A tail sequence, a posttranscriptional regulatory element (e.g., WPRE), a Rev response element (RRE), and 3’ LTR (e.g., comprising U5 and lacking a functional U3).

22. The particle of any of claims 1-7 and 10-18, wherein the particle is devoid of viral genomic nucleic acids.

23. The particle of any of claims 1-22, wherein the target molecule is ASCT2.

24. The particle of any of claims 1-22, wherein the target molecule is CD117.

25. The particle of any of claims 1-24, wherein the fusogen is a viral fusion protein.

26. The particle of any of claims 1-25, wherein the fusogen is a viral envelope protein.

27. The particle of any of claims 1-26, wherein the fusogen is a VSV-G protein or a functional variant thereof.

28. The particle of any of claims 1-26, wherein the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof.

29. A targeted lipid particle, comprising a lipid bilayer, a fusogen, wherein the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof and is exposed on the surface of the lipid bilayer.

30. The particle of claim 28 or 29, wherein the fusogen is a truncated BaEV envelope glycoprotein that comprises a cytoplasmic tail with a partial inhibitory R peptide that is less than the full length wild-type BaEV inhibitory R peptide.

31. The particle of claim 28, 29 or 30, wherein the fusogen is set forth in any of SEQ ID NO:254-260.

32. The particle of claim 28 or 29, wherein the fusogen is set forth in SEQ ID NO:261 (BaEVRLess).

33. The particle of claim 29, wherein the fusogen is set forth in SEQ ID NO:262 (BaEVTR).

34. The particle of any of claims 1-26, wherein the fusogen is a Cocal virus G protein or a functional variant thereof.

35. The particle of any of claims 1-26, wherein the fusogen is an Alphavirus fusion protein (e.g., Sindbis virus) or a functional variant thereof.

36. The particle of any of claims 1-26, wherein the fusogen is a Paramyxovirus fusion protein (e.g., a Morbillivirus or Henipavirus) or a functional variant thereof.

37. The particle of any of claims 1-26 and 36, wherein the fusogen is a Morbillivirus fusion protein (e.g., measles virus (MeV), canine distemper virus, Cetacean morbillivirus, Peste-des-petits- ruminants virus, Phocine distemper virus, or Rinderpest virus) or a functional variant thereof.

38. The particle of any of claims 1-26 and 36, wherein the fusogen is a Henipavirus fusion protein (e.g., Nipah virus, Hendra virus, Cedar virus, Kumasi virus, Mojiang virus, or Langya virus) or a functional variant thereof.

39. The particle of any of claims 1-26, 36, and 38, wherein the fusogen is a Nipah virus fusion protein or a functional variant thereof.

40. The particle of any of claims 1-26, 36, 38, and 39, wherein the fusogen comprises a Nipah virus F glycoprotein (NiV-F) or a biologically active portion thereof and a Nipah virus G glycoprotein (NiV-G) or a biologically active portion thereof.

41. The particle of claim 40, wherein the targeting agent is fused to the NiV-G protein or biologically active portion thereof.

42. The particle of claim 40 or claim 41, wherein the targeting agent is fused to the C- terminus of the NiV-G protein or biologically active portion thereof.

43. The particle of any of claims 40-42, wherein the NiV-G or biologically active portion thereof is a wild-type NiV-G protein or a functionally active variant or biologically active portion thereof.

44. The particle of any of claims 40-43, wherein the NiV-G protein or biologically active portion thereof is truncated and lacks up to 40 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein set forth in SEQ ID NO:4.

45. The particle of any of claims 40-44, wherein the NiV-G protein or biologically active portion thereof is a biologically active portion that is a truncated NiV-G that has a deletion of amino acids 2-34 at or near the N-terminus of wild-type NiV-G set forth in SEQ ID NO:4.

46. The particle of any of claims 40-45, wherein the NiV-G protein or biologically active portion thereof does not include an initial methionine.

47. The particle of any of claims 40-46, wherein the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO:42 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:42.

48. The particle of any of claims 40-47, wherein the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 42.

49. The particle of any of claims 1-48, wherein the fusogen comprises one or more modifications to reduce binding to its native receptor.

50. The particle of any of claims 40-46 and 49, wherein the NiV-G protein or biologically active portion thereof is a mutant NiV-G protein or biologically active portion thereof that exhibits reduced binding to Ephrin B2 or Ephrin B3.

51. The particle of any of claims 40-46, 49, and 50, wherein the NiV-G protein or biologically active portion thereof comprises one or more amino acid substitutions corresponding to amino acid substitutions selected from the group consisting of E501A, W504A, Q530A, and E533A with reference to numbering set forth in SEQ ID NO:4.

52. The particle of any of claims 40-46 and 49-51, wherein the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO: 17 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 17.

53. The particle of any of claims 40-46 and 49-52, wherein the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 17.

54. The particle of any of claims 40-46 and 49-51, wherein the NiV-G protein or the biologically active portion has the amino acid sequence set forth in SEQ ID NO: 18 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 18.

55. The particle of any of claims 40-46, 49-51, and 54, wherein the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 18.

56. The particle of any of claims 40-55, wherein the NiV-F protein or the biologically active portion thereof is a wild-type NiV-F protein or a functionally active variant or biologically active portion thereof.

57. The particle of any of claims 40-56, wherein the NiV-F protein or biologically active portion thereof is a truncated NiV-F that is truncated by or by at least 22 amino acids or by or by at least 20 amino acids at or near the C-terminus of wild-type NiV-F set forth in SEQ ID NO:30.

58. The particle of any of claims 40-57, wherein the NiV-F protein or biologically active portion thereof has a 22 amino acid truncation at or near the C-terminus of the wild-type NiV-F protein set forth in SEQ ID NO:30.

59. The particle of any of claims 40-58, wherein the NiV-F protein or biologically active portion thereof comprises a deletion in its cytoplasmic tail and lacks amino acid residues 525-546 of SEQ ID NO:30.

60. The particle of any of claims 40-59, wherein the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 16 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 16.

61. The particle of any of claims 40-60, wherein the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO: 16.

62. The particle of any of claims 40-59, wherein the NiV-F protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO:21 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:21.

63. The particle of any of claims 40-59 and 62, wherein the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO:21.

64. The particle of any of claims 40-46, 49-51, 54-59, 62, and 63, wherein the NiV-G protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 18, and the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO:21.

65. The particle of any of claims 1-64, wherein the targeting agent is an antibody or antibody fragment, a Design ankyrin repeat protein (DARPin), or an antigen-binding fibronectin type III (Fn3) scaffold that binds to the target molecule.

66. The particle of any of claims 1-65, wherein the targeting agent is a single domain antibody that binds to the target molecule.

67. The particle of any of claims 1-66, wherein the targeting agent is a VHH that binds to the target molecule.

68. The particle of any of claims 1-66, wherein the targeting agent is a single chain variable fragment (scFv) that binds to the target molecule.

69. The particle of any of claims 1-68, wherein the target molecule is expressed on a target cell.

70. The particle of any of claims 1-69, wherein the particle comprises an exogenous agent for delivery to a target cell.

71. The particle of any one of claims 1-69, wherein the particle comprises a fusion protein between a viral structural protein and an exogenous agent.

72. The particle of claim 71, wherein the fusion protein is a cleavable fusion protein comprising a cleavable linker positioned between the viral structural protein and the exogenous agent.

73. The particle of claim 71 or 72, wherein the fusion protein comprises one or more nuclear localization sequences.

74. The particle of claim 71 or 72, wherein the fusion protein comprises one or more nuclear export sequences.

75. The particle of claim 70 or claim 71, wherein the target molecule is expressed on the target cell.

76. The particle of any of claims 70-75, wherein the exogenous agent is present in the lumen.

77. The particle of any of claims 70-76, wherein the exogenous agent is a protein or a nucleic acid.

78. The particle of any of claims 70-77, wherein the exogenous agent is or encodes a therapeutic agent or a diagnostic agent.

79. The particle of any of claims 70-78, wherein the exogenous agent is or encodes a factor associated with gene editing.

80. The particle of any of claims 70-79, wherein the exogenous agent is or encodes a genome-modifying protein for gene editing a target gene encoding an endogenous protein.

81. The particle of claim 80, wherein the genome-modifying protein is associated with gene editing by a sequence-specific nuclease, a CRISPR-associated transposase (CAST), prime editing, or Programmable Addition via Site-specific Targeting Elements (PASTE).

82. The particle of claim 80 or claim 81, wherein the genome-modifying protein is a sequence-specific nuclease.

83. The particle of claim 81 or claim 82, wherein the sequence-specific nuclease is selected from the group consisting of an RNA-guided nuclease, a meganuclease, a transcription activator-like effector nuclease (TALEN), and a zinc-finger nuclease (ZFN).

84. The particle of any of claims 81-83, wherein the sequence-specific nuclease is an RNA- guided nuclease.

85. The particle of claim 83 or claim 84, wherein the RNA-guided nuclease is TnpB.

86. The particle of claim 83 or claim 84, wherein the RNA-guided nuclease comprises a Cas nuclease and a guide RNA (CRISPR-Cas combination).

87. The particle of claim 86, wherein the CRISPR-Cas combination is a ribonucleoprotein (RNP) complex comprising the gRNA and the Cas nuclease.

88. The particle of claim 86 or claim 87, wherein the Cas nuclease is a Type II or Type V Cas protein.

89. The particle of any of claims 80-84 and 86-88, wherein the genome-modifying protein is selected from the group consisting of Cas3, Cas4, Cas5, Cas8a, Cas8b, Cas8c, Cas9, CaslO, Casl2, Casl2a (Cpfl), Casl2b (C2cl), Casl2c (C2c3), Casl2d (CasY), Casl2e (CasX), Casl2f (C2cl0), Casl2g, Casl2h, Casl2i, Casl2k (C2c5), Casl3, Casl3a (C2c2), Casl3b, Casl3c, Casl3d, C2c4, C2c8, C2c9, Cmr5, Csel, Cse2, Csfl, Csm2, Csn2, CsxlO, Csxl l, Csyl, Csy2, Csy3, Mad7.

90. The particle of any of claims 86-89, wherein the Cas is a Cas9 or a Casl2.

91. The particle of any of claims 70-81, wherein the exogenous agent is or encodes a factor associated with base editing or prime editing (e.g., target-primed reverse transcription (TPRT)).

92. The particle of any of claims 70-81 and 91, wherein the exogenous agent is or encodes a transposase, integrase, or recombinase.

93. The particle of any of claims 70-81 and 91, wherein the exogenous agent is or encodes a DNA polymerase, RNA polymerase, or reverse-transcriptase.

94. The particle of any of claims 70-93, wherein the exogenous agent is for use in gene therapy to correct a genetic deficiency in the target cell.

95. The particle of claim 94, wherein the exogenous agent is a nucleic acid comprising a payload gene for correcting the genetic deficiency.

96. The particle of any of claims 70-78, wherein the exogenous agent is or encodes a membrane protein.

97. The particle of claim 96, wherein the membrane protein is an antigen receptor.

98. The particle of claim 97, wherein the antigen receptor binds to an antigen expressed on cells associated with a disease or condition.

99. The particle of claim 98, wherein the disease or condition is a cancer.

100. The particle of any of claims 97-99, wherein the antigen receptor binds to an antigen expressed on tumor cells.

101. The particle of any of claims 97-100, wherein the antigen receptor is a chimeric antigen receptor (CAR).

102. The particle of any of claims 97-100, wherein the antigen receptor is an engineered T cell receptor (TCR).

103. The particle of any of claims 69-102, wherein the target cell is a hematopoietic cell.

104. The particle of any of claims 69-103, wherein the target cell is CD34+.

105. The particle of any of claims 69-104, wherein the target cell is a CD34+ progenitor cell.

106. The particle of any of claims 69-105, wherein the target cell is a hematopoietic stem cell.

107. The particle of any of claims 70-106, wherein the delivery to target cells expressing the target molecule is increased by or by greater than 1.1 -fold, 1.2-fold, 1.3-fold, 1.4-fold, 1.5-fold, 1.6-fold, 1.7-fold, 1.8-fold, 1.9-fold, 2-fold, 2.5-fold, 3-fold, 3.5-fold, 4-fold, 4.5-fold, 5-fold, 5.5-fold, 6-fold, 6.5- fold, 7-fold, 7.5-fold, 8-fold, 8.5-fold, 9-fold, 9.5-fold, 10-fold, or more compared to delivery to nontarget cells not expressing the target molecule.

108. A polynucleotide comprising a nucleic acid sequence encoding a fusogen and a targeting agent that binds to a target molecule selected from the group consisting of ASCT1, ASCT2, CD105, CD110, CD117, CD133, CD146, CD164, CD34, CD46, CD49f, CD90, EPCR, and ITGA3.

109. The polynucleotide of claim 108, wherein the polynucleotide encodes a transmembrane domain that is fused to the targeting agent.

110. The polynucleotide of claim 108, wherein the targeting agent is fused to the fusogen.

111. The polynucleotide of claim 108 or claim 110, wherein the targeting agent is fused to the fusogen directly.

112. The polynucleotide of any of claims 108, 110, and 111, wherein the targeting agent is fused to the fusogen via a linker.

113. The polynucleotide of any of claims 108-112, wherein the target molecule is ASCT2.

114. The polynucleotide of any of claims 108-112, wherein the target molecule is CD117.

115. The polynucleotide of any of claims 108-114, wherein the fusogen is a viral fusion protein.

116. The polynucleotide of any of claims 108-115, wherein the fusogen is a viral envelope protein.

117. The polynucleotide of any of claims 108-116, wherein the fusogen is a VSV-G protein or a functional variant thereof.

118. The polynucleotide of any of claims 108-116, wherein the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof.

119. A polynucleotide comprising a nucleic acid sequence encoding a fusogen, wherein the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof.

120. The polynucleotide of claim 118 or 119, wherein the fusogen is a truncated BaEV envelope glycoprotein that comprises a cytoplasmic tail with a partial inhibitory R peptide that is less than the full length wild- type BaEV inhibitory R peptide.

121. The particle of any of claims 118-120, wherein the fusogen is set forth in any of SEQ ID NO:254-260.

122. The particle of claim 118 or 119, wherein the fusogen is set forth in SEQ ID NO:261 (BaEVRLess).

123. The particle of claim 118 or 119, wherein the fusogen is set forth in SEQ ID NO:262 (BaEVTR).

124. The polynucleotide of any of claims 108-116, wherein the fusogen is a Cocal virus G protein or a functional variant thereof.

125. The polynucleotide of any of claims 108-116, wherein the fusogen is an Alphavirus fusion protein (e.g., Sindbis virus) or a functional variant thereof.

126. The polynucleotide of any of claims 108-116, wherein the fusogen is a Paramyxovirus fusion protein (e.g., a Morbillivirus or Henipavirus) or a functional variant thereof.

127. The polynucleotide of any of claims 108-116 and 126, wherein the fusogen is a Morbillivirus fusion protein (e.g., measles virus (MeV), canine distemper virus, Cetacean morbillivirus, Peste-des-petits-ruminants virus, Phocine distemper virus, or Rinderpest virus) or a functional variant thereof.

128. The polynucleotide of any of claims 108-116 and 126, wherein the fusogen is a Henipavirus fusion protein (e.g., Nipah virus, Hendra virus, Cedar virus, Kumasi virus, Mojiang virus, or Langya virus) or a functional variant thereof.

129. The polynucleotide of any of claims 108-116, 126, and 128, wherein the fusogen is a Nipah virus fusion protein or a functional variant thereof.

130. The polynucleotide of any of claims 108-116, 126, 128, and 129, wherein the fusogen comprises a Nipah virus F glycoprotein (NiV-F) or a biologically active portion thereof and a Nipah virus G glycoprotein (NiV-G) or a biologically active portion thereof.

131. The polynucleotide of claim 130, wherein the targeting agent is fused to the NiV-G protein or biologically active portion thereof.

132. The polynucleotide of claim 130 or claim 131, wherein the targeting agent is fused to the C-terminus of the NiV-G protein or biologically active portion thereof.

133. The polynucleotide of any of claims 130-132, wherein the NiV-G or biologically active portion thereof is a wild-type NiV-G protein or a functionally active variant or biologically active portion thereof.

134. The polynucleotide of any of claims 130-133, wherein the NiV-G protein or biologically active portion thereof is truncated and lacks up to 40 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein set forth in SEQ ID NO:4.

135. The polynucleotide of any of claims 130-134, wherein the NiV-G protein or biologically active portion thereof is a biologically active portion that is a truncated NiV-G that has a deletion of amino acids 2-34 at or near the N-terminus of wild-type NiV-G set forth in SEQ ID NO:4.

136. The polynucleotide of any of claims 130-135, wherein the NiV-G protein or biologically active portion thereof does not include an initial methionine.

137. The polynucleotide of any of claims 130-136, wherein the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO: 42 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:42.

138. The polynucleotide of any of claims 130-137, wherein the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO:42.

139. The polynucleotide of any of claims 108-138, wherein the fusogen comprises one or more modifications to reduce binding to its native receptor.

140. The polynucleotide of any of claims 130-136 and 139, wherein the NiV-G protein or biologically active portion thereof is a mutant NiV-G protein or biologically active portion thereof that exhibits reduced binding to Ephrin B2 or Ephrin B3.

141. The polynucleotide of any of claims 130-136, 139, and 140, wherein the NiV-G protein or biologically active portion thereof comprises one or more amino acid substitutionscorresponding to amino acid substitutions selected from the group consisting of E501A, W504A, Q530A, and E533A with reference to numbering set forth in SEQ ID NO:4.

142. The polynucleotide of any of claims 130-136 and 139-141, wherein the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO: 17 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 17.

143. The polynucleotide of any of claims 130-136 and 139-142, wherein the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 17.

144. The polynucleotide of any of claims 130-136 and 139-141, wherein the NiV-G protein or the biologically active portion has the amino acid sequence set forth in SEQ ID NO: 18 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 18.

145. The polynucleotide of any of claims 130-136, 139-141, and 144, wherein the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 18.

146. The polynucleotide of any of claims 130-145, wherein the NiV-F protein or the biologically active portion thereof is a wild-type NiV-F protein or a functionally active variant or biologically active portion thereof.

147. The polynucleotide of any of claims 130-146, wherein the NiV-F protein or biologically active portion thereof is a truncated NiV-F that is truncated by or by at least 22 amino acids or by or by at least 20 amino acids at or near the C-terminus of wild-type NiV-F set forth in SEQ ID NO:30.

148. The polynucleotide of any of claims 130-147, wherein the NiV-F protein or biologically active portion thereof has a 22 amino acid truncation at or near the C-terminus of the wild-type NiV-F protein set forth in SEQ ID NO:30.

149. The polynucleotide of any of claims 130-148, wherein the NiV-F protein or biologically active portion thereof comprises a deletion in its cytoplasmic tail and lacks amino acid residues 525-546 of SEQ ID NO:30.

150. The polynucleotide of any of claims 130-149, wherein the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 16 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 16.

151. The polynucleotide of any of claims 130-150, wherein the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO: 16.

152. The polynucleotide of any of claims 130-149, wherein the NiV-F protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO:21 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:21.

153. The polynucleotide of any of claims 130-149 and 152, wherein the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO:21.

154. The polynucleotide of any of claims 130-136, 139-141, 144-149, 152, and 153, wherein the NiV-G protein or biologically active portion thereof has the sequence set forth in SEQ ID NO:18, and the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO:21.

155. The polynucleotide of any of claims 108-154, wherein the targeting agent is an antibody or antibody fragment, a Design ankyrin repeat protein (DARPin), or an antigen-binding fibronectin type III (Fn3) scaffold that binds to the target molecule.

156. The polynucleotide of any of claims 108-155, wherein the targeting agent is a single domain antibody that binds to the target molecule.

157. The polynucleotide of any of claims 108-156, wherein the targeting agent is a VHH that binds to the target molecule.

158. The polynucleotide of any of claims 108-156, wherein the targeting agent is a single chain variable fragment (scFv) that binds to the target molecule.

159. The polynucleotide of any of claims 108-158, wherein the polynucleotide is codon optimized.

160. The polynucleotide of any of claims 108-159, wherein the polynucleotide comprises at least one promoter that is operatively linked to control expression of the targeting agent and fusogen.

161. The polynucleotide of claim 160, wherein the promoter is a constitutive promoter.

162. The polynucleotide of claim 160, wherein the promoter is an inducible promoter.

163. A plasmid, comprising the polynucleotide of any of claims 108-162 or 420.

164. The plasmid of claim 163, wherein the plasmid comprises one or more nucleic acids encoding proteins for lentivirus production.

165. A vector, comprising the polynucleotide of any of claims 108-162 or 420.

166. A cell comprising the polynucleotide of any of claims 108-162 or 420, the plasmid of claim 163 or claim 164, or the vector of claim 165.

167. A cell comprising (i) a first polynucleotide comprising a nucleic acid sequence encoding a fusogen and (ii) a second polynucleotide comprising a nucleic acid sequence encoding a targeting agent that binds to a target molecule selected from the group consisting of ASCT1, ASCT2, CD105, CD110, CD117, CD133, CD146, CD164, CD34, CD46, CD49f, CD90, EPCR, and ITGA3.

168. The cell of claim 167, wherein the second polynucleotide encodes a transmembrane domain that is fused to the targeting agent.

169. A cell comprising (i) a first polynucleotide encoding a nucleic acid sequence encoding a fusogen, wherein the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof.

170. The cell of any of claims 167-169, wherein the first and / or second polynucleotide is a plasmid.

171. The cell of claim 170, wherein the plasmid of the first and / or second polynucleotide comprises one or more nucleic acids encoding proteins for lentivirus production.

172. The cell of any of claims 167-169, wherein the first and / or second polynucleotide is a vector.

173. The cell of any of claims 167-172, wherein the first and / or second polynucleotide is codon optimized.

174. The cell of any of claims 167-173, wherein: the first polynucleotide comprises at least one promoter that is operatively linked to control expression of the fusogen; and / or the second polynucleotide comprises at least one promoter that is operatively linked to control expression of the targeting agent.

175. The cell of claim 174, wherein the promoter of the first and / or second polynucleotide is a constitutive promoter.

176. The cell of claim 174, wherein the promoter of the first and / or second polynucleotide is an inducible promoter.

177. The cell of any of claims 166-176, wherein the cell is a producer cell used for producing a viral particle or virus-like particle.

178. The cell of claim 177, wherein the viral particle or virus-like particle is a retroviral particle or retrovirus-like particle.

179. The cell of claim 177 or claim 178, wherein the viral particle or virus-like particle is a lentiviral particle or lentivirus-like particle.

180. The cell of any of claims 166-179, wherein the cell is selected from the group consisting of CHO cells, BHK cells, MDCK cells, C3H 10T1 / 2 cells, FLY cells, Psi-2 cells, BOSC 23 cells, PA317 cells, WEHI cells, COS cells, BSC 1 cells, BSC 40 cells, BMT 10 cells, VERO cells, W138 cells, MRC5 cells, A549 cells, HT1080 cells, 293 cells, 293T cells, B-50 cells, 3T3 cells, NIH3T3 cells, HepG2 cells, Saos-2 cells, Huh7 cells, HeLa cells, W163 cells, 211 cells, and 211A cells.

181. The cell of any of claims 166-180, wherein the cell is a 293T cell.

182. The cell of any of claims 166-181, wherein the cell comprises a viral nucleic acid.

183. The cell of claim 182, wherein the viral nucleic acid is a retroviral nucleic acid.

184. The cell of claim 182 or claim 183, wherein the viral nucleic acid is a lentiviral nucleic acid.

185. The cell of any of claims 182-184, wherein the viral nucleic acid lacks one or more genes involved in viral replication.

186. The cell of any of claims 182-185, wherein the viral nucleic acid comprises a nucleic acid sequence encoding a viral packaging protein selected from one or more of Gag, Pol, and Rev.

187. The cell of any of claims 182-186, wherein the viral nucleic acid comprises one or more, optionally all, of the following nucleic acid sequences: 5’ LTR (e.g., comprising U5 and lacking a functional U3 domain), Psi packaging element (Psi), Central polypurine tract (cPPT) / central termination sequence (CTS) (e.g., DNA flap), Poly A tail sequence, a posttranscriptional regulatory element (e.g., WPRE), a Rev response element (RRE), and 3’ LTR (e.g., comprising U5 and lacking a functional U3).

188. The cell of any of claims 166-187, wherein the target molecule is ASCT2.

189. The cell of any of claims 166-187, wherein the target molecule is CD117.

190. The cell of any of claims 166-189, wherein the fusogen is a viral fusion protein.

191. The cell of any of claims 166-190, wherein the fusogen is a viral envelope protein.

192. The cell of any of claims 166-191, wherein the fusogen is a VSV-G protein or a functional variant thereof.

193. The cell of any of claims 166-191, wherein the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof.

194. The cell of any of claims 166-191 and 193, wherein the fusogen is a truncated BaEV envelope glycoprotein that comprises a cytoplasmic tail with a partial inhibitory R peptide that is less than the full length wild- type BaEV inhibitory R peptide.

195. The cell of claim 168 or 169, wherein the fusogen is set forth in any of SEQ ID NO:254- 260.

196. The cell of claim 168, 169, or 195, wherein the fusogen is set forth in SEQ ID NO:261 (BaEVRLess).

197. The cell of claim 168, 169 or 195, wherein the fusogen is set forth in SEQ ID NO:262 (BaEVTR).

198. The cell of any of claims 166-191, wherein the fusogen is a Cocal virus G protein or a functional variant thereof.

199. The cell of any of claims 166-191, wherein the fusogen is an Alphavirus fusion protein (e.g., Sindbis virus) or a functional variant thereof.

200. The cell of any of claims 166-191, wherein the fusogen is a Paramyxovirus fusion protein (e.g., a Morbillivirus or Henipavirus) or a functional variant thereof.

201. The cell of any of claims 166-191 and 200, wherein the fusogen is a Morbillivirus fusion protein (e.g., measles virus (MeV), canine distemper virus, Cetacean morbillivirus, Peste-des-petits- ruminants virus, Phocine distemper virus, or Rinderpest virus) or a functional variant thereof.

202. The cell of any of claims 166-191 and 200, wherein the fusogen is a Henipavirus fusion protein (e.g., Nipah virus, Hendra virus, Cedar virus, Kumasi virus, Mojiang virus, or Langya virus) or a functional variant thereof.

203. The cell of any of claims 166-191, 200, and 202, wherein the fusogen is a Nipah virus fusion protein or a functional variant thereof.

204. The cell of any of claims 166-191, 200, 202, and 203, wherein the fusogen comprises a Nipah virus F glycoprotein (NiV-F) or a biologically active portion thereof and a Nipah virus G glycoprotein (NiV-G) or a biologically active portion thereof.

205. The cell of claim 204, wherein the NiV-G or biologically active portion thereof is a wildtype NiV-G protein or a functionally active variant or biologically active portion thereof.

206. The cell of claim 204 or claim 205, wherein the NiV-G protein or biologically active portion thereof is truncated and lacks up to 40 contiguous amino acid residues at or near the N- terminus of the wild-type NiV-G protein set forth in SEQ ID NO:4.

207. The cell of any of claims 204-206, wherein the NiV-G protein or biologically active portion thereof is a biologically active portion that is a truncated NiV-G that has a deletion of amino acids 2-34 at or near the N-terminus of wild-type NiV-G set forth in SEQ ID NO:4.

208. The cell of any of claims 204-207, wherein the NiV-G protein or biologically active portion thereof does not include an initial methionine.

209. The cell of any of claims 204-208, wherein the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO:42 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:42.

210. The cell of any of claims 204-209, wherein the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO:42.

211. The cell of any of claims 166-210, wherein the fusogen comprises one or more modifications to reduce binding to its native receptor.

212. The cell of any of claims 204-208 and 211, wherein the NiV-G protein or biologically active portion thereof is a mutant NiV-G protein or biologically active portion thereof that exhibits reduced binding to Ephrin B2 or Ephrin B3.

213. The cell of any of claims 204-208, 211, and 212, wherein the NiV-G protein or biologically active portion thereof comprises one or more amino acid substitutions corresponding to amino acid substitutions selected from the group consisting of E501A, W504A, Q530A, and E533A with reference to numbering set forth in SEQ ID NO:4.

214. The cell of any of claims 204-208 and 211-213, wherein the NiV-G protein or biologically active portion thereof comprises the amino acid sequence set forth in SEQ ID NO: 17 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 17.

215. The cell of any of claims 204-208 and 211-214, wherein the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 17.

216. The cell of any of claims 204-208 and 211-213, wherein the NiV-G protein or the biologically active portion has the amino acid sequence set forth in SEQ ID NO: 18 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 18.

217. The cell of any of claims 204-208, 211-213, and 216, wherein the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 18.

218. The cell of any of claims 204-217, wherein the NiV-F protein or the biologically active portion thereof is a wild-type NiV-F protein or a functionally active variant or biologically active portion thereof.

219. The cell of any of claims 204-218, wherein the NiV-F protein or biologically active portion thereof is a truncated NiV-F that is truncated by or by at least 22 amino acids or by or by at least 20 amino acids at or near the C-terminus of wild-type NiV-F set forth in SEQ ID NO:30.

220. The cell of any of claims 204-219, wherein the NiV-F protein or biologically active portion thereof has a 22 amino acid truncation at or near the C-terminus of the wild-type NiV-F protein set forth in SEQ ID NO:30.

221. The cell of any of claims 204-220, wherein the NiV-F protein or biologically active portion thereof comprises a deletion in its cytoplasmic tail and lacks amino acid residues 525-546 of SEQ ID NO:30.

222. The cell of any of claims 204-221, wherein the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 16 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 16.

223. The cell of any of claims 204-222, wherein the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO: 16.

224. The cell of any of claims 204-221, wherein the NiV-F protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO:21 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:21.

225. The cell of any of claims 204-221 and 224, wherein the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO:21.

226. The cell of any of claims 204-208, 211-213, 216-221, 224, and 225, wherein the NiV- G protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 18, and the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO:21.

227. The cell of any of claims 166-226, wherein the targeting agent is an antibody or antibody fragment, a Design ankyrin repeat protein (DARPin), or an antigen-binding fibronectin type III (Fn3) scaffold that binds to the target molecule.

228. The cell of any of claims 166-227, wherein the targeting agent is a single domain antibody that binds to the target molecule.

229. The cell of any of claims 166-228, wherein the targeting agent is a VHH that binds to the target molecule.

230. The cell of any of claims 166-228, wherein the targeting agent is a single chain variable fragment (scFv) that binds to the target molecule.

231. A method of making a targeted lipid particle, comprising:(a) introducing into a source cell the polynucleotide of any of claims 108-162 or 420, the plasmid of claim 163 or claim 164, or the vector of claim 165;(b) culturing the source cell under conditions for producing a lipid particle comprising a lipid bilayer and the targeting agent and fusogen exposed on the surface of the lipid bilayer; and(c) separating, enriching, or purifying the lipid particle from the source cell, thereby making the lipid particle.

232. A method of making a targeted lipid particle, comprising:(a) introducing into a source cell (i) a first polynucleotide comprising a nucleic acid sequence encoding a fusogen and (ii) a second polynucleotide comprising a nucleic acid sequence encoding atargeting agent that binds to a target molecule selected from the group consisting of ASCT1, ASCT2, CD105, CD110, CD117, CD133, CD146, CD164, CD34, CD46, CD49f, CD90, EPCR, and ITGA3;(b) culturing the source cell under conditions for producing a lipid particle comprising a lipid bilayer and the targeting agent and fusogen exposed on the surface of the lipid bilayer; and(c) separating, enriching, or purifying the lipid particle from the source cell, thereby making the lipid particle.

233. The method of claim 232, wherein the second polynucleotide encodes a transmembrane domain that is fused to the targeting agent.

234. The method of claim 232 or claim 233, wherein the first and / or second polynucleotide is a plasmid.

235. The method of claim 234, wherein the plasmid of the first and / or second polynucleotide comprises one or more nucleic acids encoding proteins for lentivirus production.

236. The method of claim 232 or claim 233, wherein the first and / or second polynucleotide is a vector.

237. The method of any of claims 226-230, wherein the first and / or second polynucleotide is codon optimized.

238. The method of any of claims 226-231, wherein: the first polynucleotide comprises at least one promoter that is operatively linked to control expression of the fusogen; and / or the second polynucleotide comprises at least one promoter that is operatively linked to control expression of the targeting agent.

239. The method of claim 232, wherein the promoter of the first and / or second polynucleotide is a constitutive promoter.

240. The method of claim 232, wherein the promoter of the first and / or second polynucleotide is an inducible promoter.

241. The method of any of claims 225-234, wherein the source cell is a mammalian cell.

242. The method of any of claims 225-235, wherein the source cell is a producer cell used for producing a viral particle or virus-like particle.

243. The method of claim 236, wherein the particle is a viral particle or virus-like particle.

244. The method of claim 236 or claim 237, wherein the viral particle or virus-like particle is a retroviral particle or retrovirus-like particle.

245. The method of any of claims 236-238, wherein the viral particle or virus-like particle is a lentivirus-like particle.

246. The method of any of claims 236-238, wherein the viral particle or virus-like particle is a lentiviral particle.

247. The method of any of claims 225-240, wherein the particle is pseudotyped with the fusogen.

248. The method of any of claims 236-241, wherein the producer cell is selected from the group consisting of CHO cells, BHK cells, MDCK cells, C3H 10T1 / 2 cells, FLY cells, Psi-2 cells, BOSC 23 cells, PA317 cells, WEHI cells, COS cells, BSC 1 cells, BSC 40 cells, BMT 10 cells, VERO cells, W138 cells, MRC5 cells, A549 cells, HT1080 cells, 293 cells, 293T cells, B-50 cells, 3T3 cells, NIH3T3 cells, HepG2 cells, Saos-2 cells, Huh7 cells, HeLa cells, W163 cells, 211 cells, and 211 A cells, optionally wherein the producer cell is a 293T cell.

249. The method of any of claims 231-248, wherein the method comprises transducing the source cell with packaging plasmids encoding a Gag-pol and Rev.

250. The method of any of claims 231-249, wherein the target molecule is ASCT2.

251. The method of any of claims 231-249, wherein the target molecule is CD117.

252. The method of any of claims 231-251, wherein the fusogen is a viral fusion protein.

253. The method of any of claims 231-252, wherein the fusogen is a viral envelope protein.

254. The method of any of claims 231-253, wherein the fusogen is a VSV-G protein or a functional variant thereof.

255. The method of any of claims 231-253, wherein the fusogen is a Cocal virus G protein or a functional variant thereof.

256. The method of any of claims 231-253, wherein the fusogen is an Alphavirus fusion protein (e.g., Sindbis virus) or a functional variant thereof.

257. The method of any of claims 231-253, wherein the fusogen is a Paramyxovirus fusion protein (e.g., a Morbillivirus or Henipavirus) or a functional variant thereof.

258. The method of any of claims 231-253 and 257, wherein the fusogen is a Morbillivirus fusion protein (e.g., measles virus (MeV), canine distemper virus, Cetacean morbillivirus, Peste-des- petits-ruminants virus, Phocine distemper virus, or Rinderpest virus) or a functional variant thereof.

259. The method of any of claims 231-253 and 257, wherein the fusogen is a Henipavirus fusion protein (e.g., Nipah virus, Hendra virus, Cedar virus, Kumasi virus, Mojiang virus, or Langya virus) or a functional variant thereof.

260. The method of any of claims 231-253, 257, and 259, wherein the fusogen is a Nipah virus fusion protein or a functional variant thereof.

261. The method of any of claims 231-253, 257, 259, and 260, wherein the fusogen comprises a Nipah virus F glycoprotein (NiV-F) or a biologically active portion thereof and a Nipah virus G glycoprotein (NiV-G) or a biologically active portion thereof.

262. The method of claim 261, wherein the NiV-G or biologically active portion thereof is a wild-type NiV-G protein or a functionally active variant or biologically active portion thereof.

263. The method of claim 261 or claim 262, wherein the NiV-G protein or biologically active portion thereof is truncated and lacks up to 40 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein set forth in SEQ ID NO:4.

264. The method of any of claims 261-263, wherein the NiV-G protein or biologically active portion thereof is a biologically active portion that is a truncated NiV-G that has a deletion of amino acids 2-34 at or near the N-terminus of wild-type NiV-G set forth in SEQ ID NO:4.

265. The method of any of claims 261-264, wherein the NiV-G protein or biologically active portion thereof does not include an initial methionine.

266. The method of any of claims 261-265, wherein the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO: 42 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:42.

267. The method of any of claims 261-266, wherein the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 42.

268. The method of any of claims 231-267, wherein the fusogen comprises one or more modifications to reduce binding to its native receptor.

269. The method of any of claims 261-265 and 268, wherein the NiV-G protein or biologically active portion thereof is a mutant NiV-G protein or biologically active portion thereof that exhibits reduced binding to Ephrin B2 or Ephrin B3.

270. The method of any of claims 261-265, 268, and 269, wherein the NiV-G protein or biologically active portion thereof comprises one or more amino acid substitutions corresponding to amino acid substitutions selected from the group consisting of E501A, W504A, Q530A, and E533A with reference to numbering set forth in SEQ ID NO:4.

271. The method of any of claims 261-265 and 268-270, wherein the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO: 17 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 17.

272. The method of any of claims 261-265 and 268-271, wherein the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 17.

273. The method of any of claims 261-265 and 268-270, wherein the NiV-G protein or the biologically active portion has the amino acid sequence set forth in SEQ ID NO: 18 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 18.

274. The method of any of claims 261-265, 268-270, and 273, wherein the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 18.

275. The method of any of claims 261-274, wherein the NiV-F protein or the biologically active portion thereof is a wild-type NiV-F protein or a functionally active variant or biologically active portion thereof.

276. The method of any of claims 261-275, wherein the NiV-F protein or biologically active portion thereof is a truncated NiV-F that is truncated by or by at least 22 amino acids or by or by at least 20 amino acids at or near the C-terminus of wild-type NiV-F set forth in SEQ ID NO:30.

277. The method of any of claims 261-276, wherein the NiV-F protein or biologically active portion thereof has a 22 amino acid truncation at or near the C-terminus of the wild-type NiV-F protein set forth in SEQ ID NO:30.

278. The method of any of claims 261-277, wherein the NiV-F protein or biologically active portion thereof comprises a deletion in its cytoplasmic tail and lacks amino acid residues 525- 546 of SEQ ID NO:30.

279. The method of any of claims 261-278, wherein the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 16 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 16.

280. The method of any of claims 261-279, wherein the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO: 16.

281. The method of any of claims 261-278, wherein the NiV-F protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO:21 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:21.

282. The method of any of claims 261-278 and 281, wherein the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO:21.

283. The method of any of claims 261-265, 268-270, 273-278, 281, and 282, wherein the NiV-G protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 18, and the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO:21.

284. The method of any of claims 231-283, wherein the targeting agent is an antibody or antibody fragment, a Design ankyrin repeat protein (DARPin), or an antigen-binding fibronectin type III (Fn3) scaffold that binds to the target molecule.

285. The method of any of claims 231-284, wherein the targeting agent is a single domain antibody that binds to the target molecule.

286. The method of any of claims 231-285, wherein the targeting agent is a VHH that binds to the target molecule.

287. The method of any of claims 231-285, wherein the targeting agent is a single chain variable fragment (scFv) that binds to the target molecule.

288. A targeted lipid particle produced by the method of any of claims 231-287 and 418.

289. A targeted lentiviral particle produced by the method of any of claims 231, 241-287 and 418.

290. A composition comprising a plurality of the lipid particle of any of claims 1-8, 10-107,288 and 419.

291. A composition comprising a plurality of the lentiviral particle of any of claims 8-107,289 and 419.

292. The composition of claim 290 or claim 291, wherein the composition further comprises a pharmaceutically acceptable excipient.

293. A method of transducing a target cell, the method comprising contacting a target cell with the lentiviral particle of any of claims 8-107, 289, and 419, or the composition of claim 291 or claim 292.

294. The method of claim 293, wherein the particle comprises an exogenous agent, and the transduction introduces the exogenous agent into the target cell.

295. A method of delivering an exogenous agent to a target cell, the method comprising contacting a target cell with the particle of any of claims 70-107, 288, 289, and 419, or the composition of any of claims 290-292, the particle or plurality of particles comprising an exogenous agent for delivery to the target cell.

296. The method of any of claims 293-295, wherein the contacting is in vitro or ex vivo.

297. The method of any of claims 293-295, wherein the contacting is in vivo in a subject.

298. A method of delivering an exogenous agent to a target cell in a subject, comprising administering to a subject the particle of any of claims 70-107, 288, 289, and 419, or the composition of any of claims 290-292, the particle or plurality of particles comprising an exogenous agent for delivery to a target cell in the subject.

299. The method of any of claims 294-298, wherein the exogenous agent is or encodes a therapeutic agent or a diagnostic agent.

300. The method of any of claims 297-299, wherein the exogenous agent is for treating a disease or condition in the subject.

301. A method of treating a disease or condition in a subject, comprising administering to a subject the particle of any of claims 70-107, 288, 289, or 419, or the composition of any of claims 290- 292, the particle or plurality of particles comprising an exogenous agent for treating a disease or condition in the subject.

302. The method of any of claims 297-301, wherein the method comprises administering to the subject one or more agents that stimulate mobilization of bone marrow cells from the bone marrow to the peripheral blood.

303. The method of any of claims 297-301, wherein the subject has previously been administered one or more agents that stimulate mobilization of bone marrow cells from the bone marrow to the peripheral blood.

304. A method of treating a disease or condition in a subject, comprising administering to a subject:(a) one or more agents that stimulate mobilization of bone marrow cells from the bone marrow to the peripheral blood; and(b) a lipid particle comprising a lipid bilayer, a fusogen exposed on the surface of the lipid bilayer, and an exogenous agent for treating a disease or condition in the subject.

305. The method of claim 302 or claim 304, wherein the administration of at least one of the one or more agents that stimulate mobilization is initiated prior to the administration of the particle.

306. A method of treating a disease or condition in a subject, comprising administering to a subject a lipid particle comprising a lipid bilayer, a fusogen exposed on the surface of the lipid bilayer, and an exogenous agent for treating a disease or condition in the subject, wherein the subject has previously been administered one or more agents that stimulate mobilization of bone marrow cells from the bone marrow to the peripheral blood.

307. The method of any of claims 301-306, wherein the exogenous agent is for delivery to a target cell for treating the disease or condition in the subject.

308. The method of any of claims 295-307, wherein the particle is a viral particle or virus-like particle.

309. The method of any of claims 293-308, wherein the lipid bilayer is derived from a membrane of a producer cell used for producing a viral particle or virus-like particle.

310. The method of any of claims 293-309, wherein the particle is prepared by a method comprising transducing a producer cell with plasmids encoding the fusogen and a Gag-pol and Rev.

311. The method of any of claims 308-310, wherein the viral particle or virus-like particle is a retroviral particle or retrovirus-like particle.

312. The method of any of claims 308-311, wherein the viral particle or virus-like particle is a lentivirus-like particle.

313. The method of any of claims 308-311, wherein the viral particle or virus-like particle is a lentiviral particle.

314. The method of any of claims 293-303, 305, and 307-313, wherein the particle is pseudotyped with the fusogen.

315. The method of any of claims 309-314, wherein the producer cell is selected from the group consisting of CHO cells, BHK cells, MDCK cells, C3H 10T1 / 2 cells, FLY cells, Psi-2 cells, BOSC 23 cells, PA317 cells, WEHI cells, COS cells, BSC 1 cells, BSC 40 cells, BMT 10 cells, VERO cells, W138 cells, MRC5 cells, A549 cells, HT1080 cells, 293 cells, 293T cells, B-50 cells, 3T3 cells, NIH3T3 cells, HepG2 cells, Saos-2 cells, Huh7 cells, HeLa cells, W163 cells, 211 cells, and 211 A cells.

316. The method of any of claims 309-315, wherein the producer cell is a 293T cell.

317. The method of any of claims 293-303, 305, and 307-316, wherein the particle is replication defective.

318. The method of any of claims 293-311 and 313-317, wherein the particle comprises a viral nucleic acid.

319. The method of claim 318, wherein the viral nucleic acid is a retroviral nucleic acid.

320. The method of claim 318 or claim 319, wherein the viral nucleic acid is a lentiviral nucleic acid.

321. The method of any of claims 318-320, wherein the viral nucleic acid comprises one or more, optionally all, of the following nucleic acid sequences: 5’ LTR (e.g., comprising U5 and lacking a functional U3 domain), Psi packaging element (Psi), Central polypurine tract (cPPT) / central termination sequence (CTS) (e.g., DNA flap), Poly A tail sequence, a posttranscriptional regulatory element (e.g., WPRE), a Rev response element (RRE), and 3’ LTR (e.g., comprising U5 and lacking a functional U3).

322. The method of any of claims 295-312 and 314-317, wherein the particle is devoid of viral genomic nucleic acids.

323. The method of any of claims 293-322, wherein the fusogen is a viral fusion protein.

324. The method of any of claims 293-323, wherein the fusogen is a viral envelope protein.

325. The method of any of claims 293-324, wherein the fusogen is a VSV-G protein or a functional variant thereof.

326. The method of any of claims 293-324, wherein the fusogen is a baboon endogenous virus (BaEV) envelope glycoprotein or a functional variant thereof.

327. The method of any of claims 293-324 and 326, wherein the fusogen is a truncated BaEV envelope glycoprotein that comprises a cytoplasmic tail with a partial inhibitory R peptide that is less than the full length wild- type BaEV inhibitory R peptide.

328. The method of any of claims 293-324, 326, and 326, wherein the fusogen is set forth in any of SEQ ID NO:254-260.

329. The method of any of claims 293-324 and 326, wherein the fusogen is set forth in SEQ ID NO:261 (BaEVRLess).

330. The method of any of claims 293-324 and 326, wherein the fusogen is set forth in SEQ ID NO:262 (BaEVTR).

331. The method of any of claims 293-324, wherein the fusogen is a Cocal virus G protein or a functional variant thereof.

332. The method of any of claims 293-324, wherein the fusogen is an Alphavirus fusion protein (e.g., Sindbis virus) or a functional variant thereof.

333. The method of any of claims 293-324, wherein the fusogen is a Paramyxovirus fusion protein (e.g., a Morbillivirus or Henipavirus) or a functional variant thereof.

334. The method of any of claims 293-324 and 333, wherein the fusogen is a Morbillivirus fusion protein (e.g., measles virus (MeV), canine distemper virus, Cetacean morbillivirus, Peste-des- petits-ruminants virus, Phocine distemper virus, or Rinderpest virus) or a functional variant thereof.

335. The method of any of claims 293-324 and 333, wherein the fusogen is a Henipavirus fusion protein (e.g., Nipah virus, Hendra virus, Cedar virus, Kumasi virus, Mojiang virus, or Langya virus) or a functional variant thereof.

336. The method of any of claims 293-324, 333, and 335, wherein the fusogen is a Nipah virus fusion protein or a functional variant thereof.

337. The method of any of claims 293-324, 333, 335, and 336, wherein the fusogen comprises a Nipah virus F glycoprotein (NiV-F) or a biologically active portion thereof and a Nipah virus G glycoprotein (NiV-G) or a biologically active portion thereof.

338. The method claim 337, wherein the NiV-G or biologically active portion thereof is a wild-type NiV-G protein or a functionally active variant or biologically active portion thereof.

339. The method of claim 337 or claim 338, wherein the NiV-G protein or biologically active portion thereof is truncated and lacks up to 40 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein set forth in SEQ ID NO:4.

340. The method of any of claims 337-339, wherein the NiV-G protein or biologically active portion thereof is a biologically active portion that is a truncated NiV-G that has a deletion of amino acids 2-34 at or near the N-terminus of wild-type NiV-G set forth in SEQ ID NO:4.

341. The method of any of claims 337-340, wherein the NiV-G protein or biologically active portion thereof does not include an initial methionine.

342. The method of any of claims 337-341, wherein the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO: 42 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:42.

343. The method of any of claims 337-342, wherein the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 42.

344. The method of any of claims 293-343, wherein the fusogen comprises one or more modifications to reduce binding to its native receptor.

345. The method of any of claims 337-341 and 344, wherein the NiV-G protein or biologically active portion thereof is a mutant NiV-G protein or biologically active portion thereof that exhibits reduced binding to Ephrin B2 or Ephrin B3.

346. The method of any of claims 337-341, 344, and 345, wherein the NiV-G protein or biologically active portion thereof comprises one or more amino acid substitutions corresponding to amino acid substitutions selected from the group consisting of E501A, W504A, Q530A, and E533A with reference to numbering set forth in SEQ ID NO:4.

347. The method of any of claims 337-341 and 344-346, wherein the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO: 17 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 17.

348. The method of any of claims 337-341 and 344-347, wherein the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 17.

349. The method of any of claims 337-341 and 344-346, wherein the NiV-G protein or the biologically active portion has the amino acid sequence set forth in SEQ ID NO: 18 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 18.

350. The method of any of claims 337-341, 344-346, and 349, wherein the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 18.

351. The method of any of claims 337-350, wherein the NiV-F protein or the biologically active portion thereof is a wild-type NiV-F protein or a functionally active variant or biologically active portion thereof.

352. The method of any of claims 337-351, wherein the NiV-F protein or biologically active portion thereof is a truncated NiV-F that is truncated by or by at least 22 amino acids or by or by at least 20 amino acids at or near the C-terminus of wild-type NiV-F set forth in SEQ ID NO:30.

353. The method of any of claims 337-352, wherein the NiV-F protein or biologically active portion thereof has a 22 amino acid truncation at or near the C-terminus of the wild-type NiV-F protein set forth in SEQ ID NO:30.

354. The method of any of claims 337-353, wherein the NiV-F protein or biologically active portion thereof comprises a deletion in its cytoplasmic tail and lacks amino acid residues 525- 546 of SEQ ID NO:30.

355. The method of any of claims 337-354, wherein the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 16 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 16.

356. The method of any of claims 337-355, wherein the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO: 16.

357. The method of any of claims 337-354, wherein the NiV-F protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO:21 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:21.

358. The method of any of claims 337-354 and 357, wherein the NiV-F protein or biologically active portion thereof is set forth in SEQ ID NO:21.

359. The method of any of claims 337-341, 344-346, 349-354, 357, and 358, wherein the NiV-G protein or biologically active portion thereof has the sequence set forth in SEQ ID NO: 18, and the NiV-F protein or biologically active portion thereof has the sequence set forth in SEQ ID NO:21.

360. The method of any of claims 302-359, wherein the bone marrow cells are CD34+.

361. The method of any of claims 302-360, wherein the bone marrow cells are CD34+ progenitor cells.

362. The method of any of claims 302-361, wherein the bone marrow cells are hematopoietic stem cells.

363. The method of any of claims 302-362, wherein the one or more agents that stimulate mobilization are selected from the group consisting of stem cell factor (SCF), small molecule VLA-4 inhibitor BI05192, BOP (N-(henzenesulfonyl)-L-prolyl-L-0-(l-pyrrolidinylcarbonyl)tyrosine), heparin, granulocyte colony-stimulating factor (G-CSF), MGTA-145, and plerixafor (AMD3100).

364. The method of any of claims 302-362, wherein the one or more agents that stimulate mobilization comprise G-CSF.

365. The method of claim 364, wherein the G-CSF is administered to the subject daily on the two days, three days, four days, or five days prior to the administration of the first dose of the particle.

366. The method of claim 364 or claim 365, wherein the G-CSF is administered to the subject on the day of the administration of the first dose of the particle.

367. The method of any of claims 364-366, wherein the G-CSF is administered to the subject on the day of administration of a second or later dose of the particle.

368. The method of any of claims 302-367, wherein the one or more agents that stimulate mobilization comprise plerixafor.

369. The method of claim 368, wherein the plerixafor is administered to the subject on the day of the administration of the first dose of the particle.

370. The method of claim 368 or claim 369, wherein the plerixafor is administered to the subject on the day of the administration of a second or later dose of the particle.

371. The method of any of claims 302-370, wherein the one or more agents that stimulate mobilization are G-CSF and plerixafor.

372. The method of claim 371, wherein: the G-CSF is administered to the subject daily on the four days prior to the administration of the first dose of the particle; and the plerixafor is administered to the subject on the day of the administration of the first dose of the particle.

373. The method of claim 371, wherein: the G-CSF is administered to the subject (i) daily on the two days prior to the administration of the particle; (ii) on the day of the administration of the first dose of the particle; and (iii) on the day of administration of a second or later dose of the particle; and the plerixafor is administered to the subject on the day of administration of a second or later dose of the particle.

374. The method of any of claims 297-373, wherein the method comprises administering to the subject a transduction adjuvant.

375. The method of claim 374, wherein the transduction adjuvant is administered to the subject on the day of the administration of at least one dose of the particle.

376. The method of claim 374 or claim 375, wherein the transduction adjuvant is administered to the subject on the days of the administration of at least two doses of the particle.

377. The method of any of claims 374-376, wherein the transduction adjuvant is a cationic peptide.

378. The method of any of claims 374-377, wherein the transduction adjuvant is a histidine- rich cationic peptide.

379. The method of any of claims 374-378, wherein the transduction adjuvant is a cationic amphipathic peptide.

380. The method of any of claims 374-379, wherein the transduction adjuvant is derived from the LAH4 peptide family.

381. The method of any of claims 374-380, wherein the transduction adjuvant is Vectofusin- 1.

382. The method of any of claims 293-300, 302, 303, 305, and 307-381, wherein the target molecule is expressed on the target cell.

383. The method of any of claims 293-300, 302, 303, 305, and 307-382, wherein the target cell is a hematopoietic cell.

384. The method of any of claims 293-300, 302, 303, 305, and 307-383, wherein the target cell is CD34+.

385. The method of any of claims 293-300, 302, 303, 305, and 307-384, wherein the target cell is a CD34+ progenitor cell.

386. The method of any of claims 293-300, 302, 303, 305, and 307-385, wherein the hematopoietic cell is a hematopoietic stem cell.

387. The particle of any of claims 294-386, wherein the exogenous agent is present in the lumen.

388. The method of any of claims 294-387, wherein the exogenous agent is a protein or a nucleic acid.

389. The method of any of claims 294-388, wherein the exogenous agent is or encodes a factor associated with gene editing.

390. The method of any of claims 294-389, wherein the exogenous agent is or encodes a genome-modifying protein for gene editing a target gene encoding an endogenous protein.

391. The method of any one of claims 294-390, wherein the exogenous agent is a fusion protein between a viral structural protein and an exogenous agent.

392. The method of claim 391, wherein the fusion protein is a cleavable fusion protein comprising a cleavable linker positioned between the viral structural protein and the exogenous agent.

393. The method of claim 391 or 392, wherein the fusion protein comprises one or more nuclear localization sequences.

394. The particle of claim 391 or 392, wherein the fusion protein comprises one or more nuclear export sequences.

395. The method of any of claims 390-394, wherein the genome-modifying protein is associated with gene editing by a sequence-specific nuclease, a CRISPR-associated transposase (CAST), prime editing, or Programmable Addition via Site-specific Targeting Elements (PASTE).

396. The method of any of claims 390-395, wherein the genome-modifying protein is a sequence-specific nuclease.

397. The method of any of claims 390-396, wherein the sequence-specific nuclease is selected from the group consisting of an RNA-guided nuclease, a meganuclease, a transcription activatorlike effector nuclease (TALEN), and a zinc-finger nuclease (ZFN).

398. The method of any of claims 395-397, wherein the sequence-specific nuclease is an RNA-guided nuclease.

399. The method of claim 397 or claim 398, wherein the RNA-guided nuclease is TnpB.

400. The method of claim 397 or claim 398, wherein the RNA-guided nuclease comprises a Cas nuclease and a guide RNA (CRISPR-Cas combination).

401. The method of claim 400, wherein the CRISPR-Cas combination is a ribonucleoprotein (RNP) complex comprising the gRNA and the Cas nuclease.

402. The method of claim 400 or claim 401, wherein the Cas nuclease is a Type II or Type V Cas protein.

403. The method of any of claims 390-398 and 400-402, wherein the genome-modifying protein is selected from the group consisting of Cas3, Cas4, Cas5, Cas8a, Cas8b, Cas8c, Cas9, CaslO, Casl2, Casl2a (Cpfl), Casl2b (C2cl), Casl2c (C2c3), Casl2d (CasY), Casl2e (CasX), Casl2f (C2cl0), Casl2g, Casl2h, Casl2i, Casl2k (C2c5), Casl3, Casl3a (C2c2), Casl3b, Casl3c, Casl3d, C2c4, C2c8, C2c9, Cmr5, Csel, Cse2, Csfl, Csm2, Csn2, CsxlO, Csxl l, Csyl, Csy2, Csy3, Mad7.

404. The method of any of claims 400-403, wherein the Cas is a Cas9 or a Casl2.

405. The method of any of claims 294-395, wherein the exogenous agent is or encodes a factor associated with base editing or prime editing (e.g., target-primed reverse transcription (TPRT)).

406. The method of any of claims 294-395 and 405, wherein the exogenous agent is or encodes a transposase, integrase, or recombinase.

407. The method of any of claims 294-395 and 405, wherein the exogenous agent is or encodes a DNA polymerase, RNA polymerase, or reverse-transcriptase.

408. The method of any of claims 294-407, wherein the exogenous agent is for use in gene therapy to correct a genetic deficiency in the target cell.

409. The method of claim 408, wherein the exogenous agent is a nucleic acid comprising a payload gene for correcting the genetic deficiency.

410. The method of any of claims 294-388, wherein the exogenous agent is or encodes a membrane protein.

411. The method of claim 410, wherein the membrane protein is an antigen receptor.

412. The method of claim 411, wherein the antigen receptor binds to an antigen expressed on cells associated with a disease or condition in the subject.

413. The method of claim 412, wherein the disease or condition is a cancer.

414. The method of any of claims 411-413, wherein the antigen receptor binds to an antigen expressed on tumor cells in the subject.

415. The method of any of claims 411-414, wherein the antigen receptor is a chimeric antigen receptor (CAR).

416. The method of any of claims 411-414, wherein the antigen receptor is an engineered T cell receptor (TCR).

417. The method of any of claims 293-300, 302, 303, 305, and 307-416, wherein the delivery to target cells expressing the target molecule is increased by or by greater than 1.1 -fold, 1.2-fold, 1.3- fold, 1.4-fold, 1.5-fold, 1.6-fold, 1.7-fold, 1.8-fold, 1.9-fold, 2-fold, 2.5-fold, 3-fold, 3.5-fold, 4-fold, 4.5- fold, 5-fold, 5.5-fold, 6-fold, 6.5-fold, 7-fold, 7.5-fold, 8-fold, 8.5-fold, 9-fold, 9.5-fold, 10-fold, or more compared to delivery to non-target cells not expressing the target molecule.

418. The method of any of claims 297-417, wherein the subject is a human subject.

419. The particle of any of claims 1-22 and 25-107, wherein the target molecule is CD 133.

420. The polynucleotide of any of claims 108-112 and 115-162, wherein the target molecule is CD133.

421. The cell of any of claims 167-187 and 190-230, wherein the target molecule is CD133.

422. The method of making a targeted lipid particle of any of claims 232-249 and 252-287, wherein the target molecule is CD 133.

423. The method of any of claims 306-418, wherein the fusogen is fused to a targeting agent that binds to a target molecule selected from the group consisting of ASCT1, ASCT2, CD105, CD110, CD117, CD133, CD146, CD164, CD34, CD46, CD49f, CD90, EPCR, and ITGA3.

424. The method of claim 423, wherein the target molecule is ASCT2.

425. The method of claim 423, wherein the target molecule is CD117.

426. The method of claim 423, wherein the target molecule is CD133.

427. The particle of claim 419, the polynucleotide of claim 420, the cell of claim 421, the method of claims 422, 423 or claim 426, wherein the targeting agent is a CD 133 binding agent that is an antibody or antigen-binding fragment comprising a variable heavy (VH) chain and a variable light (VL) chain selected from a) a VH chain comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 271, 272, 273, respectively, and a VL chain comprising a CDR- Ll, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 275, 276, 277, respectively; (b) a VH chain comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 280, 281, 282, respectively, and a VL chain comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 284, 285, and 286, respectively; (c) a VH chain comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 298, 299, and 300, respectively, and a VL chain comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 302, 303, and 304, respectively; (d) a VH chain comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the aminoacid sequences of SEQ ID NOs: 307, 308, and 309, respectively, and a VL chain comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 311, 312, and 313, respectively; or (e) a VH chain comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 298, 290, and 291, respectively, and a VL chain comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 293, 294, and 295, respectively.

428. The particle of claim 419, the polynucleotide of claim 420, the cell of claim 421, the method of claims 422, 423 or claim 426, wherein the targeting agent is a CD 133 binding agent that is an antibody or antigen-binding fragment comprising a variable heavy (VH) chain and a variable light (VL) chain selected from a) a VH comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 314, 315, and 273, respectively, and a VL comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 275, 276, 277, respectively; (b) a VH comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 316, 317, and 282, respectively, and a VL comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 284, 285, and 286, respectively; (c) a VH comprising a a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 318, 319, and 300, respectively, and a VL comprising a CDR-L1, a CDR-L2, and a CDR- L3 comprising the amino acid sequences of SEQ ID NOs: 302, 303, and 304, respectively; (d) a VH comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 320, 321, and 309, respectively, and a VL comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 311, 312, and 313, respectively; and (e) a VH comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 322, 323, and 291, respectively, and a VL comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 293, 294, and 295, respectively.

429. The particle, the polynucleotide, the cell, or the method of claim 427 or claim 428, wherein the CD 133 binding agent is selected from a) a CD 133 binding agent comprising a heavy chain variable (VH) region comprising the amino acid sequence of SEQ ID NO: 270, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a light chain variable (VL) region comprising the amino acid sequence of SEQ ID NO: 274, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; b) a CD 133 binding agent comprising a VH region comprising the amino acid sequence of SEQ ID NO: 279, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a VL region comprising the amino acid sequence of SEQ ID NO: 283, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%,95%, 96%, 97%, 98%, or 99% sequence identity thereto; c) a CD133 binding agent comprising a VH region comprising the amino acid sequence of SEQ ID NO: 297, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a VL region comprising the amino acid sequence of SEQ ID NO: 301, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; d) a CD133 binding agent comprising a VH region comprising the amino acid sequence of SEQ ID NO: 306, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a VL region comprising the amino acid sequence of SEQ ID NO: 310, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; e) a CD 133 binding agent comprising a VH region comprising the amino acid sequence of SEQ ID NO: 288, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a VL region comprising the amino acid sequence of SEQ ID NO: 292, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto.

430. The particle, the polynucleotide, the cell, or the method of any of claims 427-429, wherein the CD 133 binding agent is a scFv.

431. The particle, the polynucleotide, the cell, or the method of any of claims 427-430, wherein the CD 133 binding agent comprises the amino acid sequence set forth in any one of SEQ ID NOs: 269, 278, 287, 296 or 305, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto.

432. The particle of claim 24, the polynucleotide of claim 114, the cell of claim 189, the method of claims 251 or claim 425, wherein the targeting agent is a CD117 binding agent that is a VHH comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 324-374, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto.

433. The particle, the polynucleotide, the cell or the method of claim 432, wherein the CD117 binding agent comprises the amino acid sequence set froth in any one of SEQ ID NOS: 324-374.

434. An anti-CDl 17 binding agent comprising a VHH single domain antibody comprising a CDR-H1, a CDR-H2, and a CDR-H3 contained within an amino acid sequence selected from the group consisting of SEQ ID NOs: 324-374.

435. An anti-CDl 17 binding agent comprising a VHH single domain antibody comprising an amino acid sequence selected from the group consisting of SEQ ID NOs: 324-374, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto.

436. The anti-CDl 17 binding agent of claim 434 or claim 435, wherein the VHH comprises the amino acid sequence set forth in any one of SEQ ID NOS: 324-374.

437. A viral fusion protein comprising a viral envelope protein and the anti-CDl 17 binding agent of any one of claims 434-436.

438. A viral fusion protein comprising a viral envelope protein and an anti-CDl 33 binding agent, wherein the anti-CDl 33 binding agent is an antibody or antigen-binding fragment comprising a variable heavy (VH) chain and a variable light (VL) chain selected from a) a VH comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 314, 315, and 273, respectively, and a VL comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 275, 276, 277, respectively; (b) a VH comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 316, 317, and 282, respectively, and a VL comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 284, 285, and 286, respectively; (c) a VH comprising a a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 318, 319, and 300, respectively, and a VL comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 302, 303, and 304, respectively; (d) a VH comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 320, 321, and 309, respectively, and a VL comprising a CDR- Ll, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 311, 312, and 313, respectively; and (e) a VH comprising a CDR-H1, a CDR-H2, and a CDR-H3 comprising the amino acid sequences of SEQ ID NOs: 322, 323, and 291, respectively, and a VL comprising a CDR-L1, a CDR-L2, and a CDR-L3 comprising the amino acid sequences of SEQ ID NOs: 293, 294, and 295, respectively.

439. The viral fusion protein of claim 438, wherein the CD133 binding agent is selected from a) a CD 133 binding agent comprising a heavy chain variable (VH) region comprising the amino acid sequence of SEQ ID NO: 270, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a light chain variable (VL) region comprising the amino acid sequence of SEQ ID NO: 274, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; b) a CD133 binding agent comprising a VH region comprising the amino acid sequence of SEQ ID NO: 279, or an aminoacid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a VL region comprising the amino acid sequence of SEQ ID NO: 283, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; c) a CD 133 binding agent comprising a VH region comprising the amino acid sequence of SEQ ID NO: 297, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a VL region comprising the amino acid sequence of SEQ ID NO: 301, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; d) a CD133 binding agent comprising a VH region comprising the amino acid sequence of SEQ ID NO: 306, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a VL region comprising the amino acid sequence of SEQ ID NO: 310, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; e) a CD133 binding agent comprising a VH region comprising the amino acid sequence of SEQ ID NO: 288, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto; and a VL region comprising the amino acid sequence of SEQ ID NO: 292, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto.

440. The viral fusion protein of claim 438 or claim 439, wherein the CD133 binding agent is a scFv.

441. The viral fusion protein of any of claims 438-440, wherein the CD133 binding agent comprises the amino acid sequence set forth in any one of SEQ ID NOs: 269, 278, 287, 296 or 305, or an amino acid sequence having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto.

442. The viral fusion protein of any of claims 437-441, wherein the viral envelope protein is a VSV-G protein or a functional variant thereof.

443. The viral fusion protein of any of claims 437-441, wherein the viral envelope protein is a Cocal virus G protein or a functional variant thereof.

444. The viral fusion protein of any of claims 437-441, wherein the viral envelope protein is an Alphavirus fusion protein (e.g., Sindbis virus) or a functional variant thereof.

445. The viral fusion protein of any of claims 437-441, wherein the viral envelope protein is a Paramyxovirus fusion protein (e.g., a Morbillivirus or Henipavirus) or a functional variant thereof.

446. The viral fusion protein of any of claims 437-441 and 445, wherein the viral envelope protein is a Morbillivirus fusion protein (e.g., measles virus (MeV), canine distemper virus, Cetacean morbillivirus, Peste-des-petits-ruminants virus, Phocine distemper virus, or Rinderpest virus) or a functional variant thereof.

447. The viral fusion protein of any of claims 437-441 and 445, wherein the viral envelope protein is a Henipavirus fusion protein (e.g., Nipah virus, Hendra virus, Cedar virus, Kumasi virus, Mojiang virus, or Langya virus) or a functional variant thereof.

448. The viral fusion protein of any of claims 437-441, 445 and 447, wherein the viral envelope protein is a Nipah virus fusion protein or a functional variant thereof.

449. The viral fusion protein of any of claims 437-441, 445, 447 and 448, wherein the viral envelope protein comprises a Nipah virus G glycoprotein (NiV-G) or a biologically active portion thereof.

450. The viral fusion protein of claim 449, wherein the binding agent is fused to the C- terminus of the NiV-G protein or biologically active portion thereof.

451. The viral fusion protein of claim 449 or claim 450, wherein the NiV-G or biologically active portion thereof is a wild-type NiV-G protein or a functionally active variant or biologically active portion thereof.

452. The viral fusion protein of any of claims 449-451, wherein the NiV-G protein or biologically active portion thereof is truncated and lacks up to 40 contiguous amino acid residues at or near the N-terminus of the wild-type NiV-G protein set forth in SEQ ID NO:4.

453. The viral fusion protein of any of claims 449-452, wherein the NiV-G protein or biologically active portion thereof is a biologically active portion that is a truncated NiV-G that has a deletion of amino acids 2-34 at or near the N-terminus of wild-type NiV-G set forth in SEQ ID NO:4.

454. The viral fusion protein of any of claims 449-453, wherein the NiV-G protein or biologically active portion thereof does not include an initial methionine.

455. The viral fusion protein of any of claims 449-454, wherein the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO: 42 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO:42.

456. The viral fusion protein of any of claims 449-455, wherein the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO:42.

457. The viral fusion protein of any of claims 437-456, wherein the viral envelope protein comprises one or more modifications to reduce binding to its native receptor.

458. The viral fusion protein of any of claims 449-457, wherein the NiV-G protein or biologically active portion thereof is a mutant NiV-G protein or biologically active portion thereof that exhibits reduced binding to Ephrin B2 or Ephrin B3.

459. The viral fusion protein of any of claims 449-458, wherein the NiV-G protein or biologically active portion thereof comprises one or more amino acid substitutions corresponding to amino acid substitutions selected from the group consisting of E501A, W504A, Q530A, and E533A with reference to numbering set forth in SEQ ID NO:4.

460. The viral fusion protein of any of claims 449-459, wherein the NiV-G protein or biologically active portion thereof has the amino acid sequence set forth in SEQ ID NO: 17 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 17.

461. The viral fusion protein of any of claims 449-460, wherein the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 17.

462. The viral fusion protein of any of claims 449-461, wherein the NiV-G protein or the biologically active portion has the amino acid sequence set forth in SEQ ID NO: 18 or a sequence of amino acids that exhibits at least at or about 80%, 85%, 90%, or 95% sequence identity to the sequence set forth in SEQ ID NO: 18.

463. The viral fusion protein of any of claims 449-462, wherein the NiV-G protein or biologically active portion thereof is set forth in SEQ ID NO: 18.

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