Delivery particles and uses thereof

Abstract

Provided herein are compositions of lipid delivery particles used to deliver a payload to a target cell and methods of producing the like. The lipid delivery particles can comprise a coiled-coil peptide pair, plasma membrane recruitment element, lipid membrane, envelope protein, or any combination thereof. The lipid delivery particles can be used to introduce one or more edits in a cell. Disclosed herein, in some aspects, are nucleic acid molecules encoding the same.

Description

WSGR Docket No.:62697-750.601DELIVERY PARTICLES AND USES THEREOFCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. Provisional Patent Application No. 63 / 730,837, filed December 11, 2024, U.S. Provisional Patent Application No. 63 / 730,840, filed December 11, 2024, U.S. Provisional Patent Application No. 63 / 805,885, filed May 14, 2025, and U.S. Provisional Patent Application No. 63 / 805,929, filed May 14, 2025, each of which is incorporated herein by reference in its entirety.BACKGROUND

[0002] Delivery of therapeutic payloads into the cells has been a significant challenge in drug development because payloads, such as proteins, cannot freely diffuse across the cell membrane. Although viral based constructs have been developed to deliver therapeutic payloads, these constructs often have low efficacy and / or create considerable side effects. There is a need for improved delivery of payloads into cells.SUMMARY

[0003] In some aspects, provided herein are In some aspects, provided herein are lipid delivery particles comprising: (a) a lipid membrane encapsulating a cavity; (b) an envelope protein on the lipid membrane; (c) a plasma membrane recruitment element; (d) a payload; and (e) a coiled-coil peptide pair, and wherein the coiled-coil peptide pair comprises a first coiled-coil peptide and a second coiled-coil peptide, wherein the first coiled-coil peptide and the second coiled-coil peptide comprise sequences having at least 80%, at least 85%, at least 90%, or at least 95%, or 100% sequence identity to the amino acid sequences set forth in: SEQ ID NOs: 601 and 602, respectively, or SEQ ID NOs: 602 and 601, respectively; SEQ ID NOs: 603 and 604, respectively, or SEQ ID NOs: 604 and 603, respectively; SEQ ID NOs: 605 and 606, respectively, or SEQ ID NOs: 606 and 605, respectively; SEQ ID NOs: 607 and 608, respectively, or SEQ ID NOs: 608 and 607, respectively; SEQ ID NOs: 609 and 610, respectively, or SEQ ID NOs: 610 and 609, respectively; SEQ ID NOs: 611 and 612, respectively, or SEQ ID NOs: 612 and 611, respectively; SEQ ID NOs: 613 and 614, respectively, or SEQ ID NOs: 614 and 613, respectively; SEQ ID NOs: 615 and 616, respectively, or SEQ ID NOs: 616 and 615, respectively; SEQ ID NOs: 616 and 621, respectively, or SEQ ID NOs: 621 and 616, respectively; or SEQ ID NOs: 617 and 618, respectively, or SEQ ID NOs: 618 and 617, respectively.

[0004] In some embodiments, the payload comprises a gene-editing agent. In some embodiments, the gene-editing agent comprises a zinc finger (ZF), transcription activator-like effector (TALE), and / or CRISPR-based genome editing or modulating protein; a nucleic acid encoding a zinc finger (ZF), transcription activator-like effector (TALE), and / or CRISPR-based genome editing or modulating protein, a ribonucleoprotein complex (RNP) comprising a CRISPR-based genome editing or modulating protein,WSGR Docket No.:62697-750.601 or any combination thereof. In some embodiments, the gene-editing agent comprises a base editor, a prime editor, or an epigenetic editor.

[0005] In some embodiments, the payload comprises one or more Cas proteins. In some embodiments, the one or more Cas proteins comprise a Cas9 protein, a Cas 12a protein, or any combination thereof. In some embodiments, the payload further comprises one or more guide RNA molecules (gRNAs).

[0006] In some aspects, provided herein are lipid delivery particles comprising: (a) a lipid membrane encapsulating a cavity; (b) an envelope protein on the lipid membrane; (c) a plasma membrane recruitment element; (d) a payload comprising a base editor; and (e) a coiled-coil peptide pair, and wherein the coiled-coil peptide pair comprises a first coiled-coil peptide and a second coiled-coil peptide.

[0007] In some embodiments, the coiled-coil peptide pair comprises a first coiled-coil peptide and a second coiled-coil peptide, wherein the first coiled-coil peptide and the second coiled-coil peptide comprise sequences having at least 80%, at least 85%, at least 90%, or at least 95%, or 100% sequence identity to the amino acid sequences set forth in: SEQ ID NOs: 601 and 602, respectively, or SEQ ID NOs: 602 and 601, respectively; SEQ ID NOs: 603 and 604, respectively, or SEQ ID NOs: 604 and 603, respectively; SEQ ID NOs: 605 and 606, respectively, or SEQ ID NOs: 606 and 605, respectively; SEQ ID NOs: 607 and 608, respectively, or SEQ ID NOs: 608 and 607, respectively; SEQ ID NOs: 609 and 610, respectively, or SEQ ID NOs: 610 and 609, respectively; SEQ ID NOs: 611 and 612, respectively, or SEQ ID NOs: 612 and 611, respectively; SEQ ID NOs: 613 and 614, respectively, or SEQ ID NOs: 614 and 613, respectively; SEQ ID NOs: 615 and 616, respectively, or SEQ ID NOs: 616 and 615, respectively; SEQ ID NOs: 616 and 621, respectively, or SEQ ID NOs: 621 and 616, respectively; SEQ ID NOs: 617 and 618, respectively, or SEQ ID NOs: 618 and 617, respectively; or SEQ ID NOs: 613 and 619, respectively, or SEQ ID NOs: 619 and 613, respectively.

[0008] In some aspects, provided herein are lipid delivery particles comprising: (a) a lipid membrane encapsulating a cavity; (b) an envelope protein on the lipid membrane; (c) a plasma membrane recruitment element; (d) a payload; and (e) a coiled-coil peptide pair, and wherein the coiled-coil peptide pair comprises (i) a first coiled-coil peptide comprising a heptad repeat of an amino sequence as set forth in (EXiX2X3X4X5X6)m, where Xi, X2, X3, X4, X5, and X„ are each independently any amino acid residue, and wherein m is an integer greater than or equal to 2, and (ii) a second coiled-coil peptide comprising a heptad repeat of an amino sequence as set forth in (KXyXsXgXioXnXi^n, where X7, Xs, X>, Xw, Xu, and Xi2are each independently any amino acid residue, and wherein n is an integer greater than or equal to 2.

[0009] In some embodiments, Xi or X4 is a hydrophobic amino acid residue, or Xi and X4 are each independently a hydrophobic amino acid residue. In some embodiments, Xi is isoleucine and X4 is leucine, or wherein Xi is leucine and X4 is isoleucine. In some embodiments, X7 or Xw is a hydrophobic amino acid residue, or X7 and Xw are each independently a hydrophobic amino acid residue. In some embodiments, X7 is isoleucine and Xw is leucine, or wherein X7 is leucine and Xw is isoleucine.

[0010] In some embodiments, X7, Xs, Xw, Xu, and X12 are each independently a polar amino acid residue or a charged amino acid residue. In some embodiments, X7, Xs, Xw, Xu, and X12 are each independently an amino acid residue selected from the group consisting of serine, threonine, cysteine,WSGR Docket No.:62697-750.601 asparagine, glutamine, tyrosine, glutamate, aspartate, arginine, and lysine. In some embodiments, X7, Xs, X10, Xu, and X12 are each independently a polar amino acid residue. In some embodiments, X7, Xs, X10, Xu, and X12 are each independently an amino acid residue selected from the group consisting of serine, threonine, cysteine, asparagine, glutamine, and tyrosine. In some embodiments, X7, Xs, Xw, Xu, and X12 are each independently a charged amino acid residue. In some embodiments, X7, Xs, X10, Xu, and X12 are each independently an amino acid residue selected from the group consisting of glutamate, aspartate, arginine, and lysine.

[0011] In some embodiments, m is 3 or 4. In some embodiments, n is 3 or 4. In some embodiments, m and n are each independently 3 or 4.

[0012] In some embodiments, the payload comprises a gene-editing agent. In some embodiments, the gene-editing agent comprises a zinc finger (ZF), transcription activator-like effector (TALE), and / or CRISPR-based genome editing or modulating protein; a nucleic acid encoding a zinc finger (ZF), transcription activator-like effector (TALE), and / or CRISPR-based genome editing or modulating protein, a ribonucleoprotein complex (RNP) comprising a CRISPR-based genome editing or modulating protein, or any combination thereof. In some embodiments, the gene-editing agent comprises a base editor, a prime editor, or an epigenetic editor.

[0013] In some embodiments, the payload comprises one or more Cas proteins. In some embodiments, the one or more Cas proteins comprise a Cas9 protein, a Cas 12a protein, or any combination thereof. In some embodiments, the payload further comprises one or more guide RNA molecules (gRNAs).

[0014] In some aspects, provided herein are lipid delivery particles comprising: (a) a lipid membrane encapsulating a cavity; (b) an envelope protein on the lipid membrane; (c) a plasma membrane recruitment element; (d) a payload; and (e) a coiled-coil peptide pair, and wherein the coiled-coil peptide pair comprises (i) a first coiled-coil peptide comprising one or more of a first heptad motif in tandem, the first heptad motif comprising amino acid residues of the formula (a-b-c-d-e-f-g)x, where a or d is a hydrophobic amino acid residue, or a and d are each independently a hydrophobic amino acid residue, where b, c, e, f, g, or any combination thereof is a polar-charged amino acid residue, and wherein x is an integer greater than or equal to 2, and (ii) a second coiled-coil peptide comprising one or more of a second heptad motif in tandem, the second heptad motif comprising amino acid residues of the formula (h-i-j-k-l-m-n)y, where h or k is a hydrophobic amino acid residue, or h and k are each independently a hydrophobic amino acid residue, where b, c, e, f, g, or any combination thereof is a polar-charged amino acid residue, and wherein y is an integer greater than or equal to 2.

[0015] In some embodiments, a and / or d is alanine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, valine, or any combination thereof. In some embodiments, a is isoleucine and d is leucine, or wherein d is leucine and a is isoleucine.

[0016] In some embodiments, b, c, e, f, or g are each independently a polar amino acid residue or a charged amino acid residue. In some embodiments, b, c, e, f, or g are each independently an amino acid residue selected from the group consisting of serine, threonine, cysteine, asparagine, glutamine, tyrosine, glutamate, aspartate, arginine, and lysine. In some embodiments, b, c, e, f, or g are each independently aWSGR Docket No.:62697-750.601 polar amino acid residue. In some embodiments, b, c, e, f, or g are each independently an amino acid residue selected from the group consisting of serine, threonine, cysteine, asparagine, glutamine, and tyrosine. In some embodiments, b, c, e, f, or g are each independently a charged amino acid residue. In some embodiments, b, c, e, f, or g are each independently an amino acid residue selected from the group consisting of glutamate, aspartate, arginine, and lysine.

[0017] In some embodiments, h and / or k is alanine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, valine, or any combination thereof. In some embodiments, h is isoleucine and k is leucine, or wherein k is leucine and h is isoleucine.

[0018] In some embodiments, i, j, 1, m, or n are each independently a polar amino acid residue or a charged amino acid residue. In some embodiments, i, j, 1, m, or n are each independently an amino acid residue selected from the group consisting of serine, threonine, cysteine, asparagine, glutamine, tyrosine, glutamate, aspartate, arginine, and lysine. In some embodiments, i, j, 1, m, or n are each independently a polar amino acid residue. In some embodiments, i, j, 1, m, or n are each independently an amino acid residue selected from the group consisting of serine, threonine, cysteine, asparagine, glutamine, and tyrosine. In some embodiments, i, j, 1, m, or n are each independently a charged amino acid residue. In some embodiments, i, j, 1, m, or n are each independently an amino acid residue selected from the group consisting of glutamate, aspartate, arginine, and lysine.

[0019] In some embodiments, x is an integer from 2 to 8. In some embodiments, y is an integer from 2 to 8. In some embodiments, x and y are each independently an integer from 2 to 8.

[0020] In some embodiments, the payload comprises a gene-editing agent. In some embodiments, the gene-editing agent comprises a zinc finger (ZF), transcription activator-like effector (TALE), and / or CRISPR-based genome editing or modulating protein; a nucleic acid encoding a zinc finger (ZF), transcription activator-like effector (TALE), and / or CRISPR-based genome editing or modulating protein, a ribonucleoprotein complex (RNP) comprising a CRISPR-based genome editing or modulating protein, or any combination thereof. In some embodiments, the gene-editing agent comprises a base editor, a prime editor, or an epigenetic editor.

[0021] In some embodiments, the payload comprises one or more Cas proteins. In some embodiments, the one or more Cas proteins comprise a Cas9 protein, a Cas 12a protein, or any combination thereof. In some embodiments, the payload further comprises one or more guide RNA molecules (gRNAs).

[0022] In some embodiments, a spatial orientation of the first coiled-coil peptide is parallel to a spatial orientation of the second coiled-coil peptide. In some embodiments, a spatial orientation of the first coiled-coil peptide is antiparallel to a spatial orientation of the second coiled-coil peptide.

[0023] In some embodiments, the coiled-coil peptide pair is selected from the group consisting of E4- K4, EE12RR345L-RR12EE345L, EE1234L-RR1234L, EE12345L-RR12345L, AcidPl -BasePl, P3(x3)- P4, SynZip2-SynZipl9, SynZip2-SynZipl, and N5-N6. In some embodiments, the first coiled-coil peptide is a E4 peptide, and the second coiled-coil peptide is a K4 peptide; or the first coiled-coil peptide is a K4 peptide, and the second coiled-coil peptide is a E4 peptide.WSGR Docket No.:62697-750.601

[0024] In some embodiments, the first coiled-coil peptide is coupled with the plasma membrane recruitment element. In some embodiments, the lipid delivery particle comprises a first chimeric protein comprising the first coiled-coil peptide fused with the plasma membrane recruitment element. In some embodiments, the first coiled-coil peptide is fused to the C-terminus of the plasma membrane recruitment element. In some embodiments, the first coiled-coil peptide is fused to the N-terminus of the plasma membrane recruitment element.

[0025] In some embodiments, the payload comprises a payload protein that is coupled with the second coiled-coil peptide. In some embodiments, the lipid delivery particle comprises a second chimeric protein comprising the second coiled-coil peptide fused with the payload protein. In some embodiments, the second coiled-coil peptide is fused to the C-terminus of the payload protein. In some embodiments, the second coiled-coil peptide is fused to the N-terminus of the payload protein.

[0026] In some embodiments, the plasma membrane recruitment element comprises an amino acid sequence having at least 80%, 85%, 90%, 95%, or 99% sequence identity to a sequence set forth in Table 4. In some embodiments, the plasma membrane recruitment element comprises an amino acid sequence set forth in Table 4. In some embodiments, the plasma membrane recruitment element comprises a pleckstrin homology (PH) domain. In some embodiments, the PH domain is from a protein selected from the group consisting of human phospholipase C51, human Aktl, human Aktl with E17K substitution, human 3-phosphoinositide-dependent protein kinase 1, human Daapl, mouse Grpl, human Grpl, human OSBP, human Btkl, human FAPP1, human CERT, human PKD, human PHLPP1, human SWAP70, and human MAPKAP 1. In some embodiments, the PH domain comprises a sequence set forth in any one of SEQ ID NOs: 5-14 and 23-48.

[0027] In some embodiments, the plasma membrane recruitment element comprises a gag polyprotein. In some embodiments, the gag polyprotein is a retroviral gag polyprotein. In some embodiments, the gag polyprotein is a gag polyprotein from human immunodeficiency virus (HIV), murine leukemia virus (MLV), Moloney murine leukemia virus (MMLV), Friend murine leukemia virus (FMLV), Baboon endogenous retrovirus (BaEV), Simian immunodeficiency virus (SIV), Rous sarcoma virus (RSV), human T-cell leukemia virus type-1 (HTLV), bovine leukemia virus (BLV), Feline Leukemia Virus (FeLV), Gibbon Ape Leukemia Virus (GaLV), Koala Retrovirus (KRV), Reticuloendotheliosis Virus (ReEV), Wooly Monkey Sarcoma Virus (WMSV), or a biologically active mutant thereof, or any combination thereof. In some embodiments, the gag polyprotein is a human endogenous retroviral gag polyprotein. In some embodiments, the gag polyprotein comprises a sequence having at least 80%, at least 85%, at least 90%, or at least 95%, or 100% sequence identity to the amino acid sequence set forth in any one of SEQ ID NOs: 285-305.

[0028] In some embodiments, the second chimeric protein further comprises a linker between the second coiled-coil peptide and the payload protein. In some embodiments, the linker is a cleavable linker. In some embodiments, the second chimeric protein further comprises a nuclear export signal (NES), a nuclear localization signal (NLS), or both. In some embodiments, the second chimeric protein comprises two or more NES. In some embodiments, the second chimeric protein comprises three NES. In someWSGR Docket No.:62697-750.601 embodiments, the NES is linked between the second coiled-coil peptide and the payload protein. In some embodiments, the NES and the second coiled-coil peptide are same side of the cleavage linker in the second chimeric protein. In some embodiments, the NLS and the payload protein are on same side of the cleavage linker in the second chimeric protein. In some embodiments, the NES comprises an amino acid sequence having at least 80%, 85%, 90%, 95%, or 99% sequence identity to a sequence set forth in Tables 11. In some embodiments, the NES comprises an amino acid sequence set forth in Tables 11. In some embodiments, the NLS comprises an amino acid sequence having at least 80%, 85%, 90%, 95%, or 99% sequence identity to a sequence set forth in Table 12. In some embodiments, the NLS comprises an amino acid sequence set forth in Table 12.

[0029] In some embodiments, a fusion of the coiled-coil peptide pair improves an editing efficiency of a target genetic locus in a cell contacted by the lipid delivery particle compared to an editing efficiency of the target genetic locus in the cell when contacted by an otherwise identical lipid delivery particle that does not comprise the coiled-coil peptide pair. In some embodiments, the editing efficiency of the target genetic locus in the cell contacted by the lipid delivery particle is at least about 30%. In some embodiments, the editing efficiency of the target genetic locus in the cell contacted by the lipid delivery particle is at least about 50%. In some embodiments, the editing efficiency of the lipid delivery particle is base editing percentage.

[0030] In some aspects, provided herein are lipid delivery particles comprising: (a) a lipid membrane encapsulating a cavity; (b) an envelope protein on the lipid membrane; and (c) a chimeric protein comprising a plasma membrane recruitment element and a payload protein, wherein the plasma membrane recruitment element comprises a gag polyprotein, wherein the chimeric protein comprises a nuclear export signal (NES) inside the gag protein, and wherein the payload protein does not comprise a reverse transcriptase.

[0031] In some embodiments, the chimeric protein comprises two or more NES. In some embodiments, the chimeric protein comprises three NES. In some embodiments, the NES is linked between the gag protein and the payload protein. In some embodiments, the chimeric protein further comprises a cleavage linker between the NES and the payload protein. In some embodiments, the chimeric protein further comprises a NLS, and wherein the NLS and the payload protein are both on N-terminal side or C-terminal side of the cleavage linker in the chimeric protein. In some embodiments, the chimeric protein comprises at least two NLS, optionally two NLS. In some embodiments, the at least two NLS are at both N-terminus and C-terminus of the payload protein. In some embodiments, the at least two NLS are at either N- terminus or C-terminus of the payload protein. In some embodiments, the NES comprises an amino acid sequence having at least 80%, 85%, 90%, 95%, or 99% sequence identity to a sequence set forth in Tables 11. In some embodiments, the NES comprises an amino acid sequence set forth in Tables 11. In some embodiments, the NLS comprises an amino acid sequence having at least 80%, 85%, 90%, 95%, or 99% sequence identity to a sequence set forth in Table 12. In some embodiments, the NLS comprises an amino acid sequence set forth in Table 12.WSGR Docket No.:62697-750.601

[0032] In some embodiments, the gag polyprotein is a retroviral gag polyprotein. In some embodiments, the gag polyprotein is a gag polyprotein from human immunodeficiency virus (HIV), murine leukemia virus (MLV), Moloney murine leukemia vims (MMLV), Friend murine leukemia virus (FMLV), Baboon endogenous retrovirus (BaEV), Simian immunodeficiency virus (SIV), Rous sarcoma vims (RSV), human T-cell leukemia vims type-1 (HTLV), bovine leukemia vims (BLV), Feline Leukemia Vims (FeLV), Gibbon Ape Leukemia Vims (GaLV), Koala Retrovims (KRV), Reticuloendotheliosis Vims (ReEV), Wooly Monkey Sarcoma Vims (WMSV), or a biologically active mutant thereof, or any combination thereof. In some embodiments, the gag polyprotein is a human endogenous retroviral gag polyprotein. In some embodiments, the gag polyprotein comprises a sequence having at least 80%, at least 85%, at least 90%, or at least 95%, or 100% sequence identity to the amino acid sequence set forth in any one of SEQ ID NOs: 285-305.

[0033] In some embodiments, the gag polyprotein lacks at least a fragment of a nucleocapsid protein, optionally lacks the full length of the nucleocapsid protein. In some embodiments, the plasma membrane recmitment element comprises a heterologous domain fused to the C-terminus of the gag polyprotein, optionally wherein the heterologous domain comprises a leucine zipper. In some embodiments, the heterologous domain further comprises a linker sequence flanking the N-terminus or C-terminus side of the leucine zipper. In some embodiments, the heterologous domain further comprises a linker sequence flanking on both N-terminus and C-terminus sides of the leucine zipper. In some embodiments, the linker sequence comprises at least one repeat of an amino acid sequence SGGS, the sequence of any one of SEQ ID NO: 343-346 or 673, optionally two repeats thereof.

[0034] In some embodiments, the gag polyprotein lacks at least fragment of a matrix protein, optionally lacks the full length of the matrix protein. In some embodiments, the plasma membrane recruitment element comprises a pleckstrin homology (PH) domain fused to the N-terminus of the gag polyprotein, optionally wherein the PH domain is from a protein selected from the group consisting of human phospholipase C51, human Aktl, human Aktl with E17K substitution, human 3-phosphoinositide- dependent protein kinase 1, human Daapl, mouse Grpl, human Grpl, human OSBP, human Btkl, human FAPP1, human CERT, human PKD, human PHLPP1, human SWAP70, and human MAPKAP1. In some embodiments, the PH comprises a sequence set forth in any one of SEQ ID NOs: 5-14 and 23-48.

[0035] In some aspects, provided herein are lipid delivery particles comprising: (a) a lipid membrane encapsulating a cavity; (b) an envelope protein on the lipid membrane; (c) a chimeric protein comprising a plasma membrane recruitment element and a payload protein, wherein the plasma membrane recruitment element comprises a gag polyprotein, and wherein: the gag polyprotein lacks at least a fragment of a nucleocapsid protein, and the plasma membrane recruitment element further comprises a heterologous domain fused to C-terminus of the gag polyprotein; and / or the gag polyprotein lacks at least a fragment of a matrix protein, and the plasma membrane recruitment element further comprises a pleckstrin homology (PH) domain fused to N-terminus of the gag polyprotein.

[0036] In some embodiments, the gag polyprotein comprises a matrix protein and a capsid protein, and lacks at least a fragment of a nucleocapsid protein, and the plasma membrane recruitment element furtherWSGR Docket No.:62697-750.601 comprises a heterologous domain within the gag polyprotein. In some embodiments, the gag polyprotein comprises a matrix protein and a capsid protein, and lacks at least a fragment of a nucleocapsid protein, and the plasma membrane recruitment element further comprises a heterologous domain fused to C- terminus of the gag polyprotein. In some embodiments, the heterologous domain comprises a leucine zipper. In some embodiments, the heterologous domain further comprises a linker sequence flanking the N-terminus or C-terminus side of the leucine zipper. In some embodiments, the heterologous domain further comprises a linker sequence flanking on both N-terminus and C-terminus sides of the leucine zipper. In some embodiments, the linker sequence comprises at least one repeat of an amino acid sequence SGGS, the sequence of any one of SEQ ID NO: 343-346 or 673, optionally two repeats thereof.

[0037] In some embodiments, the gag polyprotein is a retroviral gag polyprotein. In some embodiments, the gag polyprotein is a gag polyprotein from human immunodeficiency virus (HIV), murine leukemia virus (MLV), Moloney murine leukemia vims (MMLV), Friend murine leukemia virus (FMLV), Baboon endogenous retrovirus (BaEV), Simian immunodeficiency virus (SIV), Rous sarcoma vims (RSV), human T-cell leukemia vims type-1 (HTLV), bovine leukemia vims (BLV), Feline Leukemia Vims (FeLV), Gibbon Ape Leukemia Vims (GaLV), Koala Retrovims (KRV), Reticuloendotheliosis Vims (ReEV), Wooly Monkey Sarcoma Vims (WMSV), or a biologically active mutant thereof, or any combination thereof.

[0038] In some embodiments, the gag polyprotein is a human endogenous retroviral gag polyprotein. In some embodiments, the gag polyprotein comprises a sequence having at least 80%, at least 85%, at least 90%, or at least 95%, or 100% sequence identity to the amino acid sequence set forth in any one of SEQ ID NOs: 285-305.

[0039] In some embodiments, the chimeric protein further comprises a nuclear export signal (NES). In some embodiments, the chimeric protein comprises two or more NES. In some embodiments, the chimeric protein comprises three NES. In some embodiments, the NES is linked between the gag protein and the payload protein. In some embodiments, the chimeric protein further comprises a cleavage linker between the NES and the payload protein. In some embodiments, the chimeric protein further comprises a NLS, and wherein the NLS and the payload protein are on same side of the cleavage linker in the chimeric protein. In some embodiments, the chimeric protein comprises at least two NLS, optionally two NLS. In some embodiments, the at least two NLS are at both N-terminus and C-terminus of the payload protein. In some embodiments, the at least two NLS are at either N-terminus or C-terminus of the payload protein. In some embodiments, the NES comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99%, or 100% sequence identity to a sequence set forth in Tables 11. In some embodiments, the NLS comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99%, or 100% sequence identity to a sequence set forth in Table 12.

[0040] In some embodiments, the payload protein comprises a gene-editing agent. In some embodiments, the gene-editing agent comprises a zinc finger (ZF), transcription activator-like effector (TALE), and / or CRISPR-based genome editing or modulating protein; a nucleic acid encoding a zincWSGR Docket No.:62697-750.601 finger (ZF), transcription activator-like effector (TALE), and / or CRISPR-based genome editing or modulating protein, a ribonucleoprotein complex (RNP) comprising a CRISPR-based genome editing or modulating protein, or any combination thereof. In some embodiments, the gene-editing agent comprises a base editor, a prime editor, or an epigenetic editor.

[0041] In some embodiments, the payload protein comprises one or more Cas proteins. In some embodiments, the one or more Cas proteins comprise a Cas9 protein, a Cas 12a protein, or any combination thereof. In some embodiments, the lipid delivery particle further comprises one or more guide RNA molecules (gRNAs) as a payload within the cavity.

[0042] In some embodiments, the envelope protein comprises a VSV glycoprotein (VSV-G), a human immunodeficiency virus (HIV) GP160 glycoprotein, a Baboon Endogenous Retrovirus (BaEVTR) glycoprotein, a fusion protein of Vesicular stomatitis Indiana virus and Rabies virus glycoprotein (FuG- E), an ecotropic Murine Leukemia Virus envelope protein (MLV ENV ecotropic), a human T-cell lymphotropic virus type 1 (HTLV-1) glycoprotein, an amphotrophic Murine Leukemia Virus envelope protein (MLV ENV amphotropic), a Moloney murine leukemia virus 10A1 strain glycoprotein (MLV 10A1), a Baculovirus envelope glycoprotein (GP64), a pantropic MLV envelope protein, a xenotropic MLV envelope protein, a xenotropic murine leukemia virus (XMLV) envelope protein, a Moloney murine leukemia virus (MMLV) envelope protein, a Moloney murine sarcoma virus (MoMSVg) envelope protein, a simian endogenous type D retrovirus protein (RD-114), a gibbon ape leukemia virus (GALV) envelope protein, a feline leukemia virus (FLV) envelope protein, a mouse mammary tumor virus (MMTV) envelope protein, an avian leukosis virus envelope protein, a Rous Sarcoma virus envelope protein, or an endogenous feline virus envelope protein (RD114 ENV), or a mutant thereof.

[0043] In some embodiments, the envelope protein comprises a glycoprotein of a mammalian endogenous retrovirus or mutant thereof. In some embodiments, the glycoprotein of the mammalian endogenous retrovirus is a glycoprotein of a human endogenous retrovirus (hERV). In some embodiments, the glycoprotein of a hERV comprises a hENVHl, a hENVH2, a hENVH3, a hENVKl, a hENVK2, a hENVK3, a hENVK4, a hENVK5, a hENVK6, a hENVT, a hENVW, a hENVFRD, a hENVR, a hENVR(b), a hENVR(c) 1, a hENVR(c)2 or a hENVKcon, or a biologically active mutant thereof. In some embodiments, the hERV comprises a modified envelope protein. In some embodiments, the envelope protein comprises a non-viral envelope protein.

[0044] In some embodiments, the lipid delivery particle further comprises a gag / pro / pol polypeptide, wherein the gag / pro polyprotein comprises a fusion of a second polyprotein, a pro protein, and a pol polyprotein. In some embodiments, the lipid delivery particle further comprises: a first gag polyprotein, wherein the first gag polyprotein is not fused with a pro protein or a pol polyprotein; and a gag / pro polyprotein, wherein the gag / pro polyprotein comprises a fusion of a second gag polyprotein and a pro protein, and does not comprise and is not fused to a pol polyprotein. In some embodiments, the first gag polyprotein or the second gag polyprotein, or both are a retroviral gag polyprotein. In some embodiments, the first gag polyprotein or the second gag polyprotein, or both are a gag polyprotein from human immunodeficiency virus (HIV), murine leukemia virus (MLV), Moloney murine leukemia virusWSGR Docket No.:62697-750.601(MMLV), Friend murine leukemia virus (FMLV), Baboon endogenous retrovirus (BaEV), Simian immunodeficiency virus (SIV), Rous sarcoma virus (RSV), human T-cell leukemia virus type-1 (HTLV), bovine leukemia virus (BLV), Feline Leukemia Virus (FeLV), Gibbon Ape Leukemia Virus (GaLV), Koala Retrovirus (KRV), Reticuloendotheliosis Virus (ReEV), Wooly Monkey Sarcoma Virus (WMSV), or a biologically active mutant thereof, or any combination thereof.

[0045] In some embodiments, the first gag polyprotein or the second gag polyprotein, or both are a human endogenous retroviral gag polyprotein. In some embodiments, the first gag polyprotein or the second gag polyprotein, or both comprise a sequence having at least 80%, at least 85%, at least 90%, or at least 95%, or 100% sequence identity to the amino acid sequence set forth in any one of SEQ ID NOs: 285-305. In some embodiments, wherein the first gag polyprotein or the second gag polyprotein, or both lack at least a fragment of a nucleocapsid protein, optionally lack the full length of the nucleocapsid protein. In some embodiments, wherein the first gag polyprotein or the second gag polyprotein, or both are fused N-terminal to a heterologous domain, optionally wherein the heterologous domain comprises a leucine zipper.

[0046] In some embodiments, the heterologous domain further comprises a linker sequence flanking the leucine zipper on both N-terminus and C-terminus sides. In some embodiments, the gag / pro polyprotein further comprises the linker sequence flanking the pro polypeptide on its C-terminus. In some embodiments, the linker sequence comprises at least one repeat of an amino acid sequence SGGS, the sequence of any one of SEQ ID NO: 343-346 or 673, optionally two repeats thereof.

[0047] In some embodiments, the first gag polyprotein or the second gag polyprotein, or both lack at least fragment of a matrix protein, optionally lack the full length of the matrix protein. In some embodiments, the first gag polyprotein or the second gag polyprotein, or both are fused C-terminal to a pleckstrin homology (PH) domain, optionally wherein the PH domain is from a protein selected from the group consisting of human phospholipase C51, human Aktl, human Aktl with E17K substitution, human 3 -phosphoinositide-dependent protein kinase 1, human Daapl, mouse Grpl, human Grpl, human OSBP, human Btkl, human FAPP1, human CERT, human PKD, human PHLPP1, human SWAP70, and human MAPKAP1. In some embodiments, the PH domain comprises a sequence set forth in any one of SEQ ID NOs: 5-14 and 23-48.

[0048] In some embodiments, the lipid delivery particle does not comprise a pol polyprotein either standalone or in fusion. In some embodiments, the lipid delivery particle does not comprise a reverse transcriptase. In some embodiments, the lipid delivery particle does not comprise an integrase.

[0049] In some aspects, provided herein are methods for preparing a lipid delivery particle comprising a payload, the method comprising: (a) providing a producer cell that produces the lipid delivery particle described herein; and (b) collecting the lipid delivery particle from the producer cell.

[0050] In some embodiments, the method further comprises expressing in the producer cell a nucleic acid molecule encoding the plasma membrane recruitment element. In some embodiments, the producer cell comprises a nucleic acid sequence encoding the envelope protein. In some embodiments, the method further comprises expressing in the producer cell a nucleic acid molecule encoding the envelope protein.WSGR Docket No.:62697-750.601In some embodiments, the method further comprises culturing the producer cell in a medium and maintaining the producer cell under conditions sufficient to produce the lipid delivery particle. In some embodiments, the method further comprises harvesting the medium and purifying the lipid delivery particle. In some embodiments, the purifying retains the structural integrity of the lipid delivery particle.

[0051] In some aspects, provided herein are methods of introducing one or more edits in a cell, comprising contacting the cell with the lipid delivery particle described herein or the lipid delivery particle produced by the method described herein.

[0052] In some aspects, provided herein are methods of introducing two or more edits in a cell, comprising contacting the cell with the lipid delivery particle described herein or the lipid delivery particle produced by the method described herein.

[0053] In some embodiments, the two or more edits in the cell are to different gene targets.

[0054] In some aspects, provided herein are compositions comprising a nucleic acid molecule encoding the lipid delivery particle described herein.

[0055] In some embodiments, the nucleic acid molecule comprises (i) a first nucleotide sequence encoding the envelope protein; (ii) a second nucleotide sequence encoding the plasma membrane recruitment element; and (iii) a third nucleotide sequence encoding the first coiled-coil peptide; and (iv) a fourth nucleotide sequence encoding the second coiled-coil peptide.

[0056] In some aspects, provided herein are vectors comprising the composition described herein.

[0057] In some embodiments, the vector is a vector selected from a plasmid, a cosmid, a bacterial vector, a viral vector, or an artificial chromosome.

[0058] In some aspects, provided herein are cells comprising the lipid delivery particle described herein or the vector described herein.

[0059] In some aspects, provided herein are cells comprising the lipid delivery particle produced by the method described herein.

[0060] In some aspects, provided herein are pharmaceutical compositions comprising: the lipid delivery particle described herein or the composition described herein; and (b) a pharmaceutically acceptable carrier, excipient, or diluent.

[0061] In some aspects, provided herein are methods of treating a disease or condition in a subject in need thereof, the method comprising: administering to the subject in need thereof the lipid delivery particle described herein or the pharmaceutical composition described herein.

[0062] In some aspects, provided herein are kits comprising: (a) the lipid delivery particle described herein or the pharmaceutical composition described herein; and (b) instructions for use.INCORPORATION BY REFERENCE

[0063] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference.WSGR Docket No.:62697-750.601BRIEF DESCRIPTION OF THE DRAWINGS

[0064] The novel features of the disclosure are set forth with particularity in the appended claims. A better understanding of the features and advantages of the present disclosure will be obtained by reference to the following detailed description that sets forth illustrative embodiments, in which the principles of the disclosure are utilized, and the accompanying drawings of which:

[0065] FIGs. 1A-1C are schematic representations of constructs and their cleavage in various cell types. FIG. 1A (top) shows a standard cargo construct with sequences encoding a nuclear localization signal (NLS), a nuclear export signal (NES), group specific antigen (GAG), matrix (MA), pl2, capsid (CA), nucleocapsid (NC), and adenine base editor (ABE). In the standard cargo, the NES is located outside the Gag region. FIG. 1A (bottom) also shows a construct with an internal NES sequence located within the Gag region. The alternative placement of the NES leads to improved editing. FIG. IB is a schematic of different architectures that can be used to test different NLS and NES configurations. FIG. 1C is a schematic representation of cargo cleavage and transport. In producer cells, Gag-BE remains intact (the 3x NES tag is not cleaved), and the construct remains in the cytoplasm for assembly. During enveloped virus-like particle (eVLP) assembly, a protease is activated inside viral particles (optimal pH ~5) and cleaves the cargo in the same particle. The particle matures and is transduced to target cells. In recipient cells, the cleaved cargo (with no NES, only NLS), goes to the nucleus for gene editing.

[0066] FIGs. 2A-2D depict results of experiments assessing the editing efficiency of various constructs. FIGs. 2A and 2B show that having the NES (lx gRNA or 6x gRNA) placed within the Gag improves editing with an exemplary lipid delivery particle according to some embodiments of the present disclosure. FIGs. 2C and 2D show results of ELISA experiments, outlining that cas9 and p30 concentrations increase with Internal NES (lx gRNA); however, this result is not reflected with 6x gRNA.

[0067] FIGs. 3A-3C depict results of experiments assessing the editing efficiency of construct designs in HSPC. Each NVLP version on the x-axis depicts a different internal NES construct. FIGs. 3A-3B depict results with the original plasmid ratio, optimized for standard cargo. FIG. 3A and 3B show that internal NES and Kozak, as well as the combination, result in higher % B2M editing efficiencies than the standard. FIG. 3C depicts the various degrees of editing seen through ratio optimization of the Kozak + Internal NES construct.

[0068] FIGs. 4A-4C show different types of cargo loading. FIG. 4A shows the standard method of cargo loading into the exemplary lipid delivery particles by fusing the cargo directly to Gag. FIG. 4B shows the alternative coiled-coil (CC) method of cargo loading by fusing one half of the coiled-coil to Gag -Pol, and then fusing its binding partner to the cargo. FIG. 4C outlines the different combinations of cargo that can be used to achieve different editing outcomes.

[0069] FIG. 5 depicts the results of using a coiled-coiled system on loading cargo. FIG. 5 shows that using an E4-K4 CC domain CC to load less efficient ABE8e into the particles improves efficiency compared to P3-P4. The lines to the right in each condition depict 50 pL, while the ones to the left depict 5 pL.WSGR Docket No.:62697-750.601

[0070] FIGs. 6A-6B depict the results of experiments using enAsCasl2a for editing. enAsCasl2a shows higher editing with E4+K4. enAsCasl2a has lower efficacy than Cas9 with standard construct. The lines to the right in each condition depict 10 pL, while the ones to the left depict 50 pL.

[0071] FIGs. 7A-7B depict the results of loading Open CRISPR-1 (OC-1) using K3 / K4 in the exemplary lipid delivery particle. For both sites, ABE and Cas9, OC-1 performs slightly less efficiently than SpCas9. Addition of the coiled-coil increases the OC-1 performance to SpCas9 levels or higher. The lines to the right in each condition depict 50 pL, while the ones to the left depict 10 pL. FIG. 7A shows data for B2M, while FIG. 7B shows data for HEKs3.

[0072] FIGs. 8A-8D show further results confirming the trend in higher editing of the E4+K4 coiled-coil construct compared to others. ABE8e CC loading was used to compare editing efficiencies. In FIG. 8A, the lines to the right in each condition depict 50 pL, while the ones to the left depict 5 pL. In FIG. 8B, the lines to the right in each condition depict 50 pL, while the ones to the left depict 10 pL. In FIG. 8C, the lines to the right in each condition depict 50 pL, while the ones to the left depict 5 pL. In FIG. 8D, the lines to the right in each condition depict 50 pL, while the ones to the left depict 10 pL.

[0073] FIGs. 9A-9B show schematics of different Gag designs. FIG. 9A shows a Gag -Pol construct. A read-through mechanism ensures that a mixture of Gag (the majority) and Gag -Pol (roughly 5%) will be made from Gag-Pol construct. FIG. 9B shows a Gag / Gag-Pro Design with two plasmids, and without an RT / IN function.

[0074] FIG. 10 outlines the results of Gag / Gag-Pro ratio optimization. At 5% Gag-Pro content in total scaffold editing efficiency for all cargo constructs was the highest. For standard cargo, at 5% of total scaffold Gag-Pro achieved similar editing to standard scaffold (in MLV particles Gag-Pol accounts for 5% of all Gag -containing proteins). The lines to the right in each condition depict 10 pL, while the ones to the left depict 50 pL.

[0075] FIG. 11 depicts the results of experiments showing that a higher Gag -Pro ratio leads to lower Cas9 concentration.

[0076] FIGs. 12A-12D shows a schematic overview of Gag engineering to replace the nucleocapsid region. FIG. 12A shows four constructs. The top two constructs depict replacing an NC region with a leucine zipper (Zip) region in a Gag-Base Editor (BE) construct. The bottom two constructs show doing the same with Gag-Pol. Other domains include matrix (MA), pl2, capsid (CA), protease (PR), reverse transcriptase (RT / RH) and integrase (IN). FIG. 12B shows experimental results of testing the editing efficiency of various constructs. The lines to the right in each condition depict 10 pL, while the ones to the left depict 50 pL.

[0077] FIG. 12C shows the ELISA Cas9 concentration with various NC replacements. FIG. 12D shows the ELISA p30 concentration with various NC replacements. Cas9 concentration for Gag-cut site-Zip- Cargo eVLPs was similar to the standard eVLP with half the p30 amount, which could indicate improved cargo loading.

[0078] FIGs. 13A-13D show a schematic overview of Gag truncations. FIG. 13A shows a Gag-BE construct with a myristoylation signal, a PPPY domain, a protease cut site and a readthrough codon. FIG.WSGR Docket No.:62697-750.60113B shows the editing efficiencies of various Gag truncations. The lines to the right in each condition depict 10 pL, while the ones to the left depict 50 pL. FIG. 13C shows the ELISA Cas9 concentration with various truncations. FIG. 13D shows the ELISA p30 concentration with various truncations.

[0079] FIGs. 14A-14E outline the optimization of linkers in various constructs. FIG. 14A shows adding linkers to flank the leucine zipper in Gag-Zip-Pol constructs improves Cas9 processing and editing efficiency. FIG. 14A shows a schematic of the constructs with Gag-Zip-BE with / without linkers to the left, and Gag-Zip-Pol with / without linkers to the right. FIG. 14B shows the editing efficiency of different constructs; the lines to the right in each condition depict 50 pL, while the ones to the left depict 10 pL. FIG. 14C shows the Cas9 processing. Gag-dNC-2xSGGS-Zip-2xSGGS-Pol denotes the linker-containing constructs. FIG. 14D shows a schematic of the constructs with Gag-Zip with / without linkers (top), and Gag-Zip-Pro with / without linkers (middle), and Gag-Zip-Pro with an additional linker to the right of the PR (bottom). FIG. 14E shows the editing efficiency of different Gag-Zip-Pro constructs; the lines to the right in each condition depict 50 pL, while the ones to the left depict 10 pL.

[0080] In some embodiments, adding a linker at different locations within the constructs mentioned herein may improve editing efficiency. Specifically, adding a 2x SGGS linker on either side of the leucine zipper improved Cas9 processing and editing efficiency in Gag-Zip-Pol constructs (FIGs. 14A-14C) and Gag-Zip-Pro constructs (FIGs. 14D-14E).

[0081] FIG. 15 shows the results of a coiled-coil screen using OC-1 targets and BCL1 la editing. For each coiled-coil (CC) of the screen, the left bar represents the low dose (10 pl) and the right bar represents the high dose (50 pl).

[0082] FIGs. 16A-16C show single particle analysis and cargo loading. FIG. 16A shows a schematic of single particle analysis. FIG. 16B shows Cas9 loading with and without coiled-coil (CCs). FIG. 16C shows cargo loading fold change using CC with diverse cargo over CC with non-diverse cargo.

[0083] FIGs. 17A-17D show evaluation of E4 / K4 loading gene editors. Each line graph shows editing potency assessment with and without CCs for a cargo at a target site. FIG. 17A shows editing between standard delivery particle without CC (Standard) and delivery particle described herein with CC (DLVR- X-CC). Cargo was enAsCasl2a. FIG. 17B shows editing between Standard particles and DLVR-X-CC particles with ABE8.8 editor (solid lines) or ABE8.20 (dotted lines). FIG. 17C shows editing between Standard particles and DLVR-X-CC particles with CBE6b cargo. FIG. 17D shows editing between Standard particles and DLVR-X-CC particles with PE6d cargo.

[0084] FIGs. 18A-18C are line graphs of indels by dose for SpCas9 and OC-1 at various targets. FIG. 18A shows indels at BCL1 la, FIG. 18B shows indels at B2M, and FIG. 18C shows indels at HEKs3.

[0085] FIGs. 19A-19H show line graphs of indels and A>G at various targets editing with OC-1 Cas9 and base editing with and without CC. FIGs. 19A-19D show indels at BCL1 la (FIG. 19A), B2M (FIG. 19B), VEGFAs3 (FIG. 19C), and HEKs3 (FIG. 19D) between standard delivery particles without CC (Standard OC-1), delivery particles described herein with CC (DLVR-X-CC OC-1), and standard SpCas9. FIGs. 19E-19H show A>G editing at BCL1 la (FIG. 19E), B2M (FIG. 19F), VEGFAs3 (FIG. 19G), andWSGR Docket No.:62697-750.601HEKs3 (FIG. 19H) between standard delivery particles without CC (Standard OC-1 ABE), delivery particles described herein with CC (DLVR-X-CC OC-1 ABE), and standard ABE.

[0086] FIGs. 20A-20C show gene editing efficiencies of standard particles (Standard) and engineered delivery particles described herein with CC (DLVR-X-CC). Each figure shows dose responses in various primary cell types for editing with and without CC. FIG. 20A shows editing efficiency in non-human primate hepatocytes (NHP hepatocytes). FIG. 20B shows editing efficiency in human cortical neurons. FIG. 20C shows editing efficiency in hematopoietic stem and progenitor cells (HSPCs).

[0087] FIGs. 21A-21D show results of PCSK9 editing and results of cargo loading with and without CC. FIG. 21A shows editing efficiency of mPCSK9 A>G in N2A cells between standard particles (Standard) and engineered delivery particles described herein with CC (DLVR-X-CC). FIG. 21B shows cargo loading improvement, measured as ratio of Cas9 / p30, between Standard particles and DLVR-X-CC particles. FIG. 21C shows base editing in mouse liver (mPCSK9 A>G) between Standard particles and DLVR-X-CC particles. *indicates P < 0.05. FIG. 21D shows base editing in mouse spleen (mPCSK9 A>G) between Standard particles and DLVR-X-CC particles. For the results in FIGs. 21B-21D, the dose per mouse was 1.5e5 ng p30.

[0088] FIGs. 22A-22D show results of PCSK9 editing and the p30 titer using particles with minimal viral components with and without CC. FIG. 22A shows editing efficiency of mPCSK9 A>G in N2A cells between standard particles (Standard), engineered delivery particles described herein without the engineered CC (DLVR-X v2a), and engineered delivery particles described herein with the engineered CC (DLVR-X v2a CC). FIG. 22B shows comparison of p30 titer between Standard particles, DLVR-X v2a particles, and DLVR-X v2a CC particles. FIG. 22C shows base editing in mouse liver (mPCSK9 A>G) between Standard particles, DLVR-X v2a particles, and DLVR-X v2a CC particles. *indicates P < 0.05. FIG. 22D shows base editing in mouse spleen (mPCSK9 A>G) between Standard particles, DLVR- X v2a particles, and DLVR-X v2a CC particles. For the results in FIGs. 22B-22D, the dose per mouse was 1.5e5 ng p30.

[0089] FIG. 23 is a pie chart showing premature Cas9 cleavage in producer cells with various Gag -Pro: Gag ratios. The uncleaved / cleaved ratio was calculated using peak height in cleavage assay. The dark gray shows premature release, and the light gray shows unreleased. The increased Gag-Pro : Gag ratio led to greater premature release compared to that seen in standard condition.

[0090] FIG. 24 shows editing efficiency (measured as percentage of BCL1 la editing) in 293T cells with various Gag-Pro: Gag ratios. For each group of bars in the graph, the left bar represents the low rose (10 pl) and the right bar represents the high dose (50 pl).

[0091] FIGs. 25A-25D show results of varying cargo amount and gag plasmid amount on cargo / capsid ratio and editing. FIGs. 25A-25B show results of cargo / capsid ratio for delivery particles DLVR-X-v2a (FIG. 25A) and DLVR-X v2b (FIG. 25B), with increasing cargo plasmid amount and decreasing gag plasmid amount. FIGs. 25C-25D show results of editing efficiency for delivery particles DLVR-X-v2a (FIG. 25C) and DLVR-X v2b (FIG. 25D), with increasing cargo plasmid amount and decreasing gag plasmid amount. Editing efficiency was measured as percentage of B2M negative targets (% B2MWSGR Docket No.:62697-750.601 negative). For FIGs. 25C-25D, the left bar in each bar grouping represents the low dose (10 pl) and the right bar in each bar grouping represents the high dose (50 pl).

[0092] FIGs. 26A-26H show graphs of the results for editing at mPCSK9 and cargo / capsid ELISAs for the DLVR-X v2 particles as well as the DLVR-X v2a-CC particles. FIGs. 26A-26D show results of the DLVR-X v2 particles and FIGs. 26E-26H show results of the DLVR-X v2a-CC particles (e.g., comprising the coiled-coil peptides described herein). FIGs. 26A-26B and FIGs. 26E-26F show results of increasing dose on editing efficiency of mPCSK9 between tested delivery particles. FIGs. 26C-26D and FIGs. 26G-26H show results of amount of cargo and capsid between tested delivery particles, as measured via ELISA assay.

[0093] FIG. 27 shows in vivo potency in mouse liver for the DLVR-X v2 particles: mPCSK9 editing (left) and mL production (right). The dose per mouse for standard delivery particles (STD), DLVR-X v2a delivery particles, and DLVR-X v2b delivery particles was 1.5e5 ng p30. DLVR-X v2a particles were also tested at 8.5 e5 ng p30 (see farthest right bar of graph).

[0094] FIG. 28 shows editing efficiency of optimized delivery particles comprising the coiled-coil peptides (CC). Particles were tested at doses of 0.5 e5 ng, 1.5 e5 ng, and 8.5 e5 ng. Saline was used as the control.

[0095] FIGs. 29A-29B show the loading efficiency of CRISPRoff epigenetic editors using DLVR-X particles comprising the coiled-coil peptides (CC) versus standard particles. Cas9 levels (FIG. 29A) and p30 (FIG. 29B) show results of amount of cargo and capsid between tested delivery particles, as measured via ELISA assay.

[0096] FIGs. 30A-30B show results of PCSK9 epigenetic silencing using particles comprising the coiled-coil peptides (CC) vs standard particles containing CRISPRoff or KRABless CRISPRoff epigenetic silencer constructs. Expression of the tdTomato-T’CS V reporter gene was measured in cells by flow cytometry at Day 7 and Day 14 post transduction of Hep3B cells with the eVLPs containing the epigenetic silencer constructs. A CRISPRi construct using was used as a control.

[0097] FIGs. 31A-31B show the loading efficiency of CHARM epigenetic editors using DLVR-X particles comprising the coiled-coil peptides (CC) versus standard particles. Cas9 levels (FIG. 31A) and p30 (FIG. 31B) show results of amount of cargo and capsid between tested delivery particles, as measured via ELISA assay.

[0098] FIGs. 32A-32B show results of PCSK9 epigenetic silencing using particles comprising the N- or C-terminally coiled-coil peptide (CC) fusions vs standard particles containing CHARM or KRABless CHARM epigenetic silencer constructs. Expression of the tdTomato- / 'C.S' '9 reporter gene was measured in cells by flow cytometry at Day 7 and Day 14 post transduction of Hep3B cells with the eVLPs containing the epigenetic silencer constructs. A CRISPRi construct using was used as a control.

[0099] FIGs. 33A-33B show the loading efficiency of epigenetic editors using DLVR-X particles comprising the coiled-coil pro (CC-Pro) versus standard coiled-coil (CC) particles. Cas9 levels (FIG. 31A) and p30 (FIG. 31B) show results of amount of cargo and capsid between tested delivery particles, as measured via ELISA assay.WSGR Docket No.:62697-750.601

[0100] FIG. 34 shows results of CD33 epigenetic silencing in K562 cells using particles comprising the coiled-coil peptide (CC-Pro) fusions containing KRABless CHARM or KRABless CRISPRoff epigenetic silencer constructs vs standard particles coiled-coil (CC) containing KRABless CRISPRoff. A CRISPRi construct was used as a control.

[0101] FIG. 35 shows results of in vivo PCSK9 epigenetic silencing in a transgenic mice model (TgPCSK9) using particles comprising the coiled-coil peptide containing KRABless CRISPRoff or standard particles containing ABE8e. Particles were tested at a dose of 18 mg p30 per kg. Saline was used as the control.

[0102] FIGs. 36A-36B show the loading efficiency of Prime editors (PE) using particles comprising the coiled-coil peptides (CC) versus standard particles. As shown in FIG. 36A Particles were loaded with PEs (PE6d, PEMax or PE6c) by direct fusion of the Gag to a PE (shown as Gag-PE6d, Gag-PEMax, Gag- PE6d) or loaded by fusion of a CC pair (E4 / K4) to Gag (Gag-E4-Pol) and K4-PE (shown as CC-PE6d, CC-PEMax, CC-PE6c). FIG. 36B shows the Gag Average and CC Average for Gag loading and CC loading.

[0103] FIG. 37 shows evaluation of Prime editing efficiency for particles containing prime editors loaded by standard delivery particle without CC (PE6d, PEMax, or PE6c) and E4 / K4 coiled-coil (PE6d CC, PEMax CC, PE6c CC) editing the mouse DNMT1 locus of Neuro-2A cells at 12.5, 25, 50, or 100 pl dose of PE.

[0104] FIGs. 38A-38B show the Prime editing efficiency (FIG. 38A) and loading efficiency (FIG. 38B) of Prime editor (PEMax) using particles comprising the coiled-coil pro (CC-Pro PEMax) versus standard coiled-coil (K4-PEMax) particles. For K4-PEMax particle production, Gag-E4-Pol plasmid was used to transfect VPCs. For CC-Pro particle production, Gag-E4-Pro plasmid was used to transfect VPCs. Additionally, immediately after transfection, cells were treated with a protease inhibitor (Amprenavir).

[0105] FIGs. 39A-39C show the editing efficiency and loading efficiency using multiple cargoes using Coiled-Coil loading strategy vs standard particles. Particles comprising the coiled-coil (CC) or standard delivery particles without CC were loaded with EnAsCasl2, ABE8e, or EnAsCasl2 and ABE8e editors. Particles were produced in 30 mb cultures, lOOx concentrated, using a BaEV envelope.

[0106] FIGs. 40A-40C show the editing efficiency and loading efficiency using Gag-Zip-Pro particles in the presence of a protease inhibitor (Amprenavir).

[0107] FIGs. 41A-41B show the editing efficiency of B2M in Hematopoietic Stem Cells (HSCs; FIG. 41A) and HEK293 cells (FIG. 41B) using BaEV pseudotyped eVLPs particles comprising the coiled-coil pro (Gag-CC-Pro 1 and Gag-CC-Pro 2), Gag-Pro (GagPro Control), standard coiled-coil (CC standard), standard delivery particles without CC (Standard).

[0108] FIGs. 42A-42B show the editing efficiency (FIG. 42A) and loading efficiency (FIG. 42B) for MMLV Gag -HIV Protease Particles (DLVR-X v2b). Chimeric virus-like particles made from MMLV Gag / MMLV Gag-HIV Protease were compared to MMLV Gag / MMLV Gag-MMLV Protease particles when pseudotyped with VSV-G. Editing efficiency was measured as percentage of B2M negative targets (% B2M negative). 1%, 5%, 7.5%, 10% and 20% Gag-Pro were tested in the presence of a proteaseWSGR Docket No.:62697-750.601 inhibitor (5 pM Amprenavir) for MLV Gag -Pro Pseudotyped with BaEV envelopes using 3 pl or lOpl doses.

[0109] FIGs. 43A-43B show the editing efficiency of B2M in HEK293T cells (FIG. 43A) and Hematopoietic Stem and Progenitor Cells (HSPCs; FIG. 43B) transduced with the following chimeric BaEV-FeLV eVLPs: 1) Standard BaEV-FeLV eVLPs, 2) BaEV- FeLV eVLPs with Gag-MLV-Pro, 3) BaEV- FeLV eVLPs with Gag-MLV-Pro and MLV protease cleavage site (PR / RT), 4) BaEV- FeLV eVLPs with Gag-MLV-Pro and MLV protease cleavage site (RT / IN), 5) BaEV- FeLV eVLPs with Gag- HlV-Pro and HIV protease cleavage site (RT / IN), 6) BaEV- FeLV eVLPs with Gag-HIV-Pro and HIV protease cleavage site (CA / NC), or 7) BaEV- FeLV with Gag-HIV-Pro. Each figure shows dose responses in HEK293T cells (FIG. 43A) and HSPCs (FIG. 43B). DLVR-X v2a are Gag-MLV Pro and the DLVR-X V2b is Gag-HIV Pro.

[0110] FIGs. 44A-44D show the in vivo editing efficiency in Lineage -negative (Lin-) (FIG. 44A), Lineage-positive (Lin+) (FIG. 44B), and NGS (FIG. 44C) Hematopoietic Stem Cells (HSCs) in a mice model transduced using BaEV-FeLV eVLPs containing an ABE base editor targeting the B2M gene and comprising standard delivery particles without CC (FeLV-ABE-B2M), standard coiled-coil (FeLV-ABE- B2M-C-C), or the coiled-coil Pro (FeLV-ABE-C-C-Pro). Editing efficiency was measured as percentage of B2M negative targets (% B2M negative). FIG. 44D shows the production volume for each BaEV- FeLV eVLP tested.DETAILED DESCRIPTION[oni] In some aspects of the present disclosure, provided herein is a lipid delivery particle that comprises (a) a lipid membrane encapsulating a cavity; (b) an envelope protein on the lipid membrane;(c) a plasma membrane recruitment element; (d) a payload; and (e) a coiled-coil peptide pair. The coiled- coil peptide pair can comprise a first coiled-coil peptide and a second coiled-coil peptide, e.g., amino acid sequences having at least 80%, at least 85%, at least 90%, or at least 95%, or 100% sequence identity to the amino acid sequences set forth in Table 8. In some cases, the first coiled-coil peptide and the second coiled-coil peptide comprise SEQ ID NOs: 601 and 602, respectively, or SEQ ID NOs: 602 and 601, respectively; SEQ ID NOs: 603 and 604, respectively, or SEQ ID NOs: 604 and 603, respectively; SEQ ID NOs: 605 and 606, respectively, or SEQ ID NOs: 606 and 605, respectively; SEQ ID NOs: 607 and 608, respectively, or SEQ ID NOs: 608 and 607, respectively; SEQ ID NOs: 609 and 610, respectively, or SEQ ID NOs: 610 and 609, respectively; SEQ ID NOs: 611 and 612, respectively, or SEQ ID NOs: 612 and 611, respectively; SEQ ID NOs: 613 and 614, respectively, or SEQ ID NOs: 614 and 613, respectively; SEQ ID NOs: 615 and 616, respectively, or SEQ ID NOs: 616 and 615, respectively; SEQ ID NOs: 616 and 621, respectively, or SEQ ID NOs: 621 and 616, respectively; SEQ ID NOs: 617 and 618, respectively, or SEQ ID NOs: 618 and 617, respectively; or SEQ ID NOs: 613 and 619, respectively, or SEQ ID NOs: 619 and 613, respectively. In some cases, the first coiled-coil peptide comprises a heptad repeat of an amino sequence as set forth in (EXiX2X3X4X5X6)m, where Xi, X2, X3, X4, X5, and X„ are each independently any amino acid residue, and wherein m is an integer greater than or equal to 2; theWSGR Docket No.:62697-750.601 second coiled-coil peptide comprises a heptad repeat of an amino sequence as set forth in (KX7XsX9XioXnXi2)n, where X7, Xs, X9, X10, Xu, and X12 are each independently any amino acid residue, and wherein n is an integer greater than or equal to 2. In some of these cases, Xi or X4 is a hydrophobic amino acid residue, or Xi and X4 are each independently a hydrophobic amino acid residue. In some cases, Xi is isoleucine and X4 is leucine, or wherein Xi is leucine and X4 is isoleucine. In some cases, X7 or X10 is a hydrophobic amino acid residue, or X7 and X10 are each independently a hydrophobic amino acid residue. In some cases, X7 is isoleucine and X10 is leucine, or wherein X7 is leucine and X10 is isoleucine. In some cases, m can be 2, 3, 4, 5, 6, 7, 8, 9, 10, or greater than about 10. In some cases, m is 3 or 4. In some cases, n can be 2, 3, 4, 5, 6, 7, 8, 9, 10, or greater than about 10. In some cases, n is 3 or 4. In some cases, m and n can be each independently 2, 3, 4, 5, 6, 7, 8, 9, 10, or greater than about 10. In some cases, m and n are each independently 3 or 4.

[0112] The coiled-coiled peptide pair can comprise a helical wheel. One or more amino acid residues of a first coiled-coil peptide of the helical wheel can interact with one or more amino acid residues of a second coiled-coil peptide of the helical wheel. In some cases, the coiled-coil peptide described herein may comprise a format as set forth in a-b-c-d-e-f-g, wherein a, b, c, d, e, f, or g can be independently any amino acid residue. In some cases, a first coiled-coil peptide may comprise the format a-b-c-d-e-f-g and a second coiled-coil peptide may comprise the format h-i-j-k-l-m-n, wherein h, i, j, k, 1, m, or n can be independently any amino acid residue. One or more amino acid residues of the coiled-coil peptide may be hydrophobic. One or more amino acid residues of the coiled-coil peptide may be hydrophilic. One or more amino acid residues of the coiled-coil peptide may aromatic. One or more amino acid residues of the coiled-coil peptide may be polar. One or more amino acid residues of the coiled-coil peptide may be nonpolar. One or more amino acid residues of the coiled-coil peptide may be charged.

[0113] In some cases, the coiled-coil peptide (e.g., a first coiled-coil peptide) comprises a-b-c-d-e-f-g. Amino acids at positions a and / or d may be hydrophobic. For example, the amino acid residues at a and / or d may be alanine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, valine, or any combination thereof. In some cases, an amino acid residue at position a may comprise isoleucine. In some cases, an amino acid residue at position d may comprise leucine. In some cases, one or more amino acid residues at positions b, c, e, f, g, or any combination thereof may be polar. For example, one or more amino acid residues at positions b, c, e, f, g, or any combination thereof may comprise serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof. In some cases, one or more amino acid residues at positions b, c, e, f, g, or any combination thereof may be charged. For example, one or more amino acid residues at positions b, c, e, f, g, or any combination thereof may comprise glutamate, aspartate, arginine, lysine, or any combination thereof. The coiled-coil peptide (e.g., a first coiled-coil peptide) may comprise a format of a-b-c-d-e-f-g, wherein (i) an amino acid residue at position a may be a hydrophobic residue (e.g., alanine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, valine, or any combination thereof), (ii) an amino acid residue at position b may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate,WSGR Docket No.:62697-750.601 aspartate, arginine, lysine, or any combination thereof), (iii) an amino acid residue at position c may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate, aspartate, arginine, lysine, or any combination thereof), (iv) an amino acid residue at position d may be a hydrophobic residue (e.g., alanine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, valine, or any combination thereof), (v) an amino acid residue at position e may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate, aspartate, arginine, lysine, or any combination thereof), (vi) an amino acid residue at position f may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate, aspartate, arginine, lysine, or any combination thereof), (vii) an amino acid residue at position g may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate, aspartate, arginine, lysine, or any combination thereof), or any combination thereof.

[0114] The coiled-coil peptide (e.g., a first coiled-coil peptide) may comprise any number of repeat of the amino acid sequence as set forth in the a-b-c-d-e-f-g format described herein. For example, the coiled- coil peptide (e.g., a first coiled-coil peptide) may comprise (a-b-c-d-e-f-g)x, wherein x can be any integer greater than 2 (e.g., x can be 2, 3, 4, 5, 6, 7, 8, 9, 10, or greater than about 10). The coiled-coil peptide (e.g., a first coiled-coil peptide) may thus comprise an amino acid sequence comprising a repeat of a combination of hydrophobic and / or polar amino acid residues as set forth in a-b-c-d-e-f-g.

[0115] The coiled-coil peptide (e.g., the first coiled-coil peptide) can comprise a number of heptad motifs in tandem (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, or more heptad motifs in tandem). Each heptad motif may be independently in the a-b-c-d-e-f-g format described herein. In some cases, the motifs may be the same. In some cases, the motifs may be different. For example, the coiled-coil peptide (e.g., the first coiled-coil peptide) may comprise a first a-b-c-d-e-f-g motif followed by at least a second a-b-c-d-e-f-g motif. As another example, the coiled-coil peptide (e.g., the first coiled-coil peptide) may comprise a first a-b-c-d-e- f-g motif followed by a heptad motif with one or more of a, b, c, d, e, f, or g independently replaced with a different amino acid residue. In some cases, each heptad motif comprises at least one hydrophobic amino acid residue. In some cases, each heptad motif comprises at least one polar (e.g., charged) amino acid residue.

[0116] In some cases, the coiled-coil peptide (e.g., a second coiled-coil peptide) comprises h-i-j-k-l-m-n. Amino acids at positions a and / or d may be hydrophobic. For example, the amino acid residues at h and / or k may be alanine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, valine, or any combination thereof. In some cases, an amino acid residue at position h may comprise isoleucine. In some cases, an amino acid residue at position k may comprise leucine. In some cases, one or more amino acid residues at positions i, j, 1, m, n, or any combination thereof may be polar. For example, one or more amino acid residues at positions i, j, 1, m, n, or any combination thereof may comprise serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof. In someWSGR Docket No.:62697-750.601 cases, one or more amino acid residues at positions i, j, 1, m, n, or any combination thereof may be charged. For example, one or more amino acid residues at positions i, j, 1, m, n, or any combination thereof may comprise glutamate, aspartate, arginine, lysine, or any combination thereof. The coiled-coil peptide (e.g., a second coiled-coil peptide) may comprise a format of h-i-j-k-l-m-n, wherein (i) an amino acid residue at position h may be a hydrophobic residue (e.g., alanine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, valine, or any combination thereof), (ii) an amino acid residue at position i may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate, aspartate, arginine, lysine, or any combination thereof), (iii) an amino acid residue at position) may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate, aspartate, arginine, lysine, or any combination thereof), (iv) an amino acid residue at position k may be a hydrophobic residue (e.g., alanine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, valine, or any combination thereof), (v) an amino acid residue at position 1 may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate, aspartate, arginine, lysine, or any combination thereof), (vi) an amino acid residue at position m may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate, aspartate, arginine, lysine, or any combination thereof), (vii) an amino acid residue at position n may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate, aspartate, arginine, lysine, or any combination thereof), or any combination thereof.

[0117] The coiled-coil peptide (e.g., a second coiled-coil peptide) may comprise any number of repeat of the amino acid sequence as set forth in the h-i-j-k-l-m-n format described herein. For example, the coiled- coil peptide (e.g., a second coiled-coil peptide) may comprise (h-i-j-k-l-m-n)y, wherein y can be any integer greater than 2 (e.g., y can be 2, 3, 4, 5, 6, 7, 8, 9, 10, or greater than about 10). The coiled-coil peptide (e.g., a second coiled-coil peptide) may thus comprise an amino acid sequence comprising a repeat of a combination of hydrophobic and / or polar amino acid residues as set forth in h-i-j-k-l-m-n.

[0118] The coiled-coil peptide (e.g., the second coiled-coil peptide) can comprise a number of heptad motifs in tandem (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, or more heptad motifs in tandem). Each heptad motif may be independently in the h-i-j-k-l-m-n format described herein. In some cases, the motifs may be the same. In some cases, the motifs may be different. For example, the coiled-coil peptide (e.g., the second coiled- coil peptide) may comprise a first h-i-j-k-l-m-n motif followed by at least a second h-i-j-k-l-m-n motif. As another example, the coiled-coil peptide (e.g., the second coiled-coil peptide) may comprise a first h-i-j-k- l-m-n motif followed by a heptad motif with one or more of h, i, j, k, 1, m, or n independently replaced with a different amino acid residue. In some cases, each heptad motif comprises at least one hydrophobic amino acid residue. In some cases, each heptad motif comprises at least one polar (e.g., charged) amino acid residue.WSGR Docket No.:62697-750.601

[0119] In some cases, the amino acid residues of a first coiled-coil peptide and the amino acids of a second coiled-coil peptide may form one or more salt bridges between them. The polar and / or charged amino acid residues of a coiled-coil peptide (e.g., a first coiled-coil peptide) may form one or more salt bridges with the polar and / or charged amino acid residues of another coiled-coil peptide (e.g., a second coiled-coil peptide). In some cases, a coiled-coil peptide (e.g., a first coiled-coil peptide) may comprise the a-b-c-d-e-f-g format described herein and another coiled-coil peptide (e.g., a second coiled-coil peptide) may comprise the h-i-j-k-l-m-n format described herein. One or more salt bridges may form between any polar and / or charged amino acid residues of the a-b-c-d-e-f-g format described herein and h- i-j-k-l-m-n format described herein. For example, a salt bridge may form between the e amino acid residue and the g amino acid residue of the coiled-coil peptide (e.g., first coiled-coil peptide) set forth in the a-b-c-d-e-f-g format and the 1 amino acid residue and the n amino acid residue of the coiled-coil peptide (e.g., second coiled-coil peptide) set forth in the h-i-j-k-l-m-n format. Thus, a salt bridge may form between (i) the polar and / or charged e amino acid residue and the polar and / or charged 1 amino acid residue, and / or (ii) the polar and / or charged g amino acid residue and the polar and / or charged n amino acid residue.

[0120] In some aspects of the present disclosure, provided herein is a lipid delivery particle that comprises a lipid membrane encapsulating a cavity; an envelope protein on the lipid membrane; and a chimeric protein comprising a plasma membrane recruitment element and a payload protein, wherein the plasma membrane recruitment element comprises a gag polyprotein, wherein the chimeric protein comprises a nuclear export signal (NES) inside the gag protein, and wherein the payload protein does not comprise a reverse transcriptase.

[0121] In some aspects of the present disclosure, provided herein is a lipid delivery particle in which the gag polyprotein in a plasma membrane recruitment element is engineered. In some cases, the lipid delivery particle comprises a lipid membrane encapsulating a cavity; an envelope protein on the lipid membrane; and a chimeric protein comprising a plasma membrane recruitment element and a payload protein. In some of these cases, the plasma membrane recruitment element comprises a gag polyprotein, and the gag polyprotein lacks at least a fragment of a nucleocapsid protein, and the plasma membrane recruitment element further comprises a heterologous domain fused to C-terminus of the gag polyprotein. Alternatively, or additionally, the gag polyprotein lacks at least a fragment of a matrix protein, and the plasma membrane recruitment element further comprises a pleckstrin homology (PH) domain fused to N- terminus of the gag polyprotein.

[0122] In some aspects, the present disclosure also relates to methods of making a lipid delivery particle disclosed herein, the present disclosure also relates to methods of using the lipid delivery particle disclosed herein, and compositions, pharmaceutical compositions, and kits related to the lipid delivery particle disclosed herein.

[0123] The practice of some methods disclosed herein employ, unless otherwise indicated, conventional techniques of immunology, biochemistry, chemistry, molecular biology, microbiology, cell biology, genomics and recombinant DNA, which are within the skill of the art.WSGR Docket No.:62697-750.601DEFINITIONS

[0124] Certain specific details of this description are set forth in order to provide a thorough understanding of various embodiments. However, one skilled in the art will understand that the present disclosure may be practiced without these details. In other instances, well-known structures have not been shown or described in detail to avoid unnecessarily obscuring descriptions of the disclosure.

[0125] Unless the context requires otherwise, throughout the specification and claims which follow, the word “comprise” and variations thereof, such as, “comprises” and “comprising” are to be construed in an open, inclusive sense, that is, as “including, but not limited to.” Further, headings provided herein are for convenience only and do not interpret the scope or meaning of the claimed disclosure.

[0126] As used in this specification and the appended claims, the singular forms “a,” “an,” and “the” include plural referents unless the content clearly dictates otherwise. The use of the words “a” or “an” when used in conjunction with the term “comprising” herein may mean “one,” but it is also consistent with the meaning of “one or more,” “at least one,” and “one or more than one.”

[0127] It should also be noted that the term “or” is generally employed in its sense including “and / or” unless the content clearly dictates otherwise.

[0128] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present disclosure, suitable methods and materials are described below.

[0129] The term “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 in some instances ±10%, or in some instances ±5%, or in some instances ±1%, or in some instances ±0.1% from the specified value, as such variations are appropriate to perform the disclosed methods. As used herein, “about” and “approximately” generally mean an acceptable degree of error for the quantity measured given the nature or precision of the measurements. Exemplary degrees of error are within 20 percent (%), typically, within 10%, and more typically, within 5% of a given range of values.

[0130] Calculations of homology or sequence identity between sequences (the terms are used interchangeably herein) are performed as follows. To determine the percent identity of two amino acid sequences, or of two nucleic acid sequences, the sequences are aligned for optimal comparison purposes (e.g., gaps can be introduced in one or both of a first and a second amino acid or nucleic acid sequence for optimal alignment and non-homologous sequences can be disregarded for comparison purposes). In some cases, the length of a reference sequence aligned for comparison purposes is at least 30%, preferably at least 40%, more preferably at least 50%, 60%, and even more preferably at least 70%, 80%, 90%, 100% of the length of the reference sequence. The amino acid residues or nucleotides at corresponding amino acid positions or nucleotide positions are then compared. When a position in the first sequence is occupied by the same amino acid residue or nucleotide as the corresponding position in the second sequence, then the molecules are identical at that position (as used herein amino acid or nucleic acid “identity” is equivalent to amino acid or nucleic acid “homology”).WSGR Docket No.:62697-750.601

[0131] The percent identity between the two sequences is a function of the number of identical positions shared by the sequences, taking into account the number of gaps, and the length of each gap, which need to be introduced for optimal alignment of the two sequences. The comparison of sequences and determination of percent identity between two sequences can be accomplished using a mathematical algorithm. In some cases, the percent identity between two amino acid sequences is determined using the Needleman and Wunsch ((1970) J. Mol. Biol. 48:444-453 ) algorithm which has been incorporated into the GAP program in the GCG software package (available at http: / / www.gcg.com), using either a Blossum 62 matrix or a PAM250 matrix, and a gap weight of 16, 14, 12, 10, 8, 6, or 4 and a length weight of 1, 2, 3, 4, 5, or 6. In some cases, the percent identity between two nucleotide sequences is determined using the GAP program in the GCG software package (available at http: / / www.gcg.com), using a NWSgapdna. CMP matrix and a gap weight of 40, 50, 60, 70, or 80 and a length weight of 1, 2, 3, 4, 5, or 6. A particularly preferred set of parameters (and the one that should be used unless otherwise specified) are a Blossum 62 scoring matrix with a gap penalty of 12, a gap extend penalty of 4, and a frameshift gap penalty of 5.

[0132] The percent identity between two amino acid or nucleotide sequences can be determined using the algorithm of E. Meyers and W. Miller ((1989) CABIOS, 4: 11-17) which has been incorporated into the ALIGN program (version 2.0), using a PAM 120 weight residue table, a gap length penalty of 12 and a gap penalty of 4. The nucleic acid and protein sequences described herein can be used as a “query sequence” to perform a search against public databases to, for example, identify other family members or related sequences. Such searches can be performed using the NBLAST and XBLAST programs (version 2.0) of Altschul, et al. (1990) J. Mol. Biol. 215:403-10. BLAST nucleotide searches can be performed with the NBLAST program, score = 100, wordlength = 12 to obtain nucleotide sequences homologous to a nucleic acid molecule of the disclosure. BLAST protein searches can be performed with the XBLAST program, score = 50, wordlength = 3 to obtain amino acid sequences homologous to protein molecules of the disclosure. To obtain gapped alignments for comparison purposes, Gapped BLAST can be utilized as described in Altschul et al., (1997) Nucleic Acids Res. 25:3389-3402. When utilizing BLAST and Gapped BLAST programs, the default parameters of the respective programs (e.g., XBLAST and NBLAST) can be used.

[0133] It is understood that the molecules of the present disclosure may have additional conservative or non-essential amino acid substitutions, which do not have a substantial effect on their functions.

[0134] The terms “peptide,” “polypeptide,” and “protein” are used interchangeably, and refer to a compound comprised of amino acid residues covalently linked by peptide bonds. A protein or peptide must contain at least two amino acids, and no limitation is placed on the maximum number of amino acids that can comprise a protein’s or peptide’s sequence. Polypeptides include any peptide or protein comprising two or more amino acids joined to each other by peptide bonds. As used herein, the term refers to both short chains, which also commonly are referred to in the art as peptides, oligopeptides and oligomers, for example, and to longer chains, which generally are referred to in the art as proteins, of which there are many types. “Polypeptides” include, for example, biologically active fragments,WSGR Docket No.:62697-750.601 substantially homologous polypeptides, oligopeptides, homodimers, heterodimers, variants of polypeptides, modified polypeptides, derivatives, analogs, fusion proteins, among others. A polypeptide includes a natural peptide, a recombinant peptide, or a combination thereof.

[0135] The term “expression vector” refers to a vector comprising a recombinant polynucleotide comprising expression control sequences operatively linked to a nucleotide sequence to be expressed. An expression vector comprises sufficient cis-acting elements for expression; other elements for expression can be supplied by the host cell or in an in vitro expression system. Expression vectors include all those known in the art, including cosmids, plasmids (e.g., naked or contained in liposomes) and viruses (e.g., lentiviruses, retroviruses, adenoviruses, and adeno-associated viruses) that incorporate the recombinant polynucleotide.

[0136] The term “promoter” refers to a DNA sequence recognized by the transcription machinery of the cell, or introduced synthetic machinery, required to initiate the specific transcription of a polynucleotide sequence.

[0137] An example of a promoter is the immediate early cytomegalovirus (CMV) promoter sequence. This promoter sequence is a strong constitutive promoter sequence capable of driving high levels of expression of any polynucleotide sequence operatively linked thereto. In some cases, a polynucleotide comprises a CMV promoter. In some cases, a polynucleotide may comprise a promoter selected from the group consisting of, but not limited to, a simian virus 40 (SV40) early promoter, a mouse mammary tumor virus (MMTV), a human immunodeficiency virus (HIV) long terminal repeat (LTR) promoter, a MoMuLV promoter, an avian leukemia virus promoter, an Epstein-Barr virus immediate early promoter, a Rous sarcoma virus (RSV) promoter, an actin promoter, a myosin promoter, an elongation factor- la promoter, a hemoglobin promoter, and a creatine kinase promoter. In some cases, a polynucleotide comprises a sequence encoding a poly(A) tail. In some cases, a polynucleotide comprises a 3’ UTR sequence. In some cases, a polynucleotide comprises a 5’ UTR sequence. In some cases, a polynucleotide comprises multiple smaller and discrete nucleotide sequences that are typically heterologous and exhibit different and measurable function(s), a promoter sequence, a translation initiating sequence, a start codon, a polyadenylation sequence, a stop codon, and / or a linker sequence.

[0138] The term “subject” is intended to include living organisms in which an immune response can be elicited (e.g., mammals, human). Exemplary subjects include humans, monkeys, dogs, cats, mice, rats, cows, horses, camels, goats, rabbits, and sheep. In certain embodiments, the subject is a human. A “patient” is a subject suffering from or at risk of developing a disease, disorder or condition or otherwise in need of the compositions and methods provided herein.

[0139] The term “therapeutic” as used herein means a treatment. A therapeutic effect is obtained by reduction, suppression, remission, or eradication of a disease state.

[0140] The term “prophylaxis” as used herein means the prevention of or protective treatment for a disease or disease state.

[0141] The term “transfected” or “transformed” or “transduced” refers to a process by which exogenous nucleic acid is transferred or introduced into the host cell. A “transfected” or “transformed” orWSGR Docket No.:62697-750.601“transduced” cell is one which has been transfected, transformed or transduced with exogenous nucleic acid. The cell includes the primary subject cell and its progeny. The term “transformed cell” or “transfected cell” can be a transformed or transfected cell in which inserted DNA can replicate either as an autonomously replicating plasmid or as part of the host chromosome. In some cases, a transfected cell or transformed cell can express the inserted DNA or RNA transiently (e.g., for brief periods of time).

[0059] The terms “nucleic acid,” “nucleic acid sequence,” “nucleotide sequence,” or “polynucleotide sequence,” and “polynucleotide” are used interchangeably. They refer to a polymeric form of nucleotides of any length, either deoxyribonucleotides or ribonucleotides, or analogs thereof. The polynucleotide may be either single-stranded or double-stranded, and if single-stranded may be the coding strand or noncoding (antisense) strand. A polynucleotide may comprise modified nucleotides, such as methylated nucleotides and nucleotide analogs. The sequence of nucleotides may be interrupted by non-nucleotide components. A polynucleotide may be further modified after polymerization, such as by conjugation with a labeling component. The nucleic acid may be a recombinant polynucleotide, or a polynucleotide of genomic, cDNA, semisynthetic, or synthetic origin which either does not occur in nature or is linked to another polynucleotide in a non-natural arrangement.

[0142] The term “cell line” or “producer cell line”, as used herein, refers to cultured cells that can be passed (e.g., divided) more than once. In some cases, a cell like can be passed more than 2 times, more than 5 times, more than 10 times, more than 50 times, more than 100 times, or more than 150 times. In some cases, a cell line can be passed between 2-250 times. In some cases, a cell line can be passed up to 200 times.

[0143] The term “lipid delivery particle” refers to lipid vesicles secreted by the plasma membrane of a cell having a membrane or envelope that surrounds a central internal space. Lipid delivery particles can be produced via budding out of the cell’s plasma membrane, secretion from lysosomes, or export by the Golgi complex. Lipid delivery particles can promote intercellular communication in vitro and in vivo.

[0144] Exemplary lipid delivery particles can include, but are not limited to, exosomes, ectosomes (e.g., microvesicles and shedding vesicles), apoptotic bodies, microsomes, micelles, oncosomes, and liposomes.

[0062] Lipid delivery particles can have a range of size, such as a cross-sectional diameter, wherein the cross-sectional diameter has a range from about 1 nm to about 1200 nm, about 10 nm to about 1200 nm, about 25 nm to about 1200 nm, about 50 nm to about 1200 nm, about 100 nm to about 1200 nm, about 10 nm to about 1000 nm, about 25 nm to about 1000 nm, about 50 nm to about 1000 nm, about 100 nm to about 1000 nm, about 10 nm to about 500 nm, about 25 nm to about 500 nm, about 50 nm to about 500 nm or about 100 nm to about 500 nm (all inclusive). Lipid delivery particles can comprise a vesicle size of greater than 10 nm. Lipid delivery particles can comprise a vesicle size of less than 1000 nm.

[0145] The term “module” or “modification”, used interchangeably, can refer to an addition or a motif that is added to a payload or a chimeric protein within lipid delivery particles (e.g., extracellular vesicles and / or virus-like particles). The additions can enhance characteristics of particle assembly and / or payload recruitment. A module can occur in combination or alone. A module can be linked to a payload or another element (e.g., domain) in a construct.WSGR Docket No.:62697-750.601

[0146] The term “purifying,” “purified,” and “purify,” can be used interchangeably and refer to a preparation phase of a lipid delivery particle. A preparation of lipid delivery particles can comprise a known or unknown concentration of a substance. In some cases, purifying produced lipid delivery particles comprises removing, either partially removing or wholly removing, a portion of the produced lipid delivery particles from a sample containing one or more biological components (e.g., producer cells). In some cases, a composition comprising lipid delivery particles, as described herein, that have been purified are enriched as compared to a starting sample. Enrichment of the lipid delivery particles can comprise 5%, 10%, 25%, 50%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% enrichment compared to a starting sample. In some cases, purified lipid delivery particles can be free or substantially free of residual biological products. Residual biological products can include, but are not limited to, unwanted nucleic acids, proteins, lipids, and / or or metabolites or abiotic materials such as including chemicals. In some cases, lipid delivery particles free of residual biological products comprise lipid delivery particles containing no detectable producer cells in the composition.

[0147] The terms "portion" and "fragment" are used interchangeably herein and refer to a continuous element. For example, a part of a structure such as an amino acid sequence or protein refers to a continuous element of said structure. A portion, or a fragment of a structure preferably comprises one or more functional properties of said structure. In some cases, a portion, or a fragment of a structure such as an amino acid sequence comprises of at least 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 94%, 95%, 96%, 97%, 98%, 99% of the entire structure or amino acid sequence. In some cases, a portion, or a fragment of a structure such as an amino acid sequence comprises of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 85, 90, 91, 92, 94, 95, 96, 97, 98, 99, 100, 120, 130, 140, 150, 160, 170, 180, 190, 200 or more contiguous amino acid residues of the entire structure or amino acid sequence. In some cases, a portion or a fragment can be proteins having at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 85, 90, 91, 92, 94, 95, 96, 97, 98, 99, 100, 120, 130, 140, 150, 160, 170, 180, 190, 200 or more amino acids absent from the amino and / or carboxyl terminus of the amino acid sequence when compared to the reference amino acid sequence, optionally wherein the fragment has the same activity as the reference protein.

[0148] Ranges: throughout this disclosure, various aspects of the present disclosure can be presented in a range format. It should be understood that the description in range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the present disclosure. Accordingly, the description of a range should be considered to have specifically disclosed all the possible subranges as well as individual numerical values within that range. For example, description of a range such as from 1 to 6 should be considered to have specifically disclosed subranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6 etc., as well as individual numbers within that range, for example, 1, 2, 2.7, 3, 4, 5, 5.3, and 6. As another example, a range such as 95-99% identity, includes something with 95%, 96%, 97%, 98% or 99% identity, and includes subranges such as 96-99%, 96-98%, 96-97%, 97-99%, 97-98% and 98-99% identity. This applies regardless of the breadth of the range.WSGR Docket No.:62697-750.601COILED-COIL

[0149] In some aspects of the present disclosure, the lipid delivery particle utilizes coiled-coil peptide interaction to package / load payload into the particle. Coiled-coil (CC) peptides are small peptides that can tether together like Velcro. They can be attracted to each other due to hydrophobic effect and the interaction is stabilized by electrostatic interactions, causing two alpha helical domains to wind around each other. Without wishing to be bound by a certain mechanism, utilizing such mechanism, peptides that are fused to a pair of coiled-coil peptides respectively can thus be brought together due to the interaction between the coiled-coil peptide pair. Different CC pairs can exhibit unique properties, such as its oligomerization preferences, the orientation of the binding, the specificity of the binding interactions, and the strength of the binding interaction. Different combinations of payload can be used to achieve different editing outcomes. The peptide comprising the coiled-coil peptide pair may bind to each other through one or more electrostatic interactions. The electrostatic interactions between the coiled-coil peptide pair may be complimentary. The electrostatic interactions between the coiled-coil peptide pair may arise due to the amino acid composition of each peptide of the coiled-coil peptide pair.

[0150] In some aspects of the present disclosure, a lipid delivery particle provided herein comprises a first chimeric protein comprising a first coiled-coil peptide fused with a plasma membrane recruitment element, and a second chimeric protein comprising a second coiled-coil peptide fused with a payload protein, and the first coiled-coil peptide and the second coiled-coil peptide form a coiled-coil peptide pair in that they can be attracted to each other. In such configuration, the payload protein can thus be packaged into the lipid delivery particle via the interaction of the second coiled-coil peptide and the first coiled-coil peptide that is fused with the plasma membrane recruitment element.

[0151] In some cases, the first coiled-coil peptide is fused to the C-terminus of the plasma membrane recruitment element. In some cases, the first coiled-coil peptide is fused to the N-terminus of the plasma membrane recruitment element. In some cases, the second coiled-coil peptide is fused to the C-terminus of the payload protein. In some cases, the second coiled-coil peptide is fused to the N-terminus of the payload protein.

[0152] In some aspects, the coiled-coil design used herein is E4-K4 design. Using an E4-K4 CC domain can improve loading compared to other domains. In some aspects, the coiled-coil pair design used is one of E4-K4, EE12RR345L-RR12EE345L, EE1234L-RR1234L, EE12345L-RR12345L, AcidPl -BasePl, P3(x3)-P4, SynZip2-SynZipl9, SynZip2-SynZipl, and N5-N6.

[0153] In some aspects, protein payloads are loaded by fusing one half of the coiled-coil (e.g. , a member of a coiled-coil peptide pair) to gag-pol, and then fusing its binding partner (e.g., another member of the coiled-coil peptide pair) to the payload. A coiled-coiled peptide can comprise sequences having at least 80 % to 100 % sequence identity to the amino acid sequences as set forth in Table 8. A coiled-coiled peptide may comprise sequences having at least 80 % sequence identity to the amino acid sequences as set forth in Table 8. A coiled-coiled peptide can comprise sequences having at most 100 % sequence identity to the amino acid sequences as set forth in Table 8. A coiled-coiled peptide can comprise sequences having 80 % to 85 %, 80 % to 90 %, 80 % to 95 %, 80 % to 100 %, 85 % to 90 %, 85 % to 95WSGR Docket No.:62697-750.601%, 85 % to 100 %, 90 % to 95 %, 90 % to 100 %, or 95 % to 100 % sequence identity to the amino acid sequences as set forth in Table 8. A coiled-coiled peptide can comprise sequences having 80 %, 85 %, 90 %, 95 %, or 100 % sequence identity to the amino acid sequences as set forth in Table 8.

[0154] The coiled-coiled peptide pair can comprise a helical wheel. One or more amino acid residues of a first coiled-coil peptide of the helical wheel can interact with one or more amino acid residues of a second coiled-coil peptide of the helical wheel. In some cases, the coiled-coil peptide described herein may comprise a format as set forth in a-b-c-d-e-f-g, wherein a, b, c, d, e, f, or g can be independently any amino acid residue. In some cases, a first coiled-coil peptide may comprise the format a-b-c-d-e-f-g and a second coiled-coil peptide may comprise the format h-i-j-k-l-m-n, wherein h, i, j, k, 1, m, or n can be independently any amino acid residue. One or more amino acid residues of the coiled-coil peptide may be hydrophobic. One or more amino acid residues of the coiled-coil peptide may be hydrophilic. One or more amino acid residues of the coiled-coil peptide may aromatic. One or more amino acid residues of the coiled-coil peptide may be polar. One or more amino acid residues of the coiled-coil peptide may be nonpolar. One or more amino acid residues of the coiled-coil peptide may be charged.

[0155] In some cases, the coiled-coil peptide (e.g., a first coiled-coil peptide) comprises a-b-c-d-e-f-g. Amino acids at positions a and / or d may be hydrophobic. For example, the amino acid residues at a and / or d may be alanine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, valine, or any combination thereof. In some cases, an amino acid residue at position a may comprise isoleucine. In some cases, an amino acid residue at position d may comprise leucine. In some cases, one or more amino acid residues at positions b, c, e, f, g, or any combination thereof may be polar. For example, one or more amino acid residues at positions b, c, e, f, g, or any combination thereof may comprise serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof. In some cases, one or more amino acid residues at positions b, c, e, f, g, or any combination thereof may be charged. For example, one or more amino acid residues at positions b, c, e, f, g, or any combination thereof may comprise glutamate, aspartate, arginine, lysine, or any combination thereof. The coiled-coil peptide (e.g., a first coiled-coil peptide) may comprise a format of a-b-c-d-e-f-g, wherein (i) an amino acid residue at position a may be a hydrophobic residue (e.g., alanine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, valine, or any combination thereof), (ii) an amino acid residue at position b may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate, aspartate, arginine, lysine, or any combination thereof), (iii) an amino acid residue at position c may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate, aspartate, arginine, lysine, or any combination thereof), (iv) an amino acid residue at position d may be a hydrophobic residue (e.g., alanine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, valine, or any combination thereof), (v) an amino acid residue at position e may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate, aspartate, arginine, lysine, or any combination thereof), (vi) an amino acid residue atWSGR Docket No.:62697-750.601 position f may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate, aspartate, arginine, lysine, or any combination thereof), (vii) an amino acid residue at position g may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate, aspartate, arginine, lysine, or any combination thereof), or any combination thereof.

[0156] The coiled-coil peptide (e.g., a first coiled-coil peptide) may comprise any number of repeat of the amino acid sequence as set forth in the a-b-c-d-e-f-g format described herein. For example, the coiled- coil peptide (e.g., a first coiled-coil peptide) may comprise (a-b-c-d-e-f-g)x, wherein x can be any integer greater than 2 (e.g., x can be 2, 3, 4, 5, 6, 7, 8, 9, 10, or greater than about 10). The coiled-coil peptide (e.g., a first coiled-coil peptide) may thus comprise an amino acid sequence comprising a repeat of a combination of hydrophobic and / or polar amino acid residues as set forth in a-b-c-d-e-f-g.

[0157] The coiled-coil peptide (e.g., the first coiled-coil peptide) can comprise a number of heptad motifs in tandem (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, or more heptad motifs in tandem). Each heptad motif may be independently in the a-b-c-d-e-f-g format described herein. In some cases, the motifs may be the same. In some cases, the motifs may be different. For example, the coiled-coil peptide (e.g., the first coiled-coil peptide) may comprise a first a-b-c-d-e-f-g motif followed by at least a second a-b-c-d-e-f-g motif. As another example, the coiled-coil peptide (e.g., the first coiled-coil peptide) may comprise a first a-b-c-d-e- f-g motif followed by a heptad motif with one or more of a, b, c, d, e, f, or g independently replaced with a different amino acid residue. In some cases, each heptad motif comprises at least one hydrophobic amino acid residue. In some cases, each heptad motif comprises at least one polar (e.g., charged) amino acid residue.

[0158] In some cases, the coiled-coil peptide (e.g., a second coiled-coil peptide) comprises h-i-j-k-l-m-n. Amino acids at positions a and / or d may be hydrophobic. For example, the amino acid residues at h and / or k may be alanine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, valine, or any combination thereof. In some cases, an amino acid residue at position h may comprise isoleucine. In some cases, an amino acid residue at position k may comprise leucine. In some cases, one or more amino acid residues at positions i, j, 1, m, n, or any combination thereof may be polar. For example, one or more amino acid residues at positions i, j, 1, m, n, or any combination thereof may comprise serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof. In some cases, one or more amino acid residues at positions i, j, 1, m, n, or any combination thereof may be charged. For example, one or more amino acid residues at positions i, j, 1, m, n, or any combination thereof may comprise glutamate, aspartate, arginine, lysine, or any combination thereof. The coiled-coil peptide (e.g., a second coiled-coil peptide) may comprise a format of h-i-j-k-l-m-n, wherein (i) an amino acid residue at position h may be a hydrophobic residue (e.g., alanine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, valine, or any combination thereof), (ii) an amino acid residue at position i may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate,WSGR Docket No.:62697-750.601 aspartate, arginine, lysine, or any combination thereof), (iii) an amino acid residue at position) may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate, aspartate, arginine, lysine, or any combination thereof), (iv) an amino acid residue at position k may be a hydrophobic residue (e.g., alanine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, valine, or any combination thereof), (v) an amino acid residue at position 1 may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate, aspartate, arginine, lysine, or any combination thereof), (vi) an amino acid residue at position m may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate, aspartate, arginine, lysine, or any combination thereof), (vii) an amino acid residue at position n may be a polar amino acid residue (e.g., serine, threonine, cysteine, asparagine, glutamine, tyrosine, or any combination thereof) or a charged amino acid residue (e.g., glutamate, aspartate, arginine, lysine, or any combination thereof), or any combination thereof.

[0159] The coiled-coil peptide (e.g., a second coiled-coil peptide) may comprise any number of repeat of the amino acid sequence as set forth in the h-i-j-k-l-m-n format described herein. For example, the coiled- coil peptide (e.g., a second coiled-coil peptide) may comprise (h-i-j-k-l-m-n)y, wherein y can be any integer greater than 2 (e.g., y can be 2, 3, 4, 5, 6, 7, 8, 9, 10, or greater than about 10). The coiled-coil peptide (e.g., a second coiled-coil peptide) may thus comprise an amino acid sequence comprising a repeat of a combination of hydrophobic and / or polar amino acid residues as set forth in h-i-j-k-l-m-n.

[0160] The coiled-coil peptide (e.g., the second coiled-coil peptide) can comprise a number of heptad motifs in tandem (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, or more heptad motifs in tandem). Each heptad motif may be independently in the h-i-j-k-l-m-n format described herein. In some cases, the motifs may be the same. In some cases, the motifs may be different. For example, the coiled-coil peptide (e.g., the second coiled- coil peptide) may comprise a first h-i-j-k-l-m-n motif followed by at least a second h-i-j-k-l-m-n motif. As another example, the coiled-coil peptide (e.g., the second coiled-coil peptide) may comprise a first h-i-j-k- l-m-n motif followed by a heptad motif with one or more of h, i, j, k, 1, m, or n independently replaced with a different amino acid residue. In some cases, each heptad motif comprises at least one hydrophobic amino acid residue. In some cases, each heptad motif comprises at least one polar (e.g., charged) amino acid residue.

[0161] In some cases, the amino acid residues of a first coiled-coil peptide and the amino acids of a second coiled-coil peptide may form one or more salt bridges between them. The polar and / or charged amino acid residues of a coiled-coil peptide (e.g., a first coiled-coil peptide) may form one or more salt bridges with the polar and / or charged amino acid residues of another coiled-coil peptide (e.g., a second coiled-coil peptide). In some cases, a coiled-coil peptide (e.g., a first coiled-coil peptide) may comprise the a-b-c-d-e-f-g format described herein and another coiled-coil peptide (e.g., a second coiled-coil peptide) may comprise the h-i-j-k-l-m-n format described herein. One or more salt bridges may form between any polar and / or charged amino acid residues of the a-b-c-d-e-f-g format described herein and h-WSGR Docket No.:62697-750.601 i-j-k-l-m-n format described herein. For example, a salt bridge may form between the e amino acid residue and the g amino acid residue of the coiled-coil peptide (e.g., first coiled-coil peptide) set forth in the a-b-c-d-e-f-g format and the 1 amino acid residue and the n amino acid residue of the coiled-coil peptide (e.g., second coiled-coil peptide) set forth in the h-i-j-k-l-m-n format. Thus, a salt bridge may form between (i) the polar and / or charged e amino acid residue and the polar and / or charged 1 amino acid residue, and / or (ii) the polar and / or charged g amino acid residue and the polar and / or charged n amino acid residue.

[0162] In some cases, the coiled-coil peptide pair comprises (i) a first coiled-coil peptide comprising a repeat of an amino sequence as set forth in (EXiX2X3X4X5Xe)m, where Xi, X2, X3, X4, X5, and X„ are each independently any amino acid residue, and wherein m is an integer greater than or equal to 2, and (ii) a second coiled-coil peptide comprising a repeat of an amino sequence as set forth in(KX7X8X9X10X11X12)11, where X7, Xs, X9, Xw, Xu, and X12 are each independently any amino acid residue, and wherein n is an integer greater than or equal to 2, e.g. 3, 4, 5, 6, 7, 8, 9, 10, or greater than 10. In some cases, the coiled-coil peptide pair comprises (i) a first coiled-coil peptide comprising a heptad repeat of an amino sequence as set forth in (EXiX2X3X4X5Xe)m, where Xi, X2, X3, X4, X5, and Xe are each independently any amino acid residue, and wherein m is an integer greater than or equal to 2, e.g. 3, 4, 5, 6, 7, 8, 9, 10, or greater than 10, and (ii) a second coiled-coil peptide comprising a heptad repeat of an amino sequence as set forth in (KX7X8X9X10X11X12)11, where X7, Xs, X9, Xw, Xu, and X12 are each independently any amino acid residue, and wherein n is an integer greater than or equal to 2, e.g. 3, 4, 5, 6, 7, 8, 9, 10, or greater than 10. In some cases, any one of Xi, X2, X3, X4, X5, Xe, X7, Xs, X9, Xw, Xu, X12, or any combination thereof, is a hydrophobic amino acid residue. In some cases, Xi or X4 is a hydrophobic amino acid residue. In some cases, Xi and X4 are each independently a hydrophobic amino acid residue. In some cases, Xi is isoleucine and X4 is leucine, or wherein Xi is leucine and X4 is isoleucine. In some cases, X7 or Xw is a hydrophobic amino acid residue, or X7 and X are each independently a hydrophobic amino acid residue. In some cases, X7 is isoleucine and X is leucine, or wherein X7 is leucine and X is isoleucine. In some cases, m can be 2, 3, 4, 5, 6, 7, 8, 9, 10, or greater than about 10. In some cases, m is 3 or 4. In some cases, n can be 2, 3, 4, 5, 6, 7, 8, 9, 10, or greater than about 10. In some cases, n is 3 or 4. In some cases, m and n can be each independently 2, 3, 4, 5, 6, 7, 8, 9, 10, or greater than about 10. In some cases, m and n are each independently 3 or 4.Table 8. Exemplary Coiled-Coil SequencesWSGR Docket No.:62697-750.601PLASMA MEMBRANE RECRUITMENT ELEMENT

[0163] In some aspects, the lipid delivery particle provided herein comprises a plasma membrane recruitment element. The lipid delivery particle disclosed herein can comprise a membrane. In some cases, the membrane encapsulates a payload. In some cases, the lipid delivery particle comprises a plasma membrane recruitment element, for example, inside the cavity of the lipid delivery particle. The plasma membrane recruitment element can localize itself to the membrane of the lipid delivery particles. The plasma membrane recruitment element can be utilized to recruit a component (e.g., a payload) to the membrane of the lipid delivery particles via forming a chimeric protein of the plasma membrane recruitment element and a component to be localized to the membrane or other mechanisms of attachment. In some cases, the membrane encapsulates a protein core. In some cases, at least a portion of the plasma membrane recruitment element forms the basic structure of the lipid delivery particle, such as a portion of the protein core inside the lipid delivery particle. In some cases, at least a portion of the plasma membrane recruitment element binds to the membrane of the lipid delivery particle from the inside.

[0164] The plasma membrane recruitment element can play a role in the assembly of the lipid delivery particle, such as packing various components (e.g., a payload) into the lipid delivery particles. The plasma membrane recruitment element can direct budding of the lipid delivery particles from a producer cell. In some cases, expressing plasma membrane recruitment element alone or together with an envelope protein disclosed herein in a producer cell can lead to formation of the lipid delivery particle.

[0165] In some cases, the plasma membrane recruitment element has a viral origin. For instance, the plasma membrane recruitment element comprises a retroviral gag protein, e.g. , a retroviral polyprotein that comprises one or more of a matrix (MA) polypeptide, an RNA-binding phosphoprotein polypeptide, a capsid (CA) polypeptide, or a nucleocapsid (NC) polypeptide. The plasma membrane recruitment element can comprise HIV gag or a biologically active mutant thereof. The plasma membrane recruitment element can comprise a gag from murine leukemia virus (MLV) or a biologically active mutant thereof. The plasma membrane recruitment element can comprise a gag from Moloney murine leukemia virus (MMLV) or a biologically active mutant thereof. In some cases, the plasma membrane recruitment element forms structural protein that forms the protein core of the lipid delivery particles described herein. The plasma membrane recruitment element can comprise Respiratory syncytial virus (RSV) M or a biologically active mutant thereof. The plasma membrane recruitment element can comprise Human Papillomavirus (HPV) LI protein or a biologically active mutant thereof. The plasma membraneWSGR Docket No.:62697-750.601 recruitment element can comprise HPV L2 protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise Hepatitis B virus (HBV) core protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise Hepatitis C virus (HCV) core protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise hepatitis E virus (HeV) M protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise Chikungunya virus (CHIKV) C-E3-E2-6k-El or a biologically active mutant thereof. The plasma membrane recruitment element can comprise RSV NP or a biologically active mutant thereof. The plasma membrane recruitment element can comprise Human metapneumovirus (HMPV) M or a biologically active mutant thereof. The plasma membrane can comprise a glycoprotein from a flavivirus. The flavivirus can comprise Chikungunya virus, Zika virus, Dengue virus, or West Niles virus. The plasma membrane recruitment element can comprise Zika virus (ZIKV) C or a biologically active mutant thereof. The plasma membrane recruitment element can comprise ZIKV prM / M or a biologically active mutant thereof. The plasma membrane recruitment element can comprise Dengue virus (DENV) C-prM or a biologically active mutant thereof. The plasma membrane recruitment element can comprise West Nile Virus (WNV) prME protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise WNV CprME protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise Filovirus VP40 or Z protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise Baculovirus Pl protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise Rotavirus VP7 or a biologically active mutant thereof. The plasma membrane recruitment element can comprise Rotavirus VP2 protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise Rotavirus VP6 protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise Porcine Circovirus Type 2 (PCV2) capsid or a biologically active mutant thereof. The plasma membrane recruitment element can comprise baculovirus VP2 protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise baculovirus VP5 protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise baculovirus VP3 protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise or baculovirus VP7 protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise Ebola nucleocapsid or a biologically active mutant thereof. The plasma membrane recruitment element can comprise Parovirus VP1 protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise Parovirus VP2 protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise Newcastle disease virus (NDV) M protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise Human polyomavirus 2 (JCPyV) VP1 protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise Human parainfluenza virus type 3 (HPIV3) M protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise HPIV3N protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise or MumpsWSGR Docket No.:62697-750.601 virus (MuV) M proteins or a biologically active mutant thereof. The plasma membrane recruitment element can comprise SARS M protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise SARS E protein or a biologically active mutant thereof. The plasma membrane recruitment element can comprise SARS N protein or a biologically active mutant thereof.

[0166] In some cases, the plasma membrane recruitment element is a mammalian protein or part thereof. For example, the plasma membrane recruitment element can include a pleckstrin homology (PH) domain or a transmembrane domain of a mammalian protein, such as a mouse protein or a human protein. In some cases, the plasma membrane recruitment element has a human origin. Utilizing the plasma membrane recruitment element of a human origin in the lipid delivery particle can give rise to reduced immunogenicity for administration to a human subject. The plasma membrane recruitment element can include a gag from human endogenous retrovirus, such as Human Endogenous Retrovirus K (e.g., HERV- K113, HERV-K101, HERV-K102, HERV-K104, HERV-K107, HERV-K108, HERV-K109, HERV- K115, HERV- KI lp22, and HERV-K12ql3) and Human Endogenous Retrovirus-W (HERV-W) or a biologically active mutant thereof. The plasma membrane recruitment element can include a hGAGKcon or a biologically active mutant thereof. The plasma membrane recruitment element can include an endogenous gag of a mammal (e.g., human) from retrotransposons (e.g., Arc from vertebrate lineage of Ty3 / gypsy retrotransposon), which are also ancestral to retroviruses. In some cases, the plasma membrane recruitment element comprises a portion from human Arc.

[0167] The plasma membrane recruitment element can include a pleckstrin homology (PH) domain from a mammalian protein or a biologically active mutant thereof. The plasma membrane recruitment element can include a pleckstrin homology (PH) domain from a human protein or a biologically active mutant thereof. The PH domains can play a role in protein-membrane interactions via binding to phosphatidylinositol phosphate (PIP), for example PIP2 or PIP3, or other lipids or proteins within the membrane of the lipid delivery particles. PH domains with different sequences can have varied affinities and selectivity when binding different PIPs. The plasma membrane recruitment element can include a PH domain of phospholipase C51 (e.g., human phospholipase C51) or a biologically active mutant thereof. The plasma membrane recruitment element can comprise a PH domain of Aktl (e.g. , human Aktl) or a biologically active mutant thereof. The plasma membrane recruitment element can comprise a mutant PH domain of human Aktl with E17K substitution or a biologically active mutant thereof. The plasma membrane recruitment element can comprise a PH domain of 3 -phosphoinositide-dependent protein kinase 1 (e.g., human 3-phosphoinositide-dependent protein kinase 1) or a biologically active mutant thereof. The plasma membrane recruitment element can comprise a PH domain of Dappl (e.g., human Dappl) or a biologically active mutant thereof. The plasma membrane recruitment element can comprise a PH domain of Grp 1 (e.g. , mouse Grp 1) or a biologically active mutant thereof. The plasma membrane recruitment element can comprise a PH domain of human Grpl or a biologically active mutant thereof. The plasma membrane recruitment element can comprise a PH domain of OSBP (e.g., human OSBP) or a biologically active mutant thereof. The plasma membrane recruitment element can comprise a PH domain of Btkl (e.g. , human Btkl) or a biologically active mutant thereof. The plasma membrane recruitmentWSGR Docket No.:62697-750.601 element can comprise a PH domain of FAPP1 (e.g., human FAPP1) or a biologically active mutant thereof. The plasma membrane recruitment element can comprise a PH domain of CERT (e.g., human CERT) or a biologically active mutant thereof. The plasma membrane recruitment element can comprise a PH domain of PKD (e.g. , human PKD) or a biologically active mutant thereof. The plasma membrane recruitment element can comprise a PH domain of PHLPP1 (e.g., human PHLPP1) or a biologically active mutant thereof. The plasma membrane recruitment element can comprise a PH domain of SWAP70 (e.g. , human SWAP70) or a biologically active mutant thereof. The plasma membrane recruitment element can comprise a PH domain of MAPKAP1 (e.g., human MAPKAP1) or a biologically active mutant thereof.

[0168] The plasma membrane recruitment element can also include a membrane protein (e.g. , a human membrane protein), a transmembrane domain thereof, or a biologically active mutant thereof. For example, the transmembrane domain of a human protein can be a tetraspanin or a biologically active mutant thereof. In some cases, the plasma membrane recruitment element comprises a transmembrane domain of human CD9 or a biologically active mutant thereof. In some cases, the plasma membrane recruitment element comprises a transmembrane domain of human CD47 or a biologically active mutant thereof. In some cases, the plasma membrane recruitment element comprises a transmembrane domain of human CD63 or a biologically active mutant thereof. In some cases, the plasma membrane recruitment element comprises a transmembrane domain of human CD81, or a biologically active mutant thereof.

[0169] The plasma membrane recruitment element can comprise a retroviral gag or a biologically active mutant thereof. The mutant of a retroviral gag can include only a portion of the retroviral gag. The plasma membrane recruitment element can include a gag of an alpha retrovirus or a biologically active mutant thereof. The plasma membrane recruitment element can a beta retrovirus or biologically active mutant thereof. The plasma membrane recruitment element can include a gamma retrovirus or biologically active mutant thereof. The plasma membrane recruitment element can include a delta retrovirus or biologically active mutant thereof. The plasma membrane recruitment element can include an epsilon retrovirus or biologically active mutant thereof. The plasma membrane recruitment element can include a spumavirus or biologically active mutant thereof. The retroviral gag can include a gag ofHIV (e.g., HIV-1), a gag of murine leukemia virus (MLV), a gag of Moloney murine leukemia virus (MMLV), a gag of Simian immunodeficiency virus (SIV), a gag of Rous sarcoma virus (RSV), a gag of human T-cell leukemia virus type-1 (HTLV), or a gag of bovine leukemia virus (BLV), or a biologically active mutant thereof. The plasma membrane recruitment element can include a gag of HIV (e.g., HIV - 1 ) or a biologically active mutant thereof. The plasma membrane recruitment element can include a gag of MLV or a biologically active mutant thereof. The plasma membrane recruitment element can include a gag of RSV or a biologically active mutant thereof. The plasma membrane recruitment element can include a gag of Friend murine leukemia virus (FMLV) or biologically active mutant thereof.

[0170] In some cases, the envelope protein comprises one or more of the sequences set forth in Table 4 with at least one amino acid substitution, deletion, or insertion. For instance, N-terminal methionine can be absent from the envelope protein of the lipid delivery particle provided herein relative to the wild-typeWSGR Docket No.:62697-750.601 viral envelope protein. In some cases, the envelope protein comprises one or more of the sequences set forth in Table 4 and a heterologous peptide sequence fused to the N-terminal or C-terminal.

[0171] In some cases, the envelope protein comprises any one of the sequences set forth in Table 4 with at least one amino acid substitution, deletion, or insertion. For instance, N-terminal methionine can be absent from the envelope protein of the lipid delivery particle provided herein relative to the wild-type viral envelope protein. In some cases, the envelope protein comprises any one of the sequences set forth in Table 4 and a heterologous peptide sequence fused to the N-terminal or C-terminal.

[0172] In some cases, the plasma membrane recruitment element comprises the sequences set forth in Table 4 with a further truncation on the N-terminus. For example, for those amino acid sequences start with a N-terminal methionine, the N-terminal methionine can be absent. In some cases, the plasma membrane recruitment element comprises the sequences set forth in Table 4 with a further truncation on the C-terminus. In some cases, the plasma membrane recruitment element comprises the sequences set forth in Table 4 with one amino acid substitution. In some cases, the plasma membrane recruitment element comprises the sequences set forth in Table 4 with two or more amino acid substitutions. In some cases, the plasma membrane recruitment element comprises the sequences set forth in Table 4 and a heterologous peptide sequence fused to the N-terminal or C-terminal.

[0173] In some cases, the plasma membrane recruitment element comprises an amino acid sequence that has at least about 50%, 60%, 70%, 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence set forth in Table 4. In some cases, the plasma membrane recruitment element comprises an amino acid sequence that has at least about 50%, 60%, 70%, 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence set forth in any one of SEQ ID NOs: 1-48. In some cases, the plasma membrane recruitment element comprises an amino acid sequence that has at least about 50% sequence identity to a sequence set forth in any one of SEQ ID NOs: 1-48. In some cases, the plasma membrane recruitment element comprises an amino acid sequence that has at least about 60% sequence identity to a sequence set forth in any one of SEQ ID NOs: 1-48. In some cases, the plasma membrane recruitment element comprises an amino acid sequence that has at least about 70% sequence identity to a sequence set forth in any one of SEQ ID NOs: 1-48. In some cases, the plasma membrane recruitment element comprises an amino acid sequence that has at least about 75% sequence identity to a sequence set forth in any one of SEQ ID NOs: 1-48. In some cases, the plasma membrane recruitment element comprises an amino acid sequence that has at least about 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence set forth in any one of SEQ ID NOs: 1-48. In some cases, the plasma membrane recruitment element comprises an amino acid sequence that has at least about 80% sequence identity to a sequence set forth in any one of SEQ ID NOs: 1-48. In some cases, the plasma membrane recruitment element comprises an amino acid sequence that has at least about 85% sequence identity to a sequence set forth in any one of SEQ ID NOs: 1-48. In some cases, the plasma membrane recruitment element comprises an amino acid sequence that has at least about 90% sequence identity to a sequence set forth in any one of SEQ ID NOs: 1-48. In some cases, the plasma membraneWSGR Docket No.:62697-750.601 recruitment element comprises an amino acid sequence that has at least about 95% sequence identity to a sequence set forth in any one of SEQ ID NOs: 1-48. In some cases, the plasma membrane recruitment element comprises an amino acid sequence that has at least about 96% sequence identity to a sequence set forth in any one of SEQ ID NOs: 1-48. In some cases, the plasma membrane recruitment element comprises an amino acid sequence that has at least about 97% sequence identity to a sequence set forth in any one of SEQ ID NOs: 1-48. In some cases, the plasma membrane recruitment element comprises an amino acid sequence that has at least about 98% sequence identity to a sequence set forth in any one of SEQ ID NOs: 1-48. In some cases, the plasma membrane recruitment element comprises an amino acid sequence that has at least about 99% sequence identity to a sequence set forth in any one of SEQ ID NOs: 1-48.Table 4. Exemplary plasma membrane recruitment elements and their sequencesWSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601*hGAGKconis a consensus sequence derived from ten proviral GAG sequences encoded by human genomic sequences. The GAG sequences used to derive this consensus GAG sequence are from the following HERVs: HERV-K113, HERV-K101, HERV-K102, HERV-K104, HERV-K107, HERVK108, HERV-K109, HERV-K115, HERV- KI lp22, and HERV-K12ql3.Engineered Gag Polyprotein

[0174] In some aspects, disclosed herein is a recombinant protein comprising a plasma membrane recruitment element comprising a mutant gag polyprotein. Also disclosed herein is a lipid delivery particle comprising a plasma membrane recruitment element comprising a mutant gag polyprotein. The mutant gag polyprotein is a mutant of a corresponding wild-type gag polyprotein. Relative to a wild-type gag polyprotein, a mutant gag polyprotein comprises an amino acid insertion, an amino acid deletion, an amino acid substitution or any combinations thereof. In some cases, a mutant gag polyprotein comprises at least about 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, or 98% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 40% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 45% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 50% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 55% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 60% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 65% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 70% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 75% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 80% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 85% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 90% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 91% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 92% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 93% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 94% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 95% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 96% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 97% identity to a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises at least about 98% identity to a corresponding wild-type gag polyprotein.WSGR Docket No.:62697-750.601

[0175] A wild-type gag polyprotein can comprise various domains, such as a matrix domain, an RNA- binding phosphoprotein, a capsid (including a major homology region), nucleocapsid, a particle budding motif (also termed as “budding domain” or “late domain”), a spacer peptide, a gag-endogenous protease cleavage sequence. A gag -endogenous protease cleavage sequence is the cleavage sequence homologous to a mutant gag polyprotein described herein, e.g., the gag-endogenous protease cleavage sequence is naturally present in the wild-type version of the mutant gag polyprotein. For example, a gag-endogenous protease cleavage sequence for MMLV gag includes MSKLLATW.

[0176] In some aspects of the present disclosure, the retroviral gag polyprotein in a lipid delivery particle provided herein is engineered as described herein. In some cases, the retroviral gag polyprotein in the chimeric protein comprising the gag poly protein and a payload protein is engineered as described herein (or a mutant described herein). In other cases, the retroviral gag polyprotein that forms the protein core of the lipid delivery particle, e.g., standalone gag polyprotein, or gag polyprotein present in gag / pro or gag / pro / pol polyprotein, is engineered as described herein (or a mutant described herein). In still other cases, both the retroviral gag polyprotein in the chimeric protein comprising the gag poly protein and a payload protein, and the retroviral gag polyprotein that forms the protein core of the lipid delivery particle, e.g., free / standalone gag polyprotein, or gag polyprotein present in gag / pro or gag / pro / pol polyprotein, are engineered (or a mutant described herein).

[0177] In some cases, the mutant gag polyprotein is a mutant of a wild-type gag polyprotein of a retrovirus. In some cases, the retrovirus is an endogenous retrovirus. In some cases, the mutant gag polyprotein is a mutant of a retroviral gag polyprotein. The mutant of a retroviral gag protein can include only a portion of the wild-type retroviral gag protein. In some cases, the wild type retroviral gag protein is endogenous. In some cases, the retroviral gag protein can comprise a retroviral nucleocapsid domain and / or a retroviral matrix domain. In some cases, the retroviral gag protein can be a gag homology protein. In some cases, the gag homology protein can be Arc protein (e.g., Arcl), Asprvl, a Sushi-Class protein, a SCAN protein, or a PNMA protein. In some cases, the gag homology protein can be ZCC18, ZCH12, PNM8B, PNM6A, PNMA6E_i2, PMA6F, PMAGE, PNMA1, PNMA2, PNM8A, PNMA3, PNMA4, PNMA5, PNMA6, PNMA7, PNMA1, M0AP1, or CCD8. In some cases, the Gag-homology protein is an Arc protein, e.g., hARC or dARCl. In some cases, the Gag -homology protein can comprise ASPRV1. In some cases, the Gag-homology protein is PEG10, RTL3, RTL10, or RTL1. In some cases, the Gag Homology protein is a SCAN protein, for example, PGBD1. In some cases, the PEG10 Gag homology protein is PEG10_i6 or PEG10_i2. In some cases, the gag homology protein can be Arc protein (e.g., Arcl), Asprvl, PNMA1, PNMA3, PNMA4, PNMA5, PNMA6, PNMA7, PEG 10, RTL1, M0AP1, or ZCCHC12.

[0178] The plasma membrane recruitment element can include a mutant of a wild-type gag of an alpha retrovirus. The plasma membrane recruitment element can include a mutant of a wild-type gag of a beta retrovirus. The plasma membrane recruitment element can include a mutant of a wild-type gag of a gamma retrovirus. The plasma membrane recruitment element can include a mutant of a wild-type gag of a delta retrovirus. The plasma membrane recruitment element can include a mutant of a wild-type gag ofWSGR Docket No.:62697-750.601 an epsilon retrovirus. The plasma membrane recruitment element can include a mutant of a wild-type gag of a spumavirus. The retroviral gag can include a mutant of a wild-type gag of HIV (e.g., HIV-1), a mutant of a wild-type gag of murine leukemia virus (MLV), a mutant of a wild-type gag of Moloney murine leukemia virus (MMLV), a mutant of a wild-type gag of Simian immunodeficiency virus (SIV), a mutant of a wild-type gag of Rous sarcoma virus (RSV), a mutant of a wild-type gag of human T-cell leukemia virus type-1 (HTLV), or a mutant of a wild-type gag of bovine leukemia virus (BLV). The plasma membrane recruitment element can include a mutant of a wild-type gag of HIV (e.g., HIV-1). The plasma membrane recruitment element can include a mutant of a wild-type gag of MLV. The plasma membrane recruitment element can include a mutant of a wild-type gag of MMLV. The plasma membrane recruitment element can include a mutant of a wild-type gag of RSV. The plasma membrane recruitment element can include a mutant of a wild-type gag of Friend murine leukemia virus (FMLV). The plasma membrane recruitment element can include a mutant of a wild-type gag of Rous Sarcoma Virus. The plasma membrane recruitment element can include a mutant of a wild-type gag of Avian Leukosis Virus. The plasma membrane recruitment element can include a mutant of a wild-type gag of Avian Myeloblastosis Virus. The plasma membrane recruitment element can include a mutant of a wildtype gag of Avian Myeloblastosis Virus. The plasma membrane recruitment element can include a mutant of a wild-type gag of Human T cell Leukemia Virus- 1. The plasma membrane recruitment element can include a mutant of a wild-type gag of Mouse Mammary Tumor Virus. The plasma membrane recruitment element can include a mutant of a wild-type gag of Jaagsiekte sheep retrovirus. The plasma membrane recruitment element can include a mutant of a wild-type gag of Mason-Pfizer Monkey Virus. In some cases, a wild-type gag polyprotein comprises an amino acid sequence selected from any one of sequence set forth in Table 9. In some cases, a wild-type gag polyprotein comprises an amino acid sequence selected from any one of SEQ ID Nos: 285-305.

[0179] In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 50%, 60%, 70%, 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to any one of sequence set forth in Table 9. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 50% sequence identity to any one of sequence set forth in Table 9. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 60% sequence identity to any one of sequence set forth in Table 9. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 70% sequence identity to any one of sequence set forth in Table 9. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 75% sequence identity to any one of sequence set forth in Table 9. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 75% sequence identity to any one of sequence set forth in Table 9. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 80% sequence identity to any one of sequence set forth in Table 9. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 85% sequence identity to any one of sequence set forth in Table 9. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 90% sequence identityWSGR Docket No.:62697-750.601 to any one of sequence set forth in Table 9. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 95% sequence identity to any one of sequence set forth in Table 9. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 96% sequence identity to any one of sequence set forth in Table 9. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 97% sequence identity to any one of sequence set forth in Table 9. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 98% sequence identity to any one of sequence set forth in Table 9. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 99% sequence identity to any one of sequence set forth in Table 9.

[0180] In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 50%, 60%, 70%, 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to any one of sequence set forth in SEQ ID NO: 285-305. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 50% sequence identity to any one of sequence set forth in SEQ ID NO: 285-305. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 60% sequence identity to any one of sequence set forth in SEQ ID NO: 285-305. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 70% sequence identity to any one of sequence set forth in SEQ ID NO: 285-305. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 75% sequence identity to any one of sequence set forth in SEQ ID NO: 285-305. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 75% sequence identity to any one of sequence set forth in SEQ ID NO: 285-305. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 80% sequence identity to any one of sequence set forth in SEQ ID NO: 285-305. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 85% sequence identity to any one of sequence set forth in SEQ ID NO: 285-305. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 90% sequence identity to any one of sequence set forth in SEQ ID NO: 285-305. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 95% sequence identity to any one of sequence set forth in SEQ ID NO: 285-305. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 96% sequence identity to any one of sequence set forth in SEQ ID NO: 285-305. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 97% sequence identity to any one of sequence set forth in SEQ ID NO: 285-305. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 98% sequence identity to any one of sequence set forth in SEQ ID NO: 285-305. In some cases, the mutant gag polyprotein comprises an amino acid sequence that has at least about 99% sequence identity to any one of sequence set forth in SEQ ID NO: 285-305. In some cases, the mutant gag polyprotein comprises at least 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 94%, 95%, 96%, 97%, 98%, or 99% of the entire amino acid sequence of the wild type gag polyprotein that comprises an amino acid sequence selected from Table 9. In some cases, the mutant gagWSGR Docket No.:62697-750.601 polyprotein comprises at least 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 94%, 95%, 96%, 97%, 98%, or 99% of the entire amino acid sequence of the wild type gag polyprotein that comprises an amino acid sequence set forth in any one of SEQ ID NO: 285-305. In some cases, the mutant gag polyprotein comprises at least 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 85%, 90%, 91%, 92%, 94%, 95%, 96%, 97%, 98%, or 99% of the entire amino acid sequence of the wild type gag polyprotein that comprises an amino acid sequence selected from Table 9. In some cases, the mutant gag polyprotein comprises at least 1, 2, 3, 4, 5, 6, 7, 8, 9 ,10, 20, 30, 40, 50, 60, 70, 80, 90, 100 or more mutations (e.g., amino acid deletions, insertion and / or substitutions) as compared to a wild-type polyprotein. In some cases, the mutant gagpolyprotein comprises a truncation (e.g., at the N-terminus and / or at the C-terminus) of at least about 1, 2, 2, 3, 4, 5, 6, 7, 8, 9 ,10, 20, 30, 40, 50, 60, 70, 80, 90, 100 or more contiguous amino acid residues as compared to the wild type polyprotein.

[0181] In some cases, the mutant gag polyprotein comprises at least 1, 2, 3, 4, 5, 6, 7, 8, 9 ,10, 20, 30, 40, 50, 60, 70, 80, 90, 100 or more mutations (e.g., amino acid deletions, insertion and / or substitutions) as compared to a wild-type polyprotein that comprises an amino acid sequence selected from Table 9. In some cases, the mutant gag-polyprotein comprises a truncation (e.g., at the N-terminus and / or at the C- terminus) of at least about 1, 2, 2, 3, 4, 5, 6, 7, 8, 9 ,10, 20, 30, 40, 50, 60, 70, 80, 90, 100 or more contiguous amino acid residues as compared to the wild type polyprotein that comprises an amino acid sequence set forth in any one of SEQ ID NO: 285-305.

[0182] The plasma membrane recruitment element can comprise a mutant of HIV gag. The plasma membrane recruitment element can comprise a mutant of a gag from murine leukemia virus (MLV). The plasma membrane recruitment element can comprise a mutant of a gag from Moloney murine leukemia virus (MMLV).

[0183] The plasma membrane recruitment element can comprise a mutant of Respiratory syncytial virus (RSV) M. The plasma membrane recruitment element can comprise a mutant of Human Papillomavirus (HPV) LI protein. The plasma membrane recruitment element can comprise a mutant of HPV L2 protein. The plasma membrane recruitment element can comprise a mutant of Hepatitis B virus (HBV) core protein. The plasma membrane recruitment element can comprise a mutant of Hepatitis C virus (HCV) core protein. The plasma membrane recruitment element can comprise a mutant of hepatitis E virus (HeV) M protein. The plasma membrane recruitment element can comprise a mutant of Chikungunya virus (CHIKV) C-E3-E2-6k-El . The plasma membrane recruitment element can comprise a mutant of RSV NP. The plasma membrane recruitment element can comprise a mutant of Human metapneumovirus (HMPV) M. The plasma membrane recruitment element can comprise a mutant of Influenza Ml . The plasma membrane recruitment element can comprise a mutant of Zika virus (ZIKV) C. The plasma membrane recruitment element can comprise a mutant of ZIKV prM / M. The plasma membrane recruitment element can comprise a mutant of Dengue virus (DENV) C-prM. The plasma membrane recruitment element can comprise a mutant of West Nile Virus (WNV) prME protein. The plasma membrane recruitment element can comprise a mutant of WNV CprME protein. The plasma membraneWSGR Docket No.:62697-750.601 recruitment element can comprise a mutant of Filovirus VP40 or Z protein. The plasma membrane recruitment element can comprise a mutant of Baculovirus P 1 protein. The plasma membrane recruitment element can comprise a mutant of Rotavirus VP7. The plasma membrane recruitment element can comprise a mutant of Rotavirus VP2 protein. The plasma membrane recruitment element can comprise a mutant of Rotavirus VP6 protein. The plasma membrane recruitment element can comprise a mutant of Porcine Circovirus Type 2 (PCV2) capsid. The plasma membrane recruitment element can comprise a mutant of baculovirus VP2 protein. The plasma membrane recruitment element can comprise a mutant of baculovirus VP5 protein. The plasma membrane recruitment element can comprise a mutant of baculovirus VP3 protein. The plasma membrane recruitment element can comprise a mutant of or baculovirus VP7 protein. The plasma membrane recruitment element can comprise a mutant of Ebola nucleocapsid. The plasma membrane recruitment element can comprise a mutant of Parovirus VP1 protein. The plasma membrane recruitment element can comprise a mutant of Parovirus VP2 protein. The plasma membrane recruitment element can comprise a mutant of Newcastle disease virus (NDV) M protein. The plasma membrane recruitment element can comprise a mutant of Nipah virus (NIV) M protein. The plasma membrane recruitment element can comprise a mutant of Human polyomavirus 2 (JCPyV) VP1 protein. The plasma membrane recruitment element can comprise a mutant of Human parainfluenza virus type 3 (HPIV3) M protein. The plasma membrane recruitment element can comprise a mutant of HPIV3N protein. The plasma membrane recruitment element can comprise a mutant of or Mumps virus (MuV) M proteins. The plasma membrane recruitment element can comprise a mutant of SARS M protein. The plasma membrane recruitment element can comprise a mutant of SARS E protein. The plasma membrane recruitment element can comprise a mutant of SARS N protein.

[0184] In some cases, the plasma membrane recruitment element comprises a mutant gag polyprotein that lacks at least a fragment of a matrix protein as compared to a wild-type gag polyprotein. In some cases, the plasma membrane recruitment element comprises a mutant gag polyprotein that lacks at least a fragment of a RNA-binding phosphoprotein as compared to a wild-type gag polyprotein, for example, MMLV gag. In some cases, the plasma membrane recruitment element comprises a mutant gag polyprotein that lacks at least a fragment of at least a fragment of a capsid protein as compared to a wildtype gag polyprotein. In some cases, the plasma membrane recruitment element comprises a mutant gag polyprotein that lacks at least a fragment of a gag-endogenous protease cleavage sequence as compared to a wild-type gag polyprotein. In some cases, the plasma membrane recruitment element comprises a mutant gag polyprotein that lacks a gag -endogenous protease cleavage sequence as compared to a wildtype gag polyprotein. In some cases, the plasma membrane recruitment element comprises a mutant gag polyprotein that lacks at least a fragment of a gag-endogenous protease cleavage sequence as compared to a wild-type gag polyprotein. In some cases, the plasma membrane recruitment element comprises a mutant gag polyprotein that comprises a mutant gag -endogenous protease cleavage sequence as compared to a wild-type polyprotein. In some cases, the mutant gag-endogenous protease cleavage sequence comprises at least 1, 2, 3, 4, 5, 6, 7, 8, 9 ,10, 20, 30, 40, 50, 60, 70 , 80, 90 , 100 or more mutations (e.g. amino acid deletions, insertion and / or substitutions) as compared to a wild-type polyprotein. In someWSGR Docket No.:62697-750.601 cases, the mutant gag-endogenous protease cleavage sequence comprises a truncation (e.g., at the N- terminus and / or at the C-terminus) of at least about 1, 2, 2, 3, 4, 5, 6, 7, 8, 9 ,10, 20, 30, 40, 50, 60, 70 , 80, 90 , 100 or more contiguous amino acid residues as compared to a wild type polyprotein. In some cases, the plasma membrane recruitment element comprises a mutant gag polyprotein that comprises a mutant gag-endogenous protease cleavage sequence as compared to a wild-type polyprotein, wherein the mutant gag-endogenous protease cleavage sequence cannot be recognized or cleaved by the gag- endogenous protease. In some cases, the mutant gag -endogenous protease cleavage sequence is recognized or cleaved by a different protease than the gag -endogenous protease that recognizes the corresponding wild type gag-endogenous protease cleavage sequence. In some cases, the mutant gag- endogenous protease cleavage sequence exhibits improved cleavage (higher cleavage efficiency) relative to that by corresponding wild type gag -endogenous protease cleavage sequence. In some cases, the plasma membrane recruitment element comprises a mutant gag polyprotein that comprises (e.g., at least one, 2, 3, 4, 5 or more) exogenous protease cleavage sequences as compared to a wild-type polyprotein. In some cases, an exogenous protease cleavage sequence is a protease cleavage sequence that does not naturally occur in the corresponding wild type polyprotein sequence. In some cases, an exogenous protease cleavage sequence links one or more domains of the mutant gag polyprotein.

[0185] In some cases, the plasma membrane recruitment element comprises a mutant gag polyprotein that lacks at least a fragment of a nucleocapsid protein as compared to a wild-type gag polyprotein. In some cases, the plasma membrane recruitment element comprises a mutant gag polyprotein, for example, a mutant of wild-type HIV gag, a mutant of wild-type MMLV gag, a mutant of wild-type RSV gag, and a mutant of wild-type HERV gag.

[0186] In some cases, the plasma membrane recruitment element comprises a full-length capsid protein (i.e., the capsid protein of the wild-type gag). In some cases, the plasma membrane recruitment element comprises a mutant gag polyprotein comprising a fragment of a capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 20% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 25% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 30% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 35% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 40% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 45% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 50% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 55% identity to theWSGR Docket No.:62697-750.601 capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 60% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 65% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 70% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 75% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 80% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 85% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 90% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 91% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 92% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 93% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 94% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 95% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 96% identity to the capsid protein of a corresponding wildtype gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 97% identity to the capsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a capsid protein comprising at least about 98% identity to the capsid protein of a corresponding wild-type gag polyprotein.

[0187] In some cases, the plasma membrane recruitment element comprises a full-length matrix protein (i.e., the matrix protein of the wild-type gag). In some cases, the plasma membrane recruitment element comprises a mutant gag polyprotein comprising a fragment of a matrix protein of a corresponding wildtype gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 20% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 25% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 30% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 35% identity to the matrixWSGR Docket No.:62697-750.601 protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 40% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 45% identity to the matrix protein of a corresponding wildtype gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 50% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 55% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 60% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 65% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 70% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 75% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 80% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 85% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 90% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 91% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 92% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 93% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 94% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 95% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 96% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 97% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a matrix protein comprising at least about 98% identity to the matrix protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein lacks the matrix protein of a corresponding wild-type gag polyprotein.WSGR Docket No.:62697-750.601

[0188] In some cases, the plasma membrane recruitment element comprises a full-length nucleocapsid protein (i.e., the nucleocapsid protein of the wild-type gag). In some cases, the plasma membrane recruitment element comprises a mutant gag polyprotein comprising a fragment of a nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 20% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 25% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 30% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 35% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 40% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 45% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 50% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 55% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 60% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 65% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 70% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 75% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 80% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 85% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 90% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 91% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 92% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragmentWSGR Docket No.:62697-750.601 of a nucleocapsid protein comprising at least about 93% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 94% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 95% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 96% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 97% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein comprises a fragment of a nucleocapsid protein comprising at least about 98% identity to the nucleocapsid protein of a corresponding wild-type gag polyprotein. In some cases, a mutant gag polyprotein lacks the nucleocapsid protein of a corresponding wild-type gag polyprotein.

[0189] In some cases, the plasma membrane recruitment element comprises heterologous sequence. In some cases, the heterologous sequence is fused to a fragment of the matrix protein, such as the N-terminal of the fragment of the matrix protein. In some cases, the heterologous sequence is fused to a fragment of the capsid protein, such as the N-terminal of the fragment of the capsid protein. In some cases, the heterologous sequence is fused to a matrix protein or a fragment thereof. In some cases, the heterologous sequence is fused to a nucleocapsid protein or a fragment thereof. In some cases, the heterologous sequence is fused to a particle budding motif or a fragment thereof (e.g., a late domain of RSV). In some cases, the heterologous sequence is a pleckstrin homology (PH) domain or a mutant thereof, such as those listed in Table 4. In some cases, the PH domain is a human PH domain. In some cases, the fragment of the matrix protein lacks a N-terminal myristoylation motif, a N-terminal glycine, a highly basic region (HBR), a N-terminal fragment, or any combinations thereof. In some cases, the plasma membrane recruitment element comprises at least a portion of a nucleocapsid protein. In some cases, the plasma membrane recruitment element comprises two or more particle budding motifs described herein. In some cases, at least one of the two or more particle budding motifs is endogenous. In some cases, at least one of the two or more particle budding motifs is heterologous, such as RSV P2B or a mutant thereof. In some cases, at least one of the two or more particle budding motifs comprises a PTAP motif, a PPPY motif, a PPXY motif, a YXXL motif, a YPXnL motif, or any combinations thereof, and wherein n is an integer from 1 to 5. In some cases, at least one of the two or more particle budding motifs comprises a sequence listed in Table 5. In some cases, the plasma membrane recruitment element comprises the mutant gag polyprotein that is a mutant of a wild-type gag polyprotein of HIV and comprises at least a portion of a P6 domain. In some cases, the plasma membrane recruitment element comprises at least a N-terminal fragment of the P6 domain. In some cases, the plasma membrane recruitment element comprises at least a C-terminal fragment of the P6 domain. In some cases, the plasma membrane recruitment element comprises two spacer peptides (e.g., P2 domain, Pl domain, or both). In some cases, the plasma membrane recruitment element is fused to the post-translational modification motif as described herein.WSGR Docket No.:62697-750.601In some cases, the post-translational modification motif is fused to the N-terminus of the plasma membrane recruitment element. In some cases, the post-translational modification motif is fused to the C- terminus of the plasma membrane recruitment element. In some cases, the post-translational modification motif is a myristoylation motif. In some cases, the post-translational modification motif is an acetylation motif. In some cases, the post-translational modification motif is an isoprenylation motif. In some cases, the post-translational modification motif is a palmitoylation motif. In some cases, the post-translational modification motif is a famesylation motif. In some cases, the plasma membrane recruitment element is fused to the membrane penetrating peptide. In some cases, the membrane penetrating peptide is linked to the plasma membrane recruitment element. In some cases, the membrane penetrating peptide connects the plasma membrane recruitment element and a payload. In some cases, the membrane penetrating peptide comprises a sequence having at least about 80% sequence identity to a sequence listed in Table 6.

[0190] In some cases, the plasma membrane recruitment element comprises a mutant gag polyprotein of a wild-type Rous Sarcoma Virus (RSV) gag polyprotein. In some cases, the plasma membrane recruitment element lacks a capsid protein of the RSV gag. In some cases, the plasma membrane recruitment element lacks a nucleocapsid protein of the RSV gag. In some cases, the plasma membrane recruitment element comprises at least a portion of the matrix protein of the RSV gag. In some cases, the plasma membrane recruitment element comprises a matrix protein of the RSV gag and a late domain of the RSV gag. In some cases, the plasma membrane recruitment element comprises a matrix protein of the RSV gag, a late domain of the RSV gag, and a full-length nucleocapsid protein of the RSV. In some cases, the plasma membrane recruitment element comprises a matrix protein of the RSV gag, a late domain of the RSV gag, and a full- length nucleocapsid protein of the RSV arranged in order from N-terminus to C-terminus of the plasma membrane recruitment element. In some cases, the plasma membrane recruitment element comprises a matrix protein of the RSV gag, a late domain of the RSV gag, and a fragment of the nucleocapsid protein of the RSV gag. In some cases, the plasma membrane recruitment element comprises a matrix protein of the RSV gag, a late domain of the RSV gag, and a fragment of the nucleocapsid protein of the RSV gag arranged in order from N-terminus to C-terminus of the plasma membrane recruitment element. In some cases, the plasma membrane recruitment element further comprises a N-terminal myristoylation motif. In some cases, the plasma membrane recruitment element comprises a matrix protein of the RSV gag and a late domain of the RSV gag arranged in order from N-terminus to C-terminus of the plasma membrane recruitment element. In some cases, the plasma membrane recruitment element consists of a matrix protein of the RSV gag and a late domain of the RSV gag arranged in order from N-terminus to C-terminus of the plasma membrane recruitment element. In some cases, the plasma membrane recruitment element comprises a matrix protein of the RSV gag, a late domain of the RSV gag, and a fragment of a full-length nucleocapsid of the RSV gag arranged in order from N-terminus to C-terminus of the plasma membrane recruitment element. In some cases, the plasma membrane recruitment element consists of a matrix protein of the RSV gag, a late domain of the RSV gag, and a fragment of a full-length nucleocapsid of the RSV gag arranged in order from N-terminus to C-terminus of the plasma membrane recruitment element. In some cases, the plasma membrane recruitment element comprises a matrix protein of the RSV gag, a lateWSGR Docket No.:62697-750.601 domain of the RSV gag, and a fragment of a full-length nucleocapsid of the RSV gag arranged in order from N-terminus to C-terminus of the plasma membrane recruitment element. In some cases, the plasma membrane recruitment element consists of a myr group of the RSV gag, a late domain of the RSV gag, and a fragment of a full-length nucleocapsid of the RSV gag arranged in order from N-terminus to C-terminus of the plasma membrane recruitment element. In some cases, the plasma membrane recruitment element comprises a myr group of the RSV gag, a late domain of the RSV gag, and a fragment of a full-length nucleocapsid of the RSV gag arranged in order from N-terminus to C-terminus of the plasma membrane recruitment element. In some cases, the plasma membrane recruitment element consists of a myr group of the RSV gag, a late domain of the RSV gag, and a full-length nucleocapsid of the RSV gag arranged in order from N-terminus to C-terminus of the plasma membrane recruitment element. In some cases, the plasma membrane recruitment element comprises a myr group of the RSV gag, a late domain of the RSV gag, and a full-length nucleocapsid of the RSV gag arranged in order from N-terminus to C-terminus of the plasma membrane recruitment element. In some cases, a plasma membrane recruitment element is linked to a payload (e.g., a base editor), for example, via a cleavable linker, which forms a chimeric protein descried herein. In some cases, a plasma membrane recruitment element is linked to a protease, for example, with or without a cleavable linker.Table 9. Exemplary wild type gag polyprotein and the corresponding amino acid sequencesWSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601

[0191] In some cases, the lipid delivery particle disclosed herein comprises a protein core that is composed of at least a structural protein of a viral origin, for instance, a retroviral gag protein. In some of these cases, the lipid delivery particle comprises a retroviral gag-pro-pol polyprotein, e.g., a gag-pro-pol poly protein from HIV, MMLV, or FMLV, which can help assemble a protein core of the lipid delivery particle. In some of these cases, some of the gag-pro-pol polyprotein is cleaved, e.g., by pro (protease) present freely or in the gag-pro-pol polyprotein. Without wishing to be bound by any particular theory, the cleavage by pro can be inefficient, and the resultant cleavage products can include gag polyprotein, gag -pro polyprotein, free pro, and free pol (polymerase). In some cases, a retroviral gag polyprotein can be further cleaved into MA, CA, NC, and other small fragments, if any. In some other cases, the lipid delivery particle comprises a retroviral gag-pro polyprotein without the pol component, and the gag-pro polyprotein can help form a protein core of the lipid delivery particle. The gag -pro can also be cleaved by pro, in some cases, inefficiently, into separate gag and pro proteins. In some cases, there can be different plasma membrane recruitment elements in a lipid delivery particle. For instance, a gag -pro or gag-pro-pol polyprotein from one species of virus (e.g. , a retrovirus, e.g. , a HIV) can help assemble and form a protein core of the lipid delivery particle, while a chimeric protein in the lipid delivery particle, discussed infra, can comprise a payload fused with a gag protein from a different species of virus (e.g., an MMLV), or from a HERV, or a PH domain or transmembrane domain of a huma protein (e.g. , a PH domain of human Aktl with E17K substitution).

[0192] In some aspects of the present disclosure, a lipid delivery particle provided herein does not comprise a retroviral pol protein either standalone or in fusion. For instance, in a lipid delivery particle according to some embodiments of the present disclosure, there can be a retroviral gag polyprotein and gag / pro polyprotein, neither of which is fused to a retroviral pol protein, and the retroviral gag polyprotein and gag / pro polyprotein can help assemble and form a protein core of the lipid delivery particle. In some of these embodiments, such lipid delivery particle is produced by introducing into producer cells nucleic acid sequence that encodes the gag polyprotein and nucleic acid sequence that encodes the gag / pro polyprotein, without introducing into the producer cell any nucleic acid sequence encoding retroviral pol polyprotein.

[0193] In some cases, a gag polyprotein is engineered such that it lacks at least a fragment of a matrix (MA) protein. In some of these cases, the gag polyprotein lacks MA, i.e., the full length of MA. In some of these cases, the gag polyprotein that lacks MA is further modified at its N-terminus, for instance, is fused C-terminal to a heterologous sequence. In some cases, the gag polyprotein that lacks MA is fused C-terminal to a myristoylation motif, e.g., any of the myristoylation motif sequences set forth in Table 7WSGR Docket No.:62697-750.601 or Table 10. In some cases, the gag polyprotein that lacks MA is fused C-terminal to a c-Src N-terminus sequence, e.g., the sequence set forth in SEQ ID NO: 653. In some of these cases, the gag polyprotein that lacks MA is fused C-terminal to a heterologous domain, e.g., a pleckstrin homology domain, such as PH domain of Aktl with E17K mutation, the PH domain is from a protein selected from the group consisting of human phospholipase C51, human Aktl, human Aktl with E17K substitution, human 3- phosphoinositide-dependent protein kinase 1, human Daapl, mouse Grpl, human Grpl, human OSBP, human Btkl, human FAPP1, human CERT, human PKD, human PHLPP1, human SWAP70, and human MAPKAP1. In some of these cases, the PH domain comprises at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or 100% sequence identity to any amino acid sequence set forth in any one of SEQ ID NOs: 5-14 and 23-48.

[0194] In some cases, the gag polyprotein lacks at least a fragment of its pl2 protein. In some of these cases, the gag polyprotein lacks pl2, i.e., the full length of pl2 protein. In some cases, the pl2 protein in the gag polyprotein is truncated with only PPPY domain of the pl2 protein is retained.

[0195] In some cases, the gag polyprotein lacks at least a fragment of its capsid (CA) protein. In some of these cases, the gag polyprotein lacks N-terminal domain of CA protein. In some of these cases, the gag polyprotein lacks C-terminal domain of the CA protein. In some cases, the gag polyprotein lacks CA, i.e., the full length of the CA protein.

[0196] In some cases, the gag polyprotein lacks at least a fragment of a nucleocapsid (NC) protein. In some of these cases, the gag polyprotein lacks NC, i.e., the full length of NC protein. In some of these cases, the gag polyprotein that lacks NC is fused N-terminal to a heterologous domain, e.g., a leucine zipper peptide, e.g., the sequence set forth in any one of SEQ ID NOs: 311-327. In some of these cases, the gag polyprotein that lacks NC is fused N-terminal to a heterologous domain, e.g., a leucine zipper peptide, e.g., the sequence set forth in any one of the amino acid sequences of Table 16. In some cases, a leucine zipper described herein can comprise a dimer, trimer, tetramer, pentamer, hexamer, or heptamer.Table 16. Exemplary leucine zipper amino acid sequences.WSGR Docket No.:62697-750.601Leucine Zipper LQRMKQLEDKVEELLSKNYHLENEVARLKKLVGD 327

[0197] In some embodiments, the zipper motif is encoded by a nucleotide seqeunce as set forth in Table 17. The zipper motif may be encoded by a nucleotide seqeunce as set forth in any one of SEQ ID Nos: 328-342.Table 17. Exemplary nucleotide sequences encoding leucine zipper motifs

[0198] In some cases, the gag polyprotein is engineered such that it comprises a linker sequence, e.g., a cleavable linker or a non-cleavable linker, flanking a heterologous domain (e.g. , a leucine zipper peptide). The non-cleavable linker sequence can be any suitable linker sequence, such as SGGS or XTEN linker. In some cases, the linker sequence may be an amino acid sequence as set forth in SEQ ID NO: 673 or 343- 346. In some cases, the linker sequence may be an amino acid sequence as set forth in Table 18. In some cases, the gag polyprotein comprises more than one repeat of the linker sequence flanking the heterologous domain (e.g., leucine zipper peptide) on its both sides, such as 2, 3, 4, 5, or 6, or more repeats.Table 18. Exemplary linker sequencesWSGR Docket No.:62697-750.601

[0199] In some cases, the gag / pro, gag / pro / pol, or both present in the lipid delivery particle comprises one or more modifications to the gag protein as described herein. In some cases, the gag / pro, gag / pro / pol, or both present in the lipid delivery particle comprises a linker sequence flanking pro and / or pol protein on its both sides, such as, at least one repeat (e.g., 2, 3, 4, 5, or 6 or more repeats) of SGGS or XTEN linker. In some cases, the linker sequence may be an amino acid sequence as set forth in SEQ ID NO: 673 or 343-346. In some cases, the linker sequence may be an amino acid sequence as set forth in Table 18.

[0200] In some cases, the engineered gag polyprotein comprises an amino acid sequence that has at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or 100% sequence identity to any amino acid sequence set forth in SEQ ID NOs: 651, 652, 654, 656-659, 661, 663-665, 667, 669, 671, 674, and 677. In some cases, the lipid delivery particle comprises a protein that has at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, at least 99%, or 100% sequence identity to any amino acid sequence set forth in Table 10.

[0201] In some cases, the gag polyprotein is a retroviral gag polyprotein. In some cases, the gag polyprotein is a gag polyprotein from human immunodeficiency virus (HIV), murine leukemia virus (MLV), Moloney murine leukemia virus (MMLV), Friend murine leukemia virus (FMLV), Baboon endogenous retrovirus (BaEV), Simian immunodeficiency virus (SIV), Rous sarcoma virus (RSV), human T-cell leukemia virus type-1 (HTLV), bovine leukemia virus (BLV), Feline Leukemia Virus (FeLV), Gibbon Ape Leukemia Virus (GaLV), Koala Retrovirus (KRV), Reticuloendotheliosis Virus (ReEV), Wooly Monkey Sarcoma Virus (WMSV), or a biologically active mutant thereof, or any combination thereof. In some cases, the gag polyprotein is a human endogenous retroviral gag polyprotein.

[0202] In some cases, the gag polyprotein comprises a sequence having at least 80%, at least 85%, at least 90%, or at least 95%, or 100% sequence identity to the amino acid sequence set forth in any one of SEQ ID NO: 285-309. In some cases, the gag polyprotein lacks at least a fragment of a nucleocapsid protein, optionally lacks the full length of the nucleocapsid protein.

[0203] In some cases, the plasma membrane recruitment element disclosed herein comprises an engineered gag polyprotein. In some cases, the engineered gag polyprotein lacks at least a fragment of a nucleocapsid protein, and the plasma membrane recruitment element further comprises a heterologous domain fused to C-terminus of the engineered gag polyprotein. In some cases, the engineered gag polyprotein lacks at least a fragment of a matrix protein, and the plasma membrane recruitment element further comprises a pleckstrin homology (PH) domain fused to N-terminus of the engineered gag polyprotein. In some cases, the engineered gag polyprotein comprises a matrix protein and a capsid protein, and lacks at least a fragment of a nucleocapsid protein, and the plasma membrane recruitment element further comprises a heterologous domain fused to C-terminus of the engineered gag polyprotein.WSGR Docket No.:62697-750.601In some cases, the heterologous domain comprises a leucine zipper. The leucine zipper may be fused to a terminus of the gag polyprotein (e.g., the C-terminus or the N-terminus of the gag polyprotein). In some cases, the leucine zipper can be fused to a C-terminus of the gag polyprotein. In some cases, the leucine zipper can be fused to a N-terminus of the gag polyprotein. The gag polyprotein may be an intact gag polyprotein (e.g., the gag polyprotein does not lack a matrix protein, a capsid protein, or a nucleocapsid protein). In some cases, the gag polyprotein may lack a fragment of a matrix protein, a fragment of a capsid protein, a fragment of a nucleocapsid protein, or any combination thereof. In some cases, the gag polyprotein may lack a matrix protein, a capsid protein, a nucleocapsid protein, or any combination thereof.

[0204] In some cases, the plasma membrane recruitment element can comprise one or more leucine zippers (e.g., 1, 2, 3, 4, 5, or more leucine zippers). The one or more leucine zippers may be within the gag protein. For example, the one or more leucine zippers may be between any two domains of a gag protein. The one or more leucine zippers may replace one or more domains of a gag protein. For example, a leucine zipper may replace a nucleocapsid domain (e.g., a nucleocapsid fragment), a matrix domain (e.g., a matrix fragment), a pl2 domain (e.g., a pl2 fragment), a capsid domain (e.g., a capsid fragment), or any combination thereof. In some cases, a leucine zipper can be between a matrix domain and a pl2 domain. In some cases, a leucine zipper can be between a pl2 domain and a capsid domain. In some cases, a leucine zipper can be between a capsid domain and a nucleocapsid domain.Table 10. Exemplary sequences for engineered gag and gag constructsWSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601CHIMERIC PROTEIN

[0205] In aspects, the present disclosure provides a chimeric protein comprising a plasma membrane recruitment element and a payload that is a protein or a fragment thereof. In some aspects, the lipid delivery particle comprises a chimeric protein comprising a plasma membrane recruitment element and a payload that is a protein or a fragment thereof. In some cases, the plasma membrane recruitment element and the payload are fused directly in the chimeric protein. In other cases, the plasma membrane recruitment element and the payload are fused indirectly via a linker. In some cases, the linker between the plasma membrane recruitment element and the payload is a cleavable linker that is recognized by a protease.

[0206] In aspects, the present disclosure provides a chimeric protein comprising a plasma membrane recruitment element and coiled-coil peptide. In aspects, the present disclosure provides a chimeric protein comprising coiled-coiled peptide and a payload that is a protein or a fragment thereof. In some aspects, the lipid delivery particle comprises a first chimeric protein comprising a plasma membrane recruitment element and a first coiled-coil peptide, and a second chimeric protein comprising a second coiled-coil peptide and a payload that is a protein or a fragment thereof, and the first coiled-coil peptide and the second coiled-coil peptide form a coiled-coil pair. In some cases, the components of the chimeric proteins provided herein (e.g., the plasma membrane recruitment element and the coiled-coil peptide, or the payload and the second coiled-coil peptide) are fused directly in the chimeric protein. In other cases, the components of the chimeric proteins provided herein are fused indirectly via a linker. In some cases, the linker between the coiled-coil peptide and the payload is a cleavable linker that is recognized by a protease. In some embodiments, the chimeric protein comprising a coiled-coil peptide and payload protein comprises a nuclear export signal (NES), a nuclear localization signal (NLS), or both. In some of these cases, the chimeric protein comprises two or more NES, e.g., three NES. In some of these cases, the NES is linked between the coiled-coil peptide and the payload protein. In some of these cases, the chimeric protein further comprises a cleavable linker between the NES and the payload protein. In some of these cases, the chimeric protein further comprises a NLS, and the NLS and the payload protein are on same side of the cleavable linker in the chimeric protein. In some of these cases, the chimeric protein comprises at least two NLS, optionally two NLS. In some cases, the at least two NLS are at both N- terminus and C-terminus of the payload protein. In some cases, the at least two NLS are at either N- terminus or C-terminus of the payload protein. The NES can be linked between the second coiled-coil peptide and the payload protein. In some of these cases, the NES and the coiled-coil peptide are on theWSGR Docket No.:62697-750.601 same side of the cleavable linker in the chimeric protein. In some cases, the NLS and the payload protein are on the same side of the cleavable linker in the chimeric protein.

[0207] The chimeric protein provided herein (e.g., comprising a gag protein) can form at least part of a protein core of the lipid delivery particle. A lipid delivery particle can comprise two or more chimeric proteins. The chimeric protein can include a structural protein. The structural protein can comprise a plasma membrane recruitment element (e.g., retroviral gag protein). The plasma membrane recruitment element can be fused to a payload. In other cases, the plasma membrane recruitment element is coupled to a payload via a coiled-coil peptide pair as discussed above. In some cases, the two or more chimeric proteins comprise the same structural protein. In some cases, the two or more chimeric proteins comprise different structural proteins. In some cases, the two or more chimeric proteins comprise different payloads. In some cases, the chimeric protein comprises a payload that comprises a nucleic acid-binding moiety. In some cases, the payload further comprises a guide nucleic acid molecule that forms a ribonucleoprotein complex with the nucleic acid-binding moiety. In some cases, the chimeric protein is suitable for delivery by a lipid delivery particle disclosed herein.

[0208] In some cases, the lipid delivery particle of the present disclosure further comprises a protease that recognizes the cleavable linker in the chimeric protein and cuts the chimeric protein at the cleavable linker. As a result of the cleavage at the cleavable linker by the protease, the payload can be separated from the plasma membrane recruitment element or the coiled-coil peptide that it is fused with. In some cases, the payload is present as a "free" entity separate from the plasma membrane recruitment element. For instance, the payload can be free and present within an inside of the protein core of the lipid delivery particle. In some cases, the protease is part of an additional chimeric protein comprising a second plasma membrane recruitment element and the protease, where the additional plasma membrane recruitment element can be either different from or same as the plasma membrane recruitment element that is fused with the payload. In some cases, the additional chimeric protein is a gag / pro polyprotein (e.g., a fusion of a gag polyprotein and a pro polyprotein) or a gag / pro / pol polyprotein (e.g., a fusion of a gag polyprotein, a pro polyprotein, and a pol polyprotein).

[0209] In some cases, the chimeric protein disclosed herein also comprises one or more non-cleavable linkers that operably link components together. The non-cleavable linker can be any suitable linker sequence that is used for chimeric protein construction, such as peptide linkers that consist of glycine (Gly) and serine (Ser) residues. In some embodiments, the non-cleavable linker comprises an amino acid sequence selected from the group consisting of: (GS)x (SEQ ID NO: 564), (GGS)x (SEQ ID NO: 565), (GGGGS)x (SEQ ID NO: 566), (GGSG)x (SEQ ID NO: 567), and (SGGG)x (SEQ ID NO: 568), and wherein x is an integer from 1 to 50.

[0210] In some cases, the chimeric protein of the present disclosure comprises a nuclear export signal (NES) sequence that can direct transport of the chimeric protein out of the nucleus of a cell, e.g., a producer cell.

[0211] In some cases, the chimeric protein disclosed herein has one of the following configurations of components positioned in an order from N-terminus to C-terminus:WSGR Docket No.:62697-750.601[plasma membrane recruitment element] -[cleavable linker]-[payload];[plasma membrane recruitment element] -[n * NES] -[cleavable linker] -[payload];[plasma membrane recruitment element] -[cleavable linker]-[payload]-[n * NES];[plasma membrane recruitment element] -[cleavable linker]-[n * NES] -[payload];[plasma membrane recruitment element] -[cleavable linker l]-[payload]-[cleavable linker 2]-[n * NES];[payload] -[cleavable linker] -[n * NES] -[plasma membrane recruitment element];[payload]-[n * NES]-[cleavable linker] -[plasma membrane recruitment element];[n * NES]-[payload]-[cleavable linker] -[plasma membrane recruitment element];[n * NES] -[cleavable linker l]-[payload]-[cleavable linker 2] -[plasma membrane recruitment element]; and[payload]-[cleavable linker] -[plasma membrane recruitment element]; wherein n is an integer in the range of from 1 to 10, and denotes the number of repeats of the NES sequence. Non-cleavable linker sequence can be present or absent in any of the foregoing configurations between any two neighboring components. As provided herein, the payload sequence in the chimeric protein can have one or more NLS sequences, at its N-terminus, C-terminus, or both.[n * NES] -[cleavable linker l]-[payload]-[cleavable linker 2] -[plasma membrane recruitment element]; and[payload]-[cleavable linker] -[plasma membrane recruitment element]; wherein n is an integer in the range of from 1 to 10, and denotes the number of repeats of the NES sequence. Non-cleavable linker sequence can be present or absent in any of the foregoing configurations between any two neighboring components. As provided herein, the payload sequence in the chimeric protein can have one or more NLS sequences, at its N-terminus, C-terminus, or both.

[0212] In some cases, the chimeric protein comprising a plasma membrane recruitment element and a coiled-coil peptide disclosed herein has one of the following configurations of components positioned in an order from N-terminus to C-terminus:[plasma membrane recruitment element]-[coiled-coil peptide];[plasma membrane recruitment element] -[cleavable linker]-[coiled-coil peptide];[coiled-coil peptide]- [plasma membrane recruitment element]; and[coiled-coil peptide] -[cleavable linker] -[plasma membrane recruitment element];Non-cleavable linker sequence can be present or absent in any of the foregoing configurations between any two neighboring components.

[0213] In some cases, the chimeric protein comprising a payload protein and a coiled-coil peptide disclosed herein has one of the following configurations of components positioned in an order from N- terminus to C-terminus:[coiled-coil peptide]-[cleavable linker] -[payload];[coiled-coil peptide]-[n * NES]-[cleavable linker] -[payload];[coiled-coil peptide]-[cleavable linker] -[payload]-[n * NES];[coiled-coil peptide]-[cleavable linker]-[n * NES]-[payload];WSGR Docket No.:62697-750.601[coiled-coil peptide]-[cleavable linker l]-[payload] -[cleavable linker 2]-[n * NES];[payload] -[cleavable linker]-[n * NES] -[coiled-coil peptide]; [payload]-[n * NES] -[cleavable linker]-[coiled-coil peptide]; [n * NES]-[payload]-[cleavable linker] -[coiled-coil peptide];[n * NES] -[cleavable linker l]-[payload]-[cleavable linker 2] -[coiled-coil peptide]; and [payload]-[cleavable linker] -[plasma membrane recruitment element]; wherein n is an integer in the range of from 1 to 10, and denotes the number of repeats of the NES sequence. Non-cleavable linker sequence can be present or absent in any of the foregoing configurations between any two neighboring components. As provided herein, the payload sequence in the chimeric protein can have one or more NLS sequences, at its N-terminus, C-terminus, or both.

[0214] In some cases, a chimeric protein described herein may comprise one or more coiled-coil peptides. For example, the chimeric protein may comprise 1, 2, 3, 4, 5, or more coiled-coil peptides. The chimeric protein can comprise a first coiled-coil peptide and an additional coiled-coil peptide. For example, the additional coiled-coil peptide may comprise a second coiled-coil peptide, a third coiled-coil peptide, a fourth coiled-coil peptide, or a fifth coiled-coil peptide. An additional coiled-coil peptide may be present at any location in a configuration of components described herein. In some cases, an additional coiled-coil peptide may be positioned N-terminal to a cleavable linker or C-terminal to a cleavable linker. An additional coiled-coil peptide may be positioned adjacent to a cleavable linker (e.g., at the N-terminus or C-terminus of a cleavable linker). In some cases, an additional coiled-coil peptide may be positioned N-terminal to a payload or C-terminal to a payload. An additional coiled-coil peptide may be positioned adjacent to a payload (e.g., at the N-terminus or C-terminus of a payload). In some cases, an additional coiled-coil peptide may be positioned N-terminal to a nuclear export signal or C-terminal to a nuclear export signal. An additional coiled-coil peptide may be positioned adjacent to a nuclear export signal (e.g., at the N-terminus or C-terminus of a nuclear export signal).Nuclear Export Signal

[0215] Direction of nuclear transport within the cell can be governed by nuclear targeting signals within payload proteins or coupled to (e.g., fused with) the payload proteins. As used herein, the term “nuclear export signal” refers to a sequence of amino acids that targets a payload protein for export from the nucleus. In some cases, a nuclear export signal (NES) is a short target peptide sequence containing four hydrophobic residues. These residues target the protein for export from the nucleus to the cytoplasm through the nuclear pore complex. A chimeric protein provided herein can comprise 1 NES, 2 NESs, 3 NESs, 4 NESs, 5 NESs, 6 NESs, 7 NESs, 8 NESs, 9 NESs, or 10 NESs. In some cases, the NES is located at the N-terminus, C-terminus, or in an internal region of the chimeric protein. In some cases, a NES is coupled between the plasma membrane recruitment element and the payload in the chimeric protein. In some cases, there is a cleavable linker between the plasma membrane recruitment element and the payload in the chimeric protein, and one or more NESs present on the same side of the cleavable linker as the plasma membrane recruitment element.WSGR Docket No.:62697-750.601

[0216] In some aspects of the present disclosure, a chimeric protein provided herein, which comprises a gag protein and a payload protein, also comprises an NES sequence that is linked between the gag protein and the payload protein. In other aspects, a chimeric protein provided herein, which comprises a gag protein and a payload protein, also comprises an NES sequence that is present within the gag protein. For instance, the NES sequence can be inserted between a matrix protein and a pl2 protein within the gag polyprotein. Alternatively or additionally, the NES sequence can be inserted between a pl2 protein and a capsid protein, or between capsid protein and nucleocapsid protein, or both, within the gag polyprotein. Without wishing to be bound by a certain theory, in some cases, insertion of the NES sequence within the gag polyprotein can improve desirable properties of the lipid delivery particle, e.g., packaging of the payload protein into the particle, release of the payload protein from the particle or off from the chimeric protein, functionality of the payload protein once it is released into the target cell, or any combination thereof. In some cases, insertion of the NES sequence within gag polyprotein improves gene editing efficiency when the payload protein is a gene editor, e.g., a Cas protein, or part of a base editor, or a prime editor, or an epigenetic editor.

[0217] In some cases, the NES sequence that is used in the chimeric protein comprises LQLPPLERLTL (SEQ ID NO: 403) derived from HIV-1 Rev protein, or any of the sequences having at least 80% identity thereto. In some cases, the NES sequence comprises LALKLAGLDI (SEQ ID NO: 416) derived from PKIa, or any of the sequences having at least 80% identity thereto. In some cases, the NES sequence that is used in the chimeric protein comprises an amino acid sequence as set forth in Table 11. In some cases, the NES sequence comprises any one of the sequences set forth in Table 11. In some cases, the NES sequence comprises one or more of the sequences set forth in Table 11. In some cases, the NES sequence comprises more than one, more than two, more than three, more than four, more than five, more than six, more than seven, more than eight, more than nine, or more than ten of the sequences set forth in Table 11. In some cases, the NES sequences comprise multiple sequences set forth in Table 11.

[0218] In some cases, the NES sequence comprises an amino acid sequence having 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to any sequence listed in Table 11. In some cases, the NES sequence comprises an amino acid sequence having 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to any sequence set forth in SEQ ID NOs: 353-453. In some cases, the NES sequence described herein comprises a sequence with greater than 80% sequence identity to any sequence listed in Table 11. The transport of payload proteins within a cell is enabled through both NES and nuclear export receptors. In some cases, the NES described herein is associated with a nuclear export receptor (e.g., CRM-1). In some cases, the NES may be conditionally active or inactive. In some cases, the NES sequence disclosed herein comprises a sequence such as those described in T la Cour, et al., Nucleic Acids Res. 2003;31 ( 1 ) : 393 -396; and Xu D, et al. Mol Biol Cell. 2012 Sep;23(18):3673-6, each of which is incorporated herein by reference in its entirety. Any of the NES sequences described in the NES sequence database (NESdb°; prodata.swmed.edu / LRNes) or (NESbase; services.healthtech.dtu.dk / datasets / NESbase-1.0) can be used in a chimeric protein disclosedWSGR Docket No.:62697-750.601 herein, e.g., for the purpose of packaging a payload into the molecular assembly, e.g., the lipid delivery particle.

[0219] In some cases, a chimeric protein disclosed herein include a nuclear export sequence (NES). In some cases, the NES facilitates localization of the chimeric protein in the cytosol of a target cell relative to the nucleus.

[0220] In some cases, a chimeric protein disclosed herein includes at least one NES sequences, such as, 2 or more, 3 or more, 4 or more, or 5 or more NES sequences. In some cases, one or more NES sequences (2 or more, 3 or more, 4 or more, or 5 or more NES sequences) are positioned at or near (e.g. , within 50 amino acids of) the N-terminus and / or the C- terminus of the chimeric protein. In some cases, the chimeric protein disclosed herein comprises only one NES sequence. In some cases, the chimeric protein disclosed herein comprises two NES sequences. In some cases, the chimeric protein disclosed herein comprises three NES sequences. In some cases, one or more NES sequences (2 or more, 3 or more, 4 or more, or 5 or more NES sequences) are positioned at or near (e.g., within 50 amino acids of) the N- terminus of the chimeric protein. In some cases, one or more NES sequences (2 or more, 3 or more, 4 or more, or 5 or more NES sequences) are positioned at or near (e.g., within 50 amino acids of) the C- terminus of the chimeric protein. In some cases, one or more NES sequences (3 or more, 4 or more, or 5 or more NES sequences) are positioned at or near (e.g., within 50 amino acids of) both the N-terminus and the C-terminus of the chimeric protein. In some cases, an NES sequence is positioned at the N- terminus and an NES sequence is positioned at the C-terminus of the chimeric protein.

[0221] In some cases, a payload is a protein that is delivered as part of the chimeric protein disclosed herein, e.g., operably linked to a structural protein (e.g., human endogenous retroviral structural protein or a Plasma membrane recruitment element). In some embodiments, the one or more NES sequences are positioned at or near the one or both ends of the payload protein sequence inside the chimeric protein. For example, in some cases, one or more NES sequences (2 or more, 3 or more, 4 or more, or 5 or more NES sequences) are positioned at or near (e.g., within 50 amino acids of) the N-terminus and / or the C- terminus of the payload protein sequence. In some cases, one or more NES sequences (2 or more, 3 or more, 4 or more, or 5 or more NES sequences) are positioned at or near (e.g., within 50 amino acids of) the N-terminus of the payload protein sequence. In some cases, one or more NES sequences (2 or more, 3 or more, 4 or more, or 5 or more NES sequences) are positioned at or near (e.g., within 50 amino acids of) the C-terminus of the payload protein sequence. In some cases, one or more NES sequences (3 or more, 4 or more, or 5 or more NES sequences) are positioned at or near (e.g. , within 50 amino acids of) both the N-terminus and the C-terminus of the payload protein sequence. In some cases, an NES sequence is positioned at the N-terminus and an NES sequence is positioned at the C-terminus of the payload protein sequence. In some cases, the chimeric protein disclosed herein comprises only one NES sequence. In some cases, the chimeric protein comprises only one NES sequence, and the NES sequence is positioned at or near (e.g., within 50 amino acids of) the N-terminus of the payload protein.

[0222] In eukaryotic cells, transport of proteins between the nucleus and the cytoplasm can be mediated by transport factors in the karyopherin-[3 family, which are also known as importins and exportins. TheWSGR Docket No.:62697-750.601 direction of nuclear-cytoplasmic transport can be dictated by nuclear targeting signals within the payload proteins. Nuclear export sequences (NESs) can direct export of proteins from the nucleus to the cytoplasm. NESs can bind directly to the export karyopherin CRM1 (also known as exportin 1), which can escort payload proteins through the nuclear pore complex.Table 11. Exemplary NES sequencesWSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601

[0223] In some embodiments, the chimeric protein may comprise a nuclear export signal (NES), a nuclear localization signal (NLS), or both. In some cases, the second chimeric protein can comprise two or more NES. In some cases, the chimeric protein comprises two or more NES. In some cases, the chimeric protein comprises three NES. In some cases, the NES is linked between the gag protein and the payload protein. In some cases, the chimeric protein further comprises a cleavable linker between the NES and the payload protein. In some cases, the chimeric protein further comprises a NLS, and wherein the NLS and the payload protein are on same side of the cleavable linker in the chimeric protein. In some cases, the chimeric protein comprises at least two NLS, optionally two NLS. In some cases, the at least two NLS are at both N-terminus and C-terminus of the payload protein. In some cases, the at least two NLS are at either N-terminus or C-terminus of the payload protein. In some cases, the NES comprises an amino acid sequence having at least 80%, 85%, 90%, 95%, or 99% sequence identity to a sequence set forth in Tables 11. In some cases, the NES comprises an amino acid sequence set forth in Tables 11. In some cases, the NLS comprises an amino acid sequence having at least 80%, 85%, 90%, 95%, or 99% sequence identity to a sequence set forth in Table 12. In some cases, the NLS comprises an amino acid sequence set forth in Table 12. The NES can be linked between the second coiled-coil peptide and the payload protein. In some cases, the NES and the second coiled-coil peptide can be on the same side of the cleavable linker in the second chimeric protein. In some cases, the NLS and the payload protein can be on same side of the cleavable linker in the second chimeric protein.Nuclear Localization Signal

[0224] In some instances, a payload described herein comprises one or more nuclear localization sequences (NLS). As used herein, the term “nuclear localization signal” refers to a sequence of amino acids that targets a payload (e.g. , a protein or a short polypeptide), which the NLS is present within or coupled to, to localize to the nucleus. In some cases, an NLS facilitates the import of a polypeptide comprising an NLS into the cell nucleus. A polypeptide can comprise 1 NLS, 2 NLSs, 3 NLSs, 4 NLSs, 5 NLSs, 6 NLSs, 7 NLSs, 8 NLSs, 9 NLSs, or 10 NLSs. In some cases, the NLS is located at the N- terminus, C-terminus, or in an internal region of the polypeptide. In some cases, a NLS is coupled to a nucleic acid binding domain described elsewhere herein. In some cases, a NLS is coupled to a nucleic acid modifying domain described elsewhere herein. In some cases, a NLS is coupled to a guidable polypeptide domain, a deaminase domain, or a reverse transcriptase domain. In some cases, a NLS is covalently linked to a nucleic acid binding domain described elsewhere herein. In some cases, a NLS isWSGR Docket No.:62697-750.601 covalently linked to a nucleic acid modifying domain described elsewhere herein. In some cases, a NLS is covalently linked to a guidable polypeptide domain, a deaminase domain, or a reverse transcriptase domain. In some cases, a nucleic acid binding domain does not comprise an NLS. In some cases, a nucleic acid binding domain does not comprise an NLS. In some cases, a guidable polypeptide domain, a deaminase domain, or a reverse transcriptase domain does not comprise an NLS. Examples of NLS are provided in Table 12 below.

[0225] In some cases, the NLS comprises an amino acid sequence as set forth in Table 12. In some cases, the NLS comprises any one of the sequences set forth in Table 12. In some cases, the NLS comprises one or more of the sequences set forth in Table 12. In some cases, the NLS comprises more than one of the sequences set forth in Table 12. In some cases, the NLS comprises multiple sequences set forth in Table 12. In some cases, NLS sequence can comprise an amino acid sequence having 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to any sequence listed in Table 12. In some cases, the NLS sequence described herein can comprise a sequence with greater than 80% sequence identity to any sequence listed in Table 12. In some cases, NLS sequence can comprise an amino acid sequence having 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to any sequence set forth in SEQ ID NOs: 454-477.

[0226] In some cases, a chimeric protein disclosed herein includes a nuclear localization sequence (NLS). In some cases, the NLS facilitates delivery of the chimeric protein, or a payload released from the chimeric protein (for instance, released from the chimeric protein following cleavage of a cleavable linker), into the nucleus of a target cell.

[0227] In some cases, a payload is a protein and is delivered as part of the chimeric protein disclosed herein, e.g., operably linked to a structural protein (e.g., plasma membrane recruitment element). In some embodiments, the one or more NLS sequences are positioned at or near the one or both ends of the payload protein sequence of the chimeric protein. In some cases, a chimeric protein includes (e.g., is fused to) between 2 and 5 NLS sequences (e.g., 2-4, or 2-3 NLSs). Examples of NLS sequences include an NLS sequence derived from: the NLS of the SV40 virus large T-antigen, having the amino acid sequence PKKKRKV (SEQ ID NO: 468); the NLS from nucleoplasmin (e.g., the nucleoplasmin bipartite NLS with the sequence KRPAATKKAGQAKKKK (SEQ ID NO: 460); the c-myc NLS having the amino acid sequence PAAKRVKLD (SEQ ID NO: 467) or RQRRNELKRSP (SEQ ID NO: 541); the hRNPAl M9 NLS having the sequence NQSSNFGPMKGGNFGGRSSGPYGGGGQYFAKPRNQGGY (SEQ ID NO: 542); the sequence RMRIZFKNKGKDTAELRRRRVEVSVELRKAKKDEQILKRRNV (SEQ ID NO: 543) of the IBB domain from importin-alpha; the sequences VSRKRPRP (SEQ ID NO: 477) and PPKKARED (SEQ ID NO: 544) of the myoma T protein; the sequence PQPKKKPL (SEQ ID NO: 545) of human p53; the sequence SALIKKKKKMAP (SEQ ID NO: 546) of mouse c-abl IV; the sequences DRLRR (SEQ ID NO: 547) and PKQKKRK (SEQ ID NO: 548) of the influenza virus NS1; the sequence RKLKKKIKKL (SEQ ID NO: 549) of the Hepatitis virus delta antigen; the sequence REKKKFLKRR (SEQ ID NO: 550) of the mouse Mxl protein; the sequence KRKGDE VDGVDEV AKKKS KK (SEQ ID NO: 551) of the human poly(ADP-ribose) polymerase; and the sequence RKCLQAGMNLEARKTKKWSGR Docket No.:62697-750.601(SEQ ID NO: 552) of the steroid hormone receptors (human) glucocorticoid, and sequences having at least 80% identity to the foregoing. In some cases, an NLS comprises the amino acid sequence MDSLLMNRRKFLY QFKNVRWAKGRRETYLC (SEQ ID NO: 553).

[0228] Other examples of an NLS sequence include KRTADGSEFESPKKKRKV (SEQ ID NO: 462), KKTELQTTNAENKTKKL (SEQ ID NO: 554), KRGINDRNFWRGENGRKTR (SEQ ID NO: 555), RKSGKIAAIVVKRPRK (SEQ ID NO: 556), and MDSLLMNRRKFLY QFKNVRWAKGRRETYLC (SEQ ID NO: 463), SPKKKRKVEAS (SEQ ID NO: 557), encoded by AGCCCCAAGAAgAAGAGaAAGGTGGAGGCCAGC (SEQ ID NO: 558), GPKKKRKVAAA (SEQ ID NO: 559), as well as any of those described in Cokol et al., EMBO Rep., 2000, 1(5): 411-415 and Freitas et al., Current Genomics, 2009, 10(8): 550-7; Lu, J., et la., Cell Commun Signal 19, 60 (2021); international publication no. WO / 2001 / 038547, each of which is incorporated herein by reference in its entirety, and sequences having at least 80% identity to the foregoing.

[0229] In some embodiments, the chimeric protein comprises one NES sequence and two NLS sequences. In some cases of these embodiments, the NES sequence, NLS sequences, and the payload protein sequence are positioned in an order from N-terminus to C-terminus as follows: NES-NLS-payload protein-NLS. In some embodiments, the chimeric protein comprises two or more NES sequences and two NLS sequences. In some cases of these embodiments, the NES sequences, NLS sequences, and the payload protein sequence are positioned in an order from N-terminus to C-terminus as follows: n X NES (n >=2)-NLS-payload protein-NLS.Table 12. Exemplary NLS sequencesWSGR Docket No.:62697-750.601

[0230] In some cases, the chimeric protein further comprises a NLS, and wherein the NLS and the payload protein are on same side of the cleavable linker in the chimeric protein. In some cases, the NLS and the payload protein are on same side of the cleavable linker in the second chimeric protein. In some cases, the NLS comprises an amino acid sequence having at least 80%, 85%, 90%, 95%, or 99% sequence identity to a sequence set forth in Table 12. In some cases, the NLS comprises an amino acid sequence set forth in Table 12. The NLS and the payload protein are on same side of the cleavable linker in the chimeric protein. In some cases, the chimeric protein comprises at least two NLS, optionally two NLS. In some cases, the at least two NLS are at both N-terminus and C-terminus of the payload protein. In some cases, the at least two NLS are at either N-terminus or C-terminus of the payload protein.Cleavable Linker

[0231] In some cases, the chimeric protein comprises a cleavable linker in between two or more components. For instance, the chimeric protein can comprise a cleavable linker between a payload protein sequence and a plasma membrane recruitment element sequence (e.g., retroviral gag protein sequence). In some cases, the cleavable linker separates the plasma membrane recruitment element sequence from a NLS sequence, and / or a NES sequence at its N-terminus or C-terminus. The cleavable linker can separate the payload protein sequence from the plasma membrane recruitment element sequence, NLS sequence, and / or NES sequence at its N-terminus or C-terminus. Examples of cleavable linker sequences that can be used in the chimeric protein include TSTLLMENSS (SEQ ID NO: 560), PRSSLYPALTP (SEQ ID NO: 561), VQALVLTQ (SEQ ID NO: 562), and PLQVLTLNIERR (SEQ ID NO: 563), and sequences having at least 80% identity to any one of the foregoing.WSGR Docket No.:62697-750.601Motifs and Modifications in Lipid Delivery Particles

[0232] In some aspects, provided herein are short polypeptide motifs and / or modifications that can be present within the chimeric protein or the plasma membrane recruitment element as described herein. Without wishing to be bound by a certain theory, the motifs in the chimeric proteins and / or modifications to the chimeric protein disclosed herein can enhance one or multiple characteristics of particle assembly and / or payload delivery, including, but not limited to, payload expression, payload stability, payload loading and offloading, particle formation and / or budding, and payload translocation to and from the nucleus.Particle Budding Motif

[0233] A particle budding motif can be a short polypeptide fused to the payload or plasma membrane recruitment element. Without wishing to be bound by a theory, the particle budding motif as described herein can enhance the interaction with host factors that are responsible for the creation of lipid delivery particles. In some cases, the particle budding motif can be a PPXY, LPXY, PTAP, or YPXnL. In some cases, n is an integer from 1 to 5. The particle budding motif can comprise a late domain of a wild-type gag polyprotein from a retrovirus or human endogenous retroviral gag. For example, the particle budding motif includes an RSV P2B and its associated late domain. The particle budding motif can bind ESCRT complex (e.g, NEDD4, TSG101, and Alix). For example, PPXY and LPXY can bind NEDD4, PTAP can bind TSG101, YPXnL can bind Alix. In some cases, the particle budding motif promotes the efficiency of particle assembly or budding (e.g., increase particle titer). In some cases, the particle budding motif increases the particle titer by at least 1-fold, at least 2-fold, at least 3 -fold, at least 4-fold, or at least 5- fold. In some cases, the particle budding motif increase the particle titer by at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90%. The particle budding motif can comprise a late assembly (L) domain. L domains are conserved sequences that can engage the endosomal sorting complex required for transport (ESCRT) components to promote budding. The particle budding motif can be found in retroviral gag proteins which recruit members of the ESCRT complex (e.g., TSG101, ALIX, NEDD4). The ESCRT complex is located on the cytoplasmic surface of the multivesicular body and can recognize ubiquitinated proteins into vesicles. The vesicles can then bud into the multivesicular body and can be released into the extracellular space. The ESCRT also plays a role in endolysosomal membrane and plasma membrane repair, neuronal pruning, nuclear envelope maintenance, and autophagy. The ESCRT complex comprises three complexes (ESCRT-I, ESCRT-II, ESCRT-III) which are active in the cytosol and the plasma membrane to assist in assembly and fission from the membrane.

[0234] The lipid delivery particle can comprise the particle budding motif as described herein. In some cases, the particle budding motif is linked to the C-terminus of the chimeric protein. In some cases, the particle budding motif is linked to the N-terminus of the chimeric protein. In some cases, the particle budding motif is linked to the payload. In some cases, the particle budding motif is linked to the N- terminus of the payload. In some cases, the particle budding motif is linked to the C-terminus of theWSGR Docket No.:62697-750.601 payload. In some cases, the particle budding motif is linked to a plasma membrane recruitment element (e.g., a mutant gag polyprotein or a PH domain). In some cases, the particle budding motif is linked to the N-terminus of a plasma membrane recruitment element (e.g., a mutant gag polyprotein or a PH domain). In some cases, the particle budding motif is linked to the C-terminus of a plasma membrane recruitment element (e.g., a mutant gag polyprotein or a PH domain). In some cases, the particle budding motif is linked between the payload and the plasma membrane recruitment element. In some cases, the particle budding motif is linked to a nuclear export signal domain. In some cases, the particle budding motif is linked to the N-terminus of a nuclear export signal domain. In some cases, the particle budding motif is linked to the C-terminus of a nuclear export signal domain. In some cases, the particle budding motif is linked to a non-endogenous cleavage site. In some cases, the particle budding motif is linked to the N- terminus of a non-endogenous cleavage site. In some cases, the particle budding motif is linked to the C- terminus of a non-endogenous cleavage site. In some cases, the particle budding motif is linked to a post- translational modification motif. In some cases, the particle budding motif is linked to the N-terminus of a post-translational modification motif. In some cases, the particle budding motif is linked to the C- terminus of a post-translational modification motif. In some cases, the particle budding motif is linked to a multimerization motif. In some cases, the particle budding motif is linked to the N-terminus of a multimerization motif. In some cases, the particle budding motif is linked to the C-terminus of a multimerization motif. In some cases, the particle budding motif is present as tandem repeats in the chimeric protein. In some cases, the chimeric protein can comprise one, two, three, four, five, or more particle budding motifs. In some cases, flexible linkers can be present in-between the particle budding motifs. In some cases, a particle budding motif can have a linker to the N-terminus and / or the C-terminus.

[0235] In some cases, the particle budding motif is sourced from a virus. The virus can include, but is not limited to, HIV, EIAV, RSV, ALV, FUJSV, HTLV-1, MLV, MMLV, EBOV, HTLV-2, SRV-2, CHMP4A, AVISY, fMLV, influenza, zika virus, dengue virus, WNV, and Tupaia. In some cases, the particle budding motif comprises a sequence with at least 80% sequence identity to a sequence as set forth in any one of SEQ ID NOs: 106-137. In some cases, the particle budding motif comprises a sequence with at least 90% sequence identity to a sequence as set forth in any one of SEQ ID NOs: 106-137. In some cases, the particle budding motif comprises a sequence with at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to a sequence as set forth in any one of SEQ ID NOs: 106-137. In some cases, the particle budding motif comprises a sequence as set forth in any one of SEQ ID NOs: 106-137. In some cases, the particle budding motif comprises any of the sequences or a minimal sequence drawn from any of the sequences as set forth in Table 5.Table 5. Exemplary particle budding motif (BD) sequencesWSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601Membrane Penetrating Peptide

[0236] A membrane penetrating peptide (e.g., a cell penetrating peptide) can be a short polypeptide fused to the payload or plasma membrane recruitment element that can facilitate protein translocation across a phospholipid bilayer.

[0237] In some cases, the membrane penetrating peptide is fewer than 40 amino acids. In some cases, the membrane penetrating peptide is fewer than 30 amino acids. In some cases, the membrane penetrating peptide is fewer than 20 amino acids. In some cases, the membrane penetrating peptide is about 40 amino acids. In some cases, the membrane penetrating peptide is about 35 amino acids. In some cases, the membrane penetrating peptide is about 30 amino acids. In some cases, the membrane penetrating peptide is about 25 amino acids. In some cases, the membrane penetrating peptide is about 20 amino acids.

[0238] In some cases, the membrane penetrating peptide promotes recruitment of the payload to the plasma membrane. In some cases, the membrane penetrating peptide increases the efficiency of particle assembly and / or budding. In some cases, the membrane penetrating peptide functions in a pH-inducible manner. In some cases, the pH that can modulate membrane penetrating activity of the membrane penetrating peptide is between 3 and 10, such as 3, 4, 5, 6, 7, 8, 9 or 10. Without wishing to be bound by a theory, membrane penetrating peptides facilitate payload translocation via penetration of the membrane, endocytotic-mediated entry, or translocation through a transitory structure.

[0239] Membrane penetrating peptides can have amino acid compositions that either contain a high relative abundance of positively charged amino acids such as lysine or arginine or have sequences that contain an alternating pattern of polar, charged amino acids and non-polar, hydrophobic amino acids, as described in Derakhshankhah and Jafari (2018) Biomedicine and Pharmacotherapy 108: 1090-1096. Membrane penetrating peptides can also contain only apolar residues with low net charge or hydrophobic amino acid groups that are helpful for cellular uptake. As described herein, the membrane penetrating peptides can be cationic, amphipathic, or hydrophobic.

[0240] The chimeric protein can comprise the membrane penetrating peptide as described herein. In some cases, the membrane penetrating peptide is linked to the C-terminus of the chimeric protein. In some cases, the membrane penetrating peptide is linked to the N-terminus of the chimeric protein. In some cases, the membrane penetrating peptide is linked to the payload. In some cases, the membrane penetrating peptide is linked to a plasma membrane recruitment element (e.g., a mutant gag polyprotein). In some cases, the membrane penetrating peptide is linked between the payload and the plasma membrane recruitment element. In some cases, the membrane penetrating peptide is present as tandem repeats in the chimeric protein.WSGR Docket No.:62697-750.601

[0241] In some cases, the membrane penetrating peptide comprises a sequence with at least 80% sequence identity to a sequence as set forth in any one of SEQ ID NOs: 138-161. In some cases, the membrane penetrating peptide comprises a sequence with at least 90% sequence identity to a sequence as set forth in any one of SEQ ID NOs: 138-161. In some cases, the membrane penetrating peptide comprises a sequence with at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to a sequence as set forth in any one of SEQ ID NOs: 138-161. In some cases, the membrane penetrating peptide comprises a sequence as set forth in any one of SEQ ID NOs: 138-161. In some cases, the membrane penetrating peptide comprises a sequence as set forth in Table 6.Table 6. Exemplary membrane penetrating peptide sequencesPost-Translational Modifications Motif

[0242] A post-translational modification motif disclosed herein can be a short peptide present within the chimeric protein that leads the payload to be modified following translation (e.g., a post-translational modification). In some cases, the post-translational modification motif is between about 10 and about 40 amino acids in length. Without wishing to be bound by a theory, the post-translational modification alters the payload in a way that can promote recruitment to the plasma membrane. In some cases, the post- translational modification motif, upon the corresponding post-translational modification to the chimeric protein, can alter localization of the payload. The post-translational modification can be a covalentWSGR Docket No.:62697-750.601 modification of one or more amino acids within the chimeric protein following biosynthesis. The post- translational modification motif can promote plasma membrane localization of cytosolic proteins. The post-translational modification motif can be enzymatic. As described herein, a post-translational modification motif can be to one or more amino acids within the payload, the plasma membrane recruitment element, or both. In some cases, a post-translational modification comprises phosphorylation, glycosylation, ubiquitination, nitrosylation, methylation, lipidation, acetylation, or proteolysis. In some cases, a post-translational modification comprises geranylgeranylation, myristoylation, glypiation, isoprenylation, palmitoylation, or famesylation. Acetylation can comprise transfer of an acetyl group to nitrogen. Myristoylation can comprise an attachment of a myristoyl group to a N-terminal glycine residue of a protein. Isoprenylation can comprise an attachment of a prenyl group (e.g., a hydrophobic molecule) to a protein. Famesylation can comprise an attachment of a famesyl group to a C-terminal cysteine residue of a protein. Phosphorylation can comprise addition of a phosphate group and can occur on serine, threonine, or tyrosine residues. Glycosylation can comprise addition of a sugar moiety (e.g., monosaccharide, polysaccharide, oligosaccharide, or carbohydrate) to a residue. Ubiquitination can comprise the addition of a ubiquitin polypeptide to a lysine residue of a protein. Nitrosylation can comprise incorporation of a nitrosyl moiety of nitric oxide to a protein. Nitrosylation can occur on free cysteine residues to produce S-nitrothiols. Methylation can comprise the transfer of one-carbon methyl groups to a protein and can increase hydrophobicity of the protein. Lipidation can comprise incorporation of a lipid moiety to a protein. Proteolysis can comprise cleaving of peptide bonds of a protein and can assist antigen processing, apoptosis, surface protein shedding, and cell signaling. Geranylgeranylation can comprise attachment of at least one lipophilic geranylgeranyl isoprene unit from geranylgeranyl diphosphate to at least one cysteine residue. Glypiation can comprise covalent bonding of a glycosylphosphatidylinositol (GPI) anchor to a protein. Palmitoylation can comprise the attachment of at least one fatty acid (e.g., palmitic acid) to a cysteine residue of a protein.

[0243] The chimeric protein can comprise the post-translational modification motif as described herein. In some cases, the post-translational modification motif is linked to the C-terminus of the chimeric protein. In some cases, the post-translational modification motif is linked to the N-terminus of the chimeric protein. In some cases, the post-translational modification motif is linked to the payload. In some cases, the post-translational modification motif is linked to a plasma membrane recruitment element (e.g., a mutant gag polyprotein). In some cases, the post-translational modification motif is linked to both a payload and a plasma membrane recruitment element (e.g., a mutant gag polyprotein). In some cases, the post-translational modification motif is linked to some other part(s) of the chimeric protein than a payload and a plasma membrane recruitment element (e.g., a mutant gag polyprotein). In some cases, the post-translational modification motif is linked to some other part(s) of the chimeric protein besides a payload and a plasma membrane recruitment element (e.g., a mutant gag polyprotein). In some cases, the post-translational modification motif is linked between the payload and the plasma membrane recruitment element. In some cases, the post-translational modification motif is linked to a nuclear export signal domain. In some cases, the post-translational modification motif is linked to a non-endogenous cleavageWSGR Docket No.:62697-750.601 site. In some cases, the post-translational modification motif is linked to a particle budding motif sequence. In some cases, the post-translational modification motif is present as tandem repeats in the chimeric protein.

[0244] In some cases, the post-translational modification motif comprises a sequence with at least 80% sequence identity to a sequence as set forth in any one of SEQ ID NOs: 162-191. In some cases, the post- translational modification motif comprises a sequence with at least 90% sequence identity to a sequence as set forth in any one of SEQ ID NOs: 162-191. In some cases, the post-translational modification motif comprises a sequence with at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to a sequence as set forth in any one of SEQ ID NOs: 162-191. In some cases, the post-translational modification motif comprises an amino acid sequence as set forth in any one of SEQ ID NOs: 162-191, with at least one but not more than 8 nucleic acid differences. In some cases, the post-translational modification motif comprises a sequence as set forth in any one of SEQ ID NOs: 162-191 . In some cases, the post-translational modification motif comprises a sequence as set forth in Table 7.Table 7. Exemplary post-translational modification motif sequencesWSGR Docket No.:62697-750.601Multimerization Motif

[0245] The chimeric protein described herein can comprise a multimerization motif. In some cases, the multimerization motif can perform protein-protein interactions. In some cases, the multimerization motif forms a parallel homodimer, such as a coiled-coil. In some cases, the multimerization motif forms dimers, trimers, tetramers, or other oligomers. In some cases, the multimerization motif is a leucine zipper oligomerization motif. The leucine zipper oligomerization motif can comprise a series of leucines spaced at intervals of every seventh amino acid along an a-helix. In some cases, a leucine zipper oligomerization motif can mediate dimerization or oligomerization. In some cases, the coiled-coil is a leucine zipper oligomerization motif (e.g., a GCN4 leucine zipper or a mutant thereof). The coiled-coil can comprise an amino acid sequence as set forth in Table 8 or Table 16. Upon dimerization, the leucine zipper oligomerization motif can form a parallel-coiled coil. In some cases, the leucine zipper oligomerization motif (e.g., the GCN4 leucine zipper motif). In some cases, the multimerization motif is synthetic. In some cases, the multimerization motif is a dimerization zinc -finger (DZF) motif, for example, dDZFl and dDZF2. In some cases, the dDZFl is an antiparallel heterodimerization domain 1 and binds to dDZF2. In some cases, the dDZF2 is an antiparallel heterodimerization domain 2 and binds to dDZF 1. Details of synthetic multimerization motif, including DZFs, are described in Giesecke AV et al., Mol Syst Biol. 20062: 2006.0011. Exemplary sequence for dDZFl or antiparallel heterodimerization domain 1, which binds to dDZF2, can comprise: FKCEHCRILFLDHVMFTIHMGCHGFRDPFKCNMCGEKCDGPVGLFVHMARNAHGEKPFYCEHC EITFRDVVMYSLHKGYHGFRDPFECNICGYHSQDRYEFSSHIVRGEH (SEQ ID NO: 192). Exemplary dDZF2 or antiparallel heterodimerization domain 2, which binds to dDZFl, can include HHCQHCDMYFADNILYTIHMGCHSCDDVFKCNMCGEKCDGPVGLFVHMARNAHGEKPTKCVH CGIVFLDEVMYALHMSCHGFRDPFECNICGYHSQDRYEFSSHIVRGEH (SEQ ID NO: 193). Without wishing to be bound by a theory there, the multimerization motif (e.g., GCN4 leucine zipper, dDZF 1 / 2, or a mutant thereof) can be used to induce multimerization that can assist in assembly of the lipid delivery particles described herein. As such, it can complement removal of nucleic acid binding domains that are necessary for multimerization in a wild-type gag polyprotein.

[0246] In some cases, the multimerization motif (e.g., a leucine zipper oligomerization motif) is linked to the C-terminus of the chimeric protein. In some cases, the multimerization motif (e.g., a leucine zipper oligomerization motif) is linked to the N-terminus of the chimeric protein. In some cases, the multimerization motif (e.g., a leucine zipper oligomerization motif) is linked to the payload. In some cases, the multimerization motif (e.g., a leucine zipper oligomerization motif) is linked to a plasma membrane recruitment element (e.g., a mutant gag polyprotein). In some cases, the multimerization motif (e.g., a leucine zipper oligomerization motif) is linked to both a payload and a plasma membrane recruitment element (e.g., a mutant gag polyprotein). In some cases, the multimerization motif (e.g., a leucine zipper oligomerization motif) is linked to some other part(s) of the chimeric protein than a payload and a plasma membrane recruitment element (e.g., a mutant gag polyprotein). In some cases, theWSGR Docket No.:62697-750.601 multimerization motif (e.g., a leucine zipper oligomerization motif) is linked to some other part(s) of the chimeric protein besides a payload and a plasma membrane recruitment element (e.g., a mutant gag polyprotein). In some cases, the multimerization motif (e.g., a leucine zipper oligomerization motif) is linked between the payload and the plasma membrane recruitment element. In some cases, the multimerization motif (e.g., a leucine zipper oligomerization motif) is linked to a nuclear export signal domain. In some cases, the multimerization motif (e.g., a leucine zipper oligomerization motif) is linked to a heterologous protease cleavage sequence. In some cases, the multimerization motif (e.g., a leucine zipper oligomerization motif) is linked to a particle budding motif sequence. In some cases, the multimerization motif (e.g., a leucine zipper oligomerization motif) is present as tandem repeats in the chimeric protein.

[0247] In some cases, the chimeric protein comprising the plasma membrane recruitment element that comprises a mutant gag polyprotein and a payload comprises any one of the arrangements provided in Table 14A from the N-terminus to the C-terminus. Plasma membrane recruitment domain is abbreviated PMRE, particle budding motif is abbreviated BD, and a protease cleavage site cleavable sequence is abbreviated CS.Table 14A. Exemplary Construct Arrangement for Chimeric Protein with Budding Motif

[0248] In some cases, the chimeric protein comprising the plasma membrane recruitment element that comprises a mutant gag polyprotein and a payload comprises any one of the arrangements provided in Table 14B from the N-terminus to the C-terminus. Plasma membrane recruitment domain is abbreviated PMRE, membrane penetrating peptide is abbreviated MPP, and a protease cleavage sequence is abbreviated CS.Table 14B. Exemplary Construct Arrangement for Chimeric Protein with Membrane Penetrating PeptideWSGR Docket No.:62697-750.601

[0249] In some cases, the chimeric protein comprising the plasma membrane recruitment element that comprises a mutant gag polyprotein and a payload comprises any one of the arrangements provided inTable 14C from the N-terminus to the C-terminus. Plasma membrane recruitment domain is abbreviated PMRE, post-translational modification motif is abbreviated PTM, and a protease cleavage sequence is abbreviated CS.Table 14C. Exemplary Construct Arrangement for Chimeric Protein with Post-Translational Modification

[0250] In some cases, the chimeric protein comprising the plasma membrane recruitment element that comprises a mutant gag polyprotein and a payload comprises any one of the arrangements provided in Table 15 from the N-terminus to the C-terminus. Plasma membrane recruitment domain is abbreviated PMRE, particle budding motif is abbreviated BD, post-translational modification motif is abbreviated PTM, and a protease cleavage sequence is abbreviated CS.WSGR Docket No.:62697-750.601Table 15. Exemplary Construct Arrangement for Chimeric Protein with Post-Translational Modification Motif and Particle Budding MotifENVELOPE PROTEIN

[0251] In some aspects, the lipid delivery particle provided herein comprises an envelope protein. The envelope protein can be associated with the outside boundary or the surface of the lipid delivery particle, for example, the membrane or envelope of the lipid delivery particle.

[0252] The membrane of the lipid delivery particle can comprise a lipid layer, such as a single layer or a lipid bilayer. In some cases, the membrane of the lipid delivery particle is from plasma membrane, endoplasmic reticulum, or a combination thereof. In some cases, the membrane of the lipid delivery particle is from Golgi complex, ER Golgi intermediate compartment, or nuclear envelope. In some cases, the membrane of the lipid delivery particle is from plasma membrane. In some cases, the membrane of the lipid delivery particle is a phospholipid bilayer.

[0253] The envelope protein can be associated with the membrane of the lipid delivery particle in various manners. For example, the envelope protein can be anchored or attached to the external membrane of the particle or anchored or attached to the internal membrane of the particle. The envelope protein can be embedded or inserted in the membrane, spanning through the membrane, with certain portions located at the outside of the membrane, or certain portions extending to the inside of the particle, or both. The envelope protein within the lipid delivery particle described herein can be overexpressed from an exogenous source, such as plasmids or stably integrated transgenes, in the production cells.

[0254] The envelope protein can play a role in the delivery of the lipid delivery particle to a target cell and release of the components of the lipid delivery particle within the target cell. The envelope protein can contact the surface of a target cell and participate in the fusion of the lipid delivery particle and theWSGR Docket No.:62697-750.601 membrane of the target cell. The envelope protein can participate in the fusion of the lipid delivery particle with the membrane of the target cell via any appropriate mechanism, such as those described in White et al. Crit Rev Biochem Mol Biol. 2008; 43(3): 189-219. One example of the fusion mechanisms is unifying Trimer-of-Hairpins Fusion Mechanism. Membrane fusion can occur after allosteric priming by binding to a target receptor. In some cases, membrane fusion occurs after proteolysis. In some cases, membrane fusion occurs after isomerization of disulfide bridges. In some cases, membrane fusion occurs by internalization and then priming of fusion via (i) cathepsin-mediated proteolysis, or (ii) low pH / acidification. The cathepsin-mediated proteolysis can be pH dependent or pH independent. Other fusion triggering mechanisms can include low PH, binding to target cell receptors, and a receptor followed by low pH. The envelope protein can also play a role in the formation of the lipid delivery particle. The envelope protein can interact with another component within the lipid delivery particle and participate in the assembly of the lipid delivery particle, for example, in a producer cell. The envelope protein can make contact with another envelope protein and form an oligomer embedded within the membrane. The envelope protein can be a glycoprotein, for example, a transmembrane glycoprotein. In some cases, envelope protein comprises multiple membrane-spanning regions. These multiple membranespanning regions can oligomerize and form channels in the membrane.

[0255] In some cases, the envelope protein is fused with a targeting moiety. In some cases, the targeting moiety recognizes a specific molecule (e.g., antigen, receptor, or other membrane protein) on the surface of a target cell to allow targeted cell entry with more specificity. In some cases, the targeting moiety is specific for a certain cell type or is specific for a certain target cell. The targeting moiety can be fused to the envelope protein at a position that is located at an outside of the lipid delivery particle. For example, the targeting moiety includes scFvs, antibody variable regions, nanobodies, T-cell receptor variable regions, other antigen-binding fragments or their mimetics, such as DARPins. In some cases, the targeting moiety is a protein ligand from the human ligandome. The targeting moiety can be a natural peptide or a synthetic peptide. In some cases, the targeting moiety is not fused with the envelope protein and is attached to the membrane of the lipid delivery particle from the outside, for example, via a transmembrane domain.

[0256] A targeting moiety can include, e.g., an antibody or an antigen-binding fragment thereof (e.g., Fab, Fab', F(ab')2, Fv fragments, scFv antibody fragments, disulfide-linked Fvs (sdFv), a Fd fragment consisting of the VH and CHI domains, linear antibodies, single domain antibodies such as sdAb (either VL or VH), nanobodies, or camelid VHH domains), an antigen-binding fibronectin type III (Fn3) scaffold such as a fibronectin polypeptide minibody, a ligand, a cytokine, a chemokine, or a T cell receptor (TCRs). Membrane -fusion proteins can be re-targeted by non-covalently conjugating a targeting moiety to the membrane-fusion protein or targeting protein (e.g. the hemagglutinin protein). For example, the membrane -fusion protein can be engineered to bind the Fc region of an antibody that targets an antigen on a target cell, redirecting the membrane fusion activity towards cells that display the antibody’s target.WSGR Docket No.:62697-750.601

[0257] In some cases, the targeting moiety linked to the membrane-fusion protein binds a cell surface marker on the target cell, e.g., a protein, glycoprotein, receptor, cell surface ligand, agonist, lipid, sugar, class I transmembrane protein, class II transmembrane protein, or class III transmembrane protein.

[0258] In some cases, the lipid delivery particles disclosed herein display targeting moieties that are not conjugated to the membrane-fusion protein or other proteins in order to redirect the fusion activity of the lipid delivery particle towards a cell that is bound by the targeting moiety, or to affect tropism of the lipid delivery particle toward the target cell.Envelope protein of viral origin

[0259] In some cases, an envelope protein has a viral origin. For example, a suitable envelope protein is from a DNA virus, an RNA virus, or a retrovirus. The envelope protein can be envelope protein from Herpesviruses, Avian sarcoma leukosis virus, Poxviruses, Hepadnaviruses, Asfarviridae, Flaviviruses, Alphaviruses, Togaviruses, Coronaviruses, Hepatitis D, Orthomyxoviruses, Rhabdovirus, Bunyaviruses, Filoviruses, Oncoretroviruses, lentiviruses, Spumaviruses. In some cases, envelope protein can be envelope protein from lentiviruses, for example, human immunodeficiency virus (HIV), simian immunodeficiency virus (SIV), feline immunodeficiency virus (FIV) and equine infectious anemia virus (EIAV). In some cases, an envelope protein is a fusion of two different envelope proteins, wherein each comes from a different virus. Additional suitable envelope proteins that are from viral origins and their functions are described in White JM et al., Crit Rev Biochem Mol Biol. 2008 May-Jun;43(3): 189-219.

[0260] In some cases, the envelope protein is a vesicular stomatitis virus glycoprotein (VSVG) or a biologically active mutant thereof. A “biologically active mutant” disclosed herein in connection with a reference protein can refer to a mutant of the reference protein that remains displaying one or more biological activities that are of same nature as the reference protein, which are relevant to the context in which the reference protein is used in the lipid delivery particle disclosed herein, while the level of the one or more biological activities of the biologically active mutant can be either similar as or different than the reference protein. For instance, the biologically active mutant of a VSVG in the context of an envelope protein remains displaying the biological activities of an envelope protein, e.g., mediating membrane fusion, tropism of the lipid delivery particle toward a target cell, or both. Unless otherwise noted, a mutant as described in the present disclosure is equivalent to a biologically active mutant. In some cases, the envelope protein is a Human immunodeficiency virus GP160 or a biologically active mutant thereof. In some cases, the envelope protein is a Baboon Endogenous Retrovirus (BaEVTR) glycoprotein or a biologically active mutant thereof. In some cases, the envelope protein is a modified Baboon Endogenous Retrovirus (BaEVTRless) glycoprotein or a biologically active mutant thereof. In some cases, the envelope protein is the fusion protein of Vesicular stomatitis Indiana virus and Rabies virus Glycoproteins (FuG-E) or a biologically active mutant thereof. In some cases, the envelope protein pantropic murine leukemia virus envelope protein (MLV) or a biologically active mutant thereof. In some cases, the envelope protein is a modified Fusion protein of Vesicular stomatitis Indiana virus and Rabies virus Glycoproteins (FuG-E P440E) or a biologically active mutant thereof. In some cases, the envelope protein is an ecotropic Murine Leukemia Virus envelope protein (MLV ENV ecotropic) or a biologicallyWSGR Docket No.:62697-750.601 active mutant thereof. In some cases, the envelope protein is an amphotrophic Murine Leukemia Virus envelope protein (MLV ENV amphotropic) or a biologically active mutant thereof. In some cases, the envelope protein is a Moloney murine leukemia virus envelope protein (MMLV) or a biologically active mutant thereof. In some cases, the envelope protein is a Moloney murine sarcoma virus envelope protein (MoMSVg) or a biologically active mutant thereof. In some cases, the envelope protein is a Moloney murine leukemia virus 10A1 strain Glycoprotein (MLV 10A1) or a biologically active mutant thereof. In some cases, the envelope protein is a xenotropic murine leukemia virus envelope protein (MLV ENV xenotropic) or a biologically active mutant thereof. In some cases, the envelope protein is a xenotropic murine leukemia virus-related envelope protein (XMRV) or a biologically active mutant thereof. In some cases, the envelope protein is a Baculovirus envelope glycoprotein (GP64) or a biologically active mutant thereof. In some cases, the envelope protein is an endogenous feline virus envelope protein (RD114 ENV) or a biologically active mutant thereof. In some cases, the envelope protein is a mammalian endogenous retrovirus protein, or a biologically active mutant thereof. The mammalian endogenous retrovirus protein can be a koala retrovirus protein (KoRV) or a Jaagsiekte sheep retrovirus protein (enJSRV), or a biologically active mutant thereof.

[0261] In some cases, the envelope protein is a simian endogenous type D retrovirus protein (RD-114) or a biologically active mutant thereof. In some cases, the envelope protein is a gibbon ape leukemia virus envelope protein (GALV) or a biologically active mutant thereof. In some cases, the envelope protein is a feline leukemia virus envelope protein (FLV) or a biologically active mutant thereof. In some cases, the envelope protein is a mouse mammary tumor virus envelope protein (MMTV) or a biologically active mutant thereof. In some cases, the envelope protein is an avian leukosis virus envelope protein or a biologically active mutant thereof. In some cases, the envelope protein is a rous sarcoma virus envelope protein or a biologically active mutant thereof.

[0262] In some cases, the envelope protein can direct the lipid delivery particles to fuse with a certain type of target cells rather than other cells. For example, based on the specific type of envelope protein associated with the membrane of the lipid delivery particle, the lipid delivery particle can preferentially target different cell types (i.e., tropisms of the lipid delivery particles), such as liver cells, ocular cells, nerve cells, lung cells, immune cells, muscle cells, and any other cell types of interest. For example, to fuse with a target liver cells, the envelope protein can be a glycoprotein from human hepatitis viruses or a biologically active mutant thereof, e.g., Hepatitis B virus (HBV) or hepatitis C virus (HCV), VSV-G glycoprotein or a biologically active mutant thereof, a Marburg virus glycoprotein or a biologically active mutant thereof, an Ebola virus glycoprotein or a biologically active mutant thereof. To fuse with a target muscle cell, for example, a skeletal muscle cell, the envelope protein can be a Ross River virus glycoprotein or a biologically active mutant thereof, or a VSV-G or a biologically active mutant thereof. To fuse with a target ocular cell, for example, a photoreceptor cell or a retinal cell, the envelope protein can be an Ebola virus glycoprotein or a biologically active mutant thereof, a Marburg virus glycoprotein or a biologically active mutant thereof, or a VSV-G or a biologically active mutant thereof. To fuse with a target immune cell, for example, CD8+ T cell, an HTLV-1 glycoprotein or a biologically active mutantWSGR Docket No.:62697-750.601 thereof, or a VSV- G glycoprotein or a biologically active mutant thereof. To fuse with a target immune cell, for example, CD4+ T cell, the envelope protein can be a HIV-1 envelope or a biologically active mutant thereof, a HTLV-1 glycoprotein or a biologically active mutant thereof, or a VSV-G glycoprotein or a biologically active mutant thereof. To fuse with a target lung cells, the envelope protein can be a respiratory syncytial virus glycoprotein or a biologically active mutant thereof, or a SARS-CoV glycoprotein or a biologically active mutant thereof. To fuse with a target nerve cell, such as a cell from the central nervous system cell (e.g., neurons, glial cells including oligodendrocytes, astrocytes and microglia), the envelope protein can be a rabies glycoprotein or a biologically active mutant thereof, a Mokola virus glycoprotein or a biologically active mutant thereof, a Semliki Forest virus glycoprotein or a biologically active mutant thereof, a Venezuelan equine encephalitis virus glycoprotein or a biologically active mutant thereof, or a VSV-G or a biologically active mutant thereof. To fuse with a target sensory cell, such as an auditory cell, including hair cells, cochlear cells, etc., the envelope protein can be an Ebola vims glycoprotein or a biologically active mutant thereof, a Marburg virus glycoprotein or a biologically active mutant thereof, or a VSV-G or a biologically active mutant thereof.

[0263] In some cases, the envelope protein comprises the sequences set forth in Table 1. In some cases, the envelope protein comprises the sequences set forth in Table 1 with at least one amino acid substitution, deletion, or insertion. For instance, N-terminal methionine can be absent from the envelope protein of the lipid delivery particle provided herein relative to the wild-type viral envelope protein. In some cases, the envelope protein comprises the sequences set forth in Table 1 and a heterologous peptide sequence fused to the N-terminal or C-terminal.

[0264] In some cases, the envelope protein comprises one or more of the sequences set forth in Table 1 with at least one amino acid substitution, deletion, or insertion. For instance, N-terminal methionine can be absent from the envelope protein of the lipid delivery particle provided herein relative to the wild-type viral envelope protein. In some cases, the envelope protein comprises one or more of the sequences set forth in Table 1 and a heterologous peptide sequence fused to the N-terminal or C-terminal.

[0265] In some cases, the envelope protein comprises any one of the sequences set forth in Table 1 with at least one amino acid substitution, deletion, or insertion. For instance, N-terminal methionine can be absent from the envelope protein of the lipid delivery particle provided herein relative to the wild-type viral envelope protein. In some cases, the envelope protein comprises any one of the sequences set forth in Table 1 and a heterologous peptide sequence fused to the N-terminal or C-terminal.

[0266] In some cases, the envelope protein comprises an amino acid sequence that has at least about 50%, 60%, 70%, 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence set forth in Table 1. In some cases, the envelope protein comprises an amino acid sequence that has at least about 50%, 60%, 70%, 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence set forth in any one of SEQ ID NOs: 83-104. In some cases, the envelope protein comprises an amino acid sequence that has at least about 50% sequence identity to a sequence set forth in any one of SEQ ID NOs: 83-104. In some cases, the envelope protein comprises an amino acid sequence that has at least about 60% sequence identity to a sequence set forth inWSGR Docket No.:62697-750.601 any one of SEQ ID NOs: 83-104 In some cases, the envelope protein comprises an amino acid sequence that has at least about 70% sequence identity to a sequence set forth in any one of SEQ ID NOs: 83-104. In some cases, the envelope protein comprises an amino acid sequence that has at least about 75% sequence identity to a sequence set forth in any one of SEQ ID NOs: 83-104. In some cases, the envelope protein comprises an amino acid sequence that has at least about 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence set forth in any one of SEQ ID NOs: 83-104. In some cases, the envelope protein comprises an amino acid sequence that has at least about 80% sequence identity to a sequence set forth in any one of SEQ ID NOs: 83-104 In some cases, the envelope protein comprises an amino acid sequence that has at least about 85% sequence identity to a sequence set forth in any one of SEQ ID NOs: 83-104. In some cases, the envelope protein comprises an amino acid sequence that has at least about 90% sequence identity to a sequence set forth in any one of SEQ ID NOs: 83-104. In some cases, the envelope protein comprises an amino acid sequence that has at least about 95% sequence identity to a sequence set forth in any one of SEQ ID NOs: 83-104 In some cases, the envelope protein comprises an amino acid sequence that has at least about 96% sequence identity to a sequence set forth in any one of SEQ ID NOs: 83-104. In some cases, the envelope protein comprises an amino acid sequence that has at least about 97% sequence identity to a sequence set forth in any one of SEQ ID NOs: 83-104. In some cases, the envelope protein comprises an amino acid sequence that has at least about 98% sequence identity to a sequence set forth in any one of SEQ ID NOs: 83-104. In some cases, the envelope protein comprises an amino acid sequence that has at least about 99% sequence identity to a sequence set forth in any one of SEQ ID NOs: 83-104.Table 1. Exemplary envelope proteins from virus originWSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601Envelope protein of human origin

[0267] In some aspects, the envelope protein in the lipid delivery particle described herein has a human origin, e.g., has significant sequence similarity to a human wild-type protein, such as at least 90%, at least 95%, at least 98%, or at least 99%. Using an envelope protein of a human origin can have benefits such as providing a minimized immunogenicity and better tolerance in a human subject receiving the lipid delivery particles. The lipid delivery particle comprising an envelope protein of a human origin can comprise another component that is from human origin or from non-human origin (e.g., a payload or a plasma membrane recruitment element). An envelope protein that is from human origin can include, example, envelope proteins or glycoproteins of human endogenous retroviruses (HERVs), other human endogenous envelope proteins, or other human endogenous proteins that serve a similar function of recognizing and / or fusing with membrane of a target cell (e.g., clathrin adaptor protein complex- 1, CHMP4C, Proteolipid protein 1, TSAP6, immunoglobulin variable domains, or a biologically active mutant thereof).

[0268] In some cases, the envelope protein is a HERV envelope protein such as any one of those listed in Table 2. In some cases, the envelope protein is a hENVHl or a biologically active mutant thereof. In some cases, the envelope protein is a hENVH2 or a biologically active mutant thereof. In some cases, the envelope protein is a hENVH3 or a biologically active mutant thereof. In some cases, the envelope protein is a hENVKl or a biologically active mutant thereof. In some cases, the envelope protein is a hENVK2 or a biologically active mutant thereof. In some cases, the envelope protein is a hENVK3 or a biologically active mutant thereof. In some cases, the envelope protein is a hENVK4 or a biologically active mutant thereof. In some cases, the envelope protein is a hENVK5 or a biologically active mutant thereof. In some cases, the envelope protein is a hENVK6 or a biologically active mutant thereof. In some cases, the envelope protein is a hENVT or a biologically active mutant thereof. In some cases, the envelope protein is a hENVW or a biologically active mutant thereof. In some cases, the envelope proteinWSGR Docket No.:62697-750.601 is a hENVFRD or a biologically active mutant thereof. In some cases, the envelope protein is a hENVR or a biologically active mutant thereof. In some cases, the envelope protein is a hENVR(b) or a biologically active mutant thereof. In some cases, the envelope protein is a hENVR(c)2 or a biologically active mutant thereof. In some cases, the envelope protein is a hENVR(c) 1 or a biologically active mutant thereof. In some cases, the envelope protein is a hENVKcon or a biologically active mutant thereof. In some cases, the envelope protein is a truncated HERV protein.Table 2. Exemplary HERV envelope proteins

[0269] In some cases, the envelope protein comprises the sequences set forth in Table 3. In some cases, the envelope protein comprises the sequences set forth in Table 3 with at least one amino acid substitution, deletion, or insertion. For example, for those amino acid sequences start with a N-terminal methionine, the N-terminal methionine can be absent. In some cases, the envelope protein comprises the sequences set forth in Table 3 and a heterologous peptide sequence fused to the N-terminal or C-terminal.

[0270] In some cases, the envelope protein comprises one or more of the sequences set forth in Table 3 with at least one amino acid substitution, deletion, or insertion. For instance, N-terminal methionine can be absent from the envelope protein of the lipid delivery particle provided herein relative to the wild-type viral envelope protein. In some cases, the envelope protein comprises one or more of the sequences set forth in Table 3 and a heterologous peptide sequence fused to the N-terminal or C-terminal.

[0271] In some cases, the envelope protein comprises any one of the sequences set forth in Table 3 with at least one amino acid substitution, deletion, or insertion. For instance, N-terminal methionine can be absent from the envelope protein of the lipid delivery particle provided herein relative to the wild-type viral envelope protein. In some cases, the envelope protein comprises any one of the sequences set forth in Table 3 and a heterologous peptide sequence fused to the N-terminal or C-terminal.WSGR Docket No.:62697-750.601

[0272] In some cases, the envelope protein comprises an amino acid sequence that has at least about 50%, 60%, 70%, 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence set forth in SEQ ID NOs: 49-82. In some cases, the envelope protein comprises an amino acid sequence that has at least about 50% sequence identity to a sequence set forth in any one of SEQ ID NOs: 49-82. In some cases, the envelope protein comprises an amino acid sequence that has at least about 60% sequence identity to a sequence set forth in any one of SEQ ID NOs: 49-82. In some cases, the envelope protein comprises an amino acid sequence that has at least about 70% sequence identity to a sequence set forth in any one of SEQ ID NOs: 49-82. In some cases, the envelope protein comprises an amino acid sequence that has at least about 75% sequence identity to a sequence set forth in any one of SEQ ID NOs: 49-82. In some cases, the envelope protein comprises an amino acid sequence that has at least about 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence set forth in any one of SEQ ID NOs: 49-82. In some cases, the envelope protein comprises an amino acid sequence that has at least about 80% sequence identity to a sequence set forth in any one of SEQ ID NOs: 49-82. In some cases, the envelope protein comprises an amino acid sequence that has at least about 85% sequence identity to a sequence set forth in any one of SEQ ID NOs: 49-82. In some cases, the envelope protein comprises an amino acid sequence that has at least about 90% sequence identity to a sequence set forth in any one of SEQ ID NOs: 49-82. In some cases, the envelope protein comprises an amino acid sequence that has at least about 95% sequence identity to a sequence set forth in any one of SEQ ID NOs: 49-82. In some cases, the envelope protein comprises an amino acid sequence that has at least about 96% sequence identity to a sequence set forth in any one of SEQ ID NOs: 49-82. In some cases, the envelope protein comprises an amino acid sequence that has at least about 97% sequence identity to a sequence set forth in any one of SEQ ID NOs: 49-82.In some cases, the envelope protein comprises an amino acid sequence that has at least about 98% sequence identity to a sequence set forth in any one of SEQ ID NOs: 49-82. In some cases, the envelope protein comprises an amino acid sequence that has at least about 99% sequence identity to a sequence set forth in any one of SEQ ID NOs: 49-82.Table 3. Exemplary sequences for human HERV envelope proteinsWSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601WSGR Docket No.:62697-750.601PAYLOAD

[0273] A payload in a lipid delivery particle of the present disclosure can comprise a protein, a polypeptide, a nucleic acid (e.g., DNA or RNA), or any combinations thereof.

[0274] The payload can be a part of the chimeric protein disclosed herein or can comprise a part of the chimeric protein disclosed herein. Alternatively or additionally, the payload can include an entity in the lipid delivery particle separate from the chimeric protein disclosed herein. For instance, in some cases, the payload is a protein or polypeptide coupled to a plasma membrane recruitment element. In some cases, the payload comprises a first moiety (e.g., a nucleic acid-binding protein) that is fused to a plasma membrane recruitment element, and further comprises a second moiety that is coupled to the first moiety via covalent or non-covalent interaction. For instance, the first moiety can be a nucleic acid binding protein that is fused with the plasma membrane recruitment element, and the second moiety can be a nucleic acid molecule that binds to the nucleic acid binding protein.

[0275] In some cases, a payload is directly packaged within the lipid delivery particles and delivered into a target cell in its free form. In some cases, a payload can be fused to a plasma membrane recruitment element (e.g., pleckstrin homology domain) and form a chimeric protein as part of the lipid delivery particles, and then delivered into the target cell. In some cases, the plasma membrane recruitment element (e.g., pleckstrin homology domain) forms at least part of a protein core of the lipid delivery particle. In some embodiments, the payload in its free form or as part of a chimeric protein is within the inside cavityWSGR Docket No.:62697-750.601 of the protein core of the lipid delivery particles disclosed herein. In some cases, the payload in its free form derives from a cleavage of the chimeric protein comprising the payload.

[0276] In some cases, a lipid delivery particle can deliver more than one payload. Each of the payloads can independently comprise nucleic acid-binding moiety, a nucleic acid-modifying moiety, a fusion protein, or a nucleic acid, or any combinations thereof.

[0277] In some embodiments, the plasma membrane recruitment element and the payload are coupled via any suitable method. Covalent coupling between the plasma membrane recruitment element and a payload peptide can include inteins that can form peptide bonds, direct protein-protein chimeras generated from a single reading frame. In some cases, nucleic acids base pairing to other nucleic acids via hydrogen bonding interactions (e.g., DNA / RNA, DNA / DNA, or RNA / RNA hybrids), protein-protein binding, or protein-nucleic acid molecule binding can be involved for the coupling between the plasma membrane recruitment element and the payload. Examples of protein-nucleic acid molecule binding include an RNA binding protein (RBP) and an RBP binding sequence (e.g., an RNA) that binds to the RBP. In some embodiments, each of the plasma membrane recruitment element and the payload is fused to a heterologous sequence, and the two heterologous sequences dimerize or multimerize with or without the need for a chemical compound to induce the protein-protein binding, such as a single-stranded nucleic acid sequence or protein dimerization domains). In some embodiments, each of the plasma membrane recruitment element and the payload is fused to one member of a pair of binding partners (e.g., antibody and its target antigen). In some embodiments, the plasma membrane recruitment element is fused to an RBP, and the payload is fused to a RBP binding sequence. Examples of suitable protein domains or nucleic acid molecules for forming the non-covalent connections include single chain variable fragments, nanobodies, affibodies, DmrA / DmrB / DmrC, FKBP / FRB, dDZFs, Leucine zippers, proteins that bind to DNA and / or RNA, optogenetic protein domains that can dimerize or multimerize in the presence of certain light wavelengths, proteins with quaternary structural interactions, and / or naturally reconstituting split proteins. Examples of RBPs and their RBP binding sequences that can be used include a sequence having at least about 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence set forth in Table 13. Examples of RBPs and their RBP binding sequences that can be used include a sequence having at least about 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to a sequence set forth in any one of SEQ ID NOs: 478- 513. In some cases, the RBP comprises an amino acid sequence as set forth in Table 13. In some cases, the RBP comprises any one of the sequences set forth in Table 13. In some cases, the RBP comprises one or more of the sequences set forth in Table 13. In some cases, the RBP comprises more than one of the sequences set forth in Table 13. In some cases, the RBP comprises multiple sequences set forth in Table 13. In some cases, the RBP binding sequence comprises an amino acid sequence as set forth in Table 13. In some cases, the RBP binding sequence comprises any one of the sequences set forth in Table 13. In some cases, the RBP binding sequence comprises one or more of the sequences set forth in Table 13. InWSGR Docket No.:62697-750.601 some cases, the RBP binding sequence comprises more than one of the sequences set forth in Table 13.In some cases, the RBP binding sequence comprises multiple sequences set forth in Table 13.Table 13. Exemplary RNA binding proteins (RBP) and corresponding RBP binding sequencesWSGR Docket No.:62697-750.601Nucleic acid binding domains and nucleic acid modifying domains

[0278] In some cases, the payload comprises a nucleic acid-binding moiety, a nucleic acid-modifying moiety, a fusion protein, or a nucleic acid. In some cases, the payload comprises a nucleic acid-binding domain, e.g., a DNA-binding protein domain or polypeptide or an RNA-binding domain or polypeptide e.g., an RNA-binding protein (RBP). A nucleic acid-binding moiety can be capable of binding a nucleic acid. A nucleic acid-binding domain can bind to a nucleic acid in a nonspecific or a site-specific manner.

[0279] In some cases, the nucleic acid-binding moiety binds to a nucleic acid in a site-specific manner. For example, a nucleic acid-binding moiety can comprise an aptamer binding domain that selectively binds to a specific target. In some cases, a nucleic acid-binding moiety recognizes a specific recognition sequence in the target nucleic acid. In some cases, a nucleic acid-binding moiety comprises an aptamer binding domain. In some cases, a nucleic acid binding moiety selectively binds to a sequence or a structural element in a nucleic acid molecule. In some cases, an RNA-binding domain selectively binds to a specific sequence motif in an RNA molecule. In some cases, a nucleic acid-binding moiety selectively binds to a structural element in a nucleic acid molecule. For example, a nucleic acid-binding domain can bind to a stem -loop in a nucleic acid molecule.

[0280] In some cases, a nucleic acid-binding moiety is or comprises a guidable polypeptide domain, a transcriptional regulatory domain, or a nucleic acid-modifying domain. A guidable polypeptide domain can be capable of binding to a polynucleotide (e.g. an RNA guide) that can direct the guidable polypeptide domain a target site. In some cases, the guidable polypeptide domain forms a complex with the RNA guide and recognizes the target sequence through DNA-RNA base pairing. In some cases, a nucleic -acid binding moiety is or comprises a transcriptional regulatory domain. In other cases, a nucleic- binding moiety can help recruit a transcriptional repressor or activator to a target site. In some cases, a nucleic acid-binding moiety is or comprises a nucleic acid-modifying moiety. In some cases, the present disclosure uses nucleic acid-binding moieties to recruit a nucleic acid-modifying moiety to a target site. In some cases, a nucleic-acid binding moiety comprises catalytic activity. In other cases, a nucleic acidbinding moiety is catalytically inactive. In some cases, a nucleic -acid binding moiety comprising catalytic activity is modified to have a reduced level of activity compared to its wild-type counterpart.WSGR Docket No.:62697-750.601

[0281] In some cases, the payload in the present disclosure comprises a nucleic acid modifying domain. A nucleic acid-modifying domain can comprise a polypeptide domain, a nucleic acid or a combination thereof (e.g., a ribonucleoprotein complex). A nucleic acid-modifying domain can be capable of modifying nucleic acid, such as cleaving double-stranded nucleic acid; nicking a single-stranded nucleic acid; introducing a mutation, deletion, or insertion in a nucleic acid; methylating or demethylating a nucleic acid, or altering the structure of DNA (e.g., changing chromatin structure through modifying histones). For example, a nucleic acid modifying domain can comprise a nuclease domain, a nickase domain, a deaminase domain, a polymerase, reverse transcriptase domain, a recombinase domain, a transposase domain, or an epigenetic modifying domain. A nuclease domain can be capable of cleaving phosphodiester bonds between nucleotides in nucleic acids. A nuclease domain can comprise an exonuclease (e.g., a nuclease capable of cleaving nucleic acids from the ends) or an endonuclease (e.g., a nuclease capable of cleaving nucleic acids in the middle). In some cases, a nucleic acid modifying effector or nucleic acid binding domain is a nickase, which can be capable of cleaving a single-strand in a double-stranded DNA. Nucleic acid modifying domains can be useful for gene editing, or for regulating, activating, or inhibiting gene expression.

[0282] In some cases, the payload in the present disclosure comprises a guidable polypeptide domain (e.g., a CRISPR-Cas protein domain). In some cases, a guidable polypeptide domain is capable of binding to a polynucleotide (e.g., a RNA guide) that directs it to a target site. In some cases, the guidable polypeptide domain forms a complex with the polynucleotide and recognizes the target sequence through DNA-RNA base pairing. In some cases, a guidable polypeptide domain is a CRISPR / CRISPR-associated (Cas) domain. A CRISPR domain can be a natural or an engineered domain. A Cas protein or domain can be derived from a CRISPR system or share structural and / or functional similarities to a protein involved in a CRISPR system. A CRISPR system is a system encoding DNA sequence arrays known as clustered regularly interspaced short palindromic repeats (CRISPRs), which can be found in microbial genomes or phage genomes. In some cases, CRISPR systems comprise genes encoding CRISPR-associated (Cas) proteins and / or small RNA guide molecules (e.g., crRNA or tracrRNA) that assemble with the CRISPR domain. In some cases, the CRISPR-Cas domain forms a complex with one or more RNA guide molecules to form an effector ribonucleoprotein complex. The effector ribonucleoprotein complex can recognize a target sequence through sequence specific DNA-RNA base pairing with a spacer sequence in the RNA guide. In some cases, target recognition activates one or more nuclease domains (e.g., a RuvC domain or HNH domain) in the CRISPR domain to make a double -stranded cut at the target DNA. A CRISPR-Cas domain complexed with an RNA guide can be capable of inactivating target gene through a gene knockout. In some cases, the CRISPR domain is used to enable gene insertion and / or deletion, which can inactivate, modify, or restore the gene’s function.

[0283] A CRISPR system can comprise single subunit or multi-subunit effectors. In some cases, a CRISPR system is a Class 1 CRISPR system. A Class 1 CRISPR system can be a type I, type III, or a type IV system. A Class 1 type I CRISPR system can comprise a multi-subunit effector. In some cases, a Class 1 type I CRISPR system comprises a protein or domain in the Cascade-Cas3 protein complex. AWSGR Docket No.:62697-750.601Class 1 type I CRISPR system can comprise a Cas6, Cas7, Cas5, Casl l, Cas8, or Cas3 domain. A Class 1 type III CRISPR system can comprise a multi-subunit effector. In some cases, a Class 1 type III CRISPR system comprises a Csm complex or a Cmr complex. In some cases, a Class 1 type III CRISPR system comprises a Cas6, a Cas7 (Csm3 or Cmr4), a Cas7-related (Csm5, Cmrl, or Cmr6), a Cas5 (e.g., Csm4 or Cmr5), a Casl 1 (e.g., Csm2 or Cmr3), or a CaslO (e.g., Csml or Cmr2) domain. A Class 1 type IV CRISPR system can comprise a Cas6, a Cas7, a Cas5, a Casl 1, a Cas8 (e.g., Csfl), or a DinG or CysH domain.

[0284] In some cases, a CRISPR system is a Class 2 CRISPR system. A Class 2 CRISPR system can be a Class 2 type II CRISPR system, a Class 2 type V CRISPR system, or a Class 2 type VI CRISPR system. A Class 2 type II CRISPR system can comprise a Cas9 domain. A Cas9 domain can be a SpyCas9, a GeoCas9, a SauCas9, a KhuCas9, a AinCas9, an FmaCas9, a SgaCas9, a ScCas9, a SauriCas9 domain. A Cas9 domain can be a hyperactive Cas9 domain. A Class 2 type V CRISPR system can comprise a Casl 2 domain. A Casl2 domain can be a Casl2a, a Casl2b, a Casl2c, a Casl2d, a Casl2e, a Casl2f, a Casl2g, a Casl2h, a Casl2i, a Casl2j, a Casl2k, a Casl21, or a Casl2m domain. A Class 2 type VI CRISPR system can comprise a Casl 3 domain. CRISPR systems include all those known in the art, including, for example, Casl2a variants, as described in Kleinstiver et al., Nat Biotechnol 37, 276-282 (2019), which is incorporated by reference in its entirety.

[0285] In some cases, a CRISPR-Cas domain comprises one or more subdomains. For example, a Cas9 domain can comprise a Reel, a Rec2, a Rec3, a RuvC, an HNH, or a Wedge / PAM-interacting domain. A Casl2 domain can comprise a Reel, Rec2, a crRNA oligonucleotide binding domain (OBD), a Nuc domain, a PAM-interacting (PI) domain, or a RuvC domain. In some cases, the RuvC domain comprises nuclease activity. In some cases, the HNH domain comprises nuclease activity. The PAM-interacting domain can bind to a protospacer adjacent motif (PAM) sequence that is next to a target sequence in a target nucleic acid molecule. PAM recognition can help activate a nuclease domain to make a cut at the target sequence. In some cases, a CRISPR protein or domain is an engineered or mutated variant of a protein involved in a CRISPR system. An engineered or mutated CRISPR domain can comprise a truncation, a deletion of a part of one or more domains or subdomains, or a mutation of an active site (e.g., a RuvC active site or HNH active site). In some cases, a CRISPR domain with a mutation of one or more active sites is catalytically inactive (e.g., dCas9). In some cases, a CRISPR domain with one or more mutated active sites comprises less than 90%, less than 80%, less than 70%, less than 60%, less than 50%, less than 40%, less than 30%, less than 20%, less than 10%, less than 5%, or less than 1% of the nuclease activity of its wildtype counterpart. For example, a dCas9 can result from the point mutations D10A in the RuvC domain and the point mutation H840A in the HNH domain. In other cases, a mutation can result in a CRISPR nickase. A nickase can generate nick or a single -stranded cut. A nickase can generate a nick in the strand complementary to the RNA guide (e.g., the targeting strand) or in the strand on the non-targeting strand. For example, a RuvC mutation D10A in a Cas9 domain can produce a Cas9 nickase domain that nicks the targeting strand. An HNH mutation H840A in a Cas9 domain can produce a Cas9 nickase domain that nicks the non-targeting strand.WSGR Docket No.:62697-750.601

[0286] In some cases, the payload in the present disclosure comprises a gene editor. The gene editor can be an Al-created gene editor. The Al-created gene editor may be generated by one or more models (e.g., large language models (LLMs)). In some cases, the payload in the present disclosure comprises OpenCRISPR. The OpenCRISPR payload can comprise a Cas-like protein, guide RNA (gRNA), or any combination thereof. In some cases, the payload in the present disclosure comprises OpenCRISPR comprising a Cas9-like protein, gRNA, or any combination thereof.

[0287] In some cases, the payload in the present disclosure comprises a base editor.

[0288] In some cases, the payload in the present disclosure comprises a deaminase domain. The deaminase domain can be a natural or an engineered domain. A deaminase domain can be capable of carrying out deamination reactions in DNA. A deaminase domain can be capable of enabling the generation of base conversions or point mutations in a target nucleic acid. For example, a deaminase domain can be a cytidine deaminase domain or an adenosine deaminase domain. A cytidine deaminase domain can be capable of converting cytosine to uracil. A cytidine deaminase domain can be capable of enabling the conversion of a C-G base pair to a T-A base pair. For example, a cytidine deaminase can be or comprise a APOBEC1 cytidine deaminase. An adenosine deaminase domain can be capable of converting an adenosine to hypoxanthine. An adenosine deaminase domain can be capable of converting an adenosine to an inosine. An adenosine deaminase can comprise TadA or a TadA mutant. An adenosine deaminase domain can be capable of enabling the conversion of an A-T base pair to a G-C base pair. A deaminase domain can be a mutated variant. In some cases, a deaminase domain enables the conversion of C to G, A to I, or C to U.

[0289] In some cases, the payload in the present disclosure comprises a polymerase (e.g., reverse transcriptase). A polymerase can comprise a natural or an engineered domain. A polymerase can be capable of synthesizing nucleic acids. A polymerase can be a DNA polymerase or an RNA polymerase. In some cases, a polymerase is a reverse transcriptase. A reverse transcriptase can synthesize DNA from deoxyribonucleotides. In some cases, a reverse transcriptase adds deoxyribonucleotides to the 3’ end of a nucleic acid primer to synthesize DNA. In some cases, a reverse transcriptase uses an RNA template and uses base-pairing interactions to synthesize a DNA strand that is complementary to the RNA template. The reverse transcriptase domain can be a reverse transcriptase from any organism, phage, virus, or an engineered or mutated variant. The reverse transcriptase domain can be a reverse transcriptase derived from or sharing structural or sequencing similarity to a reverse transcriptase in a CRISPR system. The reverse transcriptase can be an M-MLV or HIV reverse transcriptase. The reverse transcriptase can be a human LINE-1 reverse transcriptase or a group II intron reverse transcriptase. The reverse transcriptase can be a human endogenous retrovirus reverse transcriptase.

[0290] In some cases, a nucleic-acid modifying effector or a nucleic acid-binding moiety comprises a transposase domain. A transposase domain can be a natural or an engineered domain. A transposase domain can be capable of aiding the translocation of a transposable element, a nucleic acid sequence that can change its position within a genome. In some cases, a transposase domain comprises a TnsA, a TnsB, a TnsC, or a TnsD domain. In some cases, a transposase domain comprises a TniQ domain. In someWSGR Docket No.:62697-750.601 cases, a transposase domain is derived from or shares sequence or structural similarity with a transposase in a CRISPR system (e.g., a CRISPR-associated transposase). In some cases, a transposase domain is derived from or share sequence or structural similarity with a transposase domain from a type I CRISPR- associated transposon (CAST) system. In some cases, transposase domain is derived from or share sequence or structural similarity with a transposase domain from a type V CRISPR-associated transposon (CAST) system. A transposase domain can be capable of binding to a guidable polypeptide domain. In some cases, a transposase domain is coupled to a guidable polypeptide domain. In some cases, a transposase domain is capable of binding to a type I CRISPR-Cas domain (e.g., a Cascade domain, a Cas8 domain, or a Cas5 domain). In some cases, a transposase domain is capable of binding to a type V CRISPR-Cas domain (e.g., a Casl2 domain). In some cases, a transposase domain is capable of mediating targeted insertion of a nucleic acid into a target nucleic acid. In some cases, a transposase domain is capable of mediating targeted insertion of a nucleic acid that is at least 5 kb, at least 6 kb, at least 7 kb, at least 8kb, at least 9kb, at least lOkb, at least 1 Ikb, at least 12kb, at least 13kb, at least 14kb, or at least 15 kb into a target nucleic acid.

[0291] In some cases, the payload comprises a transcriptional regulatory domain. A transcriptional regulatory domain can be a natural or an engineered domain. A transcriptional regulatory domain can be capable of regulating, activating, or inhibiting gene expression. For example, a transcriptional repressor can silence gene expression by binding to the promoter of a gene. A transcriptional activator can bind to enhancers or regulatory elements to activate expression of a gene. A transcriptional regulatory domain can comprise a transcription factor. A transcriptional regulatory domain can comprise a transcriptional activation domain or a transcriptional repression domain. For example, a transcriptional activation domain can be or comprise a CAP domain, a VP64 domain, a p65 domain, an Rta domain, a synergistic activation mediator (SAM) domain, a SunTag domain, a VPR domain, a DNA demethylase domain, a histone methyltransferase domain, a histone acetyltransferase domain, or a histone demethylase domain. A transcriptional repression domain can be or comprise a dCas9 domain, a KRAB domain, a Sin3 interacting domain (SID), or a MePC2 domain, a DNA methyltransferase domain, a histone deacetylase domain, a histone methyltransferase domain, or a histone demethylase domain. In some cases, a transcriptional regulatory domain comprises an epigenetic modifying effector domain. For example, an epigenetic modifying effector can be a DNA methyltransferase, a DNA demethylase, a histone methyltransferase, a histone demethylase, a histone acetyltransferase, or a histone deacetylase domain. A DNA methyltransferase domain can be capable of methylating a nucleic acid. A DNA demethylase domain can be capable of demethylating a nucleic acid. A histone methyltransferase domain can be capable of methylating a histone. A histone demethylase domain can be capable of demethylating a histone. A histone acetyltransferase domain can be capable of adding an acetyl group to a histone. A histone deacetylase domain can be capable of removing an acetyl group from a histone.

[0292] In some embodiments, the payload comprises a zinc finger domain. A zinc finger domain can be a natural or an engineered domain. A zinc finger domain can bind to a specific DNA sequence in a target nucleic acid. A zinc finger domain can comprise from 1 to 10, from 2 to 10, from 3 to 10, from 4 to 10,WSGR Docket No.:62697-750.601 from 5 to 10, from 6 to 10, from 7 to 10, from 8 to 10, from 9 to 10 zinc fingers, from 1 to 8, from 2 to 8, from 3 to 8, from 4 to 8, from 5 to 8, from 6 to 8, from 7 to 8, from 8 to 8, from 9 to 8 zinc fingers. In some cases, a zinc finger domain comprises a two-handed zinc finger domain. A two handed zinc finger domain can comprise two clusters of zinc finger domains that are separated by intervening amino acids. A two handed zinc finger domain can bind to two noncontiguous target DNA sequences. In some cases, the spacing between the two noncontiguous target sequences comprise from 1 to 15, from 1 to 12, from 1 to 10, from 1 to 8, or from 1 to 5 nucleotides. For example, a two handed type of zinc finger binding protein can be SIP1. A cluster of zinc finger domains in a two handed zinc finger domain can be capable of binding to a unique target nucleic acid sequence.

[0293] In some embodiments, the payload comprises a TALE domain. A TALE domain can be a natural or an engineered domain. A TALE domain can bind to a specific DNA sequence. A TALE domain can comprise one or more effector domains. A TALE effector domain can comprise a central repeat domain comprising tandem repeats. A tandem repeat can comprise repeat variable residues (RVD). One or more RVDs can detect a specific DNA base. Different TALE effector domains may have a different number of repeats and a different order of their repeats. The C-terminal repeat is usually shorter in length (e.g., about 20 amino acids). Sequential repeats and their RVDs can recognize sequential DNA bases.

[0294] A TALE domain described herein can be derived from a TALE effector from a bacterial species. The TALE domain can be engineered to target a given nucleic acid sequence based on their DNA base specificities. The TALE domain can be engineered to remove or add a TALE effector domain. In some cases, the TALE domain corresponds to a perfect match to a nucleic acid target sequence. In some cases, the TALE domain of an epigenetic effector corresponds to one or more mismatches to a target base in the target nucleic acid.

[0295] In some cases, the payload in the present disclosure comprises a fusion protein. A fusion protein can comprise two or more polypeptide domains of any of the polypeptide domains described elsewhere herein. A fusion protein can be a natural or an engineered fusion protein. In some cases, the two or more polypeptide domains are coupled together. The two or more polypeptide domains can be coupled together directly or coupled together indirectly. For example, a first polypeptide domain can be coupled directly to a second polypeptide domain. Alternatively, the first polypeptide domain can be coupled indirectly to the second polypeptide domain by coupling with a third polypeptide domain that is coupled directly to the second polypeptide domain. In some cases, a first polypeptide domain is coupled to the N-terminus of a second polypeptide domain. In some cases, a first polypeptide domain is coupled to the C-terminus of a second polypeptide domain. In some cases, a first polypeptide domain is coupled to an internal component of a second polypeptide domain. In some cases, the two or more polypeptide domains are covalently linked. In some cases, the two or more polypeptide domains are noncovalently linked. In some cases, the two or more polypeptide domains are coupled together by a linker. For example, a linker may be a peptide linker. A linker can be a rigid linker, which helps maintain a fixed distance between the polypeptide domains that it links. A linker ...

Claims

WSGR Docket No.:62697-750.601CLAIMSWHAT IS CLAIMED IS:

1. A lipid delivery particle comprising:(a) a lipid membrane encapsulating a cavity;(b) an envelope protein on the lipid membrane;(c) a plasma membrane recruitment element;(d) a payload; and(e) a coiled-coil peptide pair, and wherein the coiled-coil peptide pair comprises a first coiled-coil peptide and a second coiled-coil peptide, wherein the first coiled-coil peptide and the second coiled-coil peptide comprise sequences having at least 80%, at least 85%, at least 90%, or at least 95%, or 100% sequence identity to the amino acid sequences set forth in:(1) SEQ ID NOs: 601 and 602, respectively, or SEQ ID NOs: 602 and 601, respectively;(2) SEQ ID NOs: 603 and 604, respectively, or SEQ ID NOs: 604 and 603, respectively;(3) SEQ ID NOs: 605 and 606, respectively, or SEQ ID NOs: 606 and 605, respectively;(4) SEQ ID NOs: 607 and 608, respectively, or SEQ ID NOs: 608 and 607, respectively;(5) SEQ ID NOs: 609 and 610, respectively, or SEQ ID NOs: 610 and 609, respectively;(6) SEQ ID NOs: 611 and 612, respectively, or SEQ ID NOs: 612 and 611, respectively;(7) SEQ ID NOs: 613 and 614, respectively, or SEQ ID NOs: 614 and 613, respectively;(8) SEQ ID NOs: 615 and 616, respectively, or SEQ ID NOs: 616 and 615, respectively;(9) SEQ ID NOs: 616 and 621, respectively, or SEQ ID NOs: 621 and 616, respectively; or(10) SEQ ID NOs: 617 and 618, respectively, or SEQ ID NOs: 618 and 617, respectively.

2. The lipid delivery particle of claim 1, wherein the payload comprises a gene-editing agent.

3. The lipid delivery particle of claim 2, wherein the gene-editing agent comprises a zinc finger (ZF), transcription activator-like effector (TALE), and / or CRISPR-based genome editing or modulating protein; a nucleic acid encoding a zinc finger (ZF), transcription activator-like effector (TALE), and / or CRISPR-based genome editing or modulating protein, a ribonucleoprotein complex (RNP) comprising a CRISPR-based genome editing or modulating protein, or any combination thereof.

4. The lipid delivery particle of claim 2 or 3, wherein the gene-editing agent comprises a base editor, a prime editor, or an epigenetic editor.

5. The lipid delivery particle of any one of claims 1-4, wherein the payload comprises one or more Cas proteins.

6. The lipid delivery particle of claim 5, wherein the one or more Cas proteins comprise a Cas9 protein, a Cas 12a protein, or any combination thereof.

7. The lipid delivery particle of claim 5 or 6, wherein the payload further comprises one or more guide RNA molecules (gRNAs).

8. A lipid delivery particle comprising:WSGR Docket No.:62697-750.601(a) a lipid membrane encapsulating a cavity;(b) an envelope protein on the lipid membrane;(c) a plasma membrane recruitment element;(d) a payload comprising a base editor; and(e) a coiled-coil peptide pair, and wherein the coiled-coil peptide pair comprises a first coiled-coil peptide and a second coiled-coil peptide.

9. The lipid delivery particle of claim 8, wherein the coiled-coil peptide pair comprises a first coiled-coil peptide and a second coiled-coil peptide, wherein the first coiled-coil peptide and the second coiled-coil peptide comprise sequences having at least 80%, at least 85%, at least 90%, or at least 95%, or 100% sequence identity to the amino acid sequences set forth in:(1) SEQ ID NOs: 601 and 602, respectively, or SEQ ID NOs: 602 and 601, respectively;(2) SEQ ID NOs: 603 and 604, respectively, or SEQ ID NOs: 604 and 603, respectively;(3) SEQ ID NOs: 605 and 606, respectively, or SEQ ID NOs: 606 and 605, respectively;(4) SEQ ID NOs: 607 and 608, respectively, or SEQ ID NOs: 608 and 607, respectively;(5) SEQ ID NOs: 609 and 610, respectively, or SEQ ID NOs: 610 and 609, respectively;(6) SEQ ID NOs: 611 and 612, respectively, or SEQ ID NOs: 612 and 611, respectively;(7) SEQ ID NOs: 613 and 614, respectively, or SEQ ID NOs: 614 and 613, respectively;(8) SEQ ID NOs: 615 and 616, respectively, or SEQ ID NOs: 616 and 615, respectively;(9) SEQ ID NOs: 616 and 621, respectively, or SEQ ID NOs: 621 and 616, respectively;(10) SEQ ID NOs: 617 and 618, respectively, or SEQ ID NOs: 618 and 617, respectively; or(11) SEQ ID NOs: 613 and 619, respectively, or SEQ ID NOs: 619 and 613, respectively.

10. A lipid delivery particle comprising:(a) a lipid membrane encapsulating a cavity;(b) an envelope protein on the lipid membrane;(c) a plasma membrane recruitment element;(d) a payload; and(e) a coiled-coil peptide pair, and wherein the coiled-coil peptide pair comprises (i) a first coiled-coil peptide comprising a heptad repeat of an amino sequence as set forth in (EXiX^X XsXe)!!!, where Xi, X2, X3, X4, X5, and X„ are each independently any amino acid residue, and wherein m is an integer greater than or equal to 2, and (ii) a second coiled-coil peptide comprising a heptad repeat of an amino sequence as set forth in (KXyXsXgXioXnXn)!!, where X7, Xs, X>, X10, Xu, and X12 are each independently any amino acid residue, and wherein n is an integer greater than or equal to 2.

11. The lipid delivery particle of claim 10, wherein Xi or X4 is a hydrophobic amino acid residue, or Xi and X4 are each independently a hydrophobic amino acid residue.WSGR Docket No.:62697-750.60112. The lipid delivery particle of claim 10 or 11, wherein Xi is isoleucine and X4 is leucine, or wherein Xi is leucine and X4 is isoleucine.

13. The lipid delivery particle of any one of claims 10-12, wherein X7 or X is a hydrophobic amino acid residue, or X7 and Xw are each independently a hydrophobic amino acid residue.

14. The lipid delivery particle of any one of claims 10-13, wherein X7 is isoleucine and Xw is leucine, or wherein X7 is leucine and X is isoleucine.

15. The lipid delivery particle of any one of claims 10-12, wherein X7, Xs, Xw, Xu, and X12 are each independently a polar amino acid residue or a charged amino acid residue.

16. The lipid delivery particle of claim 15, wherein X7, Xs, Xw, Xu, and X12 are each independently an amino acid residue selected from the group consisting of serine, threonine, cysteine, asparagine, glutamine, tyrosine, glutamate, aspartate, arginine, and lysine.

17. The lipid delivery particle of claim 15, wherein X7, Xs, Xw, Xu, and X12 are each independently a polar amino acid residue.

18. The lipid delivery particle of claim 17, wherein X7, Xs, Xw, Xu, and X12 are each independently an amino acid residue selected from the group consisting of serine, threonine, cysteine, asparagine, glutamine, and tyrosine.

19. The lipid delivery particle of claim 15, wherein X7, Xs, Xw, Xu, and X12 are each independently a charged amino acid residue.

20. The lipid delivery particle of claim 19, wherein X7, Xs, Xw, Xu, and X12 are each independently an amino acid residue selected from the group consisting of glutamate, aspartate, arginine, and lysine.

21. The lipid delivery particle of any one of claims 10-20, wherein m is 3 or 4.

22. The lipid delivery particle of any one of claims 10-20, wherein n is 3 or 4.

23. The lipid delivery particle of any one of claims 10-20, wherein m and n are each independently 3 or4.

24. The lipid delivery particle of any one of claims 10-23, wherein the payload comprises a gene-editing agent.

25. The lipid delivery particle of claim 24, wherein the gene-editing agent comprises a zinc finger (ZF), transcription activator-like effector (TALE), and / or CRISPR-based genome editing or modulating protein; a nucleic acid encoding a zinc finger (ZF), transcription activator-like effector (TALE), and / or CRISPR-based genome editing or modulating protein, a ribonucleoprotein complex (RNP) comprising a CRISPR-based genome editing or modulating protein, or any combination thereof.

26. The lipid delivery particle of claim 24 or 25, wherein the gene-editing agent comprises a base editor, a prime editor, or an epigenetic editor.

27. The lipid delivery particle of any one of claims 10-26, wherein the payload comprises one or more Cas proteins.

28. The lipid delivery particle of claim 27, wherein the one or more Cas proteins comprise a Cas9 protein, a Cas 12a protein, or any combination thereof.WSGR Docket No.:62697-750.60129. The lipid delivery particle of claim 27 or 28, wherein the payload further comprises one or more guide RNA molecules (gRNAs).

30. A lipid delivery particle comprising:(a) a lipid membrane encapsulating a cavity;(b) an envelope protein on the lipid membrane;(c) a plasma membrane recruitment element;(d) a payload; and(e) a coiled-coil peptide pair, and wherein the coiled-coil peptide pair comprises (i) a first coiled-coil peptide comprising one or more of a first heptad motif in tandem, the first heptad motif comprising amino acid residues of the formula (a-b-c-d-e-f-g)x, where a or d is a hydrophobic amino acid residue, or a and d are each independently a hydrophobic amino acid residue, where b, c, e, f, g, or any combination thereof is a polar-charged amino acid residue, and wherein x is an integer greater than or equal to 2, and (ii) a second coiled-coil peptide comprising one or more of a second heptad motif in tandem, the second heptad motif comprising amino acid residues of the formula (h-i-j-k-l-m-n)y, where h or k is a hydrophobic amino acid residue, or h and k are each independently a hydrophobic amino acid residue, where b, c, e, f, g, or any combination thereof is a polar-charged amino acid residue, and wherein y is an integer greater than or equal to 2.

31. The lipid delivery particle of claim 30, wherein a and / or d is alanine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, valine, or any combination thereof.

32. The lipid delivery particle of claim 30 or 31, wherein a is isoleucine and d is leucine, or wherein d is leucine and a is isoleucine.

33. The lipid delivery particle of any one of claims 30-32, wherein b, c, e, f, or g are each independently a polar amino acid residue or a charged amino acid residue.

34. The lipid delivery particle of claim 33, wherein b, c, e, f, or g are each independently an amino acid residue selected from the group consisting of serine, threonine, cysteine, asparagine, glutamine, tyrosine, glutamate, aspartate, arginine, and lysine.

35. The lipid delivery particle of claim 33, wherein b, c, e, f, or g are each independently a polar amino acid residue.

36. The lipid delivery particle of claim 35, wherein b, c, e, f, or g are each independently an amino acid residue selected from the group consisting of serine, threonine, cysteine, asparagine, glutamine, and tyrosine.

37. The lipid delivery particle of claim 33, wherein b, c, e, f, or g are each independently a charged amino acid residue.

38. The lipid delivery particle of claim 37, wherein b, c, e, f, or g are each independently an amino acid residue selected from the group consisting of glutamate, aspartate, arginine, and lysine.WSGR Docket No.:62697-750.60139. The lipid delivery particle of any one of claims 30-38, wherein h and / or k is alanine, glycine, isoleucine, leucine, methionine, phenylalanine, proline, tryptophan, valine, or any combination thereof.

40. The lipid delivery particle of any one of claims 30-39, wherein h is isoleucine and k is leucine, or wherein k is leucine and h is isoleucine.

41. The lipid delivery particle of any one of claims 30-40, wherein i, j, 1, m, or n are each independently a polar amino acid residue or a charged amino acid residue.

42. The lipid delivery particle of claim 41, wherein i, j, 1, m, or n are each independently an amino acid residue selected from the group consisting of serine, threonine, cysteine, asparagine, glutamine, tyrosine, glutamate, aspartate, arginine, and lysine.

43. The lipid delivery particle of claim 41, wherein i, j, 1, m, or n are each independently a polar amino acid residue.

44. The lipid delivery particle of claim 43, wherein i, j, 1, m, or n are each independently an amino acid residue selected from the group consisting of serine, threonine, cysteine, asparagine, glutamine, and tyrosine.

45. The lipid delivery particle of claim 41, wherein i, j, 1, m, or n are each independently a charged amino acid residue.

46. The lipid delivery particle of claim 45, wherein i, j, 1, m, or n are each independently an amino acid residue selected from the group consisting of glutamate, aspartate, arginine, and lysine.

47. The lipid delivery particle of any one of claims 30-46, wherein x is an integer from 2 to 8.

48. The lipid delivery particle of any one of claims 30-47, wherein y is an integer from 2 to 8.

49. The lipid delivery particle of any one of claims 30-47, wherein x and y are each independently an integer from 2 to 8.

50. The lipid delivery particle of any one of claims 30-49, wherein the payload comprises a gene-editing agent.

51. The lipid delivery particle of claim 50, wherein the gene-editing agent comprises a zinc finger (ZF), transcription activator-like effector (TALE), and / or CRISPR-based genome editing or modulating protein; a nucleic acid encoding a zinc finger (ZF), transcription activator-like effector (TALE), and / or CRISPR-based genome editing or modulating protein, a ribonucleoprotein complex (RNP) comprising a CRISPR-based genome editing or modulating protein, or any combination thereof.

52. The lipid delivery particle of claim 50 or 51, wherein the gene-editing agent comprises a base editor, a prime editor, or an epigenetic editor.

53. The lipid delivery particle of any one of claims 30-52, wherein the payload comprises one or more Cas proteins.

54. The lipid delivery particle of claim 53, wherein the one or more Cas proteins comprise a Cas9 protein, a Cas 12a protein, or any combination thereof.

55. The lipid delivery particle of claim 53 or 54, wherein the payload further comprises one or more guide RNA molecules (gRNAs).WSGR Docket No.:62697-750.60156. The lipid delivery particle of any one of claims 1-55, wherein a spatial orientation of the first coiled- coil peptide is parallel to a spatial orientation of the second coiled-coil peptide.

57. The lipid delivery particle of any one of claims 1-55, wherein a spatial orientation of the first coiled- coil peptide is antiparallel to a spatial orientation of the second coiled-coil peptide.

58. The lipid delivery particle of any one of claims 1-57, wherein the coiled-coil peptide pair is selected from the group consisting of E4-K4, EE12RR345L-RR12EE345L, EE1234L-RR1234L, EE12345L- RR12345L, AcidPl -BasePl, P3(x3)-P4, SynZip2-SynZipl9, SynZip2-SynZipl, and N5-N6.

59. The lipid delivery particle of any one of claims 1-58, wherein:(i) the first coiled-coil peptide is a E4 peptide, and the second coiled-coil peptide is a K4 peptide; or(ii) the first coiled-coil peptide is a K4 peptide, and the second coiled-coil peptide is a E4 peptide.

60. The lipid delivery particle of any one of claims 1-59, wherein the first coiled-coil peptide is coupled with the plasma membrane recruitment element.

61. The lipid delivery particle of claim 60, wherein the lipid delivery particle comprises a first chimeric protein comprising the first coiled-coil peptide fused with the plasma membrane recruitment element.

62. The lipid delivery particle of claim 61, wherein the first coiled-coil peptide is fused to the C-terminus of the plasma membrane recruitment element.

63. The lipid delivery particle of claim 61, wherein the first coiled-coil peptide is fused to the N-terminus of the plasma membrane recruitment element.

64. The lipid delivery particle of any one of claims 61-63, wherein the payload comprises a payload protein that is coupled with the second coiled-coil peptide.

65. The lipid delivery particle of claim 64, wherein the lipid delivery particle comprises a second chimeric protein comprising the second coiled-coil peptide fused with the payload protein.

66. The lipid delivery particle of claim 65, wherein the second coiled-coil peptide is fused to the C- terminus of the payload protein.

67. The lipid delivery particle of claim 65, wherein the second coiled-coil peptide is fused to the N- terminus of the payload protein.

68. The lipid delivery particle of any one of claims 1-67, wherein the plasma membrane recruitment element comprises an amino acid sequence having at least 80%, 85%, 90%, 95%, or 99% sequence identity to a sequence set forth in Table 4.

69. The lipid delivery particle of any one of claims 1-67, wherein the plasma membrane recruitment element comprises an amino acid sequence set forth in Table 4.

70. The lipid delivery particle of any one of claims 1-67, wherein the plasma membrane recruitment element comprises a pleckstrin homology (PH) domain.

71. The lipid delivery particle of claim 70, wherein the PH domain is from a protein selected from the group consisting of human phospholipase C51, human Aktl, human Aktl with E17K substitution, human 3-phosphoinositide-dependent protein kinase 1, human Daapl, mouse Grpl, human Grpl, human OSBP, human Btkl, human FAPP1, human CERT, human PKD, human PHLPP1, human SWAP70, and human MAPKAP1.WSGR Docket No.:62697-750.60172. The lipid delivery particle of claim 70, wherein the PH domain comprises a sequence set forth in any one of SEQ ID NOs: 5-14 and 23-48.

73. The lipid delivery particle of any one of claims 1-67, wherein the plasma membrane recruitment element comprises a gag polyprotein.

74. The lipid delivery particle of claim 73, wherein the gag polyprotein is a retroviral gag polyprotein.

75. The lipid delivery particle of claim 73, wherein the gag polyprotein is a gag polyprotein from human immunodeficiency virus (HIV), murine leukemia virus (MLV), Moloney murine leukemia virus (MMLV), Friend murine leukemia virus (FMLV), Baboon endogenous retrovirus (BaEV), Simian immunodeficiency virus (SIV), Rous sarcoma virus (RSV), human T-cell leukemia virus type-1 (HTLV), bovine leukemia virus (BLV), Feline Leukemia Virus (FeLV), Gibbon Ape Leukemia Virus (GaLV), Koala Retrovirus (KRV), Reticuloendotheliosis Virus (ReEV), Wooly Monkey Sarcoma Virus (WMSV), or a biologically active mutant thereof, or any combination thereof.

76. The lipid delivery particle of claim 73, wherein the gag polyprotein is a human endogenous retroviral gag polyprotein.

77. The lipid delivery particle of claim 73, wherein the gag polyprotein comprises a sequence having at least 80%, at least 85%, at least 90%, or at least 95%, or 100% sequence identity to the amino acid sequence set forth in any one of SEQ ID NOs: 285-305.

78. The lipid delivery particle of any one of claims 65-77, wherein the second chimeric protein further comprises a linker between the second coiled-coil peptide and the payload protein.

79. The lipid delivery particle of claim 78, wherein the linker is a cleavable linker.

80. The lipid delivery particle of any one of claims 65-79, wherein the second chimeric protein further comprises a nuclear export signal (NES), a nuclear localization signal (NLS), or both.

81. The lipid delivery particle of claim 80, wherein the second chimeric protein comprises two or more NES.

82. The lipid delivery particle of claim 80, wherein the second chimeric protein comprises three NES.

83. The lipid delivery particle of any one of claims 80-82, wherein the NES is linked between the second coiled-coil peptide and the payload protein.

84. The lipid delivery particle of any one of claims 80-83, wherein the NES and the second coiled-coil peptide are same side of the cleavage linker in the second chimeric protein.

85. The lipid delivery particle of claim 84, wherein the NLS and the payload protein are on same side of the cleavage linker in the second chimeric protein.

86. The lipid delivery particle of any one of claims 80-85, wherein the NES comprises an amino acid sequence having at least 80%, 85%, 90%, 95%, or 99% sequence identity to a sequence set forth in Tables 11.

87. The lipid delivery particle of any one of claims 80-85, wherein the NES comprises an amino acid sequence set forth in Tables 11.WSGR Docket No.:62697-750.60188. The lipid delivery particle of any one of claims 80-87, wherein the NLS comprises an amino acid sequence having at least 80%, 85%, 90%, 95%, or 99% sequence identity to a sequence set forth in Table 12.

89. The lipid delivery particle of any one of claims 80-87, wherein the NLS comprises an amino acid sequence set forth in Table 12.

90. The lipid delivery particle of any one of claims 1-89, wherein a fusion of the coiled-coil peptide pair improves an editing efficiency of a target genetic locus in a cell contacted by the lipid delivery particle compared to an editing efficiency of the target genetic locus in the cell when contacted by an otherwise identical lipid delivery particle that does not comprise the coiled-coil peptide pair.

91. The lipid delivery particle of claim 90, wherein the editing efficiency of the target genetic locus in the cell contacted by the lipid delivery particle is at least about 30%.

92. The lipid delivery particle of claim 90 or 91, wherein the editing efficiency of the target genetic locus in the cell contacted by the lipid delivery particle is at least about 50%.

93. The lipid delivery particle of any one of claims 90-92, wherein the editing efficiency of the lipid delivery particle is base editing percentage.

94. A lipid delivery particle comprising:(a) a lipid membrane encapsulating a cavity;(b) an envelope protein on the lipid membrane; and(c) a chimeric protein comprising a plasma membrane recruitment element and a payload protein, wherein the plasma membrane recruitment element comprises a gag polyprotein, wherein the chimeric protein comprises a nuclear export signal (NES) inside the gag protein, and wherein the payload protein does not comprise a reverse transcriptase.

95. The lipid delivery particle of claim 94, wherein the chimeric protein comprises two or more NES.

96. The lipid delivery particle of claim 95, wherein the chimeric protein comprises three NES.

97. The lipid delivery particle of any one of claims 94-96, wherein the NES is linked between the gag protein and the payload protein.

98. The lipid delivery particle of claim 97, wherein the chimeric protein further comprises a cleavage linker between the NES and the payload protein.

99. The lipid delivery particle of claim 98, wherein the chimeric protein further comprises a NLS, and wherein the NLS and the payload protein are both on N-terminal side or C-terminal side of the cleavage linker in the chimeric protein.

100. The lipid delivery particle of claim 99, wherein the chimeric protein comprises at least two NLS, optionally two NLS.

101. The lipid delivery particle of claim 100, wherein the at least two NLS are at both N-terminus and C-terminus of the payload protein.

102. The lipid delivery particle of claim 100, wherein the at least two NLS are at either N-terminus or C-terminus of the payload protein.WSGR Docket No.:62697-750.601103. The lipid delivery particle of any one of claims 94-102, wherein the NES comprises an amino acid sequence having at least 80%, 85%, 90%, 95%, or 99% sequence identity to a sequence set forth in Tables 11.

104. The lipid delivery particle of any one of claims 94-102, wherein the NES comprises an amino acid sequence set forth in Tables 11.

105. The lipid delivery particle of any one of claims 94-104, wherein the NLS comprises an amino acid sequence having at least 80%, 85%, 90%, 95%, or 99% sequence identity to a sequence set forth in Table 12.

106. The lipid delivery particle of any one of claims 94-104, wherein the NLS comprises an amino acid sequence set forth in Table 12.

107. The lipid delivery particle of any one of claims 94-106, wherein the gag polyprotein is a retroviral gag polyprotein.

108. The lipid delivery particle of claim 107, wherein the gag polyprotein is a gag polyprotein from human immunodeficiency virus (HIV), murine leukemia virus (MLV), Moloney murine leukemia virus (MMLV), Friend murine leukemia virus (FMLV), Baboon endogenous retrovirus (BaEV), Simian immunodeficiency virus (SIV), Rous sarcoma virus (RSV), human T-cell leukemia virus type-1 (HTLV), bovine leukemia virus (BLV), Feline Leukemia Virus (FeLV), Gibbon Ape Leukemia Virus (GaLV), Koala Retrovirus (KRV), Reticuloendotheliosis Virus (ReEV), Wooly Monkey Sarcoma Virus (WMSV), or a biologically active mutant thereof, or any combination thereof.

109. The lipid delivery particle of any one of claims 94-106, wherein the gag polyprotein is a human endogenous retroviral gag polyprotein.

110. The lipid delivery particle of any one of claims 94-109, wherein the gag polyprotein comprises a sequence having at least 80%, at least 85%, at least 90%, or at least 95%, or 100% sequence identity to the amino acid sequence set forth in any one of SEQ ID NOs: 285-305.

111. The lipid delivery particle of any one of claims 73-110, wherein the gag polyprotein lacks at least a fragment of a nucleocapsid protein, optionally lacks the full length of the nucleocapsid protein.

112. The lipid delivery particle of claim 94-111, wherein the plasma membrane recruitment element comprises a heterologous domain fused to the C-terminus of the gag polyprotein, optionally wherein the heterologous domain comprises a leucine zipper.

113. The lipid delivery particle of claim 112, wherein the heterologous domain further comprises a linker sequence flanking the N-terminus or C-terminus side of the leucine zipper.

114. The lipid delivery particle of claim 112 or 113, wherein the heterologous domain further comprises a linker sequence flanking on both N-terminus and C-terminus sides of the leucine zipper.

115. The lipid delivery particle of claim 113 or 114, wherein the linker sequence comprises at least one repeat of an amino acid sequence SGGS, the sequence of any one of SEQ ID NO: 343-346 or 673, optionally two repeats thereof.WSGR Docket No.:62697-750.601116. The lipid delivery particle of any one of claims 73-115, wherein the gag polyprotein lacks at least fragment of a matrix protein, optionally lacks the full length of the matrix protein.

117. The lipid delivery particle of claim 116, wherein the plasma membrane recruitment element comprises a pleckstrin homology (PH) domain fused to the N-terminus of the gag polyprotein, optionally wherein the PH domain is from a protein selected from the group consisting of human phospholipase C51, human Aktl, human Aktl with E17K substitution, human 3-phosphoinositide- dependent protein kinase 1, human Daapl, mouse Grpl, human Grpl, human OSBP, human Btkl, human FAPP1, human CERT, human PKD, human PHLPP1, human SWAP70, and human MAPKAP1.

118. The lipid delivery particle of claim 117, wherein the PH comprises a sequence set forth in any one of SEQ ID NOs: 5-14 and 23-48.

119. A lipid delivery particle comprising:(a) a lipid membrane encapsulating a cavity;(b) an envelope protein on the lipid membrane;(c) a chimeric protein comprising a plasma membrane recruitment element and a payload protein, wherein the plasma membrane recruitment element comprises a gag polyprotein, and wherein:(i) the gag polyprotein lacks at least a fragment of a nucleocapsid protein, and the plasma membrane recruitment element further comprises a heterologous domain fused to C-terminus of the gag polyprotein; and / or(ii) the gag polyprotein lacks at least a fragment of a matrix protein, and the plasma membrane recruitment element further comprises a pleckstrin homology (PH) domain fused to N-terminus of the gag polyprotein.

120. The lipid delivery particle of claim 119, wherein the gag polyprotein comprises a matrix protein and a capsid protein, and lacks at least a fragment of a nucleocapsid protein, and the plasma membrane recruitment element further comprises a heterologous domain within the gag polyprotein.

121. The lipid delivery particle of claim 119, wherein the gag polyprotein comprises a matrix protein and a capsid protein, and lacks at least a fragment of a nucleocapsid protein, and the plasma membrane recruitment element further comprises a heterologous domain fused to C-terminus of the gag polyprotein.

122. The lipid delivery particle of claim 120 or 121, wherein the heterologous domain comprises a leucine zipper.

123. The lipid delivery particle of claim 122, wherein the heterologous domain further comprises a linker sequence flanking the N-terminus or C-terminus side of the leucine zipper.

124. The lipid delivery particle of claim 122, wherein the heterologous domain further comprises a linker sequence flanking on both N-terminus and C-terminus sides of the leucine zipper.

125. The lipid delivery particle of claim 123 or 124, wherein the linker sequence comprises at least one repeat of an amino acid sequence SGGS, the sequence of any one of SEQ ID NO: 343-346 or 673, optionally two repeats thereof.WSGR Docket No.:62697-750.601126. The lipid delivery particle of any one of claims 119-125, wherein the gag polyprotein is a retroviral gag polyprotein.

127. The lipid delivery particle of claim 126, wherein the gag polyprotein is a gag polyprotein from human immunodeficiency virus (HIV), murine leukemia virus (MLV), Moloney murine leukemia virus (MMLV), Friend murine leukemia virus (FMLV), Baboon endogenous retrovirus (BaEV), Simian immunodeficiency virus (SIV), Rous sarcoma virus (RSV), human T-cell leukemia virus type-1 (HTLV), bovine leukemia virus (BLV), Feline Leukemia Virus (FeLV), Gibbon Ape Leukemia Virus (GaLV), Koala Retrovirus (KRV), Reticuloendotheliosis Virus (ReEV), Wooly Monkey Sarcoma Virus (WMSV), or a biologically active mutant thereof, or any combination thereof.

128. The lipid delivery particle of any one of claims 119-127, wherein the gag polyprotein is a human endogenous retroviral gag polyprotein.

129. The lipid delivery particle of any one of claims 119-128, wherein the gag polyprotein comprises a sequence having at least 80%, at least 85%, at least 90%, or at least 95%, or 100% sequence identity to the amino acid sequence set forth in any one of SEQ ID NOs: 285-305.

130. The lipid delivery particle of any one of claims 119-129, wherein the chimeric protein further comprises a nuclear export signal (NES).

131. The lipid delivery particle of claim 130, wherein the chimeric protein comprises two or more NES.

132. The lipid delivery particle of claim 131, wherein the chimeric protein comprises three NES.

133. The lipid delivery particle of claim 130 or 131, wherein the NES is linked between the gag protein and the payload protein.

134. The lipid delivery particle of claim 133, wherein the chimeric protein further comprises a cleavage linker between the NES and the payload protein.

135. The lipid delivery particle of claim 134, wherein the chimeric protein further comprises aNLS, and wherein the NLS and the payload protein are on same side of the cleavage linker in the chimeric protein.

136. The lipid delivery particle of claim 135, wherein the chimeric protein comprises at least two NLS, optionally two NLS.

137. The lipid delivery particle of claim 136, wherein the at least two NLS are at both N-terminus and C-terminus of the payload protein.

138. The lipid delivery particle of claim 136, wherein the at least two NLS are at either N-terminus or C-terminus of the payload protein.

139. The lipid delivery particle of any one of claims 130-138, wherein the NES comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99%, or 100% sequence identity to a sequence set forth in Tables 11.WSGR Docket No.:62697-750.601140. The lipid delivery particle of any one of claims 130-139, wherein the NLS comprises an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95%, or at least 99%, or 100% sequence identity to a sequence set forth in Table 12.

141. The lipid delivery particle of any one of claims 94-139, wherein the payload protein comprises a gene-editing agent.

142. The lipid delivery particle of claim 141, wherein the gene-editing agent comprises a zinc finger (ZF), transcription activator-like effector (TALE), and / or CRISPR-based genome editing or modulating protein; a nucleic acid encoding a zinc finger (ZF), transcription activator-like effector (TALE), and / or CRISPR-based genome editing or modulating protein, a ribonucleoprotein complex (RNP) comprising a CRISPR-based genome editing or modulating protein, or any combination thereof.

143. The lipid delivery particle of claim 141 or 142, wherein the gene-editing agent comprises a base editor, a prime editor, or an epigenetic editor.

144. The lipid delivery particle of any one of claims 94-143, wherein the payload protein comprises one or more Cas proteins.

145. The lipid delivery particle of claim 144, wherein the one or more Cas proteins comprise a Cas9 protein, a Cas 12a protein, or any combination thereof.

146. The lipid delivery particle of claim 144 or 145, wherein the lipid delivery particle further comprises one or more guide RNA molecules (gRNAs) as a payload within the cavity.

147. The lipid delivery particle of any one of claims 1-146, wherein the envelope protein comprises a VSV glycoprotein (VSV-G), a human immunodeficiency virus (HIV) GP160 glycoprotein, a Baboon Endogenous Retrovirus (BaEVTR) glycoprotein, a fusion protein of Vesicular stomatitis Indiana virus and Rabies virus glycoprotein (FuG-E), an ecotropic Murine Leukemia Virus envelope protein (MLV ENV ecotropic), a human T-cell lymphotropic virus type 1 (HTLV-1) glycoprotein, an amphotrophic Murine Leukemia Virus envelope protein (MLV ENV amphotropic), a Moloney murine leukemia virus 10A1 strain glycoprotein (MLV 10A1), a Baculovirus envelope glycoprotein (GP64), a pantropic MLV envelope protein, a xenotropic MLV envelope protein, a xenotropic murine leukemia virus (XMLV) envelope protein, a Moloney murine leukemia virus (MMLV) envelope protein, a Moloney murine sarcoma virus (MoMSVg) envelope protein, a simian endogenous type D retrovirus protein (RD-114), a gibbon ape leukemia virus (GALV) envelope protein, a feline leukemia virus (FLV) envelope protein, a mouse mammary tumor virus (MMTV) envelope protein, an avian leukosis virus envelope protein, a Rous Sarcoma virus envelope protein, or an endogenous feline virus envelope protein (RD 114 ENV), or a mutant thereof.

148. The lipid delivery particle of any one of claims 1-146, wherein the envelope protein comprises a glycoprotein of a mammalian endogenous retrovirus or mutant thereof.

149. The lipid delivery particle of claim 148, wherein the glycoprotein of the mammalian endogenous retrovirus is a glycoprotein of a human endogenous retrovirus (hERV).WSGR Docket No.:62697-750.601150. The lipid delivery particle of claim 149, wherein the glycoprotein of a hERV comprises a hENVHl, a hENVH2, a hENVH3, a hENVKl, a hENVK2, a hENVK3, a hENVK4, a hENVK5, a hENVK6, a hENVT, a hENVW, a hENVFRD, a hENVR, a hENVR(b), a hENVR(c)l, a hENVR(c)2 or a hENVKcon, or a biologically active mutant thereof.

151. The lipid delivery particle of claim 149, wherein the hERV comprises a modified envelope protein.

152. The lipid delivery particle of any one of claims 1-146, wherein the envelope protein comprises a non-viral envelope protein.

153. The lipid delivery particle of any one of claims 1-152, wherein the lipid delivery particle further comprises a gag / pro / pol polypeptide, wherein the gag / pro polyprotein comprises a fusion of a second polyprotein, a pro protein, and a pol polyprotein.

154. The lipid delivery particle of any one of claims 1-152, wherein the lipid delivery particle further comprises:(a) a first gag polyprotein, wherein the first gag polyprotein is not fused with a pro protein or a pol polyprotein; and(b) a gag / pro polyprotein, wherein the gag / pro polyprotein comprises a fusion of a second gag polyprotein and a pro protein, and does not comprise and is not fused to a pol polyprotein.

155. The lipid delivery particle of claim 153 or 154, wherein the first gag polyprotein or the second gag polyprotein, or both are a retroviral gag polyprotein.

156. The lipid delivery particle of claim 155, wherein the first gag polyprotein or the second gag polyprotein, or both are a gag polyprotein from human immunodeficiency virus (HIV), murine leukemia virus (MLV), Moloney murine leukemia virus (MMLV), Friend murine leukemia virus (FMLV), Baboon endogenous retrovirus (BaEV), Simian immunodeficiency virus (SIV), Rous sarcoma virus (RSV), human T-cell leukemia virus type-1 (HTLV), bovine leukemia virus (BLV), Feline Leukemia Virus (FeLV), Gibbon Ape Leukemia Virus (GaLV), Koala Retrovirus (KRV), Reticuloendotheliosis Virus (ReEV), Wooly Monkey Sarcoma Virus (WMSV), or a biologically active mutant thereof, or any combination thereof.

157. The lipid delivery particle of claim 153 or 154, wherein the first gag polyprotein or the second gag polyprotein, or both are a human endogenous retroviral gag polyprotein.

158. The lipid delivery particle of claim 153 or 154, wherein the first gag polyprotein or the second gag polyprotein, or both comprise a sequence having at least 80%, at least 85%, at least 90%, or at least 95%, or 100% sequence identity to the amino acid sequence set forth in any one of SEQ ID NOs: 285-305.

159. The lipid delivery particle of any one of claims 153-158, wherein the first gag polyprotein or the second gag polyprotein, or both lack at least a fragment of a nucleocapsid protein, optionally lack the full length of the nucleocapsid protein.WSGR Docket No.:62697-750.601160. The lipid delivery particle of claim 159, wherein the first gag polyprotein or the second gag polyprotein, or both are fused N-terminal to a heterologous domain, optionally wherein the heterologous domain comprises a leucine zipper.

161. The lipid delivery particle of claim 160, wherein the heterologous domain further comprises a linker sequence flanking the leucine zipper on both N-terminus and C-terminus sides.

162. The lipid delivery particle of claim 161, wherein the gag / pro polyprotein further comprises the linker sequence flanking the pro polypeptide on its C-terminus.

163. The lipid delivery particle of claim 161 or 162, wherein the linker sequence comprises at least one repeat of an amino acid sequence SGGS, the sequence of any one of SEQ ID NO: 343-346 or 673, optionally two repeats thereof.

164. The lipid delivery particle of any one of claims 151-163, wherein the first gag polyprotein or the second gag polyprotein, or both lack at least fragment of a matrix protein, optionally lack the full length of the matrix protein.

165. The lipid delivery particle of claim 164, wherein the first gag polyprotein or the second gag polyprotein, or both are fused C-terminal to a pleckstrin homology (PH) domain, optionally wherein the PH domain is from a protein selected from the group consisting of human phospholipase C51, human Aktl, human Aktl with E17K substitution, human 3 -phosphoinositide-dependent protein kinase 1, human Daapl, mouse Grpl, human Grpl, human OSBP, human Btkl, human FAPP1, human CERT, human PKD, human PHLPP1, human SWAP70, and human MAPKAP1.

166. The lipid delivery particle of claim 165, wherein the PH domain comprises a sequence set forth in any one of SEQ ID NOs: 5-14 and 23-48.

167. The lipid delivery particle of any one of claims 1-166, wherein the lipid delivery particle does not comprise a pol polyprotein either standalone or in fusion.

168. The lipid delivery particle of any one of claims 1-167, wherein the lipid delivery particle does not comprise a reverse transcriptase.

169. The lipid delivery particle of any one of claims 1-168, wherein the lipid delivery particle does not comprise an integrase.

170. A method for preparing a lipid delivery particle comprising a payload, the method comprising:(a) providing a producer cell that produces the lipid delivery particle of any one of claims 1-169; and(b) collecting the lipid delivery particle from the producer cell.

171. The method of claim 170, further comprising expressing in the producer cell a nucleic acid molecule encoding the plasma membrane recruitment element.

172. The method of claim 170 or 171, wherein the producer cell comprises a nucleic acid sequence encoding the envelope protein.

173. The method of claim 172, further comprising expressing in the producer cell a nucleic acid molecule encoding the envelope protein.WSGR Docket No.:62697-750.601174. The method of any one of claims 170-173, further comprising culturing the producer cell in a medium and maintaining the producer cell under conditions sufficient to produce the lipid delivery particle.

175. The method of claim 174, further comprising harvesting the medium and purifying the lipid delivery particle.

176. The method of claim 175, wherein the purifying retains the structural integrity of the lipid delivery particle.

177. A method of introducing one or more edits in a cell, comprising contacting the cell with the lipid delivery particle of any one of claims 1-169 or the lipid delivery particle produced by the method of any one of claims 170-176.

178. A method of introducing two or more edits in a cell, comprising contacting the cell with the lipid delivery particle of any one of claims 1-169 or the lipid delivery particle produced by the method of any one of claims 170-176.

179. The method of claim 178, wherein the two or more edits in the cell are to different gene targets.

180. A composition comprising a nucleic acid molecule encoding the lipid delivery particle of any one of claims 1-169.

181. The composition of claim 180, wherein the nucleic acid molecule comprises (i) a first nucleotide sequence encoding the envelope protein; (ii) a second nucleotide sequence encoding the plasma membrane recruitment element; and (iii) a third nucleotide sequence encoding the first coiled-coil peptide; and (iv) a fourth nucleotide sequence encoding the second coiled-coil peptide.

182. A vector comprising the composition of claim 180 or 181.

183. The vector of claim 182, wherein the vector is a vector selected from a plasmid, a cosmid, a bacterial vector, a viral vector, or an artificial chromosome.

184. A cell comprising the lipid delivery particle of any one of claims 1-169 or the vector of claim 182 or 183.

185. A cell comprising the lipid delivery particle produced by the method of any one of claims 170- 176.

186. A pharmaceutical composition comprising: the lipid delivery particle of any one of claims 1-169 or the composition of claim 180 or 181; and (b) a pharmaceutically acceptable carrier, excipient, or diluent.

187. A method of treating a disease or condition in a subject in need thereof, the method comprising: administering to the subject in need thereof the lipid delivery particle of any one of claims 1-169 or the pharmaceutical composition of claim 186.

188. A kit comprising :(a) the lipid delivery particle of any one of claims 1-169 or the pharmaceutical composition of claim 186; and(b) instructions for use.

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