Methods and compositions for increased expression of integration deficient viral vectors

Abstract

The present disclosure relates generally to compositions and methods comprising recombinant integration deficient retroviral vectors where the expression of cargo(s) of interest is enhanced by the presence of elements that inhibit the silencing of viral DNA.

Description

METHODS AND COMPOSITIONS FOR INCREASED EXPRESSION OF INTEGRATION DEFICIENT VIRAL VECTORSCROSS-REFERENCE TO RELATED APPLICATION

[0001] The present application claims priority to U.S. Provisional Patent Application Serial No. 63 / 575,277, filed on April 5, 2024. The contents of the above-referenced application are herein expressly incorporated by reference in their entirety, including any drawings.BACKGROUND

[0002] With the increase in knowledge of the genome of humans and other species, intervention in the expression of genes for therapeutic or other purposes has become a common goal in human and animal medicine. Various diseases are known or postulated to be caused by deleterious mutations in specific genes. Other diseases result from over-expression, aberrant expression or silencing of particular genes. Treatment of such diseases by correcting, complementing or otherwise affecting gene expression frequently involves delivering nucleic acids into a target cell in vivo. Various approaches have been proposed for in vivo delivery.

[0003] Existing methods for delivery of nucleic acids fall into two general categories: viral vectors or non-viral vectors. One class of viral vectors that have been used for in vitro and in vivo gene transfer are retroviral vectors. Retroviruses can be broadly divided into simple and complex retroviruses. Both simple and complex retroviruses encode three major viral proteins, gag, pol and env. Complex retroviruses encode additional regulatory viral proteins that are derived from multiply spliced viral transcripts. In addition, retroviruses can be further divided into oncoretroviruses (e.g.. human T-cell leukemia virus), lentiviruses (e.g., human immunodeficiency virus), and spumaviruses (e.g.. human foamy virus). They have been shown to mediate highly efficient transgene expression in a variety of animal tissues. However, the process of vector integration into host cell chromosomes carries with it the chance of causing insertional mutagenesis. Insertional mutagenesis can be avoided by the use of gene therapy vectors that do not integrate.

[0004] Therefore, a key challenge for clinical therapies based on retroviral vectors is to achieve stable transgene expression whilst minimizing insertional mutagenesis. Retroviruses and retroviral vectors can be rendered integration-defective by mutations in the integrase coding sequence. Unfortunately, direct administration of non-integrating vectors based on human immunodeficiency virus (HIV), or feline immunodeficiency virus (FIV) has not previously resulted in significant expression in vivo. This is, in part, due to epigeneticsilencing of retroviral DNAs that occur with integration deficient vectors. This is an even larger problem with respect to vectors carrying large cargo.

[0005] The disclosure provided here provides solutions to the problems existing with previous attempts to utilize integration deficient retroviral vectors for transfer of large nucleic acid cargo and offers improved methods and compositions for the same.SUMMARY

[0006] The present disclosure relates generally to improved recombinant integration deficient retroviral vectors and systems designed to enable efficient transfer of large cargo into cells and enhanced gene expression of cargo. In particular, the present disclosure provides recombinant integration deficient retroviral vectors containing one or more elements that inhibit the epigenetic silencing of viral DNA. In another aspect, the present disclosure relates to compositions and methods of using recombinant integration deficient retroviral vectors. The foregoing summary is illustrative only and is not intended to be in any way limiting. In addition to the illustrative embodiments and features described herein, further aspects, embodiments, objects, and features of the disclosure will become fully apparent from the drawings and the detailed description and the claims.

[0007] In one aspect, the present disclosure provides a recombinant integration deficient retroviral vector comprising a nucleic acid comprising one or more virus-derived long terminal repeats (LTR); a viral packaging signal; a nucleic acid sequence encoding one or more cargos of interest; a post-transcriptional response element; and a nucleic acid sequence encoding an RNA interference molecule or molecules that targets one or more proteins involved in the epigenetic silencing of viral DNA.

[0008] In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA is part of the human silencing hub (HUSH) complex, the structural maintenance of chromosome (SMC) 5 / 6 complex, or chromatin modifiers activating transcription factor 7- interacting protein (Atf7ip) and / or its interacting partner SET domain bifurcated histone lysine methyltransferase 1 (Setdbl).

[0009] In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA is part of the HUSH complex. In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA comprises NP220, MPP8, TASOR, PPHLN1, and / or M0RC2. In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA comprises TASOR. In some embodiments, the nucleic acidsequence encoding an RNA interference molecule that targets TASOR comprises the nucleic acid sequence of SEQ ID NO: 22.

[0010] In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA is part of the SMC5 / 6 complex. In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA comprises SMC5, SMC6, NSMCE1, NSMCE2, NSMCE3, NSMCE4A. and / or SLF2.

[0011] In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA is part of the chromatin modifiers activating transcription factor 7-interacting protein (Atf7ip) and / or its interacting partner SET domain bifurcated histone lysine methyltransferase 1 (Setdbl).

[0012] In some embodiments, the RNA interference molecule is selected from a short hairpin RNA (shRNA), a small interfering RNA (siRNA), a hairpin siRNA, a microRNA (miRNA), a precursor miRNA, or an miRNA-adapted shRNA.

[0013] In some embodiments, the RNA interference molecule is positioned within the 3’ LTR.

[0014] In some embodiments, the vector comprises an RNA polymerase Ill promoter.

[0015] In some embodiments, the vector comprises an RNA polymerase II promoter. In some embodiments, the RNA polymerase II promoter is an SFFV promoter. In some embodiments, the SFFV promoter is a modified SFFV promoter. In some embodiments, the modified SFFV promoter comprises the nucleic acid sequence of SEQ ID NOs: 829-834. In some embodiments, the modified SFFV promoter comprises the nucleic acid sequence of SEQ ID NO: 831.

[0016] In some embodiments, the vector comprises an RNA poly merase II and an RNA polymerase III promoter.

[0017] In some embodiments, the vector comprises more than one RNA polymerase II and / or RNA polymerase III promoter.

[0018] In some embodiments, the one or more virus-derived LTRs are derived from murine leukemia virus (MLV), Bovine leukemia virus (BLV), Human T-lymphotropic virus 1 (HTLV-1). Human T-lymphotropic virus 2 (HTLV-2), Gibbon ape leukemia virus (GALV), Feline leukemia virus (FeLV), Porcine endogenous retroviruses (PERV), Feline Foamy Virus (FeFV), Bovine foamy virus (BFV), Mouse mammary tumor virus (MMTV), Jaagsiekte sheep retrovirus (JSRV), Mason-Pfizer monkey virus (MP MV), Avian sarcoma leukosis virus (ALV). Rous sarcoma virus (RSV), Human Endogenous Retro virus-W (HERV-W), and Human endogenous retrovirus K (HERV-K), Equine infectious anemia virus (El AV).

[0019] In some embodiments, the vector is derived from murine leukemia virus (MLV), Bovine leukemia virus (BLV). Human T-lymphotropic virus 1 (HTLV-1), Human T- lymphotropic virus 2 (HTLV-2), Gibbon ape leukemia virus (GALV), Feline leukemia virus (FeLV), Porcine endogenous retroviruses (PERV), Feline Foamy Virus (FeFV), Bovine foamy virus (BFV), Mouse mammary tumor virus (MMTV), Jaagsiekte sheep retrovirus (JSRV), Mason-Pfizer monkey virus (MP MV), Avian sarcoma leukosis virus (ALV), Rous sarcoma virus (RSV), Human Endogenous Retrovirus-W (HERV-W), and Human endogenous retrovirus K (HERV-K), Equine infectious anemia virus (ETAV).

[0020] In some embodiments, the vector is a simple retroviral vector. In some embodiments, wherein the one or more virus-derived LTRs are derived from MLV, GALV, FeLV, PERV, BFV, MMTV, JSRV. MPMV, ALV, or RSV.

[0021] In some embodiments, the vector is a complex retroviral vector. In some embodiments, the one or more virus-derived LTRs are derived from HIV-1, FIV, HIV-2, SIV, HERV-W, HERV-K, BLV, HTLV-1, HTLV-2 or EIAV.

[0022] In some embodiments, the one or more virus-derived LTRs are self-inactivating (SIN) LTRs.

[0023] In some embodiments, the vector is derived from HIV.

[0024] In some embodiments, the vector further comprises a nucleic acid sequence encoding HIV Tat and / or Rev.

[0025] In some embodiments, the post-transcriptional response element is selected from the group consisting of Woodchuck Hepatitis Virus Posttranscriptional Regulatory Element (WPRE) and Hepatitis B Virus Posttranscriptional Regulator}' Element (HP RE).

[0026] In some embodiments, the post-transcriptional response element is WPRE. In some embodiments, the WPRE comprises the sequence of SEQ ID NO: 835.

[0027] In some embodiments, the nucleic acid sequence encodes two or more cargos of interest.

[0028] In some embodiments, the nucleic acid sequence comprises coding sequences for two polypeptide cargos of interest and an IRES, protease cleavage or P2A sequence between the coding sequences.

[0029] In some embodiments, the vector further comprises a guide RNA (gRNA).

[0030] In some embodiments, the one or more cargos of interest are selected from the group consisting of gene-editing nucleases, endonuclease deficient Cas, endonuclease deficient Cas effectors, endonuclease deficient Cas enzymes, engineered DNA binding proteins, base editors, prime editors, epigenome editors, bridge editors, RNA interference(RNAi), RNA targeting systems, CRISPR DNA binding proteins. CRISPR fusion proteins, CRISPR nickases, recombinases, integrases, programmable integrases, transposons, retrotransposons, DNA polymerases, reverse transcriptases, transposases, chimeric antigen receptors (CAR), T cell receptors, HLA-independent T cell receptors, synNotch receptors, cytokine receptors, synthetic intramembrane proteolysis receptors, transcription factors, caspases, natural proteases, programmable proteases, reporter genes, and selection markers.

[0031] In some embodiments, the gene-editing nuclease is selected from the group consisting of zinc finger nucleases (ZFNs), Transcription Activator-Like Effector Nucleases (TALENs), and nucleases of the CRISPR / Cas system. In some embodiments, the geneediting nuclease is Cas9.

[0032] Also provided herein is a cell comprising the recombinant integration deficient retroviral vector according to the present disclosure.

[0033] Also provided herein is a composition comprising the recombinant integration deficient retroviral vector of the present disclosure or the cell of the present disclosure.

[0034] Also provided herein is a recombinant integration deficient retroviral particle comprising a nucleic acid comprising one or more virus-derived long terminal repeats (LTR); a viral packaging signal; a nucleic acid sequence encoding one or more cargos of interest; a post-transcriptional response element; and a nucleic acid sequence encoding an RNA interference molecule or molecules that targets one or more proteins involved in the epigenetic silencing of viral DNA.

[0035] In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA is part of the human silencing hub (HUSH) complex, the structural maintenance of chromosome (SMC) 5 / 6 complex, or chromatin modifiers activating transcription factor 7- interacting protein (Atf7ip) and / or its interacting partner SET domain bifurcated histone lysine methyltransferase 1 (Setdbl).

[0036] In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA is part of the HUSH complex. In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA comprises NP220, MPP8, TASOR, PPHLN1, and / or M0RC2. In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA comprises TASOR. In some embodiments, the RNA interference molecule that targets TASOR comprises the sequence of SEQ ID NO: 22.

[0037] In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA is part of the SMC5 / 6 complex. In some embodiments, the one or more proteinsinvolved in the epigenetic silencing of viral DNA comprises SMC5, SMC6, NSMCE1, NSMCE2, NSMCE3, NSMCE4A. and / or SLF2.

[0038] In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA is part of the chromatin modifiers activating transcription factor 7-interacting protein (Atf7ip) and / or its interacting partner SET domain bifurcated histone lysine methyltransferase 1 (Setdbl).

[0039] In some embodiments, the RNA interference molecule is selected from a short hairpin RNA (shRNA), a small interfering RNA (siRNA), a hairpin siRNA, a microRNA (miRNA), a precursor miRNA, or an miRNA-adapted shRNA.

[0040] In some embodiments, the RNA interference molecule is positioned within the 3' LTR.

[0041] In some embodiments, the vector comprises an RNA polymerase III promoter.

[0042] In some embodiments, the vector comprises an RNA polymerase II promoter. In some embodiments, the RNA polymerase II promoter is an SFFV promoter. In some embodiments, the SFFV promoter is a modified SFFV promoter. In some embodiments, the modified SFFV promoter comprises the nucleic acid sequence of SEQ ID NOs: 829-834. In some embodiments, the modified SFFV promoter comprises the nucleic acid sequence of SEQ ID NO: 831.

[0043] In some embodiments, the vector comprises an RNA polymerase II and an RNA polymerase III promoter.

[0044] In some embodiments, the vector comprises more than one RNA polymerase II and / or RNA polymerase III promoter.

[0045] In some embodiments, the one or more virus-derived LTRs are derived from murine leukemia virus (MLV), Bovine leukemia virus (BLV), Human T-lymphotropic virus 1 (HTLV-1), Human T-lymphotropic virus 2 (HTLV-2), Gibbon ape leukemia virus (GALV), Feline leukemia virus (FeLV), Porcine endogenous retroviruses (PERV), Feline Foamy Virus (FeFV), Bovine foamy virus (BFV), Mouse mammary tumor virus (MMTV), Jaagsiekte sheep retrovirus (JSRV), Mason-Pfizer monkey virus (MPMV), Avian sarcoma leukosis virus (ALV). Rous sarcoma virus (RSV), Human Endogenous Retro virus-W (HERV-W), and Human endogenous retrovirus K (HERV-K), Equine infectious anemia virus (EIAV).

[0046] In some embodiments, the vector is derived from murine leukemia virus (MLV), Bovine leukemia virus (BLV), Human T-lymphotropic virus 1 (HTLV-1), Human T- lympho tropic virus 2 (HTLV-2). Gibbon ape leukemia virus (GALV), Feline leukemia virus (FeLV), Porcine endogenous retroviruses (PERV), Feline Foamy Virus (FeFV), Bovinefoamy virus (BFV), Mouse mammary tumor virus (MMTV), Jaagsiekte sheep retrovirus (JSRV), Mason-Pfizer monkey vims (MP MV), Avian sarcoma leukosis vims (ALV), Rous sarcoma virus (RSV), Human Endogenous Retrovims-W (HERV-W), and Human endogenous retrovirus K (HERV-K), Equine infectious anemia vims (EIAV).

[0047] In some embodiments, the vector is a simple retroviral vector.

[0048] In some embodiments, the vector is a complex retroviral vector.

[0049] In some embodiments, the one or more virus-derived LTRs are derived from MLV. GALV, FeLV, PERV, BFV, MMTV, JSRV, MPMV, ALV, or RSV.

[0050] In some embodiments, the one or more virus-derived LTRs are derived from HIV-1, FIV, EIAV, HIV-2, SIV, HERV-W, HERV-K, BLV, HTLV-1, HTLV-2 or EIAV.

[0051] In some embodiments, the one or more virus-derived LTRs are self-inactivating (SIN) LTRs.

[0052] In some embodiments, the vector is derived from HIV.

[0053] In some embodiments, the vector further comprises a nucleic acid sequence encoding HIV Tat and / or Rev.

[0054] In some embodiments, the post-transcriptional response element is selected from the group consisting of Woodchuck Hepatitis Virus Posttranscriptional Regulatory Element (WPRE) and Hepatitis B Vims Posttranscriptional Regulatory' Element (HP RE).

[0055] In some embodiments, the nucleic acid sequence encodes two or more cargos of interest.

[0056] In some embodiments, the nucleic acid sequence comprises coding sequences for two polypeptide cargos of interest and an IRES or P2A sequence between the coding sequences.

[0057] In some embodiments, the recombinant integration deficient retroviral particle further comprises a guide RNA (gRNA).

[0058] In some embodiments, the one or more cargos of interest are selected from the group consisting of gene-editing nucleases, endonuclease deficient Cas, endonuclease deficient Cas effectors, endonuclease deficient Cas enzy mes, engineered DNA binding proteins, base editors, epigenome editors, prime editors. RNA interference (RNAi), RNA targeting systems, CRISPR DNA binding proteins, CRISPR fusion proteins, CRISPR nickases, recombinases, integrases, programmable integrases, transposons, retrotransposons, DNA polymerases, reverse transcriptases, transposases, chimeric antigen receptors (CAR), T cell receptors, HLA-independent T cell receptors, synNotch receptors, cytokine receptors,synthetic intramembrane proteolysis receptors, transcription factors, caspases, natural proteases, programmable proteases, reporter genes, and selection markers.

[0059] In some embodiments, the gene-editing nuclease is selected from the group consisting of zinc finger nucleases (ZFNs), Transcription Activator-Like Effector Nucleases (TALENs), and nucleases of the CRISPR / Cas system. In some embodiments, the geneediting nuclease is Cas9.

[0060] Also provided herein is a cell comprising the recombinant integration deficient retroviral particle according to the present disclosure.

[0061] Also provided herein is a composition comprising the recombinant integration deficient retroviral particle of the present disclosure or the cell of the present disclosure.

[0062] Also provided herein is a nucleic acid construct encoding a modified Gag protein comprising a Vpx interaction motif.

[0063] In some embodiments, the Vpx interaction motif is an SIV Vpx interaction motif.

[0064] In some embodiments, the Vpx interaction motif is located in the C-terminal tail of the Gag protein.

[0065] In some embodiments, the modified Gag protein is a modified HIV-1 Gag protein.

[0066] In some embodiments, the Vpx interaction motif comprises the sequence of SEQ ID NO: 839.

[0067] In some embodiments, the modified Gag protein comprises the nucleic acid sequence of SEQ ID NO: 841.

[0068] In some embodiments, the nucleic acid construct comprises the sequence of SEQ ID NO: 842.

[0069] In another aspect, the present disclosure provides a method of reducing the silencing of an integration deficient retroviral vector. The method comprises administering the recombinant integration deficient retroviral vector of the present disclosure or the recombinant integration deficient retroviral particle of the present disclosure to a cell of interest under conditions effective to reduce the silencing of the vector cargo within said cell.

[0070] In another aspect, the present disclosure provides a system comprising (i) a recombinant integration deficient retroviral vector comprising a nucleic acid comprising: one or more virus-derived long terminal repeats (LTR); a viral packaging signal; a nucleic acid sequence encoding one or more cargos of interest; a post-transcriptional response element; and (ii) one or more viral proteins capable of inhibiting the silencing of viral DNA.

[0071] In some embodiments, the one or more viral proteins is a heterologous viral protein.

[0072] In some embodiments, the one or more viral proteins capable of inhibiting the silencing of viral DNA is selected from Vpr, Vpx. a Vpr-Vpx fusion protein, VP16, ICPO, IE1, BNRF1, Hbx, pp71, and RTA. In some embodiments, the one or more viral proteins is Vpr. In some embodiments, the Vpr comprises a mutation at amino acid position 67. In some embodiments, the mutation is a substitution, deletion, or insertion. In some embodiments, the substitution is a conservative substitution. In some embodiments, the substitution is L67E.

[0073] In some embodiments, the one or more viral proteins is Vpx.

[0074] In some embodiments, the system further comprises a nucleic acid construct encoding a modified Gag protein comprising a Vpx interaction motif. In some embodiments, the Vpx interaction motif is an SIV Vpx interaction motif. In some embodiments, the Vpx interaction motif is located in the C-terminal tail of the Gag protein. In some embodiments, the modified Gag protein is a modified HIV-1 Gag protein. In some embodiments, the Vpx interaction motif comprises the sequence of SEQ ID NO: 838. In some embodiments, the modified Gag protein comprises the nucleic acid sequence of SEQ ID NO: 841.

[0075] In some embodiments, nucleic acid construct comprises the sequence of SEQ ID NO: 842.

[0076] In some embodiments, the one or more viral proteins comprises Vpr and Vpx. In some embodiments, the system further comprises a nucleic acid construct encoding a modified Gag protein comprising a Vpx interaction motif.

[0077] In some embodiments, the system further comprises a nucleic acid sequence encoding an RNA interference molecule or molecules that targets one or more proteins involved in the epigenetic silencing of viral DNA. In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA is part of the human silencing hub (HUSH) complex, the structural maintenance of chromosome (SMC) 5 / 6 complex, or the chromatin modifiers activating transcription factor 7-interacting protein ( Atf7ip) and / or its interacting partner SET domain bifurcated histone lysine methyltransferase 1 (Setdbl). In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA is part of the HUSH complex. In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA comprises NP220, MPP8, TASOR, PPHLN1, and / or MORC2. In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA comprises TASOR. In some embodiments, the RNA interference molecule that targets TASOR comprises the sequence of SEQ ID NO: 22.

[0078] In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA is part of the SMC5 / 6 complex. In some embodiments, the one or more proteinsinvolved in the epigenetic silencing of viral DNA comprises SMC5, SMC6, NSMCE1, NSMCE2, NSMCE3, NSMCE4A. and / or SLF2.

[0079] In some embodiments, the one or more proteins involved in the epigenetic silencing of viral DNA is part of the chromatin modifiers activating transcription factor 7-interacting protein (Atf7ip) and / or its interacting partner SET domain bifurcated histone lysine methyltransferase 1 (Setdbl).

[0080] In some embodiments, the RNA interference molecule is selected from a short hairpin RNA (shRNA), a small interfering RNA (siRNA), a hairpin siRNA, a microRNA (miRNA), a precursor miRNA, or an miRNA-adapted shRNA.

[0081] In some embodiments, the RNA interference molecule is positioned within the 3' LTR.

[0082] In some embodiments, the vector comprises an RNA polymerase III promoter.

[0083] In some embodiments, the vector comprises an RNA polymerase II promoter. In some embodiments, the RNA polymerase II promoter is an SFFV promoter. In some embodiments, the SFFV promoter is a modified SFFV promoter. In some embodiments, the modified SFFV promoter comprises the nucleic acid sequence of SEQ ID NOs: 829-834. In some embodiments, the modified SFFV promoter comprises the nucleic acid sequence of SEQ ID NO: 831.

[0084] In some embodiments, the vector comprises an RNA polymerase II and an RNA polymerase III promoter.

[0085] In some embodiments, the vector comprises more than one RNA polymerase II and / or RNA polymerase III promoter.

[0086] In some embodiments, the one or more virus-derived LTRs are derived from murine leukemia virus (MLV), Bovine leukemia virus (BLV), Human T-lymphotropic virus 1 (HTLV-1), Human T-lymphotropic virus 2 (HTLV-2), Gibbon ape leukemia virus (GALV), Feline leukemia virus (FeLV), Porcine endogenous retroviruses (PERV), Feline Foamy Virus (FeFV), Bovine foamy virus (BFV), Mouse mammary tumor virus (MMTV), Jaagsiekte sheep retrovirus (JSRV), Mason-Pfizer monkey virus (MPMV), Avian sarcoma leukosis virus (ALV). Rous sarcoma virus (RSV), Human Endogenous Retro virus-W (HERV-W), and Human endogenous retrovirus K (HERV-K), Equine infectious anemia virus (EIAV).

[0087] In some embodiments, the vector is derived from murine leukemia virus (MLV), Bovine leukemia virus (BLV), Human T-lymphotropic virus 1 (HTLV-1), Human T- lympho tropic virus 2 (HTLV-2). Gibbon ape leukemia virus (GALV), Feline leukemia virus (FeLV), Porcine endogenous retroviruses (PERV), Feline Foamy Virus (FeFV), Bovinefoamy virus (BFV), Mouse mammary tumor virus (MMTV), Jaagsiekte sheep retrovirus (JSRV), Mason-Pfizer monkey vims (MP MV), Avian sarcoma leukosis vims (ALV), Rous sarcoma virus (RSV), Human Endogenous Retrovims-W (HERV-W), and Human endogenous retrovirus K (HERV-K), Equine infectious anemia vims (EIAV).

[0088] In some embodiments, the vector is a simple retroviral vector.

[0089] In some embodiments, the vector is a complex retroviral vector.

[0090] In some embodiments, the one or more virus-derived LTRs are derived from MLV. GALV, FeLV, PERV, BFV, MMTV, JSRV, MPMV, ALV, or RSV.

[0091] In some embodiments, the one or more virus-derived LTRs are derived from HIV-1, FIV, EIAV, HIV-2, SIV, HERV-W, HERV-K, BLV, HTLV-1, HTLV-2 or EIAV.

[0092] In some embodiments, the one or more virus-derived LTRs are self-inactivating (SIN) LTRs.

[0093] In some embodiments, the vector is derived from HIV.

[0094] In some embodiments, the vector further comprises a nucleic acid sequence encoding HIV Tat and / or Rev.

[0095] In some embodiments, the post-transcriptional response element is selected from the group consisting of Woodchuck Hepatitis Virus Posttranscriptional Regulatory Element (WPRE) and Hepatitis B Vims Posttranscriptional Regulatory' Element (HP RE). In some embodiments, the post-transcriptional response element is WPRE. In some embodiments, the WPRE comprises the nucleic acid sequence of SEQ ID NO: 835.

[0096] In some embodiments, the nucleic acid sequence encodes two or more cargos of interest.

[0097] In some embodiments, the nucleic acid sequence comprises coding sequences for two polypeptide cargos of interest and an IRES or P2A sequence between the coding sequences.

[0098] In some embodiments, the system further comprises a guide RNA (gRNA).

[0099] In some embodiments, the one or more cargos of interest are selected from the group consisting of gene-editing nucleases, endonuclease deficient Cas, endonuclease deficient Cas effectors, endonuclease deficient Cas enzymes, engineered DNA binding proteins, base editors, epigenome editors, prime editors, bridge editors, RNA interference (RNAi), RNA targeting systems, CRISPR DNA binding proteins, CRISPR fusion proteins, CRISPR nickases, recombinases, integrases, programmable integrases, transposons, retrotransposons. DNA polymerases, reverse transcriptases, transposases. chimeric antigen receptors (CAR), T cell receptors, HLA-independent T cell receptors, synNotch receptors.cytokine receptors, synthetic intramembrane proteolysis receptors, transcription factors, caspases, natural proteases, programmable proteases, reporter genes, and selection markers.

[0100] In some embodiments, the gene-editing nuclease is selected from the group consisting of zinc finger nucleases (ZFNs), Transcription Activator-Like Effector Nucleases (TALENs), and nucleases of the CRISPR / Cas system. In some embodiments, the geneediting nuclease is Cas9.

[0101] Also provided herein is a cell comprising the system according to the present disclosure.

[0102] Also provided herein is a recombinant integration deficient retroviral particle comprising a nucleic acid comprising: (i) a recombinant integration deficient retroviral vector comprising a nucleic acid comprising: one or more virus-derived long terminal repeats (LTR); a viral packaging signal; a nucleic acid sequence encoding one or more cargos of interest; a post-transcriptional response element; and (ii) one or more viral proteins capable of inhibiting the silencing of viral DNA.

[0103] Another aspect of the present disclosure provides a method of reducing the silencing of an integration deficient retroviral vector, the method comprising administering the system of the present disclosure to a cell of interest under conditions effective to reduce the silencing of the vector cargo within said cell.

[0104] Another aspect of the present disclosure provides a method of increasing the efficiency of the delivery of one or more cargos via an integration deficient retroviral vector, the method comprising administering the system of the present disclosure to a cell of interest.BRIEF DESCRIPTION OF THE DRAWINGS

[0105] The features of the present 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. Throughout the figures, integration competent lentiviral data is used as a means of benchmarking integration deficient lentiviral data and as a control.

[0106] FIG. 1 shows integration deficient lentiviral vectors (IDLV) carrying different cargo of interest are transcriptionally silenced across different cell lines whereas integration competent lentiviral vectors (ICLV) are expressed. The only difference between IDLV and ICLV in this experiment is the IDLV are packaged with a defective viral integrase protein (Integrase D64A) while the ICLV are packaged with a wild type integrase protein.

[0107] FIG. 2 shows packaging of HIV-1 accessory protein Vpr into IDLVs enhances transgene expression in K562, Jurkat and HEK-293T cells.

[0108] FIGs. 3A-3D show packaging of HIV-1 accessory protein Vpr into IDLVs enhances transgene expression in K562, Jurkat, HEK-293T, and THP1 cells.

[0109] FIG. 4 shows packaging of wildty pe HIV-1 Vpr into IDLVs induces cellular toxicity.

[0110] FIG. 5 shows IDLVs packaged with HIV-1 Vpr-L67E mutation prevents Vpr induced cellular toxicity

[0111] FIGs. 6A-6B show IDLVs packaged with HIV-1 Vpr-L67E mutation reduce toxicity while enhancing cargo expression in K562 cells.

[0112] FIG. 7 shows efficient CRISPR epigenome editing, as measured by CRISPR interference (CRISPRi) activity, when delivered using IDLVs packaged with HIV-1 L67E across cell lines. ICLV CRISPRi activity is measured as a control.

[0113] FIG. 8 shows shRNA expression from integration deficient lentiviral vectors, when driven by RNA polymerase III based promoters (U6 promoter) is not transcriptionally silenced. ICLV shRNA knockdown data are included as a control.

[0114] FIGs. 9A-9D show knocking down NP220 / HUSH / SETDB1 or members of the SMC5 / 6 complex results in strong activation of cargo expression from IDLVs in the absence of Vpr, and further enhancement of cargo expression from IDLVs in the presence of Vpr.

[0115] FIGs. 10A-10C show knocking down HUSH / SETDB1 or members of the SMC5 / 6 complex results in strong activation of cargo expression from IDLVs in the absence of Vpr in K562, Jurkat, and HEK-293T cells.

[0116] FIGs. 11A-11B show co-packaging SIV Vpx together with HIV-1 Vpr into IDLVs results in further enhancement of cargo expression from IDLVs compared to Vpr alone.

[0117] FIGs. 12A-12B show modification of HIV-1 Gag to insert the Vpx interaction motif from SIV Gag results in successful co-packaging of Vpx. delivery into target cells, and degradation of Vpx target proteins TASOR and SAMHD1.

[0118] FIGs. 13A-13D show IDLV containing TASOR shRNA, Vpr-L67E, and Vpx enhances cargo expression to levels very’ close to that of ICLVs in K562, Jurkat, HEK-293T. and THP1 cells.

[0119] FIG. 14 shows elDLV is much more potent than protein-based VLPs for Cas9 editing.

[0120] FIG. 15 shows promoter reengineering via deletion of repressive DNA sequences that contribute to cargo silencing.

[0121] FIGs. 16A-16B show IDLV containing deletion of certain regions of the SFFV promoter, together with TASOR shRNA, Vpr-L67E and Vpx co-packaging enhances cargo expression to the same levels obtained from equal titer ICLVs.

[0122] FIGs. 17A-17B show design of an all-in-one CRISPR therapeutic vector containing Cas9 editor and gRNA, with efficient editing of the B2M locus in Jurkat cellsDETAILED DESCRIPTION OF THE DISCLOSURE

[0123] The present disclosure relates generally to improved recombinant integration deficient retroviral vectors designed to enable efficient transfer and expression of large cargo into cells. In particular, the present disclosure provides recombinant integration deficient retroviral vectors containing one or more elements that inhibit the epigenetic silencing of viral DNA. As described herein, this results in enhanced gene expression over traditional integration deficient retroviral vectors that exhibit significant silencing of cargo. Further, it is surprisingly shown that, contrary' to published literature, members of the HUSH complex can play a role in the silencing unintegrated HIV-1 retroviral DNA. Also, as shown herein, the incorporation of a L67E mutated Vpr in the retroviral particle surprisingly both enhances vector-mediated gene expression and reduces toxicity associated with wild-ty pe Vpr expression. In addition, the combination of inhibition of epigenetic silencing machinery, together with incorporation of Vpr, and Vpx using a modified HIV-1 Gag containing a SIV Vpx interaction motif result in a synergistic enhancement of cargo expression in recombinant integration deficient retroviral vectors. This effect is further enhanced by the presence of a modified SFFV promoter, which in some embodiments may be devoid of certain repressive DNA sequences. Genome editing via recombinant integration deficient retroviral delivery' of CRISPR-Cas9 cargo is 50x to 20,000x more effective per virion compared to existing Virus- Like Particles (VLPs) technologies. The increase in therapeutic efficacy will decrease the manufacturing costs relative to VLP and potentially^ reduce the toxicity associated with administering high doses of virus in vivo

[0124] The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described.

[0125] Although various features of the disclosure can be described in the context of a single embodiment, the features can also be provided separately or in any suitable combination. Conversely, although the present disclosure can be described herein in the context of separate embodiments for clarity, the present disclosure can also be implemented in a single embodiment.DEFINITIONS

[0126] Unless otherwise defined, all terms of art, notations, and other scientific terms or terminology used herein are intended to have the meanings commonly understood by those of skill in the art to which this application pertains. In some cases, terms with commonly understood meanings are defined herein for clarity and / or for ready reference, and the inclusion of such definitions herein should not necessarily be construed to represent a substantial difference over what is generally understood in the art. Many of the techniques and procedures described or referenced herein are well understood and commonly employed using conventional methodology' by those skilled in the art.

[0127] The singular form “a”, “an”, and “the” include plural references unless the context clearly dictates otherwise. For example, the term “a cell” includes one or more cells, comprising mixtures thereof. “A and / or B” is used herein to include all of the following alternatives: “A”, “B”, “A or B”, and “A and B”.

[0128] The terms “administration” and “administering”, as used herein, refer to the delivery of a retroviral vector composition or formulation by an administration route comprising, but not limited to, intranasal, transdermal. intravenous, intra-arterial, intramuscular, intranodal, intraperitoneal, subcutaneous, intramuscular, oral, intravaginal, and topical administration, or combinations thereof. The term includes, but is not limited to, administering by a medical professional and self-administering.

[0129] The term “vector” is used in accordance with its conventional meaning in the art. For example, herein, a vector can refer to a modified retrovirus as a means of transmitting nucleic acid information from one cell to another cell.

[0130] The term “retroviral vector” refers to a vector containing structural and functional genetic elements that are primarily derived from a retrovirus.

[0131] Where a range of values is provided, it is understood that each intervening value, to the tenth of the unit of the lower limit unless the context clearly dictates otherwise, between the upper and lower limit of that range and any other stated or intervening value in that stated range, is encompassed within the disclosure. The upper and low er limits of these smaller ranges may independently be included in the smaller ranges, and are also encompassed within the disclosure, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the disclosure.

[0132] Certain ranges are presented herein with numerical values being preceded by the term “about.” The term “about” is used herein to provide literal support for the exact numberthat it precedes, as well as a number that is near to or approximately the number that the term precedes. In determining whether a number is near to or approximately a specifically recited number, the near or approximating unrecited number may be a number which, in the context in which it is presented, provides the substantial equivalent of the specifically recited number. If the degree of approximation is not otherwise clear from the context, “about” means either within plus or minus 10% of the provided value, or rounded to the nearest significant figure, in all cases inclusive of the provided value. In some embodiments, the term “about” indicates the designated value ± up to 10%, up to ± 5%, or up to ± 1 %.

[0133] The term “pharmaceutically acceptable excipient” as used herein refers to any suitable substance that provides a pharmaceutically acceptable carrier, additive, or diluent for administration of a compound(s) of interest to a subject. As such, “pharmaceutically acceptable excipient” can encompass substances referred to as pharmaceutically acceptable diluents, pharmaceutically acceptable additives, and pharmaceutically acceptable carriers. As used herein, the term “pharmaceutically acceptable carrier” includes, but is not limited to, saline, solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like, compatible with pharmaceutical administration. Supplementary active compounds (e.g., antibiotics and additional therapeutic agents) can also be incorporated into the compositions.

[0134] As used herein, a “subject” or an “individual” includes animals, such as human (e.g, human individuals) and non-human animals. In some embodiments, a “subject” or “individual” is a patient under the care of a physician. Thus, the subject can be a human patient or an individual who has, is at risk of having, or is suspected of having a health condition of interest and / or one or more symptoms of the health condition. The subject can also be an individual who is diagnosed with a risk of the health condition of interest at the time of diagnosis or later. The term “non-human animals” includes all vertebrates, e.g., mammals, e.g., rodents, e.g., mice, non-human primates, and other mammals, such as e.g , sheep, dogs, cows, chickens, and non-mammals, such as amphibians, reptiles, etc.

[0135] It is understood that aspects and embodiments of the disclosure described herein include "comprising", "consisting", and "consisting essentially of aspects and embodiments. As used herein, "comprising" is synonymous with "including", "containing", or "characterized by", and is inclusive or open-ended and does not exclude additional, unrecited elements or method steps. As used herein, "consisting of excludes any elements, steps, or ingredients not specified in the claimed composition or method. As used herein, "consisting essentially of does not exclude materials or steps that do not materially affect the basic andnovel characteristics of the claimed composition or method. Any recitation herein of the term "comprising", particularly in a description of components of a composition or in a description of steps of a method, is understood to encompass those compositions and methods consisting essentially of and consisting of the recited components or steps.

[0136] All genes, gene names, and gene products disclosed herein are intended to correspond to homologs from any species for which the compositions and methods disclosed herein are applicable. Thus, the terms include, but are not limited to genes and gene products from humans and mice. It is understood that when a gene or gene product from a particular species is disclosed, this disclosure is intended to be exemplary only, and is not to be interpreted as a limitation unless the context in which it appears clearly indicates. Thus, for example, for the genes or gene products disclosed herein, which in some embodiments relate to mammalian nucleic acid and amino acid sequences, are intended to encompass homologous and / or orthologous genes and gene products from other animals including, but not limited to other mammals, fish, amphibians, reptiles, and birds. In some embodiments, the genes, nucleic acid sequences, amino acid sequences, peptides, polypeptides and proteins are human. The term “gene” is also intended to include variants thereof.

[0137] It is appreciated that certain features of the disclosure, which are, for clarity, described in the context of separate embodiments, may also be provided in combination in a single embodiment. Conversely, various features of the disclosure, which are, for brevity, described in the context of a single embodiment, may also be provided separately or in any suitable sub-combination. All combinations of the embodiments pertaining to the disclosure are specifically embraced by the present disclosure and are disclosed herein just as if each and every combination was individually and explicitly disclosed. In addition, all subcombinations of the various embodiments and elements thereof are also specifically embraced by the present disclosure and are disclosed herein just as if each and every such sub-combination was individually and explicitly disclosed herein.Retroviruses

[0138] The term “retrovirus” refers to any known retrovirus (e.g., type c retroviruses, such as Moloney murine sarcoma virus (MoMSV), Harvey murine sarcoma virus (HaMuSV), murine mammary tumor virus (MuMTV), gibbon ape leukemia virus (GaLV), feline leukemia virus (FLV), spumavirus, Friend, Murine Stem Cell Virus (MSCV) and Rous Sarcoma Virus (RSV)). “Retroviruses” also include human T cell leukemia viruses, HTLV-1 and HTLV-2, and the lentiviral family of retroviruses, such as Human ImmunodeficiencyViruses, HIV-1, HIV-2, simian immunodeficiency virus (SIV), feline immunodeficiency virus (FIV), equine immunodeficiency virus (EIV), equine infectious anemia virus (EIAV), and other classes of retroviruses. Other examples include, munne leukemia virus (MLV), mouse mammary tumour virus (MMTV), Fujinami sarcoma virus (FuSV), Moloney murine leukemia virus (Mo-MLV), FBR murine osteosarcoma virus (FBR MSV), Abelson murine leukemia virus (A-MLV), Avian myelocytomatosis virus-29 (MC29), and Avian erythroblastosis virus (AEV), Foamy virus (FMV). A detailed list of retroviruses may be found in Coffin et al. (1997) “Retroviruses”, Cold Spring Harbor Laboratory Press Eds: J M Coffin, S M Hughes, H E Varmus pp 758-763.

[0139] Retroviruses may be broadly divided into two categories: namely, “simple” and “complex.” Retroviruses may even be further divided into seven groups.

[0140] Retroviruses are RNA viruses that utilize reverse transcriptase during their replication cycle. The retroviral genomic RNA is converted into double-stranded DNA by reverse transcriptase. This double-stranded DNA form of the virus is capable of being integrated into the chromosome of the infected cell; once integrated, it is referred to as a “pro virus.” The provirus serves as a template for RNA polymerase II and directs the expression of RNA molecules which encode the structural proteins and enzymes needed to produce new viral particles.

[0141] At each end of the retroviral DNAs are structures called “long terminal repeats” or “LTRs.” The term “long terminal repeat (LTR)” refers to domains of base pairs located at the ends of retroviral DNAs which, in their natural sequence context, are direct repeats and contain U3, R and U5 regions. LTRs generally provide functions fundamental to the expression of retroviral genes (e.g., promotion, initiation, and poly adenylation of gene transcripts) and to viral replication. The LTR contains numerous regulatory signals including transcriptional control elements, polyadenylation signals and sequences needed for replication and integration of the viral genome. The viral LTR is divided into three regions called U3, R and U5. The U3 region contains the enhancer and promoter elements. The R region marks the RNA polymerase II transcription start site. The U5 region lies 3' to the R region and contains the polyadenylation sequence. The LTR composed of U3, R and U5 regions, appears at both the 5' and 3' ends of the viral genome. For the integration defective vectors of the invention, promoters that are naturally present in LTRs (and are maintained in its natural LTR sequence context) are not considered to be exogenous promoters.Simple Retroviral Vectors

[0142] Retroviruses can be divided into two groups: simple and complex retroviruses. Simple retroviruses are characterized by the presence of one coding region in addition to those encoding the common virion proteins encoded by gag, pro, pol, and env. Simple retroviruses do not encode additional proteins that directly and specifically affect viral RNA synthesis or processing. This simple pattern of RNA regulation is correlated with a simple pattern of mRNA splicing; most of the simple retroviruses make only one spliced mRNA (for env). There are no simple retroviruses that make multiply spliced mRNAs. Retroviruses which fall into this group of simple retroviruses include, for example, MLV, GALV, FeLV, PERV, BFV, MMTV, JSRV, MPMV, ALV, and RSV.Complex Retroviral Vectors

[0143] Complex retroviruses, however, have multiple splice donors in the genome, which gives rise to complex patterns of mRNA and a greater variety of gene products (up to six in addition to the gag. pro, pol, and env proteins in HIV and SIV), at least one of which has a virus-specific tram -activating function. Retroviruses that fall into this group of complex retroviruses include, for example, HIV-1, FIV, EIAV, HIV-2, SIV, HERV-W, HERV-K, BLV, HTLV-1, HTLV-2.Integration Deficient Retroviral Vectors

[0144] Integration deficient / defective retroviral vectors (IDRVs) have been produced by introducing combinations of mutations that disable the integrase protein itself or to alter the integrase recognition sequences (att) in the viral LTR (see e.g, Yanez-Munoz et al. (2006) Nat Med 12(3):348-353; Nightingale et al. (2006) Mol Ther 13(6): 1121-1 132). Studies have shown that IDRVs can mediate stable transduction in non-dividing cells and allow for measurable levels of transgene expression (see e.g., Yanez-Munoz et al. (2006) Nat Med 12(3):348-353; Nightingale et al. (2006) Mol Ther 13(6): 1121-1132); Suwanmannee et al. (2014) Mol Ther 22(3):567-74). This reduces the likelihood of insertional mutagenesis associated with integrating vectors.COMPOSITIONS OF THE DISCLOSURE

[0145] Integration-defective retroviral vectors (IDRVs) efficiently ferry large genetic cargos to dividing and non-dividing cells in vitro and in vivo. The IDRV genome is stable in non-dividing or slowly dividing cells. In dividing cells, the presence of IDRV genomes lacking origin of replication is transient. This property could in theory be employed to transiently express potentially cellular-and genotoxic proteins whose biological functions arerequired for a short time. Although IDRVs are promising, transcriptional silencing of IDRVs can significantly reduce their efficacy and their usage. Disclosed herein is the development of IDRVs that exhibit improved gene expression as well as a methodology for inhibiting the epigenetic silencing mechanism in IDRV transduced cells. As used herein, an integrationdefective lentiviral vector (IDLV) is a sub-group of IDRVs.

[0146] As described in greater detail below, one aspect of the present disclosure relates to a recombinant integration deficient retroviral vector that includes one or more virus-derived long terminal repeats (LTR), a viral packaging signal, a nucleic acid sequence encoding a cargo or cargos of interest, a post-transcriptional regulatory element and a nucleic acid sequence encoding an RNA interference molecule or molecules that target one or more proteins involved in the epigenetic silencing of viral DNA. In some embodiments, the integration deficient retroviral vector also includes an exogenous RNA polymerase II and / or III driven promoter or promoters. In some embodiments, the integration deficient retroviral vector includes a modified SFFV promoter. In some embodiments, the integration deficient retroviral vector does not comprise an exogenous promoter. In some embodiments, the integration deficient retroviral vector does not comprise a promoter between the end of the 5?LTR and the beginning of the cargo or gene of interest to be expressed.

[0147] Other aspects of the present disclosure, as described in more detail below, relate to nucleic acid constructs encoding mutant Vpr (e.g., Vpr L67E), SIV Vpx, Vpr-Vpx fusion proteins, and modified HIV-1 Gag proteins capable of recruiting both Vpr and Vpx into virions.

[0148] Also provided are cells comprising the recombinant integration deficient retroviral vector, recombinant integration deficient retroviral particles, and compositions and systems thereof.Integration Deficient Retroviral Vector Nucleic Acid Constructs

[0149] As described in greater detail below, one aspect of the present disclosure relates to a recombinant integration deficient retroviral vector that includes one or more virus-derived long terminal repeats (LTR). a viral packaging signal, a nucleic acid sequence encoding one or more cargos of interest, a post-transcriptional regulatory element, and a nucleic acid sequence encoding an RNA interference molecule or molecules that target one or more proteins involved in the epigenetic silencing of viral DNA. In some embodiments, the integration deficient retroviral vector also includes an exogenous RNA polymerase II and / or III driven promoter or promoters. In some embodiments, the integration deficient retroviralvector includes an RNA polymerase II promoter. In some embodiments, the RNA polymerase II promoter is a modified SFFV promoter. In some embodiments, the integration deficient retroviral vector includes an RNA polymerase III promoter. In some embodiments, the integration deficient retroviral vector includes an RNA polymerase II and an RNA polymerase III promoter. In some embodiments, a cargo of interest is under the control of an RNA polymerase II promoter, and the nucleic acid sequence encoding an RNA interference molecule or molecules that target one or more proteins involved in the epigenetic silencing of viral DNA is under the control of an RNA polymerase III promoter. In some embodiments, the integration deficient retroviral vector includes an RNA polymerase II and more than one RNA polymerase III promoter. For example, in some embodiments, a CRISPR cargo of interest is under the control of an RNA polymerase II promoter, an sgRNA is under the control of an RNA polymerase III promoter, and the nucleic acid sequence encoding an RNA interference molecule or molecules that target one or more proteins involved in the epigenetic silencing of viral DNA is under the control of an RNA polymerase III promoter. In some embodiments, the integration deficient retroviral vector does not comprise an exogenous promoter. For example, although the 5’ LTRs of some vectors may have an endogenous promoter, these vectors do not comprise an exogenous promoter. In some embodiments, the integration deficient retroviral vector does not comprise a promoter between the end of the 5’ LTR and the beginning of the cargo or gene of interest to be expressed.

[0150] The terms "nucleic acid molecule" and "polynucleotide" are used interchangeably herein, and refer to both RNA and DNA molecules, including nucleic acid molecules comprising cDNA, genomic DNA, synthetic DNA, and DNA or RNA molecules containing nucleic acid analogs. A nucleic acid molecule can be double-stranded or single-stranded (e g., a sense strand or an antisense strand). A nucleic acid molecule may contain unconventional or modified nucleotides. The terms “polynucleotide sequence7’ and "nucleic acid sequence” as used herein interchangeably refer to the sequence of a polynucleotide molecule. The nomenclature for nucleotide bases as set forth in 37 CFR §1.822 is used herein.

[0151] Nucleic acid molecules of the present disclosure can be of any length, including for example, between about 1.5 Kb and about 20 Kb, between about 2 Kb and about 15 Kb, between about 5 Kb and about 10 Kb, between about 7 Kb and about 9 Kb, or between about 8 Kb and about 9 Kb

[0152] The retroviral vector can be based on any suitable retrovirus which is able to deliver genetic information to eukaryotic cells. For example, the retroviral vector may be an alpharetroviral vector, a gammaretroviral vector, a lentiviral vector or a spumaretroviralvector. In some embodiments, the retroviral vector is derived from a simple retrovirus. Simple retroviruses are described in more detail above and include, without limitation, MLV, GALV, FeLV, PERV, BFV, MMTV, JSRV, MPMV, ALV, and RSV. In some embodiments, the retroviral vector of the present disclosure is derived from MLV. In some embodiments, the retroviral vector is derived from a complex retrovirus. Complex retroviruses are described in more detail above and include, without limitation, HIV-1, FIV. EIAV, HIV -2, SIV, HERV-W, HERV-K, BLV, HTLV-1, and HTLV-2. In some embodiments, the retroviral vector of the present disclosure is derived from Bovine leukemia virus (BLV), Human T- lymphotropic vims 1 (HTLV-1), Human T-lymphotropic vims 2 (HTLV-2), Gibbon ape leukemia virus (GALV), Feline leukemia vims (FeLV), Porcine endogenous retrovimses (PERV), Feline Foamy Virus (FeFV), Bovine foamy virus (BFV), Mouse mammary tumor virus (MMTV), Jaagsiekte sheep retrovirus (JSRV), Mason-Pfizer monkey vims (MPMV), Avian sarcoma leukosis virus (ALV), Rous sarcoma vims (RSV), Human Endogenous Retrovims-W (HERV-W), Human endogenous retrovirus K (HERV-K), or Equine Infectious Anemia Virus (EIAV). In some embodiments, the retroviral vector of the present disclosure is derived from HIV.

[0153] A nucleic acid can be made by any technique known to one of ordinary skill in the art. Non-limiting examples of synthetic nucleic acid, particularly a synthetic oligonucleotide, include a nucleic acid made by in vitro chemical synthesis using phosphotriester, phosphite or phosphoramidite chemistry and solid phase techniques such as described in EP 266,032, incorporated herein by reference, or via deoxynucleoside H-phosphonate intermediates as described by Froehler et al., 1986, and U.S. Pat. No. 5,705,629, each incorporated herein by reference. A non-limiting example of enzy matically produced nucleic acid include one produced by enzy mes in amplification reactions such as PCR™ (see for example, U.S. Pat. No. 4,683,202 and U.S. Pat. No. 4,682,195, each incorporated herein by reference), or the synthesis of oligonucleotides described in U.S. Pat. No. 5,645,897, incorporated herein by reference. A non-limiting example of a biologically produced nucleic acid includes recombinant nucleic acid production in living cells (see for example, Sambrook et al. 1989, incorporated herein by reference).

[0154] A nucleic acid can be purified on polyacrylamide gels, cesium chloride centrifugation gradients, or by any other means know n to one of ordinary' skill in the art (see for example, Sambrook et al. 1989, incorporated herein by reference).

[0155] The nucleic acid(s) of the present disclosure, regardless of the length of the sequence itself, can be combined with other nucleic acid sequences, including but not limitedto, enhancers, polyadenylation signals, restriction enzy me sites, multiple cloning sites, coding segments, and the like, to create one or more nucleic acid construct(s). The overall length may vary considerably between nucleic acid constructs. Thus, a nucleic acid segment of almost any length can be employed, with the total length preferably being limited by the ease of preparation or use in the intended recombinant nucleic acid protocol.

[0156] Vectors of the present invention are retroviral based as described above and in other parts of the specification. The nucleic acid molecules of the invention encode one or more cargos of interest.

[0157] The term “expression vector” refers to any type of genetic construct comprising a nucleic acid coding for an RNA capable of being transcribed. In some cases, RNA molecules are then translated into a protein, polypeptide, or peptide. In other cases, these sequences are not translated, for example, in the production of guide RNAs, antisense molecules or ribozymes. Expression vectors can contain a variety of “control sequences,” which refer to nucleic acid sequences necessary for the transcription and possibly translation of an operably linked coding sequence in a particular host cell. In addition to control sequences that govern transcription and translation, expression vectors may contain nucleic acid sequences that serve other functions as well and are described below.RNA Interference and Epigenetic Silencing

[0158] As described herein, the integration deficient retroviral vectors of the present disclosure include a nucleic acid sequence encoding an RNA interference molecule or molecules that target one or more proteins involved in the epigenetic silencing of viral DNA.

[0159] As used herein, the term “RNA interference” or “RNAi” can refer to a phenomenon in which the introduction of double-stranded RNA (dsRNA) into a diverse range of organisms and cell types causes degradation of the complementary mRNA. In the cell, long dsRNAs are cleaved into short 21-25 nucleotide small interfering RNAs, or siRNAs, by a ribonuclease known as Dicer. The siRNAs subsequently assemble with protein components into an RNA-induced silencing complex (RISC), unwinding in the process. Activated RISC then binds to complementary transcript by base pairing interactions between the siRNA antisense strand and the mRNA. The bound mRNA is cleaved and sequence specific degradation of mRNA results in gene silencing. See, for example, U.S. Pat. No. 6,506,559.

[0160] The term “siRNA” as used herein can refer to small interfering RNA, also known as short interfering RNA or silencing RNA. siRNAs can be, for example, 18 to 30, 20 to 25, 21 to 23 or 21 nucleotide-long double-stranded RNA molecules. An “shRNA” as used herein isa short hairpin RNA, which is a sequence of RNA that makes a tight hairpin turn that can also be used to silence gene expression via RNA interference. shRNA can by operably linked to the U6 promoter for expression. The shRNA hairpin structure is cleaved by the cellular machinery into siRNA. shRNA disclosed herein can comprise a sequence complementary to at least 13 nucleotides, at least 14 nucleotides, at least 15 nucleotides, at least 16 nucleotides, at least 17 nucleotides, at least 18 nucleotides, at least 19 nucleotides, at least 20 nucleotides, at least 21 nucleotides, at least 22 nucleotides, or 23 nucleotides of the mRNA a target protein.

[0161] MicroRNAs (“miRNAs”) are short, non-coding, single-stranded RNA molecules. miRNAs of the present disclosure may be naturally-occurring or synthetic (e.g., artificial). miRNAs usually induce gene silencing by binding to target sites found within the 3' UTR (untranslated region) of a targeted mRNA. This interaction prevents protein production by suppressing protein synthesis and / or by initiating mRNA degradation. Most target sites on the mRNA have only partial base complementarity7with their corresponding miRNA, thus, individual miRNAs may target 100 different mRNAs. or more. Further, individual mRNAs may contain multiple binding sites for different miRNAs. resulting in a complex regulatory' network. In some embodiments, a miRNA is 10 to 50 nucleotides in length. For example, a miRNA may be 10 to 40, 10 to 30, 10 to 20, 20 to 50, 20 to 40 or 20 to 30 nucleotides in length. In some embodiments, a miRNA is 10, 11, 12. 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34. 35. 36. 37. 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49 or 50 nucleotides in length. In some embodiments, a miRNA is 22 nucleotides in length. A "microRNA precursor" (or "pre-miRNA") refers to a nucleic acid having a stemloop structure with a microRNA sequence incorporated therein. A "mature microRNA" (or "mature miRNA") includes a microRNA that has been cleaved from a microRNA precursor (a "pre-miRNA"), or that has been synthesized (e.g., synthesized in a laboratory by cell-free synthesis), and has a length of from about 19 nucleotides to about 27 nucleotides, e.g., a mature microRNA can have a length of 19 nt, 20 nt, 21 nt, 22 nt, 23 nt, 24 nt, 25 nt, 26 nt. or 27 nt. A mature microRNA can bind to a target mRNA and inhibit translation of the target mRNA.

[0162] A microRNA-adapted shRNA consists of a shRNA stem structure with microRNA- like mismatches surrounded by the loop and flanking sequence of an endogenous microRNA. microRNA-adapted shRNAs are transcribed from RNA Polymerase 11 (Pol 11) promoters, cleaved by the endogenous RNase III Drosha enzyme in the nucleus, and then exported to the cytoplasm where they are processed by Dicer and loaded into the RISC complex.

[0163] In the compositions described herein, the nucleic acid sequence encoding the RNA interference molecule can be positioned at any location within the vector that allows for expression of the RNA interference molecule. In some embodiments, the nucleic acid sequence encoding the RNA interference molecule is positioned within the 3’ LTR of the vector. In some embodiments, the nucleic acid sequence encoding the RNA interference molecule is under the control of an RNA polymerase III promoter or an RNA polymerase II promoter.

[0164] As described above, the RNA interference molecule or molecules of the disclosure target one or more proteins involved in the epigenetic silencing of viral DNA. As used herein, epigenetic silencing can refer to the silencing or downregulation of a gene. This can involve epigenetic changes such as a modification in the gene caused by an epigenetic mechanism, such as a change in methylation status or histone acetylation for example. Frequently, the epigenetic change will result in an alteration in the levels of expression of the gene which may be detected (at the RNA or protein level as appropriate) as an indication of the epigenetic change.

[0165] Viral epigenetic silencing is mediated by the enzymatic placement, and subsequent reading, of DNA methylation marks (addition of a methyl group to position 5 of the cytosine pyrimidine [5MeCpG]) and repressive nucleosomal histone modifications. Various mechanisms are known in the art and are described in, for example, Tsai and Cullen, “Epigenetic and Epitranscriptomic Regulation of Viral Replication / ’ Nature Reviews Microbiology 18 559-570 (2020). These mechanisms can involve, without limitation, PML- NB-mediated epigenetic repression involving SplOO, ATRX, and Daxx, NP220, the human silencing hub complex (HUSH), Atf7ip, SETDB1, SMC5 / 6 complex, and IFH6-mediated repression.

[0166] In some embodiments, the RNA interference molecule is an shRNA. In some embodiments, the RNA interference molecule is an shRNA that targets NP220. In some embodiments, the RNA interference molecule is positioned in the 3’ LTR under the control of an RNA polymerase III promoter and is an shRNA that targets NP220. In some embodiments, the RNA interference molecule is an shRNA that targets TASOR. In some embodiments, the RNA interference molecule is positioned in the 3’ LTR under the control of an RNA polymerase III promoter and is an shRNA that targets TASOR. In some embodiments, the RNA interference molecule is an shRNA that targets MPP8. In some embodiments, the RNA interference molecule is positioned in the 3’ LTR under the control of an RNA polymerase III promoter and is an shRNA that targets MPP8. In someembodiments, the RNA interference molecule is an shRNA that targets PPHLN1. In some embodiments, the RNA interference molecule is positioned in the 3' LTR under the control of an RNA polymerase III promoter and is an shRNA that targets PPHLN1. In some embodiments, the RNA interference molecule is an shRNA that targets SETDB1. In some embodiments, the RNA interference molecule is positioned in the 3’ LTR under the control of an RNA polymerase III promoter and is an shRNA that targets SETDB1. In some embodiments, the RNA interference molecule is an shRNA that targets M0RC2. In some embodiments, the RNA interference molecule is positioned in the 3’ LTR under the control of an RNA polymerase III promoter and is an shRNA that targets M0RC2.HUSH complex

[0167] In some embodiments, the RNA interference molecule in the retroviral vector of the present disclosure targets one or more proteins from the HUSH complex.

[0168] The Human Silencing Hub (HUSH) complex has been shown to be involved in the epigenetic silencing of unintegrated MLV retroviral DNA, but not unintegrated HIV-1 retroviral DNA (Zhu et al., "NP220 Mediates Silencing of Unintegrated Retroviral DNA,” Nature 564(7735):278-282 (2018); Seczynska et al.. “The Sound of Silence: Mechanisms and Implications of HUSH Complex Function,” Trends in Genetics 39(4):251-267 (2023)). However, given the known contribution of the HUSH complex to silencing of integrated retroviruses as well as unintegrated MLV retroviral DNA, the present disclosure sought to determine any role the HUSH complex may play in silencing of unintegrated HIV-1 DNA. As described herein for the first time, HUSH does play a role in epigenetic silencing of unintegrated HIV-1 DNA, and therefore is a potential target for RNA interference in the vectors described herein.

[0169] The HUSH complex is a heterotrimer, composed of transcription activation suppressor (TASOR), M-phase phosphoprotein 8 (MPP8), and periphilin (PPHLN1), and is conserved from fish to mammals. To silence newly integrated retroviruses, HUSH recruits two effectors: MORC2, an ATP-dependent chromatin remodeler which compacts chromatin, and SETDB1, a histone methyltransferase which deposits histone 3 lysine 9 trimethylation (H3K9me3) at the target loci. By silencing a subset of HIV integrations, HUSH contributes to the establishment and maintenance of latent HIV (Tchasovnikarova et al. Epigenetic silencing by the HUSH complex mediates position-effect variegation in human cells. Science. 2015; 348: 1481-1485; Chougui G. Margottin-Goguet F. HUSH, a link between intrinsic immunity and HIV latency. Front. Microbiol. 2019; 10: 224; Timms et al., Position-effect Variegation Revisited: HUSHing up Heterochromatin in Human Cells,” BioEssays 2016 38:333-343),which is transcriptionally silenced but replication-competent, and therefore has the potential to reactivate.

[0170] The HUSH complex of TASOR, MPP8, and PPHLN1 localizes to its target loci where it recruits two effectors: SETDB1 promotes H3K9me3 deposition while M0RC2 is an ATP-dependent chromatin remodeler essential for transcriptional repression (Seczynska et al., “The Sound of Silence: Mechanisms and Implications of HUSH Complex Function,’" Trends in Genetics 39(4):251-267 (2023). An interaction between MPP8 and SETDB1 regulates both H3K9me3 ‘reading’ and ‘writing’ (Tchasovnikarova et al., Epigenetic silencing by the HUSH complex mediates position-effect variegation in human cells.Science. 2015; 348: 1481-1485). The MPP8 chromodomain binds both H3K9me3 (Kokura K. et al. Methyl-H3K9-binding protein MPP8 mediates E-cadherin gene silencing and promotes tumour cell motility and invasion. EMBO J. 2010; 29: 3673-3687) and methylated activating transcription factor 7 interacting protein (ATF7IP) (Tsusaka T. et al. Tri-methylation of ATF7IP by G9a / GLP recruits the chromodomain protein. EpigeneticsChromatin. 2018; 11: 56), the nuclear chaperone of the SETDB1 methyltransferase (Timms R.T. et al. ATF71P-mediated stabilization of the histone methyltransferase SETDB1 is essential for heterochromatin formation by the HUSH complex. Cell Rep. 2016; 17: 653- 659). By recognizing existing H3K9me3, MPP8 recruits ATF7IP-SETDB1 to the locus to promote further H3K9me3 deposition. However, this coupling between the MPP8 ‘read’ and SETDB1 ‘write’ functions is insufficient for repression, as TASOR, periphilin. and MPP8 protein domains with functions other than H3K9me3 binding are all essential for HUSH- mediated silencing.

[0171] An alternative means to recruit HUSH is through the sequence-specific DNA- binding protein nuclear protein 220 (NP220) as illustrated for unintegrated murine leukemia virus (MLV) (Zhu et al., “NP220 Mediates Silencing of Unintegrated Retroviral DNA,” Nature 564(7735):278-282 (2018)). This is known as the noncanonical HUSH silencing pathway. Noncanonical HUSH-mediated silencing of unintegrated MLV differs mechanistically from silencing of other targets, as it requires both the DNA-binding activity of NP220 as well as a functional MPP8 chromodomain (Zhu et al., “NP220 Mediates Silencing of Unintegrated Retroviral DNA,” Nature 564(7735):278-282 (2018)).

[0172] In some embodiments, the RNA interference molecule targets one or more proteins of the HUSH complex including NP220, MPP8, TASOR, PPHLN1, and / or MORC2.

[0173] Various methods are known in the literature for RNA interference and library construction and can be applied to the sequences described herein.

[0174] In some embodiments, the RNA interference molecule targets NP220. In some embodiments, the RNA interference molecule targets a region within NP220 (RefSeqNM_0012526I2.2) having the nucleotide sequence shown as SEQ ID NO: 1 below:GGAGGCGGTAGCGTTTTCGGCGTCGAGACTGGAGGCTGAGTGCTAAACTGTGTGGGGCGCGGATGGGATCCAGCTGTTAGTCGGTATTACAGCGTGAAATAAATTCCC AGTATTTTTCAAGATTATGTAAAAGGAGTTGTAATTACTGCTCAGCACCTAGGACAGCACCCAGTGCAACACAATGTACATGAACTCTGGCTTGAGGTTTGGGGCTTGTG TTCTGCATTCAGGAAAGAAAAGAAGAGAAAGTTTTTGGGAAATTTAAATCTTCGT AAGCCCTTACTCTAAAAGTATATGAAATGCAGAATGGTTTCATGGCTTTATTTCAAAATGTTTTACACTGGCTGTAGAACCCACTTCATGAAGTAGTTTGAGTTAAAATGATCTAATATTTGTGTTTTAACTTTCAGCTTTGTGTTATTCTTGGAAAATTTCGCAC CACTTGTGAATTCCTTGAACCTGGGCATTGCAAACCCACTTCTGTTGGGCCCATCTCCTTTGCACTTTGCTCAGATTAAGACTCAGTTGGCGCTTCAGCAGCTGAATGCC GTTGCCTCACATGGTTCAACACCACCTTATACTTTATTAAATCAGGCTTTCTTGAAAATAGCCATGTCGAGACCCAGGTTTAATCCTCGAGGAGACTTTCCACTTCAAAGG CCACGAGCACCTAACCCTTCTGGGATGAGGCCTCCAGGACCATTTATGAGGCCTG GATCTATGGGTCTCCCAAGATTTTACCCAGCAGGGAGAGCACGTGGAATTCCACACAGATTTGCTGGCCATGAATCTTATCAGAACATGGGGCCACAGAGAATGAATG TTCAGGTAACTCAACACAGAACTGATCCAAGATTGACCAAAGAAAAACTGGATTTTCATGAAGCACAACAGAAGAAGGGGAAGCCTCATGGTAGCCGGTGGGATGATG AGCCTCATATATCTGCATCAGTGGCAGTGAAACAGAGTTCTGTAACACAGGTTACAGAGCAGAGTCCCAAAGTACAGAGCCGCTATACAAAAGAGAGTGCCTCAAGTAT CTTAGCAAGTTTTGGATTATCTAATGAAGACCTAGAAGAACTTAGTCGCTATCCTGATGAACAACTAACTCCTGAAAATATGCCATTAATTTTGAGGGATATAAGAATGC GAAAAATGGGGCGCCGATTACCTAATTTACCTTCTCAGAGCAGAAATAAAGAAACACTTGGTAGTGAAGCAGTTTCAAGTAATGTGATCGATTATGGGCATGCAAGCAAATATGGCTACACAGAAGATCCACTTGAAGTACGTATTTATGATCCTGAAATTCCA ACTGATGAGGTCGAGAATGAATTTCAGTCACAGCAGAACATTTCTGCATCTGTTCCCAATCCAAATGTGATATGTAATTCTATGTTTCCTGTTGAAGACGTATTTCGCCAAATGGACTTCCCCGGTGAGTCCTCCAATAATCGGTCCTTTTTCTCAGTTGAGAGTG GAAC C AAGATGTC AGGCTTAC AC ATTTC AGGAGGAC AGTC AGTC CTTGAACC C ATAAAATCCGTCAACCAATCCATTAACCAAACAGTTAGCCAGACAATGAGTCAAT CTCTGATTCCTCCATCTATGAACCAGCAACCTTTTTCGTCGGAATTAATTTCATCTGTAAGCCAGCAAGAGCGGATCCCACATGAACCTGTGATTAATTCATCTAACGTACATGTTGGATCAAGAGGAAGTAAAAAGAATTACCAGTCACAGGCTGACATTCCCATTCGGTCTCCCTTTGGTATTGTGAAAGCATCCTGGCTACCAAAGTTTTCACATGCTGATGCCCAGAAGATGAAGAGACTTCCAACTCCTTCTATGATGAATGATTATTATGCAGCATCTCCAAGAATATTTCCACATTTGTGTTCTCTGTGTAACGTAGAATGTAGTCATTTGAAGGATTGGATTCAGCATCAAAATACATCTACTCATATTGAGAGCTGTCGACAGTTACGTCAACAGTATCCTGATTGGAATCCTGAGATCCTCCCATCGAGAAGAAATGAGGGCAATAGAAAAGAAAATGAAACTCCACGAAGACGTTCTCATTCCCCCAGTCCTAGGCGTTCTAGAAGATCAAGCTCAAGTCACAGATTCCGTCGGTCTCGAAGCCCAATGCATTACATGTATAGGCCGAGAAGTCGAAGTCCAAGAATTTGCCATCGTTTCATTTCTAGATACAGATCCAGATCCAGATCCCGTTCACCATATCGAATTAGAAATCCATTTAGAGGTAGTCCAAAATGCTTTCGATCAGTTAGCCCTGAGAGGATGTCAAGGAGATCAGTGAGATCATCAGATAGAAAAAAAGCATTAGAAGATGTAGTACAACGATCTGGGCATGGGACAGAATTTAATAAACAGAAGCATCTTGAAGCTGCTGATAAGGGACATTCACCAGCACAAAAGCCTAAAACTAGCAGTGGAACAAAACCATCAGTTAAACCTACAAGCGCTACAAAGAGTGATTCAAATCTAGGAGGACATTCTATTCGTTGTAAATCAAAGAATCTTGAAGATGACACTTTGTCAGAATGTAAACAGGTGTCTGATAAAGCTGTTTCTCTCCAGCGAAAGCTTCGGAAAGAACAGTCATTGCATTATGGTTCGGTTCTTCTTATAACTGAATTACCAGAGGATGGTTGTACTGAAGAAGATGTGAGAAAATTATTTCAACCATTTGGGAAAGTGAATGATGTCCTAATTGTTCCATATAGAAAAGAGGCTTACCTAGAAATGGAATTTAAAGAGGCAATTACTGCAATTATGAAGTACATTGAAACAACACCTCTTACGATAAAAGGAAAAAGTGTGAAAATATGTGTTCCAGGAAAGAAAAAAGCACAGAACAAAGAGGTGAAGAAAAAGACTTTAGAGTCAAAGAAAGTATCTGCATCTACCTTAAAAAGAGATGCAGATGCTTCAAAAGCTGTTGAAATTGTTACTTCAACTTCTGCTGCCAAAACTGGACAAGCCAAGGCATCTGTAGCCAAAGTAAACAAATCTACAGGGAAATCAGCAAGTTCTGTAAAATCTGTGGTAACGGTAGCTGTTAAAGGTAATAAAGCTTCAATCAAAACAGCAAAATCTGGTGGAAAGAAGTCTCTAGAAGCCAAAAAGACTGGGAATGTCAAAAACAAAGACTCTAACAAACCTGTGACTATACCAGAAAACTCTGAAATAAAGACCAGTATTGAAGTCAAAGCCACTGAAAACTGTGCTAAAGAAGCTATTTCTGATGCTGCTTTGGAGGCCACAGAGAATGAACCACTTAACAAGGAAACAGAAGAAATGTGTGTGATGCTTGTCTCTAATTTGCCTAATAAAGGATATTCTGTAGAAGAAGTTTATGACTTAGCAAAACCATTTGGTGGTTTAAAGGATATCTTGATTTTATCATCTCATAAAAAGGCATATATAGAAATAAATAGAAAAGCTGCTGAGTCTATGGTAAAATTTTATACCTGCTTCCCAGTATTGATGGATGGAAATCAACTCTCAATAAGTATGGCTCCTGAAAACATGAATATAAAAGATGAGGAAGCTATATTTATAACCTTGGTAAAAGAAAATGACCCAGAGGCAAACATAGATACAATTTATGATCGATTTGTACATCTTGATAATTTACCGGAAGATGGACTTCAGTGTGTACTTTGTGTTGGACTTCAGTTTGGAAAAGTGGATCACCATGTATTCATAAGTAATAGAAACAAGGCAATTCTTCAGTTAGATAGTCCTGAATCTGCTCAGTCAATGTATAGCTTTCTGAAACAAAATCCACAAAATATTGGTGACCATATGTTGACCTGCTCATTATCTCCAAAGATAGACTTACCAGAGGTGCAAATTGAGCATGACCCAGAATTAGAAAAAGAAAGCCCTGGCTTGAAAAACAGTCCAATTGATGAAAGTGAGGTGCAAACAGCAACTGATAGTCCCTCTGTTAAACCTAATGAGCTTGAAGAAGAAAGTACTCCCAGCATTCAAACAGAAACTTTGGTACAGCAGGAAGAGCCTTGTGAGGAAGAAGCTGAAAAAGCAACATGTGATTCTGACTTTGCTGTTGAAACTTTGGAGCTTGAAACTCAAGGAGAGGAGGTCAAAGAAGAAATTCCTCTTGTAGCATCCGCTTCAGTCAGTATTGAACAATTCACTGAAAATGCCGAGGAGTGTGCTTTAAATCAGCAGATGTTTAACAGTGACTTGGAGAAGAAAGGGGCAGAAATTATTAACCCTAAAACAGCATTGTTACCATCTGACAGTGTGTTTGCAGAAGAAAGGAACCTCAAAGGAATTCTAGAAGAATCTCCATCTGAAGCAGAAGATTTCATTTCTGGAATTACACAGACTATGGTAGAAGCTGTAGCTGAAGTAGAAAAAAATGAAACTGTTTCGGAAATATTGCCATCAACTTGTATTGTGACGTTAGTACCAGGAATTCCCACTGGGGATGAGAAGACAGTGGACAAAAAGAATATTTCTGAAAAAAAAGGTAACATGGATGAAAAGGAGGAGAAGGAATTTAATACTAAGGAAACCAGAATGGATCTTCAAATAGGAACAGAGAAGGCTGAAAAGAATGAAGGTAGGATGGATGCAGAAAAGGTGGAAAAGATGGCAGCAATGAAAGAAAAGCCTGCAGAAAACACTTTATTCAAGGCATACCCAAATAAAGGAGTGGGTCAGGCTAATAAGCCTGATGAAACTAGTAAAACTAGTATTCTGGCTGTATCAGATGTATCTAGCAGTAAACCAAGCATCAAGGCTGTTATAGTCTCTTCTCCTAAGGCAAAAGCTACAGTTTCAAAAACTGAAAATCAGAAAAGTTTTCCAAAATCTGTGCCCAGAGATCAAATAAATGCTGAAAAGAAACTTTCAGCCAAGGAATTTGGTCTGCTTAAACCCACAAGTGCCAGGTCAGGCTTGGCAGAAAGCAGCAGTAAATTCAAACCTACTCAGAGCAGTCTTACCAGAGGAGGCAGTGGAAGGATCTCAGCCCTGCAAGGCAAGCTTTCTAAACTGGATTACAGAGATATAACAAAACAATCTCAGGAAACAGAGGCTAGACCTTCCATCATGAAACGGGATGACAGCAACAATAAGACTTTGGCTGAGCAAAACACTAAGAATCCTAAAAGCACTACTGGTAGAAGTTCCAAATCTAAAGAGGAGCCATTATTTCCATTTAATTTGGATGAATTTGTTACTGTGGATGAGGTTATAGAAGAAGTGAATCCTTCTCAGGCCAAGCAGAATCCACTAAAGGGAAAAAGGAAAGAAACTCTCAAAAATGTTCCTTTCTCTGAACTTAACTTAAAGAAGAAAAAGGGGAAAACTTCCACTCCTCGTGGTGTTGAGGGAGAACTATCTTTTGTGACATTGGATGAGATTGGGGAAGAGGAAGATGCAGCTGCACATCTAGCACAAGCTCTAGTCACTGTGGATGAAGTAATTGATGAAGAAGAACTAAATAT GGAAGAAATGGTAAAAAATTCAAATTCACTTTTTACATTAGATGAATTAATTGACCAAGATGATTGCATTTCCCACAGTGAACCTAAAGATGTTACTGTTCTGTCAGTGGCTGAAGAACAAGATCTCCTCAAACAGGAACGCTTGGTAACTGTGGATGAAATTGGAGAAGTGGAAGAGCTACCTTTGAATGAGTCAGCAGACATAACTTTTGCCACTTTAAATACTAAAGGAAATGAAGGAGATACTGTAAGGGATTCCATTGGCTTCATTTCTTCTCAGGTGCCCGAAGACCCTTCTACTTTAGTTACTGTAGATGAAATACAAGATGACAGCAGTGATTTGCATTTAGTGACTTTGGATGAAGTAACTGAAGAGGATGAAGACTCTCTGGCGGATTTTAACAACCTTAAAGAAGAGCTTAATTTTGTTACTGTTGATGAAGTTGGAGAGGAGGAAGATGGAGATAATGATTTAAAAGTTGAGTTAGCACAAAGCAAAAATGACCATCCCACAGATAAAAAAGGGAATAGAAAGAAGAGAGCTGTGGACACAAAAAAGACAAAACTTGAATCCTTGTCCCAAGTGGGTCCAGTAAATGAGAATGTTATGGAAGAAGATCTAAAAACCATGATTGAAAGACACTTAACAGCTAAAACTCCAACCAAGAGAGTTAGAATTGGGAAAACTCTGCCATCAGAAAAAGCTGTTGTGACAGAACCAGCAAAAGGTGAAGAGGCCTTCCAGATGAGTGAAGTTGATGAGGAATCTGGATTAAAGGATTCAGAACCAGAGCGAAAACGCAAGAAGACTGAAGACTCTTCTTCAGGCAAATCAGTGGCGTCTGATGTCCCTGAGGAATTAGACTTTCTTGTACCTAAGGCTGGATTCTTCTGTCCAATTTGTTCCCTCTTCTACTCAGGTGAAAAAGCAATGACAAATCACTGCAAGAGTACACGTCATAAGCAAAATACTGAGAAATTCATGGCCAAGCAAAGAAAGGAAAAGGAGCAGAATGAGGCTGAAGAAAGAAGCTCTAGGTGATTGGGGGAAAGGAAAGAATTCACTAGAAATTTGTTTAGGGTCCAGTTGATTTGTGTATTTTTGTTATCATTTAATTTGTAATTTTCGTTTCAGAAGCAAATATTCGTGTTGTACAAATTTCTGATTGCCCTAAATGTAGAGAGACTGATGGGGAAAGTATGATGGGTTTGATTTTTATATCAAATCATCAGGCATGGAGAAATATCTTTTAGAAGTGTTAAAATAAATGTTCCTACTGTATATTTAAAATACCA

[0175] In some embodiments, the RNA interference molecule targets NP220 and comprises a sequence selected from SEQ ID NOs: 2-10 as shown below:CCTCAAGTATCTTAGCAAGTT (SEQ ID NO: 2)CGCTACAAAGAGTGATTCAAA (SEQ ID NO: 3)CCTGAGGAATTAGACTTTCTT (SEQ ID NO: 4)GCTATCCTGATGAACAACTAA (SEQ ID NO: 5)GCTCAGTCAATGTATAGCTTT (SEQ ID NO: 6)GCTATCCTGATGAACAACTAA (SEQ ID NO: 7)TCGTTTCAGAAGCAAATATTC (SEQ ID NO: 8)ATCCGTCAACCAATCCATTAA (SEQ ID NO: 9)CCCGTTCACCATATCGAATTA (SEQ ID NO: 10)

[0176] In some embodiments, the RNA interference molecule comprises a sequence having at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to a sequence selected from SEQ ID NOs: 2-10.

[0177] In some embodiments, the RNA interference molecule targets MPP8. In some embodiments, the RNA interference molecule targets a region within MPP8 (RefSeq NM_017520.4) having the nucleotide sequence shown as SEQ ID NO: 11 below:GATGTGGAGTAGGGCCGAGCGCGGAACGCGAGGGGCTGCTGGGGTGTTTGTCGC AGCGGGTTTTCCTCGGCGGTTTGCGGAGCTGCTAGGATGGAGCAGGTTGCGGAG GGAGCAAGGGTGACCGCAGTCCCTGTGTCAGCTGCCGACAGCACTGAGGAGTTG GCCGAAGTCGAAGAAGGAGTTGGAGTAGTGGGCGAAGATAATGACGCAGCCGC GAGAGGAGCGGAGGCCTTTGGCGACAGTGAGGAGGACGGAGAGGATGTGTTCG AGGTGGAGAAGATCCTGGACATGAAGACCGAGGGGGGTAAAGTTCTTTACAAAG TTCGCTGGAAAGGCTATACATCGGATGATGATACCTGGGAGCCCGAGATTCACCT GGAGGACTGTAAAGAAGTGCTTCTTGAATTTAGGAAGAAAATTGCAGAGAACAA AGCCAAAGCAGTCAGGAAGGATATTCAGAGACTATCCTTAAATAACGACATATT TGAGGCGAACTCTGATAGCGATCAGCAAAGTGAGACAAAAGAAGATACTTCCCC AAAGAAGAAAAAGAAAAAATTGAGGCAGAGAGAAGAGAAAAGCCCAGATGATC TGAAAAAGAAAAAAGCAAAGGCCGGGAAGCTAAAAGACAAGTCCAAACCAGAC CTGGAGAGCTCCTTGGAAAGTTTAGTTTTTGATTTAAGGACAAAGAAAAGAATTT CTGAAGCCAAAGAAGAACTAAAGGAGTCCAAAAAGCCCAAAAAAGATGAAGTA AAAGAAACAAAAGAATTAAAGAAAGTTAAAAAGGGTGAAATAAGAGATTTAAA GACGAAAACAAGAGAAGATCCCAAAGAAAATAGAAAAACAAAAAAAGAAAAA TTTGTCGAATCCCAGGTGGAATCTGAATCAAGTGTACTTAATGATTCTCCCTTTCC AGAGGATGACAGTGAAGGGCTACATTCCGACAGCAGAGAAGAGAAACAAAACA CTAAAAGTGCAAGAGAGAGAGCAGGGCAGGACATGGGGCTGGAGCATGGCTTT GAGAAGCCCCTAGACAGTGCCATGAGTGCTGAGGAGGATACCGATGTCAGAGGC AGGAGGAAAAAGAAGACCCCGAGAAAGGCTGAGGACACTAGAGAGAACAGGA AGCTAGAGAACAAGAACGCTTTCTTAGAGAAGAAAACTGTGCCTAAAAAGCAGA GGAATCAAGACAGAAGCAAAAGTGCTGCAGAGTTAGAGAAGCTGATGCCTGTAT CTGCCCAAACGCCAAAGGGCCGGAGGTTGAGCGGGGAAGAGAGAGGCCTCTGGTCCACGGACTCAGCCGAGGAGGACAAAGAAACCAAAAGAAATGAATCCAAAGAAAAATATCAGAAAAGGCATGATTCTGACAAGGAAGAAAAAGGCAGAAAAGAGCCAAAAGGATTAAAGACACTTAAGGAAATCAGAAATGCATTTGATTTATTTAAATTAACTCCAGAAGAAAAAAATGATGTTTCTGAGAATAATCGGAAAAGGGAAGAAATACCACTGGATTTTAAAACCATAGACGATCACAAAACCAAGGAAAACAAACAGTCACTTAAAGAAAGGAGAAACACCAGAGACGAAACGGATACTTGGGCATACATTGCTGCAGAAGGTGATCAGGAGGTTTTAGACAGCGTGTGCCAAGCAGATGAGAATTCAGATGGCAGGCAGCAGATTCTGAGTTTGGGCATGGACCTGCAGTTGGAATGGATGAAGTTGGAAGATTTCCAAAAGCACCTTGATGGGAAAGATGAGAATTTTGCTGCAACAGATGCAATTCCAAGTAATGTGTTAAGGGATGCTGTGAAAAATGGGGATTATATTACTGTAAAAGTTGCACTTAATTCAAATGAAGAATATAACCTGGACCAAGAGGATTCCAGTGGAATGACACTGGTGATGCTTGCCGCCGCCGGAGGGCAGGACGACCTCCTGCGACTCCTCATCACAAAAGGCGCGAAAGTGAACGGTCGGCAGAAGAACGGGACCACCGCCCTCATTCATGCTGCAGAGAAGAACTTTTTAACAACAGTGGCTATTCTTTTGGAAGCAGGAGCTTTTGTAAATGTCCAGCAAAGCAATGGTGAGACTGCACTGATGAAGGCCTGTAAAAGAGGAAATTCAGACATCGTACGACTCGTAATTGAATGTGGAGCTGACTGCAATATTTTGTCAAAGCACCAGAATAGTGCCCTGCACTTTGCGAAGCAGTCTAACAATGTGCTTGTGTACGACTTGCTGAAGAACCATTTAGAGACACTTTCAAGAGTAGCAGAAGAGACAATAAAGGATTACTTTGAAGCTCGCCTTGCTCTGCTAGAACCAGTTTTTCCAATCGCATGTCATCGACTCTGTGAGGGTCCAGATTTTTCAACAGATTTCAATTACAAACCCCCACAGAACATACCAGAAGGCTCTGGCATCCTGCTGTTTATCTTCCATGCAAACTTTTTGGGTAAAGAAGTTATTGCTCGGCTCTGTGGACCGTGTAGTGTACAAGCTGTAGTTCTGAATGATAAATTTCAGCTTCCTGTTTTTCTGGACAGTCATTTTGTTTACTCATTCAGCCCTGTTGCAGGTCCCAATAAACTCTTCATAAGGTTGACAGAAGCACCCTCTGCCAAGGTTAAGTTGCTAATAGGTGCATACAGAGTGCAGCTGCAGTGACCAAACAGAAGGGACTGGGCGGAGTTCTCTTCAGACCGATTCCTATACTCTCTTTGACAGCAGTTTGGAATTCTTCTAGCACATCTATGTAAAGTTTTGTCTGTAAACCTCTTGCAGTTAAGCCTGTTGTCTGTTGTAGTCTGTAAGATGCGACATAGCTGTGTCTGTGCCAGTATGCCGGAATCTCAGTGCAGTGTCCAGACTGCGTATTTCAGTTTTTCCACAATGTGGATAGTACATATGAGGATTATTTAAGAAAATTAAAGACTTCACTTTCTTTTTTCTGGAGACGGAGTTTCGCTCTTGTTGCCCAGGCTGGAGTGCAATGGTGCGATCTCGGCTCACTGCAAGCTCTGCCTCCTGGGTTCAAGTGATTCTCCTGCCTCAGCCTCCTGAGTAGCTGGGATTACAGACACCCGCCATGGCACCCAGCTAATTTTGTATTTTTAGTAGAGATGGGGTTTATCCATGTTGGTCAGGCTGGTCTTGAACTCCTGACCTCAGGTGATCCGCCCGCCTCGGCCTCCCAAAGTGCTTGGATTACAGGCATGAGCCACCGCACCCAGCAAGAGTTATTTTCTTAACTTGAAATTTTCTACTAGCCCTGGTGAACTTCTGTGCTTAAAAAAAAAAAAAAAAAAAGGAAAAATTCAGCTCTAAAACATTTGCTTACAGCAAGGGAGCCATGTTATATTCAAGTTACCCAAGCACCAAATAAAGTGTGGGTTTGCTGCCCTTGAAAGGGTTGGGAGGCCTGATCTTTTTCTAGTAGACAGTTGCCACACATTCCCCAAGCACAAAAGGTGGAGATGGCAGTCACTTTGTAAATATGCTGTAGTCACTTATCATCTTTTCAAAGGGTAACAAGAGAATCTAAGATGTAGTAAGAATGTAAACCAGTACACTATGAGACCTAAAAGAGAGTATTAAATGTGAACTTTTAGCAGAGCGTGGTGGCTCACACCTATAATCCCAGCGCTTTGGAGGCTGAGGTTGGAGGATTGCTTAAGTCCAGGAGTTCAAGACCAGCCTGGGTAACATGGGGTGGAACAAGCCTGTAGTCCCAGATACTCAGGAGGCTGAGGTGAAAGGATTGCTTGAGCCAGGGAGGTCAAGGCTGCAGTGAGCCGTGAAAGGCCACTGCACTCCAGCCTGAGTGACAGAATGAGACCTTGTCTCAAAAAAAAAAAAAAGTTTCTTGGAACCTATACGGTTTTTTTTTGTTTTTTTTTTTTGAAAAGCCAGACCTTGTGCCCTTGTTTTGAACACCGACTGGGAAGATGGGGCTTAGGTAACAGCCAAACCTGGCTGTCAGCTGTGTGGGAGCCACCACCCTCTCTGGGAAGAGTTCCTGCTTCTGTATGGCAAGCATAAATCAAGCTCAGTCTGGGTTATGGAGAAGTTGAAAATTGTTTTGTTCCTCATTAGTTTATAATTGTATGAAATACGATTTTAATGAAAACTTTTCAGAATTCACGTTTGTGTAGATATTTCAGAGAACCATTTTTACTTTACATCCTAAAACTGCCTTTTCCTATGGTTTTGTCAATAAAACACTATGATGTT

[0178] In some embodiments, the RNA interference molecule targets MPP8 and comprises a sequence selected from SEQ ID NOs: 12-20 as shown below:GCTAGAGAACAAGAACGCTTT (SEQ ID NO: 12)CCTCTTGCAGTTAAGCCTGTT (SEQ ID NO: 13)GCTGTTTATCTTCCATGCAAA (SEQ ID NO: 14)GCTTCTTGAATTTAGGAAGAA (SEQ ID NO: 15)CAAGCTGTAGTTCTGAATGAT (SEQ ID NO: 16)TTGCGAAGCAGTCTAACAATG(SEQ ID NO: 17)GACATCGTACGACTCGTAATT (SEQ ID NO: 18)TGGAGTAGTGGGCGAAGATAA (SEQ ID NO: 19)CAGTGTCCAGACTGCGTATTT (SEQ ID NO: 20)

[0179] In some embodiments, the RNA interference molecule comprises a sequence having at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%,at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to a sequence selected from SEQ ID NOs: 12-20.

[0180] In some embodiments, the RNA interference molecule targets TASOR. In some embodiments, the RNA interference molecule targets a region within TASOR (RefSeqNM_001112736.2) having the nucleotide sequence as shown as SEQ ID NO: 21 below:GCGCGCACGTTGAGTGGCGGGGGAAGGCAGAAGAACTGCCCGAGGGAGGAGCG GCTCCGAGGACCGGGCAGCGCATTTGGGGTGCCCGCTACAGGGCCCCCCAAGAGGCTGCC AGGTCGGGAGCGC AGC CTGAGGCC ACGGGGGGA AGGGGGTGGGTGGG CTGAGAGCAGCTCGGCGGCGAGACTGGCCCGCCGCCCCTTCCCCCCACACCCGTCGACCTTGTGGGCAGAAGCTTCCCAGAGCTCCTTAGGCCGGCGCGATGGCGACTGCTGTGGAGACGGAGGCCTGTCAGCCGACGGATGCGAGTTGGGAAAGTGGCGGCGGCGGAGACGACGAGATGAAGCAGGCGCTTCCGGAGCTTGAGTCCTCCCAACAAAATGGCGGCGGCGGCGGCCTCAACATCGCTGAGCCCAGCGGCGGCGCTGGGCGTGAGGAGAACGCGGGGGCCGAGGCCGCCCAGAGCCTCAGCCACGAGCAGCCTCAG GACTCCTCTGAGGCGGGCGCGGCCGCCCTGCCCAGAGGCCCCGAAGAGCCCGAAAGGCCTGTTAGGAGGAGTTTTCAGATCCCCAGGAAGAGCAGAGAAAAGAAAGCACTTTTCCAGCCATTAACTCCAGGCTCTCGAGAATTTGAAGATGTTGTAAATATTCTCCATTCTTCTTACCTTGAACCAACCTCAGTAACAAATTTTAACTACAGACGTGCTTGCTTGGTACACAATGAGCTTTTGGAAAAGGAGTTTACAGAAAAGCGAAGAGAACTGAAGTTTGATGGTCGTTTAGATAAGGAACTTTCAGAATCCTATGCATTTCTGATGGTTGATCGATACCAGGTTCAAACCATATGTGAAAAAGGATTACATGTGGG TCAGTCCAAAATAACAATTCTTGGCAGTCCTTCCATGGGTGTCTATCTTTCTAGGTATGCTGATTTATTACAAGCGAATCCTTTGGACACGGGGGCAATGGGTGATGTTGT TATTTTTAAAATAATGAAGGGTAAAATAAAGAGTATATATGACCCCATGGGTGTA AAAAGTTTGGAATCTATGTTAAATAAGAGTGCTTTGGACCCCACACCAAAGCATGAATGTCACGTGTCAAAGAATGCCAATCGAATTACATCACTTTTGGCTTACAGAGCCTATGAGCTTACTCAGTATTATTTTTATGAGTATGGCTTTGATGAGCTAAGGCG AAGACCAAGACACGTTTGTCCATATGCAGTTGTGTCTTTTACTTACAAAGATGATATACAAACTCCGAAGTTTGTACCTTCATCAAGATCTAACAGCTTTAATACAGATA GAAACATAGATAAATATAACTATACCTTGTGGAAAGGACAGCTTTTAAATAAAGGAAAACTTTTGTGTT ATATTTCTCTGAGGTC AGC C ACTC GTGCTTTTTTGC CTATC AAACTACCTGAGAAATTAGATGTTGAAACAGTTATGAGTATTGATCATCTGAAACAGAAAATCCCTCCAGCACTGTTTTATAAGGAAACATACTTAGGTCCAAATGAAGT TTTGAAGAATGGAATGTATTGCAGCCTTTATGAAGTTGTGGAAAAGACAAGAATTGGAAGTAACATGGAGAGTTTACTGCAAAAACTAGACAGAGAAAAACTTGTTCTTGTTAAACCTTTGGGAGACCGAGGATACCTTTTTCTTCTCTCCCCTTATCAGATGGTTCCTCCATATGAATATCAGACTGCCAAGTCTCGAGTCCTACATGCTTTGTTTCTATTTCAAGAACCTAGAAGCATAGTTACTTCACAAAAAGGTTCAACCAATGCAGCACCACAGGAGAGGCATGAGAGCATGCCAGATGTATTAAAAATAGCTCAGTTTTTACAATTTTCTTTGATTCAGTGTCGAAAGGAATTCAAAAATATAAGCGCCATAAATTTTCATTCTGTTGTTGAAAAGTATGTAAGTGAATTTTTTAAGCGAGGTTTTGGTTCAGGTAAACGAGAGTTTATTATGTTTCCATATGATTCACGATTAGATGATAAAAAATTCTTATACTCAGCTCCCAGAAATAAATCCCATATTGATACTTGTTTGCATGCCTATATTTTTCGGCCTGAAGTGTATCAGTTACCTATTTGTAAATTAAAAGAACTATTTGAAGAAAATAGAAAACTTCAGCAGTTTAGTCCACTTTCAGATTATGAAGGTCAAGAAGAAGAAATGAATGGTACAAAAATGAAATTTGGAAAACGAAATAACTCAAGAGGTGAAGCCATTATATCTGGAAAGCAAAGATCATCTCATTCTTTGGATTATGATAAGGATAGAGTCAAAGAATTGATTAATTTAATTCAGTGTAGGAAAAAGAGTGTGGGTGGGGACTCAGACACAGAAGATATGAGAAGCAAAACTGTCTTGAAGAGGAAGCTTGAGGATCTACCTGAAAATATGAGAAAGCTCGCCAAAACCAGTAATTTATCTGAAAATTGCCATCTGTATGAAGAGTCTCCACAGCCTATTGGCTCACTTGGACATGATGCTGACTTGAGGCGGCAGCAGCAGGATACCTGTAACTCCGGCATTGCTGACATCCATAGGCTGTTTAATTGGTTATCAGAAACACTAGCAAATGCGCGCCATTCTGATGCATCTCTGACAGACACAGTCAACAAAGCCTTAGGATTGAGCACTGATGATGCCTATGAAGAGCTGAGGCAAAAACATGAGTATGAGTTGAACTCTACCCCAGATAAGAAAGACTATGAGCAGCCTACTTGTGCAAAAGTTGAAAATGCACAGTTTAAGGGTACTCAGAGCTTATTACTAGAAGTTGATGCAACATCTAAGTATTCTGTTGCTATTTCTACCAGCGAAGTGGGCACTGACCATAAGCTACATTTGAAAGAAGATCCAAATTTAATTAGCGTGAATAATTTTGAAGATTGCAGTTTGTGTCCCAGTGTTCCCATTGAACATGGATTTCGTAGACAACAGTCTAAGTCAAATAATGTTGAAGAGACTGAAATACATTGGAAACTGATTCCAATTACAGGAGGGAATGCAAGAAGCCCAGAAGACCAGCTGGGGAAACATGGTGAGAAACAAACACCAGGTATGAAATCACCAGAAGAACAACTGGTGTGTGTGCCACCACAGGAGGCCTTTCCTAACGACCCCCGGGTAATAAATAGACAGAGAAGTTCTGATTACCAGTTTCCATCCTCTCCATTTACAGACACACTAAAGGGCACCACTGAGGATGACGTGTTGACAGGTCAGGTGGAGGAGCAGTGTGTGCCAGCAGCAGAGGCAGAGCCGCCTGCAGTGAGCGAAACCACAGAGAGGACAGTGTTAGGAGAGTACAATCTCTTTTCTAGGAAGATAGAAGAGATTTTGAAGCAAAAGAATGTTTCATATGTCAGTACAGTTTCCACACCTATCTTTTCAACACAAGAGAAGATGAAACGGCTTTCCGAGTTCATATATTCTAAGACTTCCAAAGCTGGTGTGCAGGAGTTTGTAGATGGTTTGCATGAGAAGCTAAATACTATTATTATTAAAGCATCAGCCAAGGGTGGGAATTTGCCACCAGTCAGTCCTAACGATTCTGGTGCTAAGATAGCATCGAATCCTCTGGAAAGGCATGTCATACCAGTTTCCTCAAGTGACTTCAACAATAAACATCTCCTTGAGCCACTGTGTAGTGATCCTTTGAAAGATACCAACTCTGATGAGCAGCATTCCACTTCGGCTTTAACTGAAGTAGAAATGAACCAGCCTCAACATGCCACAGAGTTAATGGTGACTTCTGATCATATTGTACCTGGTGATATGGCCCGGGAACCAGTAGAAGAAACAACAAAATCCCCCAGTGATGTAAACATTTCTGCTCAACCAGCTCTTTCAAATTTTATAAGCCAGTTAGAACCTGAAGTATTTAATAGTTTGGTTAAAATCATGAAAGACGTCCAGAAAAATACTGTGAAATTTTATATTCATGAAGAAGAAGAGAGTGTGCTCTGTAAAGAAATAAAGGAATATCTTATCAAATTAGGCAATACAGAATGTCATCCTGAACAGTTTTTGGAAAGAAGATCAAAATTAGATAAACTATTGATTATTATTCAAAATGAAGACATTGCAGGTTTCATTCACAAGATACCTGGCTTGGTGACTTTAAAGAAGCTCCCCTGTGTTAGTTTTGCTGGTGTTGATAGCCTGGATGATGTTAAAAATCATACATACAATGAATTATTTGTATCTGGAGGTTTTATCGTATCTGATGAATCAATTCTAAACCCAGAGGTTGTCACAGTTGAGAACCTTAAAAATTTTTTGACATTCCTTGAGGAACTTAGTACTCCAGAAGGAAAATGGCAATGGAAAGTCCACTGTAAATTTCAGAAGAAACTAAAGGAACTAGGCAGATTGAATGCTAAAGCTCTAAGTCTGTTGACGCTTCTGAATGTCTATCAGAAGAAACATCTGGTTGAAATTTTGTCATACCACAATTGTGATTCACAAACTCGAAATGCTCCAGAATTGGATTGCCTTATCAGACTTCAGGCTCAGAACATACAGCAACGACACATAGTCTTTTTAACAGAGAAGAACATCAAGATGCTTTCCAGTTATACAGATAATGGAATAGTGGTTGCAACTGCTGAAGACTTCATGCAAAACTTTAAAAATCTTGTGGGCTATCACAATTCAATCACAGAAGAAAACCTTCCACAGCTTGGTGCTAATGAGAATCTTGAGTCGCAGTCAGATGCTGTTTTGACATTAACCCCTTTGGAGCTGGGAGTTGGGATTTCCCAACATTAAACGTGAACACATTTCGCAGAAGCTTTTAGCATCGGATTATCTGGACATTCACATCTAGTGTGAGTGTTGTGACTAAGTGAACTTGTGGCAGAAGACCAAGCTTTAAGGGATTGTGGACTTGCAGACCCTGACGCGTTATATTGTGTGAGTAGTGTATAATTTTAAAACTTAAACAAACTGTGTATTTCAATAGAGGAGGACTTTTTGTTTTTTAAATGTAGCTCTTTTAGAAAACGATGAAAAGGATGAAGAGGATATGTCTCTGGATTCAGGGGATGAAATCTCACATATAGAAGTATGCAGCAATTTTCATTCAGAAATATGGGAGAAAGAGACCAAAGGATCACGTGGAACAGATCAAAAAAAGAATACTCAAATTGAGTTGCAATCGTCTCCTGATGTGCAAAACAGTTTATTAGAAGATAAGACTTACCTTGATTCTGAAGAGAGAACTTCTATTGATATAGTATGCTCTGAAGGAGAGAACAGCAATTCAACAGAACAAGATTCATATAGTAACTTTCAGGTTTATCATAGTCAATTAAATATGTC CCATCAGTTTAGTCATTTTAATGTTCTCACTCATCAGACATTTTTGGGGACACCAT ATGCCCTTTCATCAAGTCAGTCTCAAGAAAATGAGAATTACTTCTTATCTGCTTAT ACTGAAAGCTTGGATAGAGATAAATCTCCACCTCCCTTAAGTTGGGGGAAAAGT GATTCTTCCAGGCCATATTCACAAGAGAAATAACTGTAGTAACTTTTTTTTTAAG AGATTGTTGTGGACTTTGTTTATTAACAATTTATATTTCATTCTCTAAACAAAAGG TTCTTGTTCTTTCTCAAATGTTTTTTCTTTTATTTAAATCATGATGGCCTGTAACAG TTGAAGCATCTAAAAATTGAAATAAATATATATTTTTAACATATATTGTACTTGA ATTATCTCATGTTGGCACTTCTAAGTTTTACTTTTTGTATGTTACCTGTACTTAGGC TTCAAATTGAAGCCTGTTTTATGTGTAAAATCAGATGCTTGCAGGAGGACGGTAA ACAGCAAAGGCTTTACTTCTTTTTCTCTAATGTCACCTGATATAAAGTACTTTTGT TGAAGTGAATTATTTTGATGTAGGCTTTTGTATACTGTGTTTACACAGGTATAAGT CAGTAATAGCAGTGTTAATATTTACAGGATTTTGGCTCGATACCTCAAGATTTAT CTAACAGTGCAGTTTCATTTAGATAATAAACTATGTACATAGGACACATTTTTTA TCTCTAGGCTTAGTCATGTAAAAACTGCCTATTGCTAAAATTCTGGTAACTTTTTG AGAGTATTGGATAATTTTCAGTACCTGCGCAATCCTTGATTTAATGAAATTAAAT TTTTT ATTTC AAAAGATAGGCTTCTGTTTATC AAAGGC GTTGAAC AATTTGATTTT TAAATTAGTTTTAGCAAAATGGAAGTATAATGGGAATATATTTTTGAGGTGGTCT TAGTAGTAATTACCATTTGTTGAACACATTAATGCTATACTAGACAGATACAGTGGAGATAGCTTTCAAACCCGTATCTATTCCTAACTACTGCCTCTAATACTAATTACT AACTTGTTACTAATGCTAACTTTTAAAAATGTTTTTGAAAATTGCAATTTCTACAA ATAAATGTCATAGTGCAAATAAATGTCAGGATTTAGAGAAGGAGCCTAAACTGA ATTTGTGTTATTTCAGTAAAAGTTATGGAGT ATGGGTCTAGACGTGTTAATAAG TTAGACAGAACTTTGGCACTTTAGAACAAATGCATGAGTGGTATTTCAGTTCCTA AGTTACATAAAAAGTGTGAAAGAACACTGTGAGGTCCCAGCAACGCCAGACTAT ATTAAGGTAAGTAGAAAGTGTTTTTATAGGGCTTCAATACCCAGGTGGTGACAGA GCAGAAGAACCGGTTTTTTTTTTTTTTTTTTTACTACTAAGCTTTTACAAAAGAAG TATCTGTTTTATTGTATAGAACAAGTACAGCATTTTACTACATAGTATACAAGTTT TTAATGAGCATTTAAAAAAATAAGTAAATCTAGGCTATTTGAAAAATACAGTTGA GCCAGTGAGCACATTTTATAATTTGGAAGACACAAATCAAATGTGAAGGATTTG ATTTTCTACATTTAAAATGAAGAACCAAAACTCTCTTCTTGATTTTCAGCTAAAG GCAGGAGACTACTTTCCAACTCCTTTTGCTTCTGGAGAAGGCCCTGAAATCACAT TGATATATGTTTGTTAGTAAAATGTGCACACCTGACTTGCAATGTTGTGTTAAACT AGAATTATATTCACTTGGAAAACATATCGCTTAGGATAAAATTTTGTCATTCAAAAATTTTTTCAGAAATGCTTGTACTAGACCACCTCATTTGGATATATAGCTATCACA TTTGTAAGATTCTGTTTTTAGCATAAAAATCAATAGGGATGAAAATTTTCTTAGA ACTCAAGACTCCATAACTATGGGGGTAAATGAAGTCATCCCTTCTCCCCCAGTCC ACCCCCAAGTGCTCTTAGATACTGAAGGAGTCTCCCTTAACATACAGTGTTTTTTT TTTGTTGTTGTTGTTTTGTTTTTTTTTGGAGATGGAGTCTCGCTCTGTCGCCCAAGC TGGAGTACAGTGGTGCAATCTCACTGCAAGCTCTGCCTCCCAGGTTCACACCATT CTCCTGCCTCAACCTCCCGAATGGCTGGGACTACAGGCACCCACCACCACGCCTG GCTAACTAACTAACTAACTTTTTGTATTTTTAGTAGAGACAGGGTTTCACTGTGTTAGCCAGGATGGTCTTGATCTCCTGACCTCATGATCTGCCTGCCTCAGCCTCCCAA AGTGCTGGGATTACAGGCTTGAGCCACTGCTCCTGGCCAACATCCAGTATTTTAT CTTGACTGTCCTAACCTTACCTTAGATGCTAACAGAAGGGTCCTGCTGAAATAACACTGGGTGCTATATTGATGGGTAAATGTGTACATCCTATTCCTTCCTCTTTATCTC ACAATTTTTGTCTCCACTAAGCAAGAAGTAAACTAACACTTCGTCACTCTAAAGA AATAACTTATGTAAAACTCTTAGTAACCCTGTTTGTCTTCAAATGAGTAAATAGACCAAAGTGGGGGGACAATTTTCTAGTTCTGTAGAGGGAAAAACATCTGAGTCAA CATTTTGAAATGCAGAGGGTATTGGTACATGACGACATGGAAAAGGGCACTTTT AAACACAGCTTACTCTTCCTCAAGTACAGAGAGTATATAGTGAATCAAAACTAACTACAGCCATTCTTTTTAAAGCCCAAGGGATGGAGCAAAGGTGTAAGGATGTTA CCTGTTTGTTTTAATCAGAGAGCAAAAAGAAGTCACAATAGTTTGGGAGAAAAA GTAGTATGGTGAGTAAGGTTATGCGTATAATTTCATACTGAATTTATTACTATTTGGGATGTACGTCAATGTTCTAACAAACACTGCCAACACGTCAATTTTTTAAAAAGC GTGGGCCACATTGCTAAGAATTTGTTAAAGCATAACTGTATTTTTTGTTTTAGGGC CTTATTGATGTTTTGCCGTTCCAATGTATGCATTTTTTTACTCAATAAACTTGTCTT AATTTTA

[0181] In some embodiments, the RNA interference molecule targets TASOR. In some embodiments, the RNA interference molecule targets TASOR and comprises a sequence selected from SEQ ID NOs: 22-30 as shown below:CAATGGAAAGTCCACTGTAAA (SEQ ID NO: 22)GCAGGTTTCATTCACAAGATA (SEQ ID NO: 23)GCAATTCAACAGAACAAGATT (SEQ ID NO: 24)CGCCAGACTATATTAAGGTAA (SEQ ID NO: 25)GCCCTGAAATCACATTGATAT (SEQ ID NO: 26)GTTCCCATTGAACATGGATTT (SEQ ID NO: 27)AGCATTCCACTTCGGCTTTAA (SEQ ID NO: 28)TCAGGTTTATCATAGTCAATT (SEQ ID NO: 29)GTCCCATCAGTTTAGTCATTT (SEQ ID NO: 30)

[0182] In some embodiments, the RNA interference molecule comprises a sequence having at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to a sequence selected from SEQ ID NOs: 22-30.

[0183] In some embodiments, the RNA interference molecule targets TASOR and comprises the sequence of SEQ ID NO: 22. In some embodiments, the RNA interference molecule targets TASOR and consists of the sequence of SEQ ID NO: 22.

[0184] In some embodiments, the RNA interference molecule targets PPHLN1. In some embodiments, the RNA interference molecule targets a region within PPHLN1 (RefSeq NM_016488.7) having the nucleotide sequence shown as SEQ ID NO: 31 below:ACCTGGGATAACGGCGGCGAGCGGACGGCTGCATTTACGGGGTCTCCCGGAGGG CCAGAGTCGTGGCTTACAGAAGAGACGAAATGTGGTCTGAGGGACGATATGAAT ATGAAAGAATTCCGAGAGAACGAGCACCTCCTCGAAGTCATCCCAGTGATGGCT ACAATAGACTAGTTAATATTGTGCCAAAGAAACCACCACTGCTAGACAGACCTG GTGAAGGAAGCTACAATAGATATTACAGTCATGTTGATTACCGAGACTATGACG AGGGCCGCAGTTTTTCTCATGATCGAAGAAGTGGTCCACCTCACAGAGGAGATG AATCTGGTTATAGATGGACAAGAGACGATCATTCTGCAAGCAGGCAACCTGAAT ACAGGGACATGAGAGATGGCTTTAGAAGAAAAAGTTTCTACTCTTCCCATTATGC GAGAGAGCGGTCTCCTTATAAAAGGGACAATACTTTTTTCAGAGAATCACCTGTT GGCCGAAAGGATTCTCCACACAGCAGATCTGGTTCCAGTGTCAGTAGCAGAAGC TACTCTCCAGAAAGGAGCAAATCATACTCTTTCCATCAGTCTCAACATAGAAAGT CCGTGCGTCCTGGTGCCTCCTACAAACGGCAGAATGAAGGAAATCCTGAAAGAG ATAAAGAGAGGCCTGTCCAGTCTTTGAAAACATCAAGAGATACTTCACCCTCAA GTGGTTCAGCAGTTTCTTCATCAAAGGTGTTAGACAAACCCAGTAGGCTAACTGA AAAGGAACTTGCTGAGGCTGCAAGCAAGTGGGCTGCTGAAAAGCTAGAGAAATC AGATGAAAGTAACTTGCCTGAAATTTCTGAGTATGAGGCGGGATCCACAGCACC ATTGTTTACTGACCAGCCAGAGGAACCTGAGTCAAACACAACACATGGGATAGA ATTATTTGAAGATAGTCAGCTAACCACTCGCTCTAAAGCAATAGCATCAAAAACC AAAGAGATTGAACAGGTTTACCGACAAGACTGTGAAACTTTCGGGATGGTGGTG AAAATGCTGATTGAAAAAGATCCTTCATTAGAAAAGTCTATACAGTTTGCATTGA GGCAGAATTTACATGAAATAGAGTCTGCAGGACAGACTTGGCAGCAGGTACCCC CTGTGAGGAACACTGAAATGGATCACGACGGAACCCCCGAAAACGAAGGCGAAGAGACTGCGCAGTCAGCCCCGCAGCCCCCGCAGGCCCCGCAGCCCCTGCAGCCCCGGAAGAAACGGGTTCGTCGGACGACACAGCTTCGTCGGACGACAGGAGCCCCAGACATTACCTGGGGGATGCTGAAGAAGACAACTCAGGAGGCTGAGAGGATCCTGCTCCGAACACAGACACCATTCACTCCAGAAAATTTGTTCCTTGCTATGCTCTCTGTTGTACATTGCAACTCACGCAAGGATGTAAAGCCAGAAAACAAGCAGTAACTGCTATGCCTGACAAAACTGTTGCTGCACACATCTGTACTCGTCAATCAACAAAACCTGATGCAAAAAACAGAAAAGGGGTGATGTAGGAGATGGTCAGGTTGGTAGGAGAAGCTATAAGGAAAGACGCAATTGGAAGGTCGGGAGGTTTTCCAAAGCTTCAGGAGAGAATAAAGCTGAAGGCAGCTTTATTAATTAATTCTCTTACCCTGAGGCTGAGGGCGAACAGTAGGTAGCAAGGGAGTGTAAAGGAATTTATCTAGATAAGTTTGTTTACTTATGCCCTCCGGAAATCATGCAAGACTGCTCCCTGCAAAGGGGGGCGACAATGTTCATTACTCACAAATTGTGTTGGCTTCAGGCCTTTGGTATTCTGTCTCTACTGAATAAATACAAATGGTTCCAGCCTATCAGGACTGCACTCTCTTCTCGGCTGCACTAAAGCTGGCACTCCCCCAGCCGTTCTCATGCAAAATACCTGTGTCAGAATACTCCTTTCATCCATCACTCAGCCAGAGTCTTCAGGACAGACTCCGCATGGGACTTGTCCAAAAAAACTCTAATCAAAAGAGGAAAATTTTGGAATATGCCAGGAATAGTGGAATTTTATTTTTTAAATTTTTTTATAGGCCCATATGCTCTATCTCAAGAAACAAGATGATTGTAACATGTCCATGATTAAACTATTGGCAGATTATTGCTGTGTTAATCTCTGTAGTCTAATGAGTTCTTTGTTCTGTTCTGCTGCCTTTTACGTTTTCTTGTCCTTTCAAAAGTGTTCTTGAAAGAAACAAAGCGAATAGGCAGTTAGCACAGCACAGCTACCCCTTACCAAGCAGTCTATGGAAACAACCCCTCATCCAAATCATGGGTTAGTTAAGAATCTAACTGGGGCAATTAAGATGAATTCCACTCACTTCCTGGTCACTTCAGCAGCCCAGCGGCATTGAGCCAAAATATACAATTCTGTGTTATTAGTGAGGAAACTTTAAAACTCATGTTTGTTATTACTTACTACCCAATTTCATTATCCTCCCTTCCTCTTTCCATTTCTATTCTCTCTCACTTGAATTCTGGCATTATTTTTAGTGGCCTCTACTGATAATACCTACCCTAGAGTACATAAAAATTATATTAAAAGAGGAAGTAGCAGTATGCATAATTTTAACAGATTCTATAATGGGTGCCTCAAAATATGTATTGTGCCATTCCGCAAATTTAAAAGCTAATTGAGGACAATTTTTTTTTAATTTCCTAAATGAGACCACCTTGGATTTTTATTTTTGCCATTTAGATGTTTATACTTATTTAGCTTTTATAAAACATAAGCCAAGCTAAATCCCACATAACAACTCTGGTATTCTTCCCTCATATGAGCAGTGATTTTATTTGTTACCCACCTTAGATAGACTAAGAAAGTTCTAGTCTTGTTTCTCCTTCTCCCCGCTTCCCTGGGGTTTTTCCTTACCATAAGTATTCTGGTCCGAGGGTTCAGTTCCTTTAGTCAAGATGTCACAAGTTTAAAAACAAAACTTGAGAAACTACCAAAGGCTCAGGAGTTGTCCACTTTGTTGAAATCCATTAAATTAGAGAAGTCTCACTAACAGATGTATTTAAATATAGGTACAACAAATAATTTCTTTTTCTCCCCTTCCCCAAA TTACAGTCAGCATTTAAAGCTGTTTATGGCTTGCCATCAGCATTATTCTGGTAGGC TTGTTAGTGTTAAAATCTATTTGATTTTTTTTTTTTTTTTTTGCCTCTTAAAGTCTA ATTTTAGGATGGATGAATTCAGATGTTTACCAGAGTGTGTATTTTACATAATGTTC TTGATTAAAAAGACTTGTTTGTAAATTATCCGTTGTTTTTGCATATGCCCAGTTGA TGTGATAAAATTTTCATTGTCTTGCCATATAAAGCCTTGGTTATCAACAGGTGGA ATGTAGATATTGTAAAGCTTTTTGTGAATTAAAAGTGCAAAATAAAGCAACCACA TTTAAGTA

[0185] In some embodiments, the RNA interference molecule targets PPHLN1. In some embodiments, the RNA interference molecule targets PPHLN1 and comprises a sequence selected from SEQ ID NOs: 32-45 as shown below:GAGATGAATCTGGTTATAGAT (SEQ ID NO: 32) GATGGACAAGAGACGATCATT (SEQ ID NO: 33) GATGAAAGTAACTTGCCTGAA (SEQ ID NO: 34) CCAAAGAGATTGAACAGGTTT (SEQ ID NO: 35) CCTGGTGAAGGAAGCTACAAT (SEQ ID NO: 36) CGGCAGAATGAAGGAAATCCT (SEQ ID NO: 37) CTGAAATGGATCACGACGGAA (SEQ ID NO: 38) GCAGAAGCTACTCTCCAGAAA (SEQ ID NO: 39) GCAACCTGAATACAGGGACAT (SEQ ID NO: 40) CAAGTGGTTCAGCAGTTTCTT (SEQ ID NO: 41) CCTGAGTCAAACACAACACAT (SEQ ID NO: 42) CATGTTGATTACCGAGACTAT (SEQ ID NO: 43) GATGGCTACAATAGACTAGTT (SEQ ID NO: 44) GCTCTAAAGCAATAGCATCAA (SEQ ID NO: 45)

[0186] In some embodiments, the RNA interference molecule comprises a sequence having at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to a sequence selected from SEQ ID NOs: 32-45.

[0187] In some embodiments, the RNA interference molecule targets MORC2. In some embodiments, the RNA interference molecule targets a region within MORC2 (RefSeq NM_014941.3) having the nucleotide sequence shown as SEQ ID NO: 46 below:GGGAACCAAAAAAGGCTGGAAGAACATGAAGATGGAGCAGTCATAAACCACCC ACTCAAGGACCATCTCCTTCAGGACCATCCACACGAGACTCAGATTGTCTGAATT GAGCTATCGCAACTTAATGCTAAAAGCTCCTTAAAGCTACAGATTTATGACATAGTTCCTTCCAAAATATTACATCATAAATCATTGAGAAGATTAAAAAAAAACACTTGAAGAAATTGTAGTTTTAAACATCTCTGCATATATTTTGGATAGCTACTAGGTTACTTTAACTGTCATTAAGGAGCACAGACTTACTGAAGCTTTACTGGACAGAATCCTGGGAAATCGATATCATTATAAGGTTATATTTCCCAGTTAGCGGGTGAAGGGCTGGAGACCTTATTGCAGTCATGGCTTTCACAAATTACAGCAGTCTGAATCGAGCTCAGCTAACCTTTGAATATCTGCACACAAATTCGTAAGTATCCTCTAGGTGCCACTGAGGTAACCAGTAACTCGTTCCTTGATATTATATGGAAATCGTTTCCCCAGAAAATTTTGCTTTTTCACTT^TGAGATGTATCCCACTGGAGTGAAATGTGTCACTGGATATCTTGAGCTCTGTATTGAAGAACTGAGATCAGTGAAATACTTGTTGCTAATCCAGAAGAATCTGATTTTTGTTTATTGGATCAAAATTTTCTAAATGCAAACTTTAGTTATTTGAAGTCAATATGTTGAGTTGTTTCATTCAAGTGTTTATAGGAATCCAACAAATACTGCTCTATTGGATCGCCAAATGTTGGACTATTTTAGTATCAACCGTTTCCCCTCTGTAGTGACAACGTCCTAAACAGTTAGGTTTATAACAAGTGTTTACTTTCTAACAAGAAAACAGAAGACATTTAAATGACAACTTTCAAGAAGAAAATTTTTATTTTTTCAGAAGTTGGCATTATCTTCCTGGCAGATTGCTCACATCCAATATTATTTGTATATGCTAAACAGGAAACGGCAACTTGTTTATATCTCTATTTAGATAGTCTTTCCCCAAAATTTCCACAGAAACATACAGTGTTCATGGTTCTTGAGTTCATGAAGGAGTAATCTAATCACTCCAACATGGTCTGGAATGTTTCAGGTTTAATCCATATGCCCACTCTCTTGGAGGCTGTCCAGTAGCGTCAAAACTTTAGTGTTTTAATACATTCACCTGTTACTTTTGAGATGAAGTTCACCTTTCTTGGATCACATGCAAAGGATGTTTAGGTCTGTGAAGAAAAGAATTTCTAGGCCGGGTGCTGTGGCTCACGCCTGTAATCCCAGCACTTTGGGAGGCCGAGAACCACTCACGAATTCTTGTTTGGTGCCCTTGCTGAACTGGTTGATAATGCAAGAGATGCTGATGCCACCAGAATAGATATTTATGCAGAAAGACGAGAGGACCTTCGAGGAGGATTTATGCTTTGCTTTTTGGATGATGGAGCAGGAATGGATCCAAGTGATGCTGCCAGTGTGATCCAGTTTGGGAAGTCGGCCAAGCGAACACCTGAGTCTACTCAGATTGGGCAGTACGGGAATGGGTTAAAATCGGGCTCAATGCGCATTGGGAAGGATTTTATCCTGTTCACCAAGAAGGAAGACACCATGACCTGCCTCTTCCTGTCTCGCACGTTTCATGAGGAAGAAGGCATTGATGAAGTGATAGTCCCACTGCCCACCTGGAATGCTCGGACCCGGGAACCTGTCACAGACAATGTAGAGAAATTTGCCATTGAGACAGAACTCATCTATAAGTACTCTCCATTCCGCACTGAGGAGGAAGTGATGACCCAGTTTATGAAGATTCCTGGGGACAGCGGAACATTGGTGATCATCTTCAATCTCAAACTCATGGATAATGGAGAGCCAGAACTAGACATAATCTCAAATCCAAGAGATATCCAGATGGCAGAGACGTCCCCAGAGGGCACGAAGCCAGAGCGGCGCTCGTTCCGTGCCTATGCCGCTGTGCTCTATATTGATCCCCGGATGAGGATCTTCATCCATGGGCACAAGGTGCAGACCAAGAGGCTCTCCTGCTGCCTGTACAAGCCCAGGATGTACAAGTACACGTCAAGCCGTTTCAAGACCCGTGCGGAGCAGGAGGTGAAGAAAGCAGAGCACGTAGCAAGGATTGCTGAAGAGAAGGCGCGGGAGGCAGAGAGCAAAGCTCGGACATTAGAAGTACGCCTAGGTGGAGACCTCACGCGGGACTCCAGGGTGATGTTGCGACAGGTCCAGAACAGAGCCATCACTCTGCGCAGAGAAGCCGATGTCAAGAAGAGGATCAAGGAGGCCAAGCAGCGAGCACTTAAAGAACCTAAGGAACTGAATTTTGTTTTTGGTGTCAACATTGAACACCGGGATCTGGATGGCATGTTCATCTACAACTGTAGCCGACTGATCAAAATGTATGAGAAAGTGGGCCCACAGCTGGAAGGGGGCATGGCATGTGGCGGGGTTGTTGGGGTTGTTGATGTGCCCTACCTGGTCCTGGAGCCTACACACAACAAACAGGACTTTGCTGATGCCAAGGAGTACCGGCACCTGCTCCGAGCAATGGGGGAGCACCTGGCGCAGTATTGGAAGGATATTGCCATCGCCCAGAGGGGAATCATCAAGTTCTGGGATGAGTTTGGCTACCTCTCTGCCAACTGGAACCAGCCCCCATCCAGTGAGCTGCGTTACAAACGCCGGAGAGCTATGGAAATCCCCACCACCATCCAGTGCGATTTGTGTCTGAAATGGAGAACCCTCCCCTTCCAGCTGAGTTCTGTGGAAAAAGATTACCCTGACACCTGGGTTTGCTCCATGAACCCTGATCCTGAACAGGACCGGTGTGAGGCTTCTGAACAAAAGCAGAAGGTTCCCCTGGGAACATTCAGAAAGGACATGAAGACGCAGGAAGAGAAGCAGAAACAACTGACAGAGAAAATTCGCCAGCAGCAGGAGAAGCTGGAGGCCCTTCAGAAAACCACACCCATCCGCTCCCAAGCAGACCTGAAGAAATTGCCCTTGGAAGTGACCACCAGACCTTCCACTGAGGAACCTGTGCGTAGACCTCAGCGTCCTCGGTCGCCCCCTTTACCTGCTGTGATCAGGAACGCCCCCAGCAGACCCCCTTCTTTGCCAACTCCTAGACCAGCCAGCCAGCCCCGAAAGGCTCCTGTCATCAGCAGTACCCCAAAGCTCCCTGCTTTGGCAGCCCGGGAGGAGGCCAGCACATCTAGGCTGCTCCAGCCACCTGAGGCACCCCGAAAGCCTGCCAACACTCTCGTCAAGACTGCATCCCGACCTGCCCCTCTGGTGCAGCAACTGTCACCATCTTTACTGCCCAACTCCAAGAGCCCTCGGGAGGTTCCTTCTCCCAAAGTCATCAAGACTCCAGTGGTGAAGAAGACAGAGTCACCCATCAAACTCTCCCCGGCTACCCCTAGTCGGAAGCGGAGTGTCGCAGTTTCTGATGAGGAAGAAGTTGAGGAGGAAGCTGAGAGGAGGAAGGAGAGGTGCAAGCGGGGCAGATTTGTTGTGAAGGAGGAAAAGAAGGACTCGAATGAGCTCTCAGACAGTGCTGGGGAAGAGGACTCGGCTGACCTCAAGAGAGCTCAGAAAGATAAAGGGCTGCACGTGGAGGTGCGTGTGAACAGGGAGTGGTACACGGGCCGTGTCACAGCCGTGGAGGTGGGCAAGCATGTGGTGCGGTGGAAGGTGAAGTTTGACTACGTGCCCACAGACACGACACCAAGAGACCGCTGGGTGGAGAAAGGCAGTGAGGATGTGCGGCTGATGAAACCCCCTTCTCCGGAACATCAGAGCCTTGATACACAACAGGAGGGCGGGGAGGAGG AGGTGGGC CCTGTGGCCC AGC AGGC C ATAGCTGTCGC AGAGC CCTCC ACTTCCGAATGCCTCCGCATTGAGCCTGACACCACTGCCCTGAGCACCAATCACGAGACCATCGACCTGCTTGTCCAGATCCTCCGGAATTGTTTACGGTACTTCCTGCCTCCAAGTTTCCCCATCTCCAAGAAGCAGCTGAGTGCTATGAATTCAGATGAGCTAATATCTTTTCCTCTGAAGGAGTACTTCAAGCAATATGAAGTAGGGCTCCAAAACCTGTGCAATTCCTACCAGAGCCGTGCTGACTCCCGGGCCAAGGCCTCCGAGGAAAGCCTGCGCACCTCCGAGAGGAAGCTCCGCGAGACGGAGGAGAAGCTGCAGAAGCTGAGGACCAACATCGTGGCACTCCTGCAAAAGGTGCAGGAGGACATAGACATCAACACAGATGATGAGCTGGACGCCTACATTGAGGACCTCATCACCAAGGGGGACTGAAGGCAGGAGAGAGAGCAGCTCCCCTGCCCACCTGCCCCTCAACCCTGTAGCTGCAGGGGGAGGGGACTTCATTCATGGGTTGGTGGTCGCACCTTGGTTTGACTTACACGGGACATTTGTGTTTTTGGAGGAAAAGATACCCTGATTCTTTGAATCTTCCTTAAGTTTATAAATATTTATTTTTTAAAAGAAGATGCTGTGCCTGTGAGACCATACTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTGGTGACTGCAAAGGACAGAGAACCTTTCCACTTTGGCCATACTGGGTTGCTAAGCCGGAGCCATTTCAGCTCCTGGCTCCTCAAGATAACGGCGAGTCCAGTGCCATCTTGGAGAAGCTCCAGGGGCAGGGCTGACTTTTCTCCTACAGGAGGAACAATGTGGGGATCTGAGGGATGGGAGGGAGACTTCCCCCTAGAGTGGTGGTCCTGCTGGGGGCTCATATCCAGGGACCCAAAAGGGGGGCTGTGTAGGAGGTTCCACATTGGAGGGGCTCTCTCTCTCGCAGCTGTCAGAGTTGGTCCTGGCTGTGGCGTCCAAACAGCTTGAGGGAAAAAGATCCTGTCTAACCACCTCATCTACTACTCAAGTTCTTTCTGAAGGAGGGATTTCTTCAGTTAACCATGGACAGTGAGGTTTCTCACCACAGTAACTTGAGTCCAGGTTGAGGGGGAGACAGATCTGTGGTAAATCTCTGACTTGGGCAGCACACTGAGTGTGGAACCCCACAGGACTCCTTAGGGAAGGAGCTTGTGTGTGAAAGAACCCCTGGGGCTGAGCTGGTGACCTCCATGTGTGGGTGCAGCAGGGCCTTGGATGGTGCCAATTGATCTGGACAGCCTGTTGATGCTTTTCTACTTCCACCCTTCGGCCTGGCCCCACTGAGCCCCATCAAGGTGCCTGAAGAGGGGGCCAGTGAGATCCCGTGGCCACAGGGACTCCAGAGGCATCTCTGCAGGAAGCACACCATGCCTTCCCTCCATGTTCCATCACGGCGCCACAATCTGTGTCCCTTAACTTCTCAGGGTCAAAGACAAAGGCAAGCGTTGCTGAATTTTCTCTTTAATGGCCATTGGAAGAGTCTGGTTCAGTTTTTCCCAACTCTCTTCCCACTCGTATTTGGGGCCTCTGGGTTTTCTGAAGGCACAAGGACTGTGACTTTGTACCACTAACCTGTGGTTCAAACCTGGGTGTGTTTCTGGCATCTCCCTAACCCAGATCAGCAATGGCCACCCTGCTCCTCTGAGGTCAGCAGAAGACTGGGAAGCAGAGGTGAGGGATGCAGGCCACACTGGAATGGGAAGTCTTTTCCCCACTGGATGTGCCTGTCTGGTGGGTTTTGGACCTTCCCAGACTACATCTCTAGGAGAGGCCTGTTTGGAGTAGTACACTGAGA GACCCTGGCCTCTTCTGCTGGAAGACTGTCCAAGTCTTGGGGTTTCTTGAGCTGG TGATTCCTCTGCCATCCTGCTCTCTCTCTTCATCTCAGGCAGCATGGGGTCCACTT TTGTCCCCAAACCTAATGTTTTAATCCAAATGCAAATTGGTTCCACATTTTTACTG GAGGGTAATCAGTTTTTAAAATTCACACACATGTAATCGAAACGCAATACTATAA AGTTCTACTGGTGAGACTCCTGAAAAAAAAAAAAAAAAAA

[0188] In some embodiments, the RNA interference molecule targets MORC2. In some embodiments, the RNA interference molecule targets MORCs and comprises the sequence GCAGTTTCTGATGAGGAAGAA (SEQ ID NO: 826).

[0189] In some embodiments, the RNA interference molecule comprises a sequence having at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 826.SMC5 / 6 Complex

[0190] In some embodiments, the RNA interference molecule targets one or more proteins from the SMC5 / 6 complex.

[0191] Human cells express 3 conformationally similar structural maintenance of chromosome (SMC) complexes called cohesin, condensin and the SMC5 / 6 complex (Aragon L. The Smc5 / 6 Complex: New and Old Functions of the Enigmatic Long-Distance Relative. Annu Rev Genet. 2018;52:89-107). In addition to SMC5 and SMC6, the SMC5 / 6 complex contains 6 other proteins: nonstructural maintenance of chromosomes element 1 through 4 (NSMCE1-4A) and SMC5 / 6 localization factors 1 and 2 (SLF1 and 2).

[0192] The SMC5 / 6 complex plays an important role in chromosomal DNA repair, including repair of double-stranded DNA breaks via homologous recombination, and maintenance of replication fork stability. The SMC5 / 6 has also been shown be involved in silencing of unintegrated HIV-1 DNA (Dupont et al., "‘The SMC5 / 6 Complex Compacts and Silences Unintegrated HIV-1 DNA and is antagonized by Vpr,” Cell Host Microbe 29(5):792-805 (2021). By compacting viral chromatin, the SMC5 / 6 complex creates a repressive chromatin structure which therefore silences viral gene expression. By degrading SLF2, HIV-1 Vpr prevents recruitment of the SMC5 / 6 complex to unintegrated viral genomes and antagonizes this silencing. Therefore, the SMC5 / 6 complex was identified as a target for RNA interference in the vectors described herein.

[0193] In some embodiments, the RNA interference molecule that targets one or more proteins of the SMC5 / 6 complex targets SMC5, SMC6, NSMCE1, NSMCE2, NSMCE3. NSMCE4A, and / or SLF2. In some embodiments, the RNA interference molecule targets SMC5. In some embodiments, the RNA interference molecule targets SMC6. In some embodiments, the RNA interference molecule targets NSMCE1. In some embodiments, the RNA interference molecule targets NSMCE2. In some embodiments, the RNA interference molecule targets NSMCE3. In some embodiments, the RNA interference molecule targets NSMCE4A. In some embodiments, the RNA interference molecule targets SLF2.

[0194] In some embodiments, the RNA interference molecule targets SMC5. In some embodiments, the RNA interference molecule targets a region within SMC5 (RefSeq NM_015110.4) having the nucleotide sequence as shown as SEQ ID NO: 47 below:AGTTCGCGCGGGAGCGGGGCGCCTGGGTGGATGGGCGCTTGGGCGCCTGGGCTG CCGGACGGTGGGAACGGAAGTCGCTGTGGGACGCTGAGGAAGCCAGGATGGCG ACTCCGAGCAAGAAGACGTCAACTCCAAGCCCCCAGCCTTCCAAGAGAGCTCTC CCGAGAGACCCTTCGTCGGAGGTCCCGAGCAAGAGGAAGAATTCGGCCCCGCAG CTGCCGCTGTTGCAGTCGTCCGGGCCTTTCGTGGAAGGCTCTATCGTCCGCATCT CGATGGAGAACTTCCTAACATATGATATTTGTGAAGTATCTCCTGGACCCCACTT GAATATGATCGTTGGAGCCAATGGAACAGGGAAGTCGAGCATTGTGTGTGCCAT TTGCCTTGGTTTAGCTGGAAAACCTGCTTTCATGGGACGAGCAGATAAGGTTGGGTTTTTTGTGAAGAGAGGATGTTCTAGAGGCATGGTTGAAATTGAATTGTTCAGGGCTTCTGGAAATCTTGTAATCACCCGTGAGATTGATGTGGCAAAAAATCAGTCCTT TTGGTTCATCAACAAAAAATCTACAACCCAGAAAATAGTGGAAGAGAAAGTTGC AGCCTTAAATATTCAAGTGGGGAATCTTTGCCAGTTTCTCCCTCAGGACAAAGTT GGAGAATTTGCTAAACTCAGCAAAATTGAACTCCTCGAAGCCACTGAAAAGTCA ATTGGTCCCCCAGAAATGCACAAATATCACTGTGAACTCAAAAACTTAAGGGAG AAAGAAAAACAGCTCGAGACCTCATGCAAAGAGAAAACTGAGTATCTACAGAA AATGGTTCAGAGGAATGAAAGATATAAACAAGATGTGGAGAGGTTCTATGAACG GAAGCGACATTTAGATTTAATTGAGATGCTTGAAGCAAAAAGGCCATGGGTGGA ATATGAAAATGTTCGTCAGGAATATGAAGAAGTAAAACTAGTTCGTGACCGAGT GAAGGAAGAGGTCAGAAAACTTAAAGAAGGGCAGATTCCTGTAACATGTCGAAT TGAAGAAATGGAAAACGAGCGTCACAATTTGGAGGCTCGAATCAAAGAAAAGG CAACAGATATTAAGGAGGCATCTCAAAAATGCAAACAGAAGCAAGATGTTATAG AAAGGAAAGATAAACATATTGAGGAACTTCAGCAGGCTTTAATAGTAAAGCAAA ATGAAGAGCTTGACCGACAGAGGAGAATAGGTAATACCCGCAAAATGATAGAG GATTTGCAAAATGAACTAAAGACCACGGAAAACTGCGAGAATCTTCAGCCCCAG ATTGATGCCATTACAAATGATCTGAGACGGATTCAGGATGAAAAGGCATTATGT GAAGGCGAAATAATTGATAAGCGAAGAGAGAGGGAAACTCTAGAGAAGGAGAAAAAGAGTGTGGACGATCATATTGTACGTTTTGACAATCTTATGAATCAGAAGGAA GATAAGCTAAGACAGAGATTCCGTGACACGTATGATGCTGTTTTATGGCTAAGAA ATAACAGAGACAAATTTAAACAAAGAGTCTGTGAGCCCATAATGCTCACGATCA ATATGAAAGATAATAAAAATGCCAAATATATTGAAAATCATATTCCATCAAATG ACTTAAGAGCCTTTGTATTTGAAAGTCAAGAAGATATGGAGGTTTTCCTCAAAGA GGTTCGTGACAATAAAAAATTAAGAGTAAATGCTGTTATTGCTCCCAAGAGTTCA TATGCAGACAAAGCACCTTCAAGATCTTTGAATGAACTTAAACAATACGGATTTT TCTCTTATTTGAGAGAATTATTTGATGCACCTGATCCTGTAATGAGTTACCTTTGC TGTCAGTATCATATTCATGAAGTTCCTGTAGGAACTGAAAAGACCAGAGAAAGA ATTGAACGGGTAATACAAGAAACCCGATTAAAACAGATTTATACAGCAGAAGAA AAGTATGTGGTGAAAACTTCTTTTTATTCAAACAAAGTTATTTCTAGTAACACAT CTCTAAAAGTAGCGCAGTTTCTCACTGTCACTGTGGACCTAGAGCAGAGAAGAC ACTTAGAAGAACAGCTAAAGGAAATTCATAGAAAATTGCAAGCAGTGGATTCAG GGTTGATTGCCTTACGTGAAACAAGCAAACATCTGGAGCACAAAGACAATGAAC TTAGACAAAAGAAGAAGGAGCTTCTTGAGAGAAAAACCAAGAAAAGACAACTG GAACAAAAAATCAGTTCCAAACTAGGAAGTTTAAAGCTGATGGAACAGGATACT TGCAATCTTGAAGAGGAAGAGCGAAAAGCAAGTACCAAAATCAAAGAAATAAA TGTTCAAAAAGCGAAACTTGTTACCGAATTAACAAACCTAATAAAGATTTGTACT TCTTTGCATATACAAAAAGTAGATTTAATTCTCCAAAATACTACAGTGATCTCTG AGAAGAACAAATTAGAATCAGATTATATGGCCGCATCTTCACAACTCCGTCTTAC AGAGCAACATTTCATTGAATTGGATGAAAATAGACAGAGATTATTGCAGAAATG CAAGGAACTTATGAAAAGAGCTAGGCAAGTATGTAACCTGGGTGCAGAGCAGACTCTTCCTCAAGAATACCAGACACAAGTACCCACCATTCCAAATGGACACAACTCC TC ACTCC CC ATGGTTTTCC AAGAC CTTC C AAAC AC ATTGGATGAAATTGATGCTT TATTAACTGAAGAAAGATCAAGAGCTTCCTGCTTCACGGGACTGAATCCTACAAT TGTTCAGGAATATACAAAAAGAGAAGAAGAAATAGAACAGTTAACTGAGGAACT AAAGGGAAAGAAAGTTGAACTAGATCAATACAGGGAAAACATTTCACAGGTAA AAGAAAGGTGGCTTAATCCTTTAAAAGAGCTGGTAGAAAAAATTAATGAAAAAT TCAGCAATTTTTTTAGTTCCATGCAGTGTGCTGGTGAAGTTGATCTCCATACAGA AAATGAGGAAGATTATGATAAATATGGAATTCGAATTAGAGTCAAATTTCGAAG TAGTACTCAACTGCATGAATTAACTCCTCATCATCAAAGTGGAGGTGAAAGAAG TGTTTCTACCATGTTATACTTGATGGCACTTCAGGAGCTAAATAGATGTCCATTCA GAGTAGTTGATGAAATCAATCAGGGAATGGACCCAATCAATGAACGGAGAGTGT TTGAAATGGTTGTAAATACTGCCTGTAAAGAAAATACATCTCAATACTTTTTCAT AACACCAAAGCTCCTGCAAAATCTTCCTTATTCTGAAAAGATGACAGTTTTGTTT GTCTACAATGGCCCTCATATGCTGGAACCAAACACATGGAATTTAAAGGCTTTCC AAAGGCGGCGGCGCCGTATTACATTCACTCAACCTTCTTAATAAAAGTAAAGAG AGGGAACTTGGGAATTTTTTTTGTTAAATTCTGTTTATAAGTATGGCTCAACTGAATAAAAGGAGATTCACTAAAACGAAAAGCAGTTATTTTTGGAAACCTGCTTTTAAATACAAATAGGTTGATAATGGAAACTATAATGACCTTTCCAAAATAGCAGCTGGTAGTAAAAGTTAAGTCTTCTTCAGTCTTGGTTGAACTTGAGTTCTTGGCACTCTGACCATGAGTCATTCAGTTCTCATGTTAAAATGTACTTAATATTACAAATCAAAGGTACAGTGGAAGAAGGGTTAATCACAAGAAGTTACTTATATGGTAGCCCTGAGCTTTAATTGCAGAGTAACTTTAATTACTTTTAGAGCCTAAAGATGACTCTAGAGCCTAAGTCCTAGTTTCTCCCAATGTTATATTTAATTTTAAAAAATTGATATGAAAATGTCTAATGTATAGTAATAATTTATGACAGATCTAGTCATTTCTTCCTATTAAAAAAGATTACCTTATCTCCAGTAGGAAATGGAATTTTATGGGCCTTTAAAAGAAAGTTTTATGAAACTTGATGCTATAATTTTATTGGTATTTCAAGGGGAAAAAAGCACTGGGGTTCAAAAATGGTAGCAGAACTGCTTTGAAATGCTGCAAGGTGGCCACTAGATGATGCAAAATACAACCAAAAGATTGACTGAGAATAAAATTAGGTGACAAGGGTTTTTAAAGAATAACCTTTTAAAGTGTGGGGGCAGGGGTTGCTTTTTTTTATTTTATTTAAAGTCAATTATATTTTACATCTTACATTTCTAAAAGCATTTTATAATTATTTTTAGTAAGATTTTTCTTAAAATTTCATATACTGGTTTCTACAATTTATATTTGAAATTTCTCAGTGTTATGTAAAGAGTGATGGAAAAGCATTGATTTCTTTAAAACCGTAATGTTTTTAGAACTTAAGCCTATAGGGCCTTTCTTACAATGTTGATGTACCCATTATCTTAGAAAATCTAGTTTAAACTGTTTTCTTTCACCGCAAAAGAATTAAATGGGAAAATCATTTGTTTATCTCTAAGTTATACTAATTAGTAGAACCAAACAAATTATCTTCTTTTAAAAAATAAATCTTATAGGAAAATAGACAGTCCAAAGTCATGTCTTTGAACAGTGGATTGGATCTGTGCCAGTAATGACAAAATTATTTTTTTGACTTGCTTGCCTGAATAAATTGAAGAATTGCTTTCAGTTTGGGTTTTGTATATTCTTAAGTAGCCATTGAAATTTATATTCTTAACTAGGTCAAAAAATAATGAGCCATAAGTTTATGTCCTCTCACTTAGACATTTTCTCTTTAAAAAGGTATTTTCTTCTTTATAAACATTTTAAAAGAGCCTTCCCTTCTTAAACTAACTCCAGTGCATGAAGTGTGAAAATATTTTAAAATGACATTTTTACTAATATGAGCAAGTCATGTAAACATTGAAGAACTTGGTAACATATTAGTAAATGGATATTACCAAATGTTTTCATCGTTAATTACTTTGCGTTCCACCAAAATATCTTTACTAAAATGTGCTTGGTGTAGTTTGTTTATTGTCTAAATTAGTACCAGTCATCTTATTTCTGCAAAATGAGTATCAATGTGAAAAAGACACGTGAAGATTAAGCATGTTTGAAAATAAAATGGTCAATTACATTTCAATTTACATAGGCCAACAACTGTTCCATACTTTGTTTGTAAACATTTAATTTCTCTACTGGACAAAATTAATATTTGGCTTTACATTGAATTTTGAGCTGTGAAGAATAAATTATGTATCATTTTAGCATATTAAACAGTAGTAAGTCTAGCACATAGTCTCAGCCACTTAAAACAAAAGTTTTTTTGTTTGTTTGTTTGTTTGTTTTTTTGAGATGGAGTCTCACTCTGTTGCCCAGGCTGGAGTGCAGTGGCGTGATCTCGGCTTACTGCAACCTCCGCCTCCCGGGTTCAAGCGATTCTCCTGCCTCAGCCTCCCAAGTAACTGGGACAACAGGCGCGTCCCACCACACCCAGCTAATTTTTTATACTTTTAGTAGAGATGGGGTTTCAGCATATTGGCCAGGCTGGTCTCGAACTCCTGACCTTGTGATCCACCCGCCTCGGCCTCCCAAAGTGCTGGGATTATAGGCGTGAGCCCCTGCACCCGGCCAAAAGTTGATTTTTAATTACATAAAAATCG TAAAAACTTCTAGTAAAAACTTGATTTGGTGAATACAGTTATATTTAAAAACCTT AAGGTGACAAGCATTTTCTATGCCTAAATCTTCATTGGTTTGCCTGGAAAGAGTCTCTGTTAAAAGATTTTCCATATTCAAAGTAAAAGGAAAGATTTCTTGCTTTCTAAT TGTCTTTTGGACACATGCCTATTTTCTTTGAGGTATAAACCTTTAGATGTGAAAAA TGTAATTTCATTCTGCTATTGTGTGTGCTTGTGTGTGTGTAATTGAAAAAACTGGGAAATCCTGCTTTGTTGGTAATAAATCAATATTTTTATATTC

[0195] In some embodiments, the RNA interference molecule targets SMC6. In some embodiments, the RNA interference molecule targets a region within SMC6 (RefSeq NM_001142286.2) having the nucleotide sequence shown as SEQ ID NO: 48 below:GGTTAGTACCGCGGTGGGCGCCGGGGCTCCCGGGAATCTACCTTCTCCTGCGGCC GGCACGCGGTTCCCAGGGGGCCAGCGGCGGTCAGCCGAGGTCGAGACGCCCGCA GGGTGGCCTTAGCGGCCGGTCGTACCACGGCAGCCCCGCCGATCAGGTTCCTTTGGGAGACTTCGACTTGTTGGCGAAATGAACCGGAGAAGAATCCCAATTGGGAATT GCGGAAAACAGGACTCTAGGGTAGAGAAAGGTTGTAGAACCAATAGGGTTTGAG ACCTGATGGCCAAAAGAAAGGAAGAAAATTTTTCCTCTCCTAAAAATGCCAAAAGGCCAAGACAAGAAGAATTGGAGGATTTTGATAAAGATGGTGACGAAGACGAAT GTAAAGGTACTACTTTGACTGCAGCAGAAGTTGGAATAATTGAGAGTATTCACCT AAAAAACTTCATGTGTCATTCAATGCTTGGACCTTTTAAGTTTGGTTCTAATGTCA ACTTTGTTGTTGGCAACAATGGAAGTGGGAAGAGTGCAGTACTCACAGCTCTCAT AGTCGGTCTTGGTGGAAGAGCAGTTGCTACTAATAGAGGATCCTCTTTAAAAGGT TTTGTGAAAGATGGACAGAACTCTGCAGATATCTCAATAACATTGAGGAACAGA GGAGATGATGCCTTTAAAGCCAGTGTGTATGGTAACTCTATACTTATACAGCAAC ACATCAGCATAGATGGAAGTCGATCTTATAAACTTAAAAGTGCAACAGGCTCCG TGGTTTCCACGAGGAAAGAAGAGCTGATTGCAATTCTTGATCATTTTAACATCCA GGTGGATAATCCAGTTTCTGTTTTAACACAAGAAATGAGCAAGCAGTTCTTACAG TCTAAAAATGAAGGAGACAAATACAAATTCTTCATGAAAGCAACGCAACTTGAA CAGATGAAGGAAGATTATTCATACATTATGGAAACGAAAGAAAGAACAAAGGA GCAGATACATCAAGGAGAAGAGCGGCTTACTGAACTAAAGCGCCAGTGTGTAGAGAAAGAGGAACGTTTTCAAAGTATTGCTGGTTTAAGTACAATGAAGACTAATTTA GAGTCCTTGAAACATGAAATGGCTTGGGCAGTGGTCAATGAAATTGAAAAACAA TTGAATGCCATCAGAGATAATATCAAAATTGGAGAAGATCGTGCTGCTAGACTTGACAGGAAAATGGAAGAACAGCAGGTCAGACTTAATGAGGCAGAACAAAAGTAC AAGGATATTCAAGACAAACTAGAAAAGATTAGTGAAGAGACAAATGCACGAGCACCAGAATGTATGGCATTGAAAGCAGATGTTGTTGCTAAGAAAAGGGCCTATAA TGAAGCTGAGGTTTTATATAACCGATCCTTAAACGAATATAAAGCATTAAAGAA AGATGATGAGCAGCTTTGTAAACGAATTGAAGAGCTGAAAAAAAGTACTGACCA ATCTTTGGAACCTGAACGGTTGGAAAGACAAAAAAAAATATCTTGGTTAAAAGA GAGAGTAAAGGC CTTTC AAAATC AAGAAAATTC AGTC AATC AAGAGATC GAAC A GTTTCAGCAAGCCATAGAAAAGGACAAAGAAGAACATGGCAAAATTAAGAGAG AAGAATTAGATGTGAAGCATGCACTGAGCTACAATCAGAGGCAACTGAAAGAAT TGAAAGATAGTAAAACTGATCGACTCAAAAGATTTGGCCCTAATGTTCCAGCTCT TCTTGAAGCCATAGATGATGCTTATAGACAAGGACATTTTACCTATAAACCTGTA GGCCCTTTAGGAGCTTGCATTCATCTTCGGGACCCAGAACTTGCTTTGGCTATTG AATCTTGCTTAAAAGGGCTTCTGCAGGCCTATTGTTGCCATAATCATGCTGATGA AAGGGTCCTTCAGGCACTCATGAAAAGGTTTTATTTACCAGGGACCTCACGGCCA CCGATAATAGTTTCTGAGTTTCGGAATGAGATATATGATGTAAGACACAGAGCTG CTTATCATCCAGACTTTCCAACAGTTCTGACAGCTTTAGAAATAGATAATGCGGT TGTGGCAAATAGCCTAATTGACATGAGAGGCATAGAGACAGTGCTACTAATCAA AAATAATTCTGTAGCTCGTGCAGTAATGCAGTCCCAAAAGCCACCCAAAAATTGT AGAGAAGCTTTTACTGCTGATGGTGATCAAGTTTTTGCAGGACGTTATTATTCAT CTGAAAATACAAGACCTAAGTTCCTAAGCAGAGATGTGGATTCTGAAATAAGTG ACTTGGAGAATGAGGTTGAAAATAAGACGGCCCAGATATTAAATCTTCAGCAAC ATTTATCTGCCCTTGAAAAAGATATTAAACACAATGAGGAACTTCTTAAAAGGTG CCAACTACATTATAAAGAACTAAAGATGAAAATAAGAAAAAATATTTCTGAAAT TCGGGAACTTGAGAACATAGAAGAACACCAGTCTGTAGATATTGCAACTTTGGA AGATGAAGCTCAGGAAAATAAAAGCAAAATGAAAATGGTTGAGGAACATATGG AGCAACAAAAAGAAAATATGGAGCATCTTAAAAGTCTGAAAATAGAAGCAGAA AATAAGTATGATGCAATTAAATTCAAAATTAATCAACTATCGGAGCTAGCAGAC CCACTTAAGGATGAATTAAACCTTGCTGATTCTGAAGTGGATAACCAAAAACGA GGGAAACGACATTATGAAGAAAAACAAAAAGAACACTTGGATACCTTAAATAA AAAGAAACGAGAACTGGATATGAAAGAGAAAGAACTAGAGGAGAAAATGTCAC AAGCAAGACAAATCTGCCCAGAGCGTATAGAAGTAGAAAAATCTGCATCAATTC TGGACAAAGAAATTAATCGATTAAGGCAGAAGATACAGGCAGAACATGCTAGTC ATGGAGATCGAGAGGAAATAATGAGGCAGTACCAAGAAGCAAGAGAGACCTATCTTGATCTGGATAGTAAAGTGAGGACTTTAAAAAAGTTTATTAAATTACTGGGAG AAATCATGGAGCACAGATTCAAGACATATCAACAATTTAGAAGGTGTTTGACTTT ACGATGCAAATTATACTTTGACAACTTACTATCTCAGCGGGCCTATTGTGGAAAAATGAATTTTGACCACAAGAATGAAACTCTAAGTATATCAGTTCAGCCTGGAGAAGGAAATAAAGCTGCTTTCAATGACATGAGAGCCTTGTCTGGAGGTGAACGTTCTTTCTCCACAGTGTGTTTTATTCTTTCCCTGTGGTCCATCGCAGAATCTCCTTTCAGATGCCTGGATGAATTTGATGTCTACATGGATATGGTTAATAGGAGAATTGCCATGGACTTGATACTGAAGATGGCAGATTCCCAGCGTTTTAGACAGTTTATCTTGCTCACACCTCAAAGCATGAGTTCACTTCCATCCAGTAAACTGATAAGAATTCTCCGAATGTCTGATCCTGAAAGAGGACAAACTACATTGCCTTTCAGACCTGTGACTCAAGAAGAAGATGATGACCAAAGGTGATTTGTAACTTAACATGCCTTGTCCTGATGTTGAAGGATTTGTGAAGGGAAAAAAAATTCTGGACTCTTTGATATAATAAAATGAGACTGGAGGCATTCTGAAATGAAAGAAACTCCTTTATATATCCAACCACAATCAAACATATAAATAAGCCTGGAAAACCAACTACAACCTGCAATTTAAGATTACTATTACTTTAAGAAAATCAATTTCATAGTATTGGTTTTAAATCTTTTTAAGTTTTTTTAATACGATCTATTTTTATAGGTTCTTTTTCAGAAGTAAAATTTTGTACATATATACATGTACATATCTGTTTAGTTTGGGTTCATTTCTATAACATTTTGTAAGAAAATAAAAGTTTGAGCACCTGATTATATTTAGTTTTGCTTTTCCAGATATTACATTCTATAGTTACCAAAAATGGTTGAAGGGAGGGATTTCTCATTGCAGAGGGTGGGGTGCAAGGGAATAAGACACTTGTACGGAACACTGAAGCTTTGCCAACTTCTACACATGCCTTTTTTGCAGTCCTTTAACTGTCCACCCTACCAAGAGCTTATAACCAGTATCAGAACTGGATAATGACGCAGTTTTTCACTCTGACCTCCATCATGCTTGCCTGATTTAAAAGCCCTCAGTTTGCAGTCCAGGGACTGTTCAGGCTTGTCCTCAGCTGAGAGGACACAGGCTAGAGGGACTGTGCAGAACCAGGCTGGGAGAAGGGCTGGGAAAACTGGGAGTGGAGGGTGGATCCTCATGGAGCAGGAGAGTAGCTCATGGCTCCAGGAGCCTGAGGCCATGCAGTTGATGGTGAGCTGACATCAATTCTAAGACTCATCCTAATTGAGGGGTGTTAAAAAGTGTGCTGCTTAGAATGACCAAATATAGTTATTGTAAAAAATGATATTTATGAACTTTTTATTTTAGAAAACATGAATTTTATTGCTCCCTGTATTATTTGTTTGATACTAGGATTCATGCTAAACTTTTTAAGAATGTATTGGATATCAAGAAGCATTCCTTACATTAGTAGCAATAAATATTAGAATAAATATGAAATTGAACTATTTTCAGAAAAAGGGCAGTATATTAAGAGCAGGGACTGTTCTCTAGTTATTGAGGAAAACTGGACTTTGTTTGTGTTTTTGGTGGAGGAAGAAGTTTAAGATACTTTAGTCTTAAATTGAGGTTTGCCAAATGAGAAGTTCAAAAACTTGGGCTTTCTAATCAGAATTTCCAGGAGGAGGAAAGTGTGTGCTGAATATTTTAAACATTTCCCACTGATCATACAAAGTCTGATTTTTAAATTTACACTTATAATGCCTTTGTATTAAAATTATTTTTAACATGTGCTTTTCCAAATTAAAAATGAAGTAGAGTATACCAAATGCATAAACTTTCATTAGCTAAGGAACTCATGTCTGAATTTTGTTGTAGTTTTGAATGTTGTGCTCTTTCATACAGAATGGGAAACATAATCCTCAGGTATCCCAGCATCTCTTGTTGAATTGAAGATTAT TCATTGCTTTGGCCTCACAAAGTTTTGATTTCAACTATCATAAGTGAAAATATCTT CCTTTAATGTTCTAAGTAGTGATAATATTACTAGAATGAAAGAATAAAAAGAAAT TGTTCTTTTAAAATATGTGTAACTTCTAAAAATAAAACTTAAAATTTATATGTA

[0196] In some embodiments, the RNA interference molecule targets NSMCE1. In some embodiments, the RNA interference molecule targets a region within NSMCE1 (RefSeq NM_145080.4) having the nucleotide sequence shown as SEQ ID NO: 49 below:AAAGGAAGAAGAGTCCACCTTGCGACCGTATCCGCTAGCGCGGCCTGGGATGCG CTTGGGCTCCCTGTTCGTTCCCACATGCAGGGCAGCACAAGGAGAATGGGCGTC ATGACTGATGTCCACCGGCGCTTCCTCCAGTTGCTGATGACCCATGGCGTGCTAG AGGAATGGGACGTGAAGCGCTTGCAGACGCACTGCTACAAGGTCCATGACCGCAATGCCACCGTAGATAAGTTGGAGGACTTCATCAACAACATTAACAGTGTCTTGGA GTCCTTGTATATTGAGATAAAGAGAGGAGTCACGGAAGATGATGGGAGACCCAT TTATGCGTTGGTGAATCTTGCTACAACTTCAATTTCCAAAATGGCTACGGATTTTG CAGAGAATGAACTGGATTTGTTTAGAAAGGCTCTGGAACTGATTATTGACTCAGA AACCGGCTTTGCGTCTTCCACAAACATATTGAACCTGGTTGATCAACTTAAAGGC AAGAAGATGAGGAAGAAGGAAGCGGAGCAGGTGCTGCAGAAGTTTGTTCAAAA CAAGTGGCTGATTGAGAAGGAAGGGGAGTTCACCCTGCACGGCCGGGCCATCCT GGAGATGGAGCAATACATCCGGGAGACGTACCCCGACGCGGTGAAGATCTGCAA TATCTGTCACAGCCTCCTCATCCAGGGTCAAAGCTGCGAAACCTGTGGGATCAGG ATGCACTTACCCTGCGTGGCCAAGTACTTCCAGTCGAATGCTGAACCGCGCTGCC CCCACTGCAACGACTACTGGCCCCACGAGATCCCAAAAGTCTTCGACCCTGAGA AGGAGAGGGAGTCTGGTGTCTTGAAATCGAACAAAAAGTCCCTGCGGTCCAGGC AGCATTAGCCATCGTGCCCTGCTGAGGGGCTGGCTGCCTTGAGTGGCCTGATCGC CACAGCCCTTCTTGGAAGAAAGGCGTCCGTGTTTCAGGTTCCACGCGAGTCACCT CTTTCGTCTTAATGTTCACCGTCCACAGCTTTGGAATAAACCATCCTGGGAAGTT GCTGCA

[0197] In some embodiments, the RNA interference molecule targets NSMCE2. In some embodiments, the RNA interference molecule targets a region within NSMCE2 (RefSeq NM_001349486.2) having the nucleotide sequence shown as SEQ ID NO: 50 below:CTCACTTTTCAGCGGCAGGCGAAGGGGGCTGAGGAAAGGAGGTGGGTCTAGGCA GGGGAAATTGGGGTGCCACCAGACGGAGACAGCTTGGACTACCAGGCTTTTAAG AACTGTGCTATTGTTGACTTCTTCACAGAATCAAGCACTCTTTTGGAAGAGGGTA ATCTCTCTCCAAAAACTGAGGACACTTACCTTCCCCATATATTGAGTCCAGCTGTGTTTGGTGGCCCAGGTACTAATTTCAAGATGCCAGGACGTTCCAGTTCAAATTCA GGTTCAACTGGTTTCATCTCCTTCAGTGGTGTAGAGTCTGCTCTCTCCTCCTTGAA AAACTTCCAAGCCTGTATCAACTCTGGTATGGACACAGCTTCTAGTGTTGCTTTG GATCTTGTGGAAAGTCAGACTGAAGTGAGTAGTGAATATAGTATGGACAAGGCA ATGGTTGAATTTGCTACATTGGATCGGCAACTAAACCATTATGTAAAGGCTGTTCAATCTACAATAAATCATGTGAAAGAAGAACGTCCAGAAAAAATACCAGATTTAA AATTATTGGTAGAGAAGAAATTTTTGGCTTTACAGAGCAAGAATTCTGATGCAGA CTTTCAAAATAATGAAAAATTTGTACAGTTTAAACAACAGCTGAAAGAACTAAAGAAGCAATGTGGTCTTCAAGCTGACAGAGAAGCTGACGGAACAGAAGGAGTGG ATGAAGATATAATTGTGACCCAAAGTCAGACCAACTTCACCTGCCCCATTACAAA GGAGGAAATGAAGAAGCCAGTGAAAAATAAAGTGTGTGGCCACACCTATGAAGAGGACGCCATTGTTCGCATGATTGAGTCCAGGCAAAAGCGGAAGAAAAAGGCCT ATTGCCCTCAAATTGGCTGTAGCCACACGGATATAAGAAAGTCAGATCTTATCCA GGATGAAGCACTTAGAAGGGCAATTGAGAACCATAACAAGAAAAGACATCGTCATTCCGAGTAGGAAAAGCCACCTGCCTGCAGGGACACCAGCAGCCTACCTCCTA CCCCAGCTGTCTGTTGAGAGCAGTGCTGACCCCAGCAGTTAGGGACTGGCTGCAT AGCATACTTGTTGGGGGTAAAACTTGTTGCTTTTATGTGTGCTTGAAAACATTTTTCAAAGTTACACAACAGAAATGCAATCATATTGTTTATTTTTAAGTGTTCTATAAT GTTAAATAAAACTTTGATCATCTGCA

[0198] In some embodiments, the RNA interference molecule targets NSMCE3. In some embodiments, the RNA interference molecule targets a region within NSMCE3 (RefSeq NM_138704.4) having the nucleotide sequence shown as SEQ ID NO: 51 below:AGTCTCAGCCGACACTGCGCGCGCCTCCAGGCACCGGCGTTAGCGGGTCGCCGA CCCGCAATCCCCGCCGCGGCTGCTTGCCTACCGGAGTGTGCGCCGGCACCTGCCG CCGGAGACATGTTGCAAAAACCGAGGAACCGGGGCCGCTCTGGCGGCCAGGCCG AGAGGGACAGAGACTGGAGCCATAGCGGAAACCCCGGGGCTTCGCGGGCCGGG GAAGACGCCCGGGTTCTCAGAGACGGCTTTGCCGAGGAGGCCCCGAGCACGTCC CGCGGGCCGGGCGGCTCGCAGGGGTCGCAGGGCCCCTCGCCTCAGGGCGCCCGC CGGGCCCAGGCCGCCCCCGCCGTGGGGCCCAGGAGCCAGAAGCAGCTGGAGCTG AAAGTGTCCGAGCTGGTGCAGTTCTTGCTGATTAAAGACCAGAAGAAGATTCCG ATCAAGCGGGCCGACATACTGAAGCACGTCATCGGGGACTACAAGGACATCTTC CCCGACCTCTTCAAACGGGCCGCCGAGCGCCTCCAGTACGTCTTCGGGTATAAGCTGGTGGAACTTGAACCCAAGAGCAACACTTACATCCTCATCAACACCCTGGAGC CTGTGGAGGAGGATGCCGAGATGAGGGGTGACCAAGGCACGCCCACTACGGGCCTCCTGATGATCGTCTTAGGGCTCATCTTTATGAAGGGCAACACCATCAAGGAAACTGAAGCCTGGGACTTTCTGCGGCGCTTAGGGGTCTACCCCACCAAGAAGCATTTAATTTTCGGAGATCCAAAGAAACTCATTACTGAGGACTTTGTGCGACAGCGTTACCTGGAATACCGGCGGATACCCCACACCGACCCCGTCGACTACGAATTCCAGTGGGGCCCGCGAACCAACCTGGAAACCAGCAAGATGAAAGTTCTTAAGTTTGTGGCCAAGGTCCATAATCAAGACCCCAAGGACTGGCCAGCGCAGTACTGTGAGGCTTTGGCAGATGAGGAGAACAGGGCCAGACCTCAGCCTAGTGGCCCAGCTCCATCCTCTTGAAAGGTGGATTCAGAGGGACCCCCGGGACAAGGGTCTGAGACCCAAAGGCACAGTTTAGAGGATTGGGGGAAGGGAGAACGAACCCAGGGAGCATATTGCTGTAAACGCTTCAATGTGTGTAGCTTTAGGATGTGTTTGCAAAGTTTTGTTTTTTTAATGTTGTGTTATTTTGCTCCAGATTTTCATCTATAAACAAAGGAGCATTTGTTTTGATTTTACTCTTTTTGGTATAAAAAATTTTGCTAGCTTAGTAAAACGAATTGGAAAACTTGACTATGATCTGGAACAGATAATGCAAGAAGGAACACATAAGTAAGTTGCTTTGGTGCCAAGAAAATAAAAAAGCTATTATCAGGTCTCCTAACTACCCCAGTTTGTAAGGAAAAATAAAAGTTTTTATAAAATTAAAAAAAATAATTGCCTATATCCTTATTATGTTAACCTATTTTATTTTTCTATATTTCGTACATATATAAGTATTATTCATGGTTCCAAGCAATGAGTGTAATGTATTTTCTCCCAATAGTAATTGTTTATATTCATAATTATTCAGCTCTGCTGCTAAACACGTAGTTCCACTTCGTTTTTTTCTTCTTGCTACTGTAGTTATTACAATAAATACTGTATTTTTGAAGCACTTAAGCATTGTGTGTTTTTTAAATCGTGGGTCATGAGAGAATTTCATGACTGGAAATCAACATCTTAAAGTTTTTGCTACATATTAAGGTCAATGCTGTTTTGTAGAAGTTGTTTCACTTTTGTGTGAAACAAAATCAGGATCATGAGGTGAAATATATTTCTCACTGTGGATCTCAAAAAGGCTTATAAAACAATAGCGTAACCATTAACCATGCCTGTTTGGGAATCGGACTGTCTGGGTTGAGATCCTGGCCCCACCATAACTAGTATATGAATTTACATAGGTTCCTTAGTTTGTATGCAGATGTTTCTTGACCTGTAAAATGGTTTTAGTCCGTGATGTGGAGAATTTTATTTGTTATTCTTTAATGTTTTTATTTTTTATTCTCCTTGGTCTGGAGCTGCCACTTTTTAAATATAACTTTTCTTTATCTTTTTTTCCAAATCATTCCTAGGAGAGCAATTCCTGGTTCAAAAGATAGTCCTCTAGTCTCACTCTATTGTATGGGGAAATATAACGTGTATGGTGAGTGTGCCCCACATAGAACATAAGAGCAGGACATAAGATCTACCTCCTAGGGAGTTAGGTATAGGGAAAAGGGTATGGGTCTTGGGGCTGTCTGGCAAACTGAATTTGAATTACGGCTCATGCTAACTTGTCAGCATGGTGGTTGTGGGGGTAGGTTTTAAGCTCTCTGAATCTCAATTTATCTGTAAAATGAGAAAAATAATAGCTATATAATAGTTTTATGCACTAAATGAACTAGAAGAGTTCATAGTCCCATTGTGTTCCTCCATTACGTGTAGCAGATGCTGTTTTTAATGCAGAGATTCAACACGTTTTATTCAGTGGGCAAATAAGTTAATTGTTTTAGGGGGAGGTGCTGCAAATTAACCTTGAAATAAGGCAAATATTTTTAACTCGGGGAAAAAAGTGAACTTCATCAACACTGGACGGAATGTCATTTAAAAGAGTCTATAGTGAGTATGGAGGAATGTGATAGGTTACTGGGTTTTATTTCATTCTGTTGAATTGGGATTTGCATTTGTTTCATCAGTGGAATATGTTTGGCCCTTACAGGCATTTCTTCTTGATCTGCATTTTATTTCAGTGTATAAACTCCTTCCTGCCTCCTGGTCCCCTTACCTCCCTGACTCAGCTTTCTCCTCCTCATGCCCTCTGATACTCGCATTCTCACCTGTATAATAAAAATTATTACCCAGCAGTCACCACCAGTTATATCTGAGGCAAAATTCCCCCCGCTTTTCTCAGAGGTGTTAGCAATTTACCATCGATAATCTTGTTTCAAATTTTAAAATATGAAACATCAGAGGCCACATCAAGGCCTTATATCTTTTCCTTCTCACCCTCCCCAGAGGTAACAAAAGTCTGTGGCTATCGGGCTGTTGTATGACTTTATAATTTTACTATACATGTATATATACATAACCAATATAGAGTTTTATTTTGTGGGCTTTCATGATTTATATAAGCGGTGTTACGTGTGTCCTTATGCAACTTGCTGTACTCACTCACCATTATGTATTTGAGATTCAGCCATGTGGATATATGTAGACCTGATTCACTCATTTTAATTGTTCTTTTATGGGGGAAAAATAACTATCCATTCCTTAATGTCTGGACATTTAAGGTCTTTCCAGTTCTTTGAGATTTGCAGGCATTGTACAGTACTTCTTTAGGCACATGTGTGAGTTTCTCCAGCCAGAAACAAATGGAGCAATCAAGGACTATTGCCAAGCTGCTCTTGAGCTTGTGTCAAGTTACACTCCCATCAACATTTTATACAGTTTTGTTTTGTTTTTGTTTACCAATCCCTCCACCTTCACCCCCTGTGAACACTTGGCCTTTTCAGTATCTTTTCAGCCATTCTGGTACAGTGTGAAATGATAATTTATTACGGTATTAACTTATACCTGTTGCTTACTAGTAATGTTAAGTATCTGCCTATCTTTTGCTCTCACTCCTTTTTGAGTGTCAGATTTCATGTCTTAGACCTATTTCTATTGATTTGTAAGACATCTTTATATGTTCTACATGTTAATCCATGGTCTGCTCTATCTATCTAGTGCAAATACCTTCTCCCACATTTTCACTTCTTTTGGTCATACAGATCATTTGACCTAAATGTGTTACGATCATTAGTGTTTATAAGAGCTACATTTGTGTATAGTGCTTTAAAAAATTCCCCTAATCCTAAAATTGTAAATATGTTTTCTGCCTTTTTTCCCTAAGTGTTCAAAGTTTTGATCTTCACATTTTATGCTTTTAATCTGCTTTTGTGTATGATGTGAGGTATGGAAATAGTTTTTGTATTTTTCCTATATGGATAGCCTAGCACCAGTTTTTCATCATTTATACTTTCCACACTGATTTATGACACTCAGTATTTTGTAGCAATGTCCCATGTATAGATAAATTTATGGCTAGGACTGAATTGGTTTATTTGCTATCCATTTGCAAATACCATACTGTAATTCTTTTTTTCTTTTTGAGACGGAGTCTTGCTCTTGTCGCCCAGGCTGGAGTGCATTGGCACAATCTTGGCTCAGTGCAATCTCTGCCTCCTGGATTCAAGCGATTCTCCTGCCTCAGCCTCCTGAGTAGCTGGGATTACAGGTGCCCGCCACCACGTCCGGCTAATTTTTGTACTTTTAGTAGAGACGGGGTTTTGCCATGTTGGCCAGGGTGGTCTCAAACTCCTGACTTTAGGTAATCTGCCCGCCTCGGCCTCCTAAAGTGCTAGGATTACAGGCATGAGCCACCTCGCCCAGCCTA TAATTCTTTAGCATCATATGCCTGTCTTCTTTTTCAATTCTTAAAGCCTTTTATTTT AGTCTTTTTTGCATTGGCTGAAAATGCAATATATTATTGAAAAGAAGCAGTTACAGCAGGCTCTAAGGATGTTTCTGATTCCTTATTCTTGATTTTAAAGGAAATAATTTT AAGTTTTGCCATAGAGTGTGATGTTTGCTGTTAAGAGTTTTTTTCCCCTATAAATA CCTTTTATTGGGCTATTTAGTTACCCCTTCAGTCCTAGCTTGCTTAGAGATGTTCAGAAAAAAAACTATGAATGACTTTGGATGTATATATGCTCTTTATTTACCTGCTAA TATGATTAAAATGTCTTATTTACTCTGTAAA

[0199] In some embodiments, the RNA interference molecule targets NSMCE4A. In some embodiments, the RNA interference molecule targets a region within NSMCE4A (RefSeq NM_017615.3) having the nucleotide sequence shown as SEQ ID NO: 52 below:AGTTTCTGGCGCGAACTTCCGCCGTTCCGAAGTTGCACGGTGAATTGGCGCTATG TCTGGGGACAGCAGCGGCCGCGGGCCAGAGGGCCGGGGCCGGGGCCGCGACCC GCATCGGGATCGCACCCGCTCCCGCTCCCGCTCGCGGTCCCCTTTGTCGCCCAGGTCCCGCCGCGGCTCTGCGCGGGAGCGCAGAGAGGCCCCAGAGCGCCCGAGCCTG GAGGACACAGAGCCGTCGGATTCCGGGGACGAGATGATGGACCCGGCCAGCTTG GAGGCGGAGGCCGACCAAGGCCTGTGCCGCCAGATCCGCCATCAGTACCGGGCGCTCATCAACTCCGTCCAACAAAACCGTGAGGACATACTGAATGCCGGTGACAAA TTAACAGAGGTCCTTGAAGAGGCTAACACTCTGTTTAATGAAGTGTCCCGAGCAA GAGAAGCAGTCCTGGATGCCCACTTTCTTGTTTTGGCTTCAGATTTGGGCAAAGAGAAAGCAAAGCAGCTGCGCTCAGACCTGAGCTCCTTTGACATGTTAAGATATGTT GAAACTCTACTCACACATATGGGTGTAAATCCGCTAGAAGCTGAAGAACTCATC CGTGATGAAGATAGTCCTGATTTTGAATTCATAGTCTATGACTCCTGGAAGATAACAGGCAGAACAGCAGAAAACACCTTTAATAAAACCCATACATTCCACTTTCTGTTGGGTTCAATATACGGAGAGTGCCCTGTGCCAAAGCCACGAGTTGATCGTCCAAG AAAAGTTCCTGTGATACAAGAGGAGAGGGCAATGCCTGCCCAGTTAAGAAGAAT GGAAGAATCTCATCAAGAAGCAACAGAGAAAGAAGTAGAAAGAATCTTGGGATTGTTGCAGACATATTTTCGAGAAGATCCTGATACCCCAATGTCCTTCTTTGACTTT GTGGTTGATCCTCATTCTTTCCCCCGTACAGTGGAAAACATCTTTCATGTTTCCTT CATTATACGGGATGGTTTTGCAAGAATAAGACTTGACCAAGACCGACTGCCAGTAATAGAGCCTGTTAGTATTAATGAAGAAAATGAGGGATTTGAACATAACACACA AGTTAGAAATCAAGGAATTATAGCTTTGAGTTACCGTGACTGGGAGGAGATTGT GAAGACCTTTGAGATTTCAGAGCCTGTGATTACTCCAAGTCAGAGGCAGCAGAAGCCAAGTGCTTGATGCTAGCTGAAGGACTCAAATGGATAGTGAAGTCCAAAACGGAAAGCGGCATGTATCGTACATATTGTATGATTCAACATTTTTAAAGGCAGATTGTTTTTAGTAAAATGTAGCTTTTGATAGTTAATAAATT GTCATGGTTGTCTTTGATTAAAGGAAACTCACCGCCATATTCACAAA

[0200] In some embodiments, the RNA interference molecule targets SLF2. In some embodiments, the RNA interference molecule targets a region within SLF2 (RefSeqNM_018121.4) having the nucleotide sequence shown as SEQ ID NO: 53 below:ACTTCCGAAGAGAGAACCGCCATGAAGAGAGAAGGGGGTGCCGCCCACCTCTGCTCCGACAGCCTCCCGGAGTCCCAGCAGCAAGACGGCAACCACGCACCCAACTTCTCCAGCCACGGCTCATGCCGCCGTCGCCAGCGGCGCCGACATGACAAGGCGCTGCATGCCCGCTAGGCCAGGTTTCCCCTCATCCCCAGCCCCGGGGTCGTCGCCCCCGCGCTGCCATCTGAGACCCGGTAGTACCGCCCATGCTGCAGCGGGAAAGAGAACAGAGAGTCCTGGGGACAGGAAGCAGTCAATTATAGATTTCTTCAAACCAGCTTCAAAACAAGACAGACACATGTTGGATTCACCACAAAAATCAAACATCAAATATGGAGGAAGTAGATTGTCTATCACTGGGACAGAGCAGTTTGAAAGGAAACTATCCTCACCAAAAGAATCTAAACCCAAAAGGGTGCCACCAGAAAAGAGCCCTATTATAGAAGCTTTCATGAAAGGTGTTAAAGAGCACCATGAAGATCATGGTATACATGAGTCACGTCGGCCTTGTCTGTCACTAGCCTCCAAATATTTAGCCAAAGGAACAAATATCTATGTTCCTTCTTCATATCACTTGCCAAAGGAGATGAAGTCACTAAAGAAAAAACATCGATCCCCAGAGAGAAGGAAGTCACTATTCATTCATGAAAATAATGAGAAGAATGATAGAGATCGAGGCAAAACCAATGCAGACTCCAAAAAGCAGACCACAGTGGCAGAAGCTGACATCTTCAATAACAGCTCCAGAAGCCTTAGCAGCAGGAGCAGCCTGTCCAGGCACCACCCGGAAGAAAGCCCACTGGGAGCTAAATTCCAGTTGTCACTAGCTTCTTACTGCAGAGAACGAGAACTAAAGAGGTTGAGAAAGGAGCAAATGGAGCAGAGAATCAACTCCGAGAATTCTTTCTCAGAAGCAAGCAGTCTTTCCTTAAAATCTAGTATAGAAAGAAAATATAAACCAAGGCAGGAACAAAGGAAACAGAATGACATCATACCTGGAAAAAATAATCTGTCAAATGTGGAAAATGGACATCTCTCAAGAAAAAGATCCTCTTCTGATTCATGGGAACCTACTTCAGCAGGCTCTAAGCAGAATAAATTCCCTGAAAAAAGAAAAAGGAACTCTGTGGACTCAGATCTGAAAAGCACAAGAGAATCTATGATACCAAAAGCAAGAGAGTCCTTCCTTGAGAAGCGTCCTGATGGACCACATCAGAAAGAAAAATTTATAAAACATATTGCACTGAAGACACCTGGTGATGTGTTGCGCTTAGAAGATATATCCAAGGAACCGAGTGATGAAACTGATGG CTCTTCTGCAGGCTTGGCACCTTCAAATTCTGGCAATTCTGGCCACCATTCTACCA GGAATAGTGACCAAATCCAAGTGGCAGGTACCAAGGAGACTAAGATGCAGAAACCCCACTTACCTTTATCTCAGGAAAAGTCTGCAATTAAAAAAGCTAGCAACCTTCAGAAAAATAAAACCGCTAGCTCCACGACAAAGGAGAAGGAGACAAAACTACCTTTACTTTCCCGTGTTCCAAGTGCTGGTTCCTCTCTAGTACCATTAAATGCTAAAAATTGTGCTCTTCCAGTTTCTAAAAAAGATAAAGAGCGTTCCTCATCTAAAGAATGTTCTGGGCATTCTACAGAATCCACCAAACACAAGGAACACAAAGCAAAGACTAATAAGGCCGATTCTAATGTATCTTCAGGGAAAATTTCTGGGGGACCTTTGCGCTCAGAATATGGCACTCCTACAAAGTCTCCCCCTGCTGCTTTGGAAGTTGTGCCATGTATCCCAAGCCCTGCAGCACCTTCAGATAAAGCCCCTTCAGAAGGAGAGAGTTCAGGAAATTCCAATGCAGGTAGCAGTGCACTGAAAAGAAAACTAAGGGGTGATTTTGATAGTGATGAAGAAAGTTTAGGTTACAACCTAGACAGTGATGAGGAAGAGGAAACATTAAAGTCACTGGAAGAAATAATGGCTTTGAACTTCAATCAGACTCCTGCAGCTACAGGAAAGCCTCCTGCTCTTTCCAAGGGGCTTAGATCTCAGTCATCAGACTATACAGGACATGTTCATCCTGGAACTTACACAAATACCTTAGAACGTCTAGTGAAGGAAATGGAAGACACACAAAGGCTAGATGAACTGCAGAAGCAACTACAAGAAGACATAAGGCAAGGCCGAGGCATTAAATCCCCAATCAGAATTGGAGAAGAAGACAGTACAGATGATGAGGATGGCCTCTTAGAAGAGCACAAGGAATTTCTAAAGAAATTTTCAGTTACAATTGATGCTATTCCTGATCATCATCCAGGTGAAGAAATATTTAATTTCCTCAATTCTGGAAAAATTTTCAATCAGTATACCTTGGATTTAAGAGACTCTGGTTTTATTGGACAAAGTGCTGTAGAAAAACTTATTCTTAAATCGGGAAAAACAGATCAGATTTTTTTGACAACACAAGGTTTCCTTACGTCTGCTTATCACTATGTCCAGTGTCCTGTCCCTGTGTTAAAGTGGCTGTTTCGGATGATGTCAGTTCATACAGACTGTATTGTGTCAGTGCAGATTTTAAGTACATTGATGGAAATAACAATTAGAAATGATACCTTCAGTGACTCACCAGTTTGGCCATGGATCCCATCATTGTCTGATGTAGCAGCTGTGTTTTTCAATATGGGGATTGATTTTAGATCTTTGTTTCCCCTGGAGAATCTTCAGCCAGACTTTAATGAAGACTATCTAGTTTCTGAAACACAGACAACATCAAGGGGGAAAGAAAGTGAAGATTCATCTTATAAGCCAATTTTTTCAACACTTCCTGAAACCAACATTTTAAATGTGGTTAAGTTTCTAGGCTTGTGTACATCTATACATCCAGAAGGTTACCAGGATCGTGAAATAATGTTGCTGATTTTAATGTTATTTAAAATGAGTTTGGAAAAACAGCTGAAACAGATTCCTTTAGTAGACTTTCAAAGCCTCCTGATAAACCTGATGAAAAACATCAGAGATTGGAACACAAAGGTGCCTGAACTCTGTCTGGGCATAAATGAACTCTCCAGTCATCCCCACAACCTCCTGTGGTTGGTACAGCTGGTCCCTAATTGGACATCACGTGGAAGGCAACTGAGACAGTGCCTCAGTCTAGTGATTATTTCAAAGCTTTTGGATGAGAAACACGAAGATGTTCCTAATGCCAGTAATCTGCAGGTATCAGTCCTACATCGCTATCTTGTGCAGATGAAGCCTTCTGATTTGTTAAAGAAAATGGTCTTGAAGAAAAAGGCTGAACAACCAGATGGCATTATTGATGACAGTCTTCATTTAGAACTTGAAAAGCAGGCATATTACCTGACCTACATTCTTCTTCATTTAGTCGGTGAAGTTAGTTGTTCTCATTCTTTTTCTTCTGGACAACGGAAACACTTTGTGCTACTCTGTGGGGCTTTGGAAAAGCATGTTAAATGTGATATTAGGGAAGATGCAAGACTTTTTTACAGAACTAAGGTGAAAGACTTGGTCGCCAGGATACATGGAAAATGGCAGGAAATAATCCAGAACTGTCGGCCTACTCAGGGGCAGCTTCATGACTTCTGGGTACCAGATTCTTAATAGGAGTTGCAGCAGCAAAAATATGAACCAAGAGAAATTCAATAAGAGCCTTTCATAGAGGAGTAGAAAGGATTATTACAGAATCCAATGAATGCCAAGAAAATGTACAGCAAATGTGCCACTTGAATATCTAGTATGAAGCTGGTAATGAAGAAATTGCCATTTCTGAAGCAGATATGAAATATGATCTGCTTAATTGTTAAGGCAACTGACCTTTCAAAAGTGCAGAGTCTTATTAAAAGAGGGGAGGGGTAGAAGCAGAATAATAGTCATATGTCTAACCTGCCCCAGTTAACTCCTCTTGTTAAATTATAAGCCAGTTATCTTTTTTAGATAGTATTTTTGTCACTTGGATAATCACAGGAAATATATAAGAAAAGAGCTTGGACTAACTTGAGAAGTTGGACATGGAAAGCAAGACCAAGTTCCAGTTGGGTTTAATTTTCCCTCTTGGTTATTTTCGGACACAAAGGGAATGCTTAAAACTGAGTTTAGTAATAAAAAGCATAAATCTCTTCTGTAACTTTTATAAACCACAGGGAGGTTTCAATCCATGCATTTTCCTTCATTACTCAAGATTATAAATCTGTTTTTAAAATACATCTAAACAAACAGTTGAGAAACAAAAGTTTGGCATGTTGTCAGATCCCCTTAAGAGGAAGAGGTTAAGCTGTAAAGTAGTGGCCCTGTTTTGATGCCAGAACATTCATATGCTGTTTGTTCTGGATTTCTTTTAAATGCATGTATTTTAAATACTGGTTAAATCTTAGAATCTTGGCTATATCTTAGAATTCTGGCTCTTGGTAACCATATTACAGAAGTCTATATTGTAAAAGCCTAAAGATCTGGACAAGTTTCCAACAACCTTGTTTCCATGAGTATATAATTTTGGCAGACACCTGAATTCTTTGGGACACTGTGTCTGTGTATGTGTGTGTGTGTGTGTGTGTGTGTGTGCGCTTGTATCTTGAAGTTGTTGCACTTCAAAACCACAGCTGCTGTAAATTCTTAAACACTGGAGAGCCATCCTTTGGTTTAAATGGTAGAGGGTTAGTGGAAGTGCACACGTTGTTTTTAGCCCAGGGGTAGTAACAAACTCTTACTAGCCAACTAGAAAATTATTTTGTTGACGAGATGTCCCTTTTAGGAGGGTTTGTTTCCTGAAAGAATGCCAACTTTTTTTGTTGTTCTTTCAGTATTGGCTTGAGTGACCTGTTCTCCTGAGTGCTCTAGTGTCTCCAGTTGTGGGGGGGAAAGATGATGGAGGGGAACAGAAACTGGACTTGATGTTTGCGGTTTGAGAGGCAAGAAAATAAAATAACTTTCTACCTCTAAATTGAGGCTTAGGAGTAAAAAGCATTTTGTCCTAAATTTATCATTTAAAATAGCATCAGTAACTTTTGAGCTCATGTCAATCAAGCATTGGCAGTCAGAGATTTTATAGGGAAGACTAAGTAAATCCAGTTTCCAAGAACCTAAACTGATTGAGGCTCCAAGAGTCAGACCAACAAAAGTTTTATTCTGTGTTGTTTACTGGTAAGAATATTATTATCTTGATACTACCTCTCAAGGGTATTGTTACAAAATGCCACTTATGGTTAAAGAGATAGATACAAAGAGTTCTATTTGACAGAA GCTTGAAACTCTGGCATCTATCTGCCCAACGATGGGGGCTTTCGTTCTGTAATTT AATCCTTTGTAGATACATTATTTGTGTGTAATTTTATACGTGTTCATATTTTTCTCATTTTGCATTGTGTAAAGTGTACAAAATCTCAAAGTATAAAATACTGCTTATATTG CTTGTAATTTACAGTGTGTAAATATTTTCTAATTGTGTACATTGATGGGGGGGAC AAGTGGGTTATTCAGGTTTTTTTTTAATGACCCTTTTGTATTGCAGTTTCAACAGATAACTGTCCATCAAATTTAAAACCACTTTGATACATTTTTATTTAACAGTTCCAAT AAGAAAAAAATCTCTATTTTTAATTATATTTCTCCTTTAGAAAAAAAATACTTCAT GGTCCCTCTAAAATAAATGGCCAGACCTCAGTTTAGGCCCTTGTATTTATGACTCTGGTAGAGGTCTTCAGACTCCAGGTCATAGTCCAGTCAGTATTTGCATTTGGGTT CTCATGAAAACTTTGTGACTCTCTTTTAGCACAAGTAGCCCATGGTTTTTCTTCCA AATCAAGTATTTTACTTCTCCCTTGAGTCCCCATGCTTTGTCTCCCTCCTGCTGCT ATTGCCCGGTGAATGGGATGGTAGAGAGGGAGAAATGTTCATTGCACAGAGAAT GTCAGGCCACTTTGGGGACTTGGCAAACGAAGCTTGCACTGAGTGGTGGTGTGTT TGGCAATATTACTGTGCCAAAAATCACCTTGTCTAATTTTATGGATATGTATGGC AAACTTATTACCCTTCTTGCAAGCTGCACATTAAGGTACTTGTAAGTGTTTATGCTTTTGTGTGTAACTGTTTGCCTTTTCTAACTGCTTTTCTTGTTACTGAAAATATAATT CTACTAGTTTTATCACATTAAGAGGCTCTCCATGCACAAAATGCATTGATGTAGC CGTAAAGTAACAGCATTTGTACATTTTCTATCTCCTGATGGCAGTGGTGCCTTTGTCACCTTTTGGAAGGTTTGCATTTTGTTTCTGCTTTCAAGCGCAAATAGTAGCTTGC TTTCAGCTTCACAAAAATCTCCTCAAATGTAAAAGATACAAAATGGCGTGTTATC ATCCAGGCTTAGTTGGAGTATTTGCATTTTTATTTTTATCAAAACAAATATAATTGGTGGGGACTGGCCAGTAATAATGTGTGGCGTCTTTAAGCCAAGAGTAATTTTTAA ATTAAAAATGAAAATTTTAAGAGGTAGCCCATTAAACAAGCTACTGAGTTGGAG AATTAGGGGATGACTGTGGTGGTTTGTCGCTAAGGAGGCAACAGTAGGGTCCAGGCGGCGGGGCACGTAGAGCAGTTAGCATGATCCATGGTTATTCCTTACTCATGAA CAGTGTCGGCGCACCTCATTATCTGTGCATTTGTTTTTCCTGGGCAGTCCTGTCAG CCAGTTAATCCACCAGCTCTTAGGAAGTAAATACAGATTTTTTTCTTTCCTTTTTTTTTTTTTTAAAGATTTTATACTACATCTTGTTAAAGTCCTATGAATGTGTGTGTGTGTGTGCTATTAAAACTGCTTTTTCTCAGTTTGAAAtpip / Setdbl

[0201] As described above, ATF7IP and SETDB1 are also known to be involved in epigenetic silencing mechanisms. ATF7IP (Activating Transcription Factor 7 Interacting Protein; also known as MBD1 -containing chromatin-associated factor (MCAF) or mouseATFa associated Modulator (mAM)) is an epigenetic and transcriptional regulator, which recruits transcriptional factors and histone modification enzymes to the general transcription apparatus, thereby modulating chromatin formation and gene expression. Evidence of ATF7IP in epigenetic regulation is provided by studies showing that it binds SET domain and bifurcated 1 protein (SETDB1) to mediate transcriptional repression through recruiting MBD (methyl CpG-binding protein). SETDB1 is a lysine methyltransferase (KMT) that catalyzes the trimethylation of histone H3 at lysine 9 (H3K9me3) for silencing genes and endogenous retroviruses (ERVs) in embryonic stem cells (ESCs) or during embryogenesis. In cultured human cells, ATF7IP regulates SETDB1 nuclear localization and ubiquitination that facilitates the H3K9 methyltransferase activity.

[0202] In some embodiments, the RNA interference molecule targets SETDB1. In some embodiments, the RNA interference molecule targets a region within SETDB1 RefSeq NM_001393958.1) having the nucleotide sequence as shown in SEQ ID NO 54 below: GTTTCTTTTGCTTCCCCTTCCCTCTTTCACGCTTCCTCCCCTCCCCCTCCTCCCTTA TCCCTTCGCTTTCGCTCTTTTCCGTCGAGGCCGACCCCTGAGTTGTGAGTCTGGGG TCTGGTTGGTGAAAAAGAGCCCTTGAAGCTGGAAGACGGGAGGTGCGGGGAATA CTGTTGAGTTATTTGAGGACAAAAGCATGTCTTCCCTTCCTGGGTGCATTGGTTTG GATGCAGCAACAGCTACAGTGGAGTCTGAAGAGATTGCAGAGCTGCAACAGGCA GTGGTTGAGGAACTGGGTATCTCTATGGAGGAACTTCGGCATTTCATCGATGAGG AACTGGAGAAGATGGATTGTGTACAGCAACGCAAGAAGCAGCTAGCAGAGTTAG AGACATGGGTAATACAGAAAGAATCTGAGGTGGCTCACGTTGACCAACTCTTTG ATGATGCATCCAGGGCAGTGACTAATTGTGAGTCTTTGGTGAAGGACTTCTACTC CAAGCTGGGACTACAATACCGGGACAGTAGCTCTGAGGACGAATCTTCCCGGCC TACAGAAATAATTGAGATTCCTGATGAAGATGATGATGTCCTCAGTATTGATTCA GGTGATGCTGGGAGCAGAACTCCAAAAGACCAGAAGCTCCGTGAAGCTATGGCT GCCTTAAGAAAGTCAGCTCAAGATGTTCAGAAGTTCATGGATGCTGTCAACAAG AAGAGCAGTTCCCAGGATCTGCATAAAGGAACCTTGAGTCAGATGTCTGGAGAA CTAAGCAAAGATGGTGACCTGATAGTCAGCATGCGAATTCTGGGCAAGAAGAGA ACTAAGACTTGGCACAAAGGCACCCTTATTGCCATCCAGACAGTTGGGCCAGGG AAGAAATACAAGGTGAAATTTGACAACAAAGGAAAGAGTCTACTGTCGGGGAA CCATATTGCCTATGATTACCACCCTCCTGCTGACAAGCTGTATGTGGGCAGTCGG GTGGTCGCCAAATACAAAGATGGGAATCAGGTCTGGCTCTATGCTGGCATTGTA GCTGAGACACCAAACGTCAAAAACAAGCTCAGGTTTCTCATTTTCTTTGATGATG GCTATGCTTCCTATGTCACACAGTCGGAACTGTATCCCATTTGCCGGCCACTGAAAAAGACTTGGGAGGACATAGAAGACATCTCCTGCCGTGACTTCATAGAGGAGTATGTCACTGCCTACCCCAACCGCCCCATGGTACTGCTCAAGAGTGGCCAGCTTATCAAGACTGAGTGGGAAGGCACGTGGTGGAAGTCCCGAGTTGAGGAGGTGGATGGCAGCCTAGTCAGGATCCTCTTCCTGGATGACAAAAGATGTGAGTGGATCTATCGAGGCTCTACACGGCTGGAGCCCATGTTCAGCATGAAAACATCCTCAGCCTCTGCACTGGAGAAGAAGCAAGGACAGCTCAGGACACGTCCAAATATGGGTGCTGTGAGGAGCAAAGGCCCTGTTGTCCAGTACACACAGGATCTGACCGGTACTGGAACCCAGTTCAAGCCAGTGGAACCCCCACAGCCTACAGCTCCACCTGCCCCACCTTTCCCACCTGCTCCACCTCTATCCCCCCAAGCAGGTGACAGTGAAAGCTTGGAAAGCCAGCTTGCCCAGTCACGGAAGCAGGTAGCCAAAAAGAGCACGTCCTTTCGACCAGGATCTGTGGGCTCTGGTCATTCCTCCCCTACATCTCCTGCACTCAGTGAAAATGTCTCTGGTGGGAAACCTGGGATCAACCAGACATATAGATCACCTTTAGGCTCCACAGCCTCTGCCCCAGCACCCTCAGCACTCCCGGCCCCTCCAGCACCCCCAGTCTTCCATGGCATGCTGGAGCGGGCCCCAGCAGAGCCCTCCTACCGTGCTCCCATGGAGAAGCTTTTCTACTTACCTCATGTCTGCAGCTATACCTGTCTGTCTCGAGTCAGACCTATGAGGAATGAGCAGTACCGGGGCAAGAACCCTCTGCTGGTCCCGTTACTATATGACTTCCGGCGGATGACAGCCCGGCGTCGAGTTAACCGCAAGATGGGCTTTCATGTTATCTATAAGACACCTTGTGGTCTCTGCCTTCGGACAATGCAGGAGATAGAACGCTACCTTTTCGAGACTGGCTGTGACTTCCTCTTCCTGGAGATGTTCTGTTTGGATCCATATGTTCTTGTGGACCGAAAGTTTCAGCCCTATAAGCCTTTTTACTATATTTTGGACATCACTTATGGGAAGGAAGATGTTCCCCTATCCTGTGTCAATGAGATTGACACAACCCCTCCACCCCAGGTGGCCTACAGCAAGGAACGTATCCCGGGCAAGGGTGTTTTCATTAACACAGGCCCTGAATTTCTGGTTGGCTGTGACTGCAAGGATGGGTGTCGGGACAAGTCCAAGTGTGCCTGCCATCAACTAACTATCCAGGCTACAGCCTGTACCCCAGGAGGCCAAATCAACCCTAACTCTGGCTACCAGTACAAGAGACTAGAAGAGTGTCTACCCACAGGGGTATATGAGTGTAACAAACGCTGCAAATGTGACCCAAACATGTGCACAAACCGGTTGGTGCAACATGGACTACAAGTTCGGCTACAGCTATTCAAGACACAGAACAAGGGCTGGGGTATCCGCTGCTTGGATGACATTGCCAAAGGCTCTTTTGTTTGTATTTATGCAGGCAAAATCCTGACAGATGACTTTGCAGACAAGGAGGGTCTGGAAATGGGTGATGAGTACTTTGCAAATCTGGACCATATCGAGAGCGTGGAGAACTTCAAAGAAGGATATGAGAGTGATGCCCCCTGTTCCTCTGACAGCAGTGGTGTAGACTTGAAGGACCAGGAAGATGGCAACAGCGGTACAGAGGACCCTGAAGAGTCCAATGATGATAGCTCAGATGATAACTTCTGTAAGGATGAGGACTTCAGCACCAGTTCAGTGTGGCGGAGCTATGCTACCCGGAGGCAGACCCGGGGCCAGAAAGAGAACGGACTCTCTGAGACAACTTCCAAGGACTCCCACCCCCCAGATCTTGGACCCCCACATATTCCTGTTCCTCCCTCAATCCCTGTAGGTGGCTGCAATCCACCTTCCTCCGAAGAGACACCCAAGAACAAGGTGGCCTCATGGTTGAGCTGCAATAGTGTCAGTGAAGGTGGTTTTGCTGACTCTGATAGCCATTCATCCTTCAAGACTAATGAAGGTGGGGAGGGCCGGGCTGGGGGAAGCCGAATGGAGGCTGAGAAGGCCTCCACCTCAGGACTAGGCATCAAGGATGAGGGAGACATCAAACAGGCCAAGAAAGAGGACACTGACGACCGAAACAAGATGTCAGTAGTTACTGAAAGCTCTCGAAATTACGGTTACAATCCTTCTCCTGTGAAGCCTGAAGGACTTCGCCGCCCACCTAGTAAGACTAGTATGCATCAAAGCCGAAGACTCATGGCTTCTGCTCAGTCCAACCCTGATGATGTCCTGACACTGTCCAGCAGCACAGAAAGTGAGGGGGAAAGTGGGACCAGCCGAAAGCCCACTGCTGGTCAGACTTCGGCTACAGCGGTTGACAGTGATGATATCCAGACCATATCCTCTGGCTCTGAAGGGGATGACTTTGAGGACAAGAAGAACATGACTGGTCCAATGAAGCGTCAAGTGGCAGTAAAATCAACCCGAGGCTTTGCTCTTAAATCAACCCATGGGATTGCAATTAAATCAACCAACATGGCCTCTGTGGACAAGGGGGAGAGCGCACCTGTTCGTAAGAACACACGCCAATTCTATGATGGCGAGGAGTCTTGCTACATCATTGATGCCAAGCTTGAAGGCAACCTGGGCCGCTACCTCAACCACAGTTGCAGCCCCAACCTGTTTGTCCAGAATGTCTTCGTGGATACCCATGATCTTCGCTTCCCCTGGGTGGCCTTCTTTGCCAGCAAAAGAATCCGGGCTGGGACAGAACTTACTTGGGACTACAACTACGAGGTGGGCAGTGTGGAAGGCAAGGAGCTACTCTGTTGCTGTGGGGCCATTGAATGCAGAGGACGTCTTCTTTAGAGGACAGCCTTCTTCCCAACCCTTCTTGAACTGTCGTTTCCTCAGGAACTGGGTCTTCCTGATTGTTGAACCCTGACCCGAAGTCTCTGGGCTAGCTACTCCCCCCAGCTCCTAGTTGATAGAAATGGGGGTTCTGGACCAGATGATCCCTTCCAATGTGGTGCTAGCAGGCAGGATCCCTTCTCCACCTCCAAAGGCCCTAAAGGGTGGGGAGAGATCACCACTCTAACCTCGGCCTGACATCCCTCCCATCCCATATTTGTCCAAGTGTTCCTGCTTCTAACAGACTTTGTTCTTAGAATGGAGCCTGTGTATCTACTATCTCCAGTTTGTATTATTTCTTGAAAGTCTTTTAACAATATGATAAAACTAA

[0203] In some embodiments, the RNA interference molecule targets SETDB1 and comprises the sequence selected from SEQ ID NOs: 55-63 as shown below:AGTTAGAGACATGGGTAATAC (SEQ ID NO: 55)GCTCAGATGATAACTTCTGTA (SEQ ID NO: 56)CCAGACATATAGATCACCTTT (SEQ ID NO: 57)CAGTGACTAATTGTGAGTCTT (SEQ ID NO: 58)CGTGACTTCATAGAGGAGTAT (SEQ ID NO: 59)ATCCCTCCCATCCCATATTTG (SEQ ID NO: 60)CGTGACTTCATAGAGGAGTAT (SEQ ID NO: 61)GCTCAGATGATAACTTCTGTA (SEQ ID NO: 62)CAGTGACTAATTGTGAGTCTT (SEQ ID NO: 63)

[0204] In some embodiments, the RNA interference molecule comprises a sequence having at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%. at least 97%, at least 98%, at least 99%, or 100% sequence identity to a sequence selected from SEQ ID NOs: 55-63.

[0205] In some embodiments, the RNA interference molecule that targets ATF7IP. In some embodiments, the RNA interference molecule targets a region within ATF7IP (RefSeq NM_001388183.1) having the nucleotide sequence as shown in SEQ ID NO: 64 below: GTAGGCTAAGACATAACAGCTTCCCTTTGTTGTTTCATTGCTATGATCTCTTGCTAGAAAGAAGTGGGTGGAGGGGGATGAAGCAGAGAAGAGAGTAATGTTTTCATTTA AAGAAGAAGCTCGTTGGGCTTTCCTTAAAAATATTTTAAAAATAATTTACCAAAA ATTTGAAAAAAAAATTTCCAAGCCAGAGAGACTTATTTGCAGGTAACATACCCTT CTCTGGTTATTTAAATATAAATTTCTATAGAAAAAGGTTTTATTAGTGATATTATT TTCCCAGAGTTCTTGAATATACAGAGACTTAAAAAACATTATTTTGACTCTCATC CTCATGAAAAAAATACTTTCAGGAGAAACTAGTTCATCATGTTTTGCAGTTACTT TTAGACATGGCAGACATCTTCTGCCGTAAAATACCAATGCTTTTTTCTGTGAAAA TGAATTACAGTCATGTGAAAGCTTTCTGATTTGAGTTGCCACCTAAAATACTAAC ATTTATTTTATAGAATAGTTTTATGAATGGCATCTTAGTTCTAGGCGACAGTACTA AAATGATCCTTGGAAGTGTTTTGCACTGGTATTACAGCCATGGAAACTTGTTTGC AGATTCAGAATGGACAGTTTAGAAGAACCTCAGAAAAAAGTCTTTAAGGCTCGA AAAACGATGAGAGTGAGTGATCGTCAGCAACTTGAAGCAGTGTACAAGGTCAAA GAAGAACTGTTGAAAACTGATGTCAAGCTGTTAAATGGCAACCATGAAAATGGA GATTTGGACCCAACCTCACCTTTGGAAAACATGGATTACATTAAAGACAAGGAA GAGGTGAATGGCATTGAAGAGATTTGTTTTGATCCTGAAGGAAGTAAAGCAGAA TGGAAGGAAACACCCTGTATCCTAAGTGTTAATGTAAAAAACAAGCAGGATGAT GATTTAAATTGTGAACCTTTGTCTCCCCATAATATAACTCCAGAACCAGTCTCTA AACTGCCTGCTGAACCAGTTTCTGGTGATCCAGCCCCTGGTGATCTGGATGCCGG AGATCCAGCCTCCGGAGTACTGGCCTCTGGTGATTCCACCTCTGGTGATCCCACC TCTAGCGAGCCCTCCTCTAGTGATGCTGCCTCTGGTGATGCAACCTCTGGTGATG CCCCTTCTGGTGATGTGTCCCCTGGTGATGCCACCTCTGGTGATGCCACTGCTGAT GATCTCTCCTCTGGTGATCCCACCTCTAGTGATCCCATCCCAGGTGAACCGGTCCCTGTTGAACCCATTTCTGGTGATTGTGCCGCTGATGATATAGCCTCTAGTGAAATAACTTCTGTTGATCTGGCTTCTGGAGCACCAGCTTCCACTGATCCAGCCTCTGATGATCTGGCCTCTGGTGATCTATCCTCTAGTGAACTGGCCTCTGATGATCTGGCCACTGGTGAACTGGCCTCTGATGAGCTGACTTCTGAATCAACCTTTGATCGTACCTTTGAACCAAAGTCTGTACCAGTTTGTGAACCAGTTCCTGAAATTGACAATATAGAACCAAGTAGCAATAAAGATGATGATTTTCTTGAAAAAAATGGAGCTGATGAAAAATTAGAGCAAATTCAGAGTAAAGACTCATTGGATGAGAAAAATAAAGCTGATAATAATATTGATGCTAATGAAGAAACTCTAGAAACAGATGATACAACTATTTGTTCAGATCGACCTCCTGAAAATGAAAAGAAGGTAGAGGAAGATATTATCACAGAGCTTGCTCTTGGAGAAGATGCTATATCTAGCAGTATGGAAATTGACCAAGGTGAAAAGAATGAAGATGAAACTTCTGCAGATCTTGTAGAAACGATTAATGAAAATGTTATTGAAGATAACAAAAGTGAGAATATCTTAGAAAATACAGACTCTATGGAGACAGATGAAATCATTCCTATTTTGGAAAAGCTTGCACCTTCTGAGGATGAACTTACTTGCTTTTCTAAAACATCTCTCCTTCCAATCGATGAGACAAATCCAGATTTGGAAGAGAAAATGGAAAGTTCTTTTGGTTCACCATCTAAACAAGAAAGTAGTGAGAGTTTGCCAAAAGAAGCCTTTCTGGTCCTCTCTGATGAAGAGGATATTTCGGGTGAAAAAGATGAGTCTGAAGTTATATCGCAAAATGAAACGTGCTCTCCAGCAGAAGTAGAAAGTAATGAAAAGGACAACAAACCTGAGGAAGAAGAGCAAGTAATACATGAAGATGATGAAAGACCTTCTGAGAAAAATGAATTTTCTAGACGAAAACGTTCTAAATCAGAAGACATGGACAATGTACAGTCTAAACGTCGTCGATATATGGAAGAAGAATATGAGGCAGAATTTCAAGTAAAGATTACAGCCAAAGGAGACATTAACCAGAAACTTCAAAAGGTTATACAGTGGTTGCTGGAAGAAAAATTGTGTGCGCTGCAGTGTGCTGTATTTGATAAGACTTTGGCAGAATTGAAAACACGAGTGGAAAAGATTGAATGTAACAAGAGGCATAAAACAGTTCTCACTGAACTACAGGCCAAGATAGCCAGGTTAACCAAACGCTTTGAAGCAGCCAAAGAAGATCTTAAGAAAAGACATGAACATCCACCCAACCCACCAGTATCACCAGGAAAAACTGTAAATGATGTCAACAGCAATAATAACATGTCTTACAGAAATGCAGGCACAGTGAGACAGATGCTGGAGTCCAAAAGAAATGTAAGCGAGAGTGCACCACCATCCTTTCAAACTCCTGTGAATACAGTATCTTCAACCAATCTTGTCACTCCTCCAGCAGTTGTCAGTAGTCAACCTAAATTGCAGACTCCAGTGACTTCGGGTTCCCTCACAGCAACGTCAGTTCTTCCTGCACCCAATACAGCTACTGTAGTTGCTACTACTCAGGTGCCTAGTGGAAATCCCCAGCCTACAATCTCTTTACAGCCTTTGCCAGTGATTTTGCATGTACCTGTTGCAGTATCCTCCCAGCCTCAGCTTCTACAGAGCCATCCAGGGACTTTGGTGACTAATCAACCATCTGGCAATGTTGAATTCATTTCTGTGCAAAGCCCACCTACAGTGAGTGGTCTTACCAAAAATCCAGTATCCTTGCCATCCTTGCCAAATCCCACTAAACCAAACAACGTTCCTTCTGTGCCCAGTCCTAGTATTCAAAGGAACCCTACTGCCAGTGCTGCACCATTGGGAACAACACTTGCTGTGCAGGCTGTTCCAACAGCACACTCTATTGTACAAGCCACAAGGACTTCTTTACCCACAGTGGGCCCATCAGGACTCTATAGTCCATCAACTAATCGAGGTCCTATACAGATGAAAATTCCAATTTCTGCATTTAGTACTTCGTCTGCTGCAGAACAGAACAGCAATACCACCCCAAGAATTGAAAACCAGACAAACAAAACAATAGATGCTTCTGTCAGTAAGAAAGCAGCTGATAGCACATCACAGTGTGGAAAAGCCACTGGCAGTGATTCAAGTGGTGTCATTGATCTCACAATGGATGATGAAGAGAGTGGAGCTTCACAAGACCCCAAAAAACTAAATCACACTCCTGTATCAACCATGAGTTCTTCTCAGCCTGTGTCACGACCATTGCAACCCATACAACCAGCACCGCCTCTTCAACCATCTGGGGTGCCAACAAGTGGACCATCTCAGACCACCATACACTTACTACCTACAGCTCCAACTACCGTGAATGTAACACATCGTCCAGTAACTCAGGTGACCACAAGACTCCCTGTACCAAGAGCTCCTGCAAACCACCAGGTGGTTTATACAACTCTTCCTGCACCACCAGCTCAGGCTCCCTTGCGAGGAACTGTTATGCAGGCTCCTGCTGTTCGGCAGGTCAATCCCCAAAATAGTGTTACAGTTCGAGTGCCTCAAACAACCACATATGTTGTAAACAATGGACTAACCCTGGGATCAACAGGACCTCAGCTCACAGTGCATCACCGACCACCACAAGTGCATACTGAGCCCCCACGCCCCGTGCACCCAGCACCCTTACCAGAAGCTCCACAACCACAGCGTCTGCCCCCAGAAGCTGCCAGCACATCTCTGCCTCAGAAGCCACACTTGAAGTTAGCACGCGTTCAGAGTCAAAATGGCATAGTACTGTCATGGAGTGTCCTGGAGGTGGATCGAAGCTGTGCCACTGTTGATAGCTACCATCTCTATGCTTACCATGAGGAACCCAGTGCCACTGTGCCCTCACAATGGAAAAAGATTGGGGAAGTCAAGGCACTTCCCTTGCCCATGGCATGTACTCTCACCCAGTTTGTATCTGGTAGCAAATACTACTTTGCAGTACGAGCCAAGGATATTTATGGACGTTTTGGGCCTTTCTGTGATCCTCAGTCAACAGATGTGATCTCTTCTACCCAGAGCAGTTAAACCTTGGAGCCTTTATATTTTCCTCTTTTAAAATTTCCACCTTTTGGTCTTGTTTTTAATCTTGTGCATGATACCCCATGTAAAATCCACCTTGTGCAAGATTTCTTGGACAGATGTGTGTATACACTACATTTGTTTATAACCAGAAGCAAAATAAACTCAGCCCACAAAGCTAGAATCTTTTCCTGGACAGTTTAGGCTTTGGGGTTTGGAAATGTAAATGTGTACCTTGCTTTAGTTTTGAGGCTGGGGAATATGTGTGGGTGTTTATGTGTGTTTTTCCTTATGTAGGTGTTATTGCATTGGAGTCTCCCATTTTCATTCTCAAATTTACCTCTTAAAGTACGAAGTAAGTAGATCAAAGGATTTGAGATGTGTAACTGGCATGATTCTGCTTTTGAAGGATCTATAGTATCATTTTAGTTAAGTGGGTCAAACAGAATCAAAACAAAACCCAAAGAAATAAATAAAAAACAAAATGGCTAAATAGTTTAAAATAGGTTAATTCGAACACAGGAAAGGATCTATTTGTTGTTTCTTTTGTCTGGTCTCCTGAGTTGTTAATTAGGTGAAAAAAGATCTGCAATGGCCCCCTCCCTTTCCTAATCTGGCTTTTACATTTATTTTGTGCCTTAAAGATTAACTACAAAGATAAACATGGCCAAAAATAAATAAATAAATATGGCCATATGTCCGTTGTTGCTTAGTCTTCCCTTGCAGCCTTTTACCCTTGATTTCTCCTTCATCTCTACCAAATATAGCACAACTCCTCAAGTAATTTTTTTTTTTTAAGATGGAGTTTTGCTCTTGTTGCCCAGGCTAGAGTGCGGTGGCACAATCTTGGCTCACTGCAACCTCTGCCTCCTGGGTTCAAGCAATTCTTCTGTCTCAGCCCCAGAGTAGCTGGGATTACAGGCACCTGTCACCATGCCCGGCTAATTTTTGTATTTTTAGTAGAGATGGGGTTTCACCATGTTGGCCAGGCTGGTCTTGAACTCCTGACCTCAGGTGATCCACCCGCCTCGGCCTCCCAGAGTGCTGCGATTACAGTTGTGAGCCACTGTATCCAGCCTACTCAAGTGATTTTTAAACCAAGGTGTGTGTATGTACATGGATGTGTATGTACACACACACACAGGTGCATGCATAGTCTCATCTTAGCCGGGCATGGTGGCGACAATCACATTTAACTCCAAAAATTTGACGTGCTATTTTCTTCTAAAACTATAGATGCTTTTATTTTTGGTCACTATTTAACTTTGTGTAAAGTGGACCAAGAGAAAACCATATTGTGGAAGACAGTTTTTCACACATTTTAGTATCCGTGTTGCACACTGTGTTAGAGATTATGAAAACCATTCTAGTTCAGTTTATCACCCAAAGCTATCACCCAACACTAACTCCTTAGTATTCTTTCAAAGGAAGCAAATAGTAAGATTACAATATAGAATGAGCTAAAATATCAAGGGGAAAGCTTCATTTCCTCTTTATTCTCTTTATCGTCCTTCCCATTTTAACCTTTTAACTTTGTTTTGATGGTGCCAAGTTGAGTCTGATGTACCCATTGTTAGCTTTGGATAGAATTTAAAGTATATCCCAGAATTAGCTGGGCATGTTGGTGCATGCCTGTAATCCCAGCTACTCAGGAGGCAAGGCTGGAGAATCGCTTGAACCCGGGAGGCGGAGGTTGCAGTGAGCTGAGATTGCGCCATTGCACTCCATCCTGGGAGACAAGAGCGAAACTCTGCCTCAAAAAAAAAAGAATTTAAAGTATATCCTTTGAGGTCATTTTATGTAACAATTAGATATAACCTCCTGTGTATCTCTCTGTTCCTCTTCAAAGTTATTAAAATTCAGCTTAGGTCATGGATTTTTAATATGAGGGCCAGCTGTACTGGTGTCAGGTAGGTGTTTTGGGTGTAGATAATTTAGGACTGGAGGGCTGAGTTAAGTTTTAGGAGGAGAATGTGGTTGAGAATATTTTGAAAGGAATTTATGGAGTTTGTGAAATAACTTCGAAGTCCTCTTCCTTTACAATATTTGAATTCATATTTGTACCTTCTCAAAATAGTGATTCATTTTTCCTAGAATTACAGGAGGGAGCTCTTTTACTAATGTTGTTTTGTTTGCAACTTTGATGGCTTATAATAGGAAGTATTCTAGTTGTAAAGAAAACTCTTTAGAGACTTTTGACTGGTCAGTATACTGAGGTGTGAGATTTGATTCATGATGAAGAAAGCCTATAGATTGCCAAAAAATTAATTCTCCAAACCACCTTTCACTCTCAGAAAATGAGACCACAAAGGAGTATGCTATAAATCAAATTTGCCAACCAATTATGTAGATATTACTCATTCTAGGACTAATGATGATGGTAAAGAAGTTGCCAGTGTTATGGCAATGAAAATTTCAGAAAGGAGGAGTTGATGATCTTCTAGATGTATATGAACACCTGTCTATATCTGCATGTATATGTTTTGACCTGCAGTGGTTGCAATGTTGATATGTGTTCAAGATTATTCCTGTCTACAAAACTGAAGGCCCATGTTCAAATTGTTCTTTATTGGGTGTTTTTATGGTCACGTGGTAACAATTTTCTTACCTAACCTACAAAAGGTTCTCTTGATGAACATTTTTATTTATATTTACTAATCTTTTTTAAAAAAAGCTTTCATAGCATTATATAATCAGATGAAGAAAGCCCAGTAGAATAAAAAAAAAATTCATTAGCCTAGCCTATATTATGTTTTCTGTCAAAGGAAAACAAATTCTCAAATAGGAATTCTAAAAATATTTACTAAAGTAAAATAACTACTTAAAATGTTTTATTCCAGTTGGAAGGAAGGTACAGGGAGAAATCGCAATTATTTAGGGGAGAAGTATATTTATTATAAGATGGTGTCCTCAAATTAGCCTACCATGGCACGTAGGGGCAGCAGCTATATTAGATTTACTAGAGGTGCTAAGTTAGAACACTAGGCTTTTATTGAGGCAGGTTTTAATATTGATAGATGCTTTTTGTTTGGTTTGTTTCTTCTGGGAGAGAATGGAGGACTTAAGTAGAAGTAGCTACTGATAACAGACTTTCTAGTAGCAGTTTCCACTCCACGGTTACCTTTTTAGTTTCATAGTATCTTTTCACAAAGTATTACAAATAAGCTAGATTCTCCCAGTTTGGGAATGCAAGTTTGCTACATTTTTAGCCTGGCAATATTTGTGTAGGTATTGCCTTATTGGAAATTCTGGAAACCTGATACTGCAACCTGCAATGTAGGATGTTTGTATGGCATTTAAAGGTAATGGTGATGTTTATTATTCTATACTTTGCATTCTGTGAGAGTAATTTTCACTCTGTCTTAAGTGTGAGTAAGCCTCTTCTAAAAATCTTGTTCTTGCCAAGAAATTTATAAATCACATACGAAGACGTCTGTTGCTAACAGTTAACTTTATGAGGTAACTATATCCTTCTATTTCTCTGGACTCATTTTTAAAAAATATGCCGAATACTGCATACTGTTTAAGGTAGTATATAAGTTTATGAGAGAAGTGGAGAGCTTTCTTCCTTGAAAAGTCGGTATTTGTTGAGATACCATTTGCCTCACAGAGAGGTGTTCCCCACTCCCATCCCCATTGCCAGATAATAAATATTTTGAGAAAAGTGACCTAAAACAGCTGAAATCTTAGGTGCATCTGTCTGCAGACCTCCTTAAGCAGGCTGTATCTTACAATTCCCTTACTGCACTGGGTAAGTGTTAACTTAGTTTTTGTTGTTTGCTCTTTTGCTTTAAATATTCTCCAAATTACCATTTATGCAACATGGTTAGGGTTAATACTGCATGGTATTCATTTATCTTGTTTCATGAACTTTCCAGTACTGTACAAGGTCAACAAAGTAATGCCTGTGGTATCCTCATCTCTCACTTTTTTACTCTGTGATTTTAGCACAGTAAGGTACTGCAAAGACCTTCCTTCCAAATGTTCTCCTTGACTTTATTTCTTGGGCCAATTCAGTATCCTCAACATCCTAAGATTTTGTTGTTTTATCACTGACCTGTGGTTGGCCTGTTTTATTCTAATTTCCAGAAAAGTCAAGTCCCAGTATTTGCAATATCAAATAACTCTAAAACCGATGTGTGATTCTACCTTCCTTACTATTTTTACTGGGCAAATGCCCTATTTTTTTAATTATTATTATTTTTAACTTTTGGGACACACAAAAATCAGCAATTCTCATGAAGCGTTTGTTAGTGTGGCAGACTTGTCTAATTCCTGAAACTCATTCATCCCCTTGAGCCAGCCAATGGGGAGGAATAGGATAATGCAAACACATGTTTTGTTTTCTCATTTTCAAATAATTTACCATGTTAAAATAAACTTTTCTTTGTTTTTTATTTGTAGAGTCAGCTAAGTACCCATATTTAAATGCCGTCTTTATTATTTTTTTGAGGTCTTTGTTTTTGTCTGTTTTTGTTTTGTTTTGTTTTGTAAATAAGGTAACTGGGCAATCAAACACCTTTTGGGGATTCTGGCTTTAGTATTTTATCAGCCATTTTAAAATTAAATATAAAAATCCTTTGTAAGAAACTTGCATCCTAATTTTTCTTTATTGCAATTGAAAGTGTAAATAATAAGACAATGTAAGTAAGACCTTCCTAATGTCTAATACAAACTGGGCTCCAGCAAGTGGCCCTATTTTTATTAGGGTTTTGAAGGTTTGTGTGTGTGTGTGCGTGCGTGTGTGTGTTTTTCTTTTTTAAATGTATAGTAGAGTGGTGTCTGTATAAGTGTTACCTGTAGTGGGGTTTTGTCCAGCAAG CCTGAAATTTATACTTTGAAATAAAACTACTGGGTTTTTAA

[0206] In some embodiments, the RNA interference molecule targets Atf7ip and comprises the sequence of CCCTGTATCCTAAGTGTTAAT (SEQ ID NO: 827).

[0207] In some embodiments, the RNA interference molecule comprises a sequence having at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NO: 827.Long Terminal Repeats

[0208] As described herein, the retroviral vectors of the present disclosure also include one or more LTRs. An LTR can be, for example, a sequence of about 100 to 1400 base pairs, which is repeatedly present at both ends of the retroviral DNA, DNA of retrotransposons or the like. Functional LTR sequences less than 100 or greater than 1400 may also be used. The LTR is composed of U3, R, and U5 regions which are involved in transcription of a viral gene, reverse transcription from a viral genome, and integration of a double-stranded DNA synthesized via the reverse transcription into a host DNA. IR sequences (inverted repeat regions) located at the 5' and 3' ends of the provirus are 4 to 20 base pairs in length. The U3 region contains an enhancer sequence and a promoter sequence for transcription. Herein, promoters that naturally occur in LTRs are not considered to be “exogenous’' promoters in the vector constructs of the invention that comprise one or more LTRs but no exogenous promoters.

[0209] The one or more LTR sequences used in the present disclosure can include any sequence as long as they are derived from retrovirus and capable of producing a retrovirus that contains an RNA having these sequences as a genome. These sequences may be derived from the same virus or may be derived from different viruses to the extent that virus particles can be formed. In some embodiments, the retroviral vector of the present disclosurecomprises a 3’ LTR and 5‘ LTR derived from the same virus. In some embodiments, the retroviral vector of the present disclosure comprises a 3’ LTR derived from one vims and 5’ LTR derived from a different vims. Exemplary LTR sequences that can be used in the compositions herein include, without limitation, LTR sequences from derived from Bovine leukemia virus (BLV), Human T-lymphotropic virus 1 (HTLV-1), Human T-lymphotropic virus 2 (HTLV-2), Gibbon ape leukemia virus (GALV). Feline leukemia virus (FeLV), Porcine endogenous retrovimses (PERV). Feline Foamy Virus (FeFV). Bovine foamy virus (BFV), Mouse mammary tumor vims (MMTV), Jaagsiekte sheep retrovims (JSRV), Mason- Pfizer monkey virus (MPMV), Avian sarcoma leukosis vims (ALV), Rous sarcoma vims (RSV), Human Endogenous Retro virus-W (HERV-W), Human endogenous retrovirus K (HERV-K). In some embodiments, the one or more LTRs are derived from MLV. In some embodiments, the one or more LTRs are derived from HIV. In some embodiments, the retroviral vector of the present disclosure comprises a 3’ LTR and a 5’ LTR that are both derived from HIV.

[0210] In some embodiments, LTR sequences comprising a mutation(s) can also be used. A retroviral vector thus obtained in which the U3 region of the 3' LTR sequence is mutated is called a self-inactivating (SIN) vector. The term “self-inactivating vector’ refers to vectors in which the right (3') LTR enhancer-promoter region, known as the U3 region, has been modified (e.g., by deletion or substitution) to prevent viral transcription beyond the first round of viral replication. Consequently, the vectors are capable of infecting and then integrating into the host genome only once and cannot be passed further. This is because the right (3') LTR U3 region is used as a template for the left (5') LTR U3 region during viral replication and, thus, the viral transcript cannot be made without the U3 enhancer-promoter. If the viral transcript is not made, it cannot be processed or packaged into virions, hence the life cycle of the virus ends. Accordingly, SIN vectors greatly reduce risk of creating unwanted replication-competent virus since the right (3') LTR U3 region has been modified to prevent viral transcription beyond the first round of replication, hence eliminating the ability of the virus to be passed.

[0211] In some embodiments, only a portion of one or more LTRs is used. For example, a portion of an LTR sequence can include at least about 20%, for example, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, about 95% of the full length LTR sequence. In some embodiments, the portion of the LTR that is used comprises a naturally occurring promoter of the LTR. Naturally occurring promoters of LTRs are known to the person of ordinary skill in the art. For example, the HIV-1 LTR core promoter consistsof a series of three tandem Spl binding sites, a non-canonical TATA box element, and an initiator element.

[0212] Exemplary LTR sequences for use in the vectors described herein are shown below.Human Immunodeficiency Virus 1 (HIV-1) LTRTGGAAGGGCTAATTTGGTCCCAAAAAAGACAAGAGATCCTTGATCTGTGGATCTACCACACACAAGGCTACTTCCCTGATTGGCAGAACTACACACCAGGGCCAGGGATCAGATATCCACTGACCTTTGGATGGTGCTTCAAGTTAGTACCAGTTGAACCAGA GCAAGTAGAAGAGGCCAAATAAGGAGAGAAGAACAGCTTGTTACACCCTATGA GCCAGCATGGGATGGAGGACCCGGAGGGAGAAGTATTAGTGTGGAAGTTTGACAGCCTCCTAGCATTTCGTCACATGGCCCGAGAGCTGCATCCGGAGTACTACAAAGACTGCTGACATCGAGCTTTCTACAAGGGACTTTCCGCTGGGGACTTTCCAGGGAGGTGTGGCCTGGGCGGGACTGGGGAGTGGCGAGCCCTCAGATGCTACATATAAGCAGCTGCTTTTTGCCTGTACTGGGTCTCTCTGGTTAGACCAGATCTGAGCCTGGGAGCTCTCTGGCTAACTAGGGAACCCACTGCTTAAGCCTCAATAAAGCTTGCCTTGAG TGCTCAAAGTAGTGTGTGCCCGTCTGTTGTGTGACTCTGGTAACTAGAGATCCCT CAGACCCTTTTAGTC AGTGTGGAAAATCTCTAGC A (SEQ ID NO: 65)Moloney Murine Leukemia Virus (MLV) LTRAATGAAAGACCCCACCTGTAGGTTTGGCAAGCTAGCTTAAGTAACGCCATTTTGCAAGGCATGGAAAAATACATAACTGAGAATAGAGAAGTTCAGATCAAGGTCAGGAACAGATGGAACAGCTGAATATGGGCCAAACAGGATATCTGTGGTAAGCAGTTCCTGCCCCGGCTCAGGGCCAAGAACAGATGGAACAGCTGAATATGGGCCAAACAGGATATCTGTGGTAAGCAGTTCCTGCCCCGGCTCAGGGCCAAGAACAGATGGTCCCCAGATGCGGTCCAGCCCTCAGCAGTTTCTAGAGAACCATCAGATGTTTCCAGGGTGCCCCAAGGACCTGAAATGACCCTGTGCCTTATTTGAACTAACCAATCAGTTCGCTTCTCGCTTCTGTTCGCGCGCTTCTGCTCCCCGAGCTCAATAAAAGAGCCCACAACCCCTCACTCGGCGCGCCAGTCCTCCGATTGACTGAGTCGCCCGGGTACCCGTG TATCCAATAAACCCTCTTGCAGTTGCATCCGACTTGTGGTCTCGCTGTTCCTTGGG AGGGTCTCCTCTGAGTGATTGACTACCCGTCAGCGGGGGTCTTTCA (SEQ ID NO: 66)Bovine leukemia virus (BLV) LTRTGTATGAAAGATCATGCCGACCTAGGAGCCGCCACCGCCCCGTAAACCAGACAG AGACGTCAGCTGCCAGAAAAGCTGGTGACGGCAGCTGGTGGCTAGAATCCCCGTACCTCCCCAACTTCCCCTTTCCCGAAAAATCCACACCCTGAGCTGCTGACCTCACCTGCTGATAAATTAATAAAATGCCGGCCCTGTCGAGTTAGCGGCACCAGAAGCGTTCTTCTCCTGAGACCCTCGTGCTCAGCTCTCGGTCCCGAGCTCTCTTGCTCCCGAGACCTTCTGGTCGGCTATCCGGCAGCGGTCAGGTAAGGCAAGCCACGGTTTGGAGGGTGGTTCTCGGCTGAGACCACCGCGAGCTCTATCTCCGGTCCTCTGACCGTCTCCACGTGGACTCTCTCCTTTGCCTCCTGACCCCGCGCTCCAAGGGCGTCTGGCTTGCACCCGCGTTTGTTTCCTGTCTTACTTTCTGTTTCTCGCGGCCCGCGCTCTCTCCTTCGGCGCCCTCTAGCGGCCAGGAGAGACCGGCAAACA (SEQ ID NO: 67)Human T-lymphotropic virus 1 (HTLV-1) LTRTGACAATGACCATGAGCCCCAAATATCCCCCGGGGGCTTAGAGCCTCCCAGTGAAAAACATTTCCGAGAAACAGAAGTCTGAAAAGGTCAGGGCCCAGACTAAGGCTCTGACGTCTCCCCCCGGAGGGACAGCTCAGCACCGGCTCAGGCTAGGCCCTGACGTGTCCCCCTGAAGACAAATCATAAGCTCAGACCTCCGGGAAGCCACCGGAACCACCCATTTCCTCCCCATGTTTGTCAAGCCGCCCTCAGGCGTTGACGACAACCCCTCACCTCAAAAAACTTTTCATGGCACGCATATGGCTGAATAAACTAACAGGAGTCTATAAAAGCGTGGAGACAGTTCAGGAGGGGGCTCGCATCTCTCCTTCACGCGCCCGCCGCCCTACCTGAGGCCGCCATCCACGCCGGTTGAGTCGCGTTCTGCCGCCTCCCGCCTGTGGTGCCTCCTGAACTGCGTCCGCCGTCTAGGTAAGTTTAGAGCTCAGGTTGAGACCGGGCCTTTGTCCGGCGCTCCCTTGGAGCCTACCTAGACTCAGCCTGCTCTCCACGCTTTGCCTGACCCTGCTTGCTCAACTCTGCGTCTTTGTTTCGTTTTCTGTTCTGCGTCGCTACAGATCGAAAGTTCCACCCCTTTCCCTTTCATTCACGACTGACTGCCGGCTTGGCCCACGGCCAAGTACCGGCGACTCCGTTGGCTCGGAGCCAGCGACAGCCCATTCTATAGCACTCTCCAGGAGAGAAACTTAGTACACA (SEQ ID NO: 68)Human T-lymphotropic virus 2 (HTLV-2) LTRTGACAATGGCGACTAGCCTCCCGAGCCAGCCACCCAGGGCGAGTCATCGACCCAAAAGGTCAGACCGTCTCACACAAACAATCCCAAGTAAAGGCTCTGACGTCTCCCCCTTTTTTTAGGAACTGAAACCACGGCCCTGACGTCCCTCCCCCCCAGGAACCAGGAACAGCTCTCCAGAAAAATAGACCTCACCCTTACCCACTTCCCCTAGCGCTGAAAAACAAGGCTCTGACGATTACCCCCTGCCCATAAAATTTGCCTAGTCAAAATAAAAGATGCCGAGTCTATAAAAGCGCAAGGACAGTTCAGGAGGTGTCTCGCTCCCTCACCGACCCTCTGGTCACGGAGACTCACCTTGGGGATCCATCCTCTCCAAGCGGCCTCGGTTGAGACGCCTTCCGTGGGACCGTCTCCCGGCCTCGGCACCTCCTGAACTGCTCCTCCCAGGGTAAGTCTCCTCTCAGGTCGAGCTCGGCTGCCCCTTAGGTAGTCGCTCCCCGAGGGTCTTTAGAGACACCCGGGTTTCCGCCTGCGCTCGGCTAGACTCTGCCTTAAACTTCACTTCCGCGTTCTTGTCTCGTTCTTTCCTCTTCGCCGTCACTGAAAACGAAACCTCAACGCCGCCCTCCTGACAGGCTTGGCCCGGGGCCAACATACTGCCGTGGAGGCGCAGCAAGGGCTAGGGCTTCCTGAACCTCTCCGGGAGAGGTCTATTGCTATAGGCAGGCCCGCCCCAGGAGCATTTGTCTTCCCGGGGAAGACAAAC A (SEQ ID NO: 69)Gibbon ape leukemia virus (GALV) LTRTGAAAGAAGTGTTTTTTTTTAGCCAACTGCAGTAACGCCATTTTGCTAGGCACACCTAAAGGATAGGAAAAATACAGCTAAGAACAGGGCCAAACAGGATATCTGTGGTCATGCACCTGGGCCCCGGCCCCAGGCCAAGGACAGAGGGTTCCCAGAAATAGATGAGTCAACAGCAGTTTCCAGGGTGCCCCTCAACCGTTTCAAGGACTCCCATGACCGGGAATTCACCCCTGGCCTTATTTGAACTAACCAATTACCTTGCCTCTCGCTTCTGTACCCGCGCTTTTTGCTATAAAATAAGCTCAGAAACTCCACCCGGCGCGCCAGTCCTTAGAGAGACTGAGCCGCCCGGGTACCCGTGTGTCCAATAAAACCTCTTGCTGATTGCATCCGGAGCCGTGGTCTCGTTGTTCCTTGGGAGGGTTTCTCCTAACTATTGACCGCCCACTTCGGGGGTCTCACA (SEQ ID NO: 70)Feline leukemia virus (FeLV) LTRTGAAAGACCCCCTACCCCAAAATTTAGCCAGCTATTGCAGTAATGCCATTTCACAAGGCATGGAAAATTACCCAAGCATGTTCCCATGAGATATAAGGAAGTCAGAGGCTAAAACAGGATATCTGTGGTTAAGCACCTGGGCCCCGGCTTGAGGCCAAGAACAGTTAAGCCTCGGATATAGCTGAAACAGCAGAAGTTTCAAGGCCACTGCCAGCAGTCTCCAGGCTCCCCAGTTGACCAGAGTCCGACCTTCCGCCTCATTTAAACTAACCAATCCCTACGCCTCTCGCTTCTGTGCGCGCGCTTTCTGCTATAAAACGAGCCATCAGCCCCCAACCGGCGCGCAAGTCTTTGCTGAGACTTGACCGCCCCGGGTACCCGTGTACGAATAAACCTCTTGCTGTTTGCATCTGACTCGTGGTCTCGGTGTTCTGTGGGCACGGGGTCTCATCGCCGAGGAAGACCTAGTTTGGGGGTCTTTCAT (SEQ ID NO: 71)Porcine endogenous retroviruses (PERV) LTRTGAAATGGAGTTTTAACTAAAGAACAACCCCCCCCCCCCTTGGCTAAGAAGGCAGCTGCAAGCCTTTCCAACTGTGCTAATCACTATGCCACCCTTCTTGTAATAACATAACAACTGTGTGCTCTCTGTCCAAGCCAGCAGCCCTGCTAATTAGGTACCCAGCATTGCTTATCAGTTCTGCTTCTGTAACCTTGCTTGCTCAATTTCTTAGGGAGGAACACTGTCTGCTTGGGGATGGGGGAGGTCCAGGATGCTTACATGGGAAAATCAAGACCTTGTAGTGTATAAAAGTTACTGAGAACAAAGACCTGGTGCTCAGACTCTGGAGGTGACTCCTCTGAGCCCACACCAGTGCTGAATAAACCTTGCTTTTCCATATCTCTGAGTGCTGTTTGTCTCCTTCTGTTGCCACAGTTTTTGTGGTTTCTGCAACA (SEQ IDNO: 72)Feline Foamy Virus (FeFV) LTRTGTCATGGGCCAAAGAGAATTCTCACAGAGGAGAATACTCTCTGCTGCCATCTAGTGACGATGAGGAAGAAGAAATGTCAGAAAGAGAGGAATTATTGTGCCATATAAATCAGTGTCAACAAAAGCTCTTTTATCCCGGAGGGACGACTGATGTCCTTGGAATGGAAAGCAATGTTTGGCTCACTAAATTTGTTAATATTAAATTTCCTAAAGGAACAAAAGTGATACTTCCTGATGGAAGAAAATTCATAGCCTGTGATCCTGAGCTAAAACCATTATTGCAGGAATTGAAATTCTTGGATAGGGCAACATCTGAGTCATCTGACTCTGAATAGAAAGCCTGAATTTACCTGGATTATGCAACTTTGTCCGAGGTGGCAGAGTGGTTATGTATCTGTCATACTCGGGGAAAGTTTTGTCTTTACATGTTCAAGACATATAAAGGGTGGAAAAATATATTCCTGACTAAACTTCCTGGGGACTAGAGGTGTGGAAACTTTGCTGCCTCTGCTTCACGGGAAGTTTTTGGTTCGAATCCTTTTTTAGGTACTTAGTTAAGATAAGTAGTGAATAAATTACTCTCGTTCATGTATTCATATCGAAACTATGTATCCTTTAAAACCATGTATTCTTTAGTCATCTAGATACTTAGAGTATGAAAAAAGAAACTGCAATAGTAACTATCAATGTTAGTAAATAAAGTACAGCTTAGTCATCTGATGATGTCACGAGAAAAGAACCTAGAAGAGAAGAACAACTTTCGGCATGCAACAGAGCGGGAGCTTGGTGTAGGAGCTAAGTCACCGTCTTACATCTAGAGCCTACTCTTCTTGAACTGTTCGAATCCTATTTTTGGAACTCTTACATCACCTTTAAGAGACTGAAAAGCATGACTCGTGCACAGGAAGCTCCTTTAGGGTAGAGGAAATGTTCTAATCTCCTATCTTAAAGGGTTGCTTCATTTAAGGTTCGAAACTGTGTACTGGAAGTAGATTTTGCATAACTTTAAACTTTTAGTTGCATGTTTCTGCTATTAGCAGCATATAAAAGGGTTATGGTAGATTGTACGGGAGCTCTTCTCACAGACTTGGCTGCGTCCAGGGTGAGATTGAGACTCTCCAGCTTGGGTAAGATTTTGATATGTATTTTGCTTGAATATTATTTGCCTTGCTCAAAATTAAATAAATTGGCTTTTCTTTCACTCAATTGAAGCTTCATATAATTATATTATTGTCTGAAGCCAGAACTCACATGAGTGGTGTTTCTCTATTCTTGGGGAAAAGTGTTCTTCTATTTGAAAGTGTTAGAGCTACTAAGTGAAGAACTAATCTATCCCAGGTATAGGCCACGACA (SEQ ID NO: 73)Bovine foamy virus (BFV) LTRTGTGGTGGGAAGACCACCCGGAAATAAGCAAGGGCCAGCCCCCCTTTACTTGCCCGGAGGATTGGCTGGAGTGTGAGGAAGATAGTGAGAATGAGGAACTCAGTGTTTCTCCCCTCACAGGCTGGGAGCCTCATAGAGACTTAAAAAAGTGGGTAAATTTCGCCCTACCTTATGGATGGAGCCTTATGGACCCGCTAGGTAATCGTTTCAGGTCCCAGAGGAAAGATGATTCCGATGAATCCACTAGTACGGATTCTGAAGAGGAGTTACAAGAAATACAGCAGATCAACCCCGCTGAAGTAGGGCCAAGCAGGCCTCCCGGCGGGTACTCTAAGCTGCGCAGGCGCCGTCGGGTTCCAGCTGCTCGACTTTACGCAACTACAGACAGTAGTGAAGAAGACCTGGATCTGAAGACATCTCAATAAGGAGCTGGACTGTTTGAGATCTGTGTGTGATTACATTGTATTAAGATAGCCTGTTACTGGGGTTCGGAGGATGGCTCATCAAGCTAGTAAGGAGCTCCCTGAAGCCATATCCGAGGCTCAGTAGAAAGACAGTACCTCGCCTGTGTGGGTTCGAGGTCAGGCGGTATGCTTTCTACTTTTTAGATGTATAACTAGAAGAATAAGGTTAAGGAGATGCTTTTTAAATAGATAGCTTAGGGAGTAAGTTAGATAGCTTAAAGAACAAGTTAACTGCAAGTATCGCTTATGCTCAGAATCAGGAAGTAGCTTGCGTCACCAGTTATAGAGAAATAAGAGCTTGAAAGTTAACCGCAACATAGGATGTGAGTCATAGCTATTTTAGTAAGTTAGCGCTTGGGAACTGAGAGAACAAGGTTCGGATCCCATATCATTATAATAAAATATATCTTGTATAAGTAGTTAGCTTACTGTAAAAGGGGGGAATTATTGGGAATCCCATATTGTAGGAAAAGTCAGTTGGATAAGGATATAAAACCCTTAGACTGTAAGGAAGGAGGAGCTCTTGAATCGGCTGCGCCCGTGAGAGATCTGACTTCCAGGCTGTGGTAAGACCTAATTTGGACTTGACTCTCTTAACTCGCTCTGCTTAGAACAAGCTTGTTTTTAATATTATGAGATTATAATGCCTGTAATGTTAATCAATAAATGGTGAAGGAAACTCGAGAATCTCCATCTCTATTATTGTTATCCCTATACTCCGAGTGAGTGGGAACCTGTCTCCAACCTGGGCTTACCTTCCAGATCACTAATAAGGGTGGTATCTACTGGAACCTTATAGAGTTATCTGGAATCCACTAAAGATTGGAGAAGGTCACAACAA (SEQ ID NO: 74)Mouse mammary tumor virus (MMTV) LTRTGCCGCGCCTGCAGCAGAAATGGTTGAACTCCCGAGAGTGTCCTACACTTAGGAGAGAAGCAGCCAAGGGGTTGTTTCCCACCAAGGACGACCCGTCTGCGTGCACGCGGATGAGCCCATCAGACAAAGACATACTCATTCTCTGCTGCAAACTTGGCATAGCTCTGCTTTGCCTGGGGCTATTGGGGGAAGTTGCGGTTCGTGCTCGCAGGGCTCTCACCCTTGATTCTTTTAATAACTCTTCCGTGCAAGATTACAATCTAAACGATTCGGAGAACTCGACCTTCCTCCTGGGGCAAGGACCACAGCCAACTTCCTCTTACAAGCCACACCGACTTTGTCCTTCAGAAATAGAAATAAGAATGCTTGCTAAAAATTATATTTTTACCAATGAGACCAATCCAATAGGTCGATTATTAATCATGATGTTAAGAAATGAATCTTTGTCTTTTAGCACTATATTTACTCAAATTCAAAGGTTAGAAATGGGAATAGAAAATAGAAAGAGACGCTCAACCTCAGTTGAAGAACAGGTGCAAGGACTAAGGGCCTCAGGCCTAGAAGTAAAAAGGGGAAAGAGGAGTGCGCTTGTCAAAATAGGAGACAGGTGGTGGCAACCAGGGACTTATAGGGGACCTTACATCTACAGACCAACAGACGCCCCGCTACCATATACAGGAAGATATGATTTAAATTTTGATAGGTGGGTCACAGTCAACGGCTATAAAGTGTTATACAGATCCCTCCCCTTTCGTGAAAGGCTCGCCAGAGCTAGACCTCCTTGGTGTGTGTTGTCTCAAGAAGAAAAAGACGACATGAAACAACAGGTACATGATTATATTTATCTGGGGACAGGAATGATACATTGGAAAGTATTTTATAACAGTAGAGAGGAGGCCAAAAGACATATAATAGAACATATTAAGGCATTGCCCTTAGCTTTCTAAAGTTTGCTTGCGGCTCCCAAGGTTTAAGTAAG TTCATGGTCACAAACTGTTCTTAAAACAAGGATGTGAGACAAGTGGTTTCCTGAC TTGGTTTGGTATCAAATGTTTTGATCTAAGCTCTGAATGTTCTATTCTCCTATGTTCTTTTGGAACTTATCCAAGTCTTATGTAAATGCTTATGTAAACCATGATATAAAAGAGTGCTGATTTTTTGAGTAAACTTGCAACAGTCCTAACATACACGTCTCGTGTGTTTGTGTCTGTTCGCCATCCCGTCTCCGCTCGTCACTTATCCTTCACTTTCCAGAG GGTCCCCCCGCAGACCCCGGTGACCCTCAGGTCGGCCGACTGCGGCA (SEQ ID NO: 75)Jaagsiekte sheep retrovirus (JSRV) LTRTGGGAGCTCTTTGGCAAAAGCCAAAGCCTAGGACAAGTACCTAAGCTCCCTGTCCCGCCACCCTCAAGAATTTTTAAAAGCTCTTAAGGCTCGGATGTTTGCTTTTGGCACTGCTTCACAGAAATATCAGGAAATCTGATTATATAAGAATCCGGTGATTGTGTAAGAATCCGGTGGGTGTAAGTGAATAATGAATAAACAAGTTATGTTGTGTACTTTATAAATATAGCATTGTAATAAAGCAGAGTATCAGCCATTTTGGTCTGATCCTCTC AACCCCATCTTTTGTCTCTCTCTTATTTTCTTAGCGAAGACGCTCCGTTCTCTCCCT GTGCAGGTGCGACTCTTGCTTGTGCTGGCCGCGGCAGG (SEQ ID NO: 76)Mason-Pfizer monkey virus (MP MV) LTRTGTCCGGAGCCGTGCTGCCCGGATGATGTCTTGGCCTCTGTTTGCTCTAGCTCCATGTTATGAATTTAAGATGGCGTATTTCCTGGTTCTTCTCCGTCTTACTTTCCCGCCG GCGCGAATGTTTCCCGCTCTTGGGCTTACGTGGCTTTCCTTGCTCTGCTACTGAGC ATGCGCCCAGTATCTTTCCCCTCCCACTTGCTGCCTGTGTATATAAGGCAACACATTGCCACCATTAAATGAGACTTGATCAGAACACTGTCTTGTCTCCATTTCTTGTGTCTCTTGTTCCCTTCAATTCCCACTCCCTCCTCCAGGTTCCTACTGTTGATCCCGCGGGTCGGGACAGT (SEQ ID NO: 77)Avian sarcoma leukosis virus (ALV) LTRTGTAGTCTTATGCAATACTCTTGTAGTCTTGCAACATGGTAACGATGAGTTAGCAACATGCCTTACAAGGAGAGAAAAAGCACCGTGCATGCCGATTGGTGGAAGTAAGGTGGTACGATCGTGCCTTATTAGGAAGGCAACAGACGGGTCTGACATGGATTGGACGAACCACTGAATTCCGCATTGCAGAGATATTGTATTTAAGTGCCTAGCTCGATACAATAAACGCCATTTGACCAT (SEQ ID NO: 78)Rous sarcoma virus (RSV) LTRTGTAGTCTTATGCAATACTCCTGTAGTCTTGCAACATGCTTATGTAACGATGAGTTAGCAATATGCCTTACAAGGAAAGAAAAGGCACCGTGCATGCCGATTGGTGGTAGTAAGGTGGTACGATCGTGCCTTATTAGGAAGGTATCAGACGGGTCTAACATGGATTGGACGAACCACTGAATTCCGCATCGCAGAGATATTGTATTTAAGTGCCTAGCTCGATACAATAAACGCCATTTTACCATTCACCACATTGGTGTGCACCTGGGTTGATGGCCGGACCGTCGATTCCCTAACGATTGCGAACACCTGAATGAAGCAGAAGGCTT CATT (SEQ ID NO: 79)Human Endogenous Retrovirus-W (HERV-W) LTRTGTTGAGATGGGGGACTGAGAGACAGGACTAGTTGGATTTCCTAGGCCGACTAAGAATCCCTAAGCCTAGCTGGGAAGGTGACTGCATCTACCTTTAAACACGGGGCTTGCAACTTAGCTCACACCCGACCAATCAGTAAAGAGAGCTCACTAAAAATCTAATTAGGCAAAAACAGGAGGTAAAGAAATAGCCAATCATCTAATGCCTGAGAGCACAGTGGGAGGGACGATGATTGGGATATAAACCCAGGCATTCGAGCCGGCAACAGCAACCCTCTTTGGGTCCCCTCCCTTTGTATGGGATCTCTGTTTTCACTCTATTTCACTCTTTTGCAACTGCACTCTT (SEQ ID NO: 80)Human endogenous retrovirus K (HERV-K) LTRTGTGGGGAAAAGCAAGAGAGATCAGATTGTTACTATGTCTGTGTAGAAAGAAGTAGACATAGGAGACTCCATTTTGTTCTGTACTAAGAAAAATTCTTCTGCCTTGAGGTTCTGTTAAACTATGACCTTACCCCCAACCCCGTGCTCTCTGAAATATGTGCTGTGTCAAACTCAGGGTTAAATGGACTAAGTGTTGTGCAAGATGTGCTTTTTAAACAGATGCTTGAAGGCAGCATGCTCCTTAAGAGTCATCACCACTCCCTAATCTCAAGTAC TCAGGGACACAAAAACTGCGGAAGGCCGCAGGGACCTCTGCCTAGGAAAGCCA GGTATTGTCCAAGGTTTCTCCCCATGGGATAGTTTGAAATATGGCCTCGTGGGAA GGGAAAGACCTGATTGTCCCCCAGCCTGACACCTGTAAATGGTCTGTGCTGAGG AGGATTAGTATAAGAGGAAGGCATGCCTCTTGCAGTTGAGACAAGAGGAAGGCA TCTGTCTCCTGCCCGTCCATGGGCAATGGAATGTCTCGGTATAAAACCCGATTGT ACGTTCCATCTACTGAGATAGGGAAAAACCGCCTTAGGGCTGGAGGTGGGACAT GCGGGCAGCAATACTGCTTTGTAAAGCATTGAGATGTTTATGTGTATGCATATCT AAAAGCACAGCACTTGATTCTTTACCTTGTCTATGATGCAAAGACCTTTGTTCAC GTGTTTGTCTGCTGACCCTCTCCCCACTATTGTCTTATGACCCTGACACATCACCC TCTCGGAGAAACACCCACAAATGATCAATAAATACTAAGGGAACTCAGAGGCTG GCGGGATCCTCCATATGCTGAACGCTGGTTCCCTGGGTCCCCTTATTTCTTTCTCT ATACTTTGTCTCTGTGTCTTTTTATTTTCCAAGTCTCTCGTTCCACCTAATGAGAA ACACCCACAGGTGTGGAGGGGCAACCCACCCCTTCA (SEQ ID NO: 81)

[0213] In some embodiments, the nucleic acid constructs of the disclosure include one or more LTR sequences having at least 50%. at least 60%. at least 70%. at least 80%. at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the sequence of SEQ ID NO: 65 or SEQ ID NO: 66.Packaging Signals

[0214] As described herein, the integration deficient retroviral vectors of the present disclosure also include a packaging signal. As used herein, the term "packaging signal" or "packaging sequence" can be a sequence located within the retroviral genome required for insertion of viral RNA into a viral capsid or particle.

[0215] In some embodiments, the packaging signal is a minimal packaging signal.

[0216] In some embodiments, the packaging signal is a psi [VP] sequence.

[0217] Exemplary7packaging signals include, without limitation, packaging signals derived from HIV-1, MLV, BLV, HTLV 1, HTLV 2, FeLV, PERV, MMTV, MPMV, ALV, and RSV. Exemplary sequences of packaging signals are shown below.Human Immunodeficiency virus 1 (HIV-1) packaging sequenceCTCTCTCGACGCAGGACTCGGCTTGCTGAAGCGCGCACGGCAAGAGGCGAGGGG CGGCGACTGGTGAGTACGCCAAAAATTTTGACTAGCGGAGGCTAGAAGGAGAGA GATGGGTGCGAGAGCGTC (SEQ ID NO: 82)Moloney murine leukemia virus (MLV) packaging sequenceAAGCTGGCCAGCAACTTATCTGTGTCTGTCCGATTGTCTAGTGTCTATGACTGATTTTATGCGCCTGCGTCGGTACTAGTTAGCTAACTAGCTCTGTATCTGGCGGACCCGTGGTGGAACTGACGAGTTCGGAACACCCGGCCGCAACCCTGGGAGACGTCCCAGGGACTTCGGGGGCCGTTTTTGTGGCCCGACCTGAGTCCAAAAATCCCGATCGTTTTGGACTCTTTGGTGCACCCCCCTTAGAGGAGGGATATGTGGTTCTGGTAGGAGACGAGAACCTAAAACAGTTCCCGCCTCCGTCTGAATTTTTGCTTTCGGTTTGGGACCGAAGCCGCGCCGCGCGTCTTGTCTGCTG(SEQ ID NO: 83)Bovine leukemia virus (BLV) packaging sequenceTTGATCACCCCGGAACCCTAACAACTCTCTGGACCCACCCCCTCGGCGGCATTTTGGGTCTCTCCTTCAAATTATATCATGGGAAATTCCTCCTCCTATAACCCCCCCGCTGGTATCTCCCCCTCAGACTGGCTCAACCTTCTGCAAAGCGCGCAAAGGCTCAATCCGCGACCCTCTCCTAGCGATTTTACCGATTTAAAGAATTACATCCATTGGTTTCATAAGACCCAGAAAAAACCATGGACTTTCACTTCTGGTGGCCCCACCTCATGTCCACCCGGGAGATTCGGCCGGGTTCCCCTTGTCTTGGCCACCCTAAACGAAGTGCTCTCAAACGAAGGGGGCGCCCCGGGTGCATCGGCCCCAGAAGAACAACCCCCCCCTTATGACCCCCCCGCCGTTTTGCCAATCATATCTGAAGGGAATCGCAACCGCCATCGTGCTTGGGCACTCCGAGAATTACAAGATATCAAAAAAGAAATTGAAAATAAGGCACCGGGTTCGCAAGTATGGATACAAACACTACGACTTGCAATCCTGCAGGCCGACCCTACTCCGGCTGACCTAGAACAACTTTGCCAATATATTGCTTCCCCGGTCGACC AAACGG (SEQ ID NO: 84)Human T-lymphotropic virus 1 (HTLV-1) packaging sequenceGCAATGGGCCAAATCTTTTCCCGTAGCGCTAGCCCTATCCCGCGGCCGCCCCGGGGGCTGGCCGCT (SEQ IDNO: 85)Human T-lymphotropic virus 2 (HTLV-2) packaging sequenceTATGGGACAAATCCACGGGCTTTCCCCAACTCCAATACCCAAAGCCCC (SEQ ID NO: 86)Feline leukemia virus (FeLV) packaging sequenceGCGCAAGTCTTTGCTGAGACTTGACCGCCCCGGGTACCCGTGTATGAATAAACCTCTTGCTGTTTGCATCTGACTCGTGGTCTCGGTGTTCTGTGGGCACGGGGTCTCATCGCCGAGGAAGACCTAGCTCAGGGGTCTTTCATTTGGGGACTCGTCCGGGATAGAGACCCCCAACCCCCGGGACCACCGACCCACCATCAGGAGGTAAGCTGGCCGGCGATCATATCTGTTGTCCTTGTATGAGTGTCTCTGTCATTTGATCTGATTTTGGCGGTGGAGCCGAAGGAGCTGACGAGCTCGTACTTCGCCCCCGCAACCCTGGAAGACGTTCCACGGGTGTCTGATGTCTGGAGCCACTAGTGGGACAGCCATTGGGGCTCATCTGTTTGGGGTCTCACCTGAATACAGGGTGTTGATCGGAGACGAGGGAGCCGGACCCTCAAAGTCTCTTTCTGAGGTTTCA (SEQ ID NO: 87)Porcine endogenous retroviruses (PERV) packaging sequenceTTTTGGTGCATTGGCTGGGAAGCCGACAACCGAGCTGGCCTCTTGTGCCACCGTTGTCTGGGAACTGGGGGAGGCCACGGTCACCGGCCTGGTGACAGGTGGATAGCACACACCGGCCATTTTGCCGGATCCAGACCCTCACTTGGGCCATGGCCTTGGCCACACCGCTTCCCGAATGGACTCAGAAGGAGAGTGGAAACAGGTTCCTGGACAGGACGTCAGACTGGTGAGTGCCCCGGGGACATCTGCCCAAGTCAGGACCCACCATATGGTGAAGGAGAGACTGATCACCTCTCAGTGACCAAAGGGTCACTCAGGTCAGGGTTTGCTCCGTTGGCATCGGGAGGCTTGTCAATTCTGGTGTGTGTGTGTGAGTGACTGGAGAGAACAAGAGCCCATGCTCCACCATCTTTGGACTACGGAGTCAGAGTGAGTCCTAACCTGCAGTTCCATGGCGACCCTCATGCGGCTTATGGTGGCCCTCTTCAGGGGGATCCTGATGTTGGGGTTTATACAGTCCCACCTATGCTAAGAGGCGCCTGAAATATCTCCTCCAGGGGAGAGGTCAGACGGACGAAGAGATTGTTCGTCTTGCGTCACCGGCACAGGGTGGGACTTCACACAATGGGTAAATCGGCTTCCAAGCCCACTGTCTTAGAGCATATGGTTAAGAATTTTAAGAAAGGATTTTCTAGAGATTATGGTGTTAAACTGAGTCCTGGAAAACTGTGTACCCTCTGTACC (SEQ ID NO: 88)Mouse mammary tumor virus (MMTV) packaging sequenceGCAACAGTCCTAACATTCACCTCTCGTGTGTTTGTGTCTGTTCGCCATCCCGTCTCCGCTCGTCACTTATCCTTCACTTTCCAGAGGGTCCCCCCGCAGACCCCGGTGACCCTCAGGTCGGCCGACTGCGGCAGCTGGCGCCCGAACAGGGACCCTCGGATAAGTGACCCTTGTCTCTATTTCTACTATTTTGTGTTCGTCTTGTTTTGTCTCTATCTTATCTGGCTATTATCACAAGAGCGGAACGGACTCACCACAGGGAACTGCAGTCTCGCCTACAGAGAAGAGGTAGGTTACGGTGAGCCATTGGAAATGGGGGTCTCGGGATCAAAAGGGCAGAAACTCTTTGTTTCTGTTCTACAAAGACTCCTCTCAGAGAGGGGTCTTCATGTGAAAGAGAGCAGTGCAATAGAGTTTTATCAGTTCCTAATA (SEQ ID NO:Mason-Pfizer monkey virus (MP MV) packaging sequenceGCCACCATTAAATGAGACTTGATCAGAACACTGTCTTGTCTCCATTTCTTGTGTCTCTTGTTCCCTTCAATTCCCACTCCCTCCTCCAGGTTCCTACTGTTGATCCCGCGGGTCGGGACAGTTGGCGCCCAACGTGGGGCACGAACCCACGACCCTGGGATTAAGAGTCCCATGCTCTACCGACTGGATGATGTCTTGGCCTCTGTTTGCTCTAGCTCCATGTTATGAATTTAAGATGGCGTATTTCCTGGTTCTTCTCCGTCTTACTTTCCCGCCGGCGCGAATGTTTCCCGCTCTTGGGCTTACGTGGCTTTCCTTGCTCTGCTACTGAGCATGCGCCCAGTATCTTTCCCCTCCCACTTGCTGCCTGTGTATATAAGGCAACACATTGCCACCATTAAATGAGACTTGATCAGAACACTGTCTTGTCTCCATTTCTTGTGTCTCTTGTTCCCTTCAATTCCCACTCCCTCCTCCAGGTTCCTACTGTTGATCCCGCGGGTCGGGACAGTTGGCGCCCAACGTGGGGCTGGATACGAGGGAATTTCGTGAGGAAGACGACGCGTTCGCCGGCCGGCGATTAAAAGTGAAAGTAAACTCTCTTGGCCGCCGCGGGAACCTGCCGCGTTGGACCTGAAAGTAAGTGTTGCGCTCGGATATGGGGCAAGAATTAAGCCAGCATGAACGTTATGTAGAACAATTGAAGCAGGCTTTAAAGACACGGGGAGTAAAGGTTAAATATGCTGATCTTTTGAAATTTTTTGATTTTGTGA AGGAT (SEQ ID NO: 90)Avian sarcoma leukosis virus (ALV) packaging sequenceGCCATTTTACCTCCCACCACATTGGTGTGCACCTGGGTTGATGGCCGGACCGTCGATTCCCTGACGACTACGAGCACCTGAATGAAGCAGAAGGCTTCATCTGGTGACCCCGACGTGATAGTTAGGGAATAGTGGTCGGCCACAGACGGCGTGGCGATCCTGTCCTCATCCGTCTCGCTTATTCGGGGAGCGGGCGATGACCCTAGTAGAGGGGGCTGCGGCTTAGGAGGGCAGAAGCTGAGTGGCGTCGGAGGGAGCTCTACTGCAGGGAGCCCACGTACCCTACCGAGAACTCAGAGAGTCGTTGGAAGACGGGAAGGAAGCCCGACGACTGAGCGGTCCGCCCCAGACGTGGTTCTGGTTGCCTGGAGGATCAAGTATGGAAGCCGTCATAAAGGTGATTTCGTCCGCGTGTAAAACCTATTGCGGGAAAACCTCTCCTTCTAAGAAGGAAATAGGGGCCATGCTGTCCCTGCTACAAAAGGAAGGGTTGCTTGCGTCTCCCTCAGACTTGTACTCCCCGGGGTCCTGGGATCCCATTACCGCGGCGCTCTCCCAGCGAACAATGGTACTTGGAAAATCGGGGGAGTTAAAAACCTGGGGATTGGTTTTGGGGGCACTGAAGG (SEQ ID NO: 91)Rous sarcoma virus (RSV) packaging sequenceGATCCTGCCCTCATCCGTCTCGCTTATTCGGGGAGCGGACGATGACCCTAGTAGA GGGGGCTGCGGCTTAGGAGGGCAGAAGCTGAGTGGCGTCGGAGGGAGCTCTACT GCAGGGAGCCCAGATACCCTACCGAGAACTCAGAGAGTCGTTGGAAGACGG (SEQ ID NO: 92)

[0218] In some embodiments, the packaging signal is located downstream of the 5’ LTR. In some embodiments, the packaging signal is located downstream of the 5‘ LTR and upstream of the cargo of interest.

[0219] In some embodiments, the nucleic acid constructs of the disclosure include a packaging signal having at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the sequence of SEQ ID NO: 82 or SEQ ID NO: 83.Post-transcriptional Regulatory Element

[0220] As described herein, the integration deficient retroviral vectors of the present disclosure also include a post-transcriptional regulatory element. In some embodiments, the integration deficient retroviral vectors of the present disclosure can include more than one post transcriptional regulatory element. Post-transcriptional regulatory elements (PRE), as used herein, refer to exogenous nucleic acid sequences that can enhance expression of a cargo of interest. As used herein. PREs do not include any nucleic acid sequences derived from naturally occurring LTR elements that can enhance expression of a cargo of interest. Furthermore, in some embodiments, a PRE element is in the same 5’ to 3’ orientation relative to the 5’ to 3’ orientation of the one or more LTRs and the cargo of interest.

[0221] One ty pe of PRE is an intron positioned within the expression cassette, which can stimulate gene expression. However, introns can be spliced out during the life cycle events of a retrovirus.

[0222] Posttranscriptional regulatory elements that do not rely on splicing events offer the advantage of not being removed during the viral life cycle. Some examples are the posttranscriptional processing element of herpes simplex virus, the posttranscriptional regulatory element of the hepatitis B virus (HPRE) and the woodchuck hepatitis virus (WPRE). The WPRE is characterized and described in U.S. Pat. No. 6,136,597, incorporated herein by reference. WPRE is a tripartite regulatory7element with gamma, alpha, and beta components. HPRE shares two domains with WPRE (PREa and PRE[3) but is missing the PREgamma domain found in WPRE. As demonstrated in the Examples presented herein, theWPRE element is a useful tool for stimulating and enhancing gene expression of desired transgenes in the context of integration deficient retroviral vectors.

[0223] In some embodiments, the post-transcriptional regulatory element comprises a WPRE. In some embodiments, the WPRE is a tripartite WPRE (i.e., comprising the alpha, beta, and gamma components). In some embodiments, the WPRE comprises only the alpha component. In some embodiments, the WPRE comprises only the beta component. In some embodiments, the WPRE comprises only the gamma component. In some embodiments, the WPRE comprises any combination of two different components selected from the alpha, beta, and gamma components. In some embodiments, the WPRE comprises multiple copies of one or more of the alpha, beta, and gamma components. In some embodiments, any combination of alpha, beta, and / or gamma, whether as a single copy or multiple copies of each component.

[0224] The alpha component of WPRE has a nucleic acid sequence corresponding to SEQ ID NO: 93 below.GGTGTGCACTGTGTTTGCTGACGCAACCCCCACTGGTTGGGGCATTGCCACCACC TGTCAGCTCCTTTCCGGGACTTTCGCTTTCCCCCTCCCTATTGCCACGGCGGAACT CATCGCCGCCTGCCTTGCCCGCTGCTGGACAGGGGCTCGGCTGTTGGGCACTGAC AATTCCGTGGTGTTGTCGGGGAAGCTGACGTCCTTTCCAT

[0225] The beta component of WPRE has a nucleic acid sequence corresponding to SEQ ID NO: 94 below.GGCTGCTCGCCTGTGTTGCCACCTGGATTCTGCGCGGGACGTCCTTCTGCTACGT CCCTTCGGCCCTCAATCCAGCGGACCTTCCTTCCCGCGGCCTGCTGCCGGCTCTG CGGCCTCTTCCGCGTCTTCGCCTTCGCCCTCAGACGAGTCGGATCTCCCTTTGGGC CGCCTCCCCGC

[0226] The gamma component of WPRE has a nucleic acid sequence corresponding to SEQ ID NO: 95 below.AATCAACCTCTGGATTACAAAATTTGTGAAAGATTGACTGGTATTCTTAACTATG TTGCTCCTTTTACGCTATGTGGATACGCTGCTTTAATGCCTTTGTATCATGCTATT GCTTCCCGTATGGCTTTCATTTTCTCCTCCTTGTATAAATCCTGGTTGCTGTCTCTT TATGAGGAGTTGTGGCCCGTTGTCAGGCAACGTGGCGT

[0227] In some embodiments, the WPRE comprises a nucleic acid sequence as shown in SEQ ID NO: 96 below: AATCAACCTCTGGATTACAAAATTTGTGAAAGATTGACTGGTATTCTTAACTATG TTGCTCCTTTTACGCTATGTGGATACGCTGCTTTAATGCCTTTGTATCATGCTATT GCTTCCCGTATGGCTTTCATTTTCTCCTCCTTGTATAAATCCTGGTTGCTGTCTCTTTATGAGGAGTTGTGGCCCGTTGTCAGGCAACGTGGCGTGGTGTGCACTGTGTTTG CTGACGCAACCCCCACTGGTTGGGGCATTGCCACCACCTGTCAGCTCCTTTCCGG GACTTTCGCTTTCCCCCTCCCTATTGCCACGGCGGAACTCATCGCCGCCTGCCTTG CCCGCTGCTGGACAGGGGCTCGGCTGTTGGGCACTGACAATTCCGTGGTGTTGTC GGGGAAGCTGACGTCCTTTCCATGGCTGCTCGCCTGTGTTGCCACCTGGATTCTG CGCGGGACGTCCTTCTGCTACGTCCCTTCGGCCCTCAATCCAGCGGACCTTCCTTC CCGCGGCCTGCTGCCGGCTCTGCGGCCTCTTCCGCGTCTTCGCCTTCGCCCTCAGA CGAGTCGGATCTCCCTTTGGGCCGCCTCCCCGCCTG

[0228] In some embodiments, the nucleic acid constructs of the disclosure include a WPRE having at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the sequence of SEQ ID NO: 93, 94, 95, or, 96.

[0229] In some embodiments, the WPRE comprises a nucleic acid sequence as shown in SEQ ID NO: 835 below: AATCAACCTCTGGATTACAAAATTTGTGAAAGATTGACTGGTATTCTTAACTATG TTGCTCCTTTTACGCTATGTGGATACGCTGCTTTAATGCCTTTGTATCATGCTATT GCTTCCCGTATGGCTTTCATTTTCTCCTCCTTGTATAAATCCTGGTTGCTGTCTCTTTATGAGGAGTTGTGGCCCGTTGTCAGGCAACGTGGCGTGGTGTGCACTGTGTTTG CTGACGCAACCCCCACTGGTTGGGGCATTGCCACCACCTGTCAGCTCCTTTCCGG GACTTTCGCTTTCCCCCTCCCTATTGCCACGGCGGAACTCATCGCCGCCTGCCTTG CCCGCTGCTGGACAGGGGCTCGGCTGTTGGGCACTGACAATTCCGTGGTGTTGTC GGGGAAATCATCGTCCTTTCCTTGGCTGCTCGCCTGTGTTGCCACCTGGATTCTGCGCGGGACGTCCTTCTGCTACGTCCCTTCGGCCCTCAATCCAGCGGACCTTCCTTCC CGCGGCCTGCTGCCGGCTCTGCGGCCTCTTCCGCGTCTTCGCCTTCGCCCTCAGAC GAGTCGGATCTCCCTTTGGGCCGCCTCCCCGC

[0230] In some embodiments, the nucleic acid constructs of the disclosure include a WPRE having at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the sequence of SEQ ID NO: 835.

[0231] In some embodiments, the post-transcriptional regulatory element comprises an HPRE. In some embodiments, the HPRE comprises only the alpha component. In some embodiments, the HPRE comprises only the beta component. In some embodiments, the HPRE comprises multiple copies of one or more of the alpha and beta components. In some embodiments, the HPRE comprises a nucleic acid sequence as shown in SEQ ID NO: 97below (single underline indicates the HPRE alpha component sequence; the remainder is the HPRE beta component sequence):TTGCTCGGCAACGGCCTGGTCTGTGCCAAGTGTTTGCTGACGCAACCCCCACTGG TTGGGGCTTGGCCATAGGCCATCAGCGCATGCGTGGAACCTTTGTGTCTCCTCTG CCGATCCATACTGCGGAACTCCTAGCCGCTTGTTTTGCTCGCAGCAGGTCTGGAG CAAACCTCATCGGGACCGACAATTCTGTCGTACTCTCCCGCAAGTATACATCGTT TCCATGGCTGCTAGGCTGTGCTGCCAACTGGATCCTGCGCGGGACGTCCTTTGTT TACGTCCCGTCGGCGCTGAATCCCGCGGACGACCCCTCCCGGGGCCGCTTGGGGC TCTACCGCCCGCTTCTCCGTCTGCCGTACCGTCCGACCACGGGGCGCACCTCTCTT TACGCGGACTCCCCGTCTGTGCCTTCTCATCTGCCGGACCGTGTGCACTTCGCTTC ACCTCTGCACGTCGCATGGAGACCACCGTGAACGCCCACCGGAACCTGCCCAAG GTCTTGCATAAGAGGACTCTTGGACTTTCAGCAATGTC

[0232] In some embodiments, the alpha component of HPRE has a nucleic acid sequence corresponding to SEQ ID NO: 827 below.TTGCTCGGCAACGGCCTGGTCTGTGCCAAGTGTTTGCTGACGCAACCCCCACTGG TTGGGGCTTGGCCATAGGCCATCAGCGCATGCGTGGAACCTTTGTGTCTCCTCTG CCGATCCATACTGCGGAACTCCTAGCCGCTTGTTTTGCTCGCAGCAGGTCTGGAG CAAACCTCATCGGGACCGACAATTCTGTCGTACTCTCCCGCAAGTATACATCGTT TCCATGGCTGCTAGGCTGTGCTGCCAACTGGATCCTGCGCGGG

[0233] In some embodiments, the beta component of HPRE has a nucleic acid sequence corresponding to SEQ ID NO: 828 below.ACGTCCTTTGTTTACGTCCCGTCGGCGCTGAATCCCGCGGACGACCCCTCCCGGG GCCGCTTGGGGCTCTACCGCCCGCTTCTCCGTCTGCCGTACCGTCCGACCACGGG GCGCACCTCTCTTTACGCGGACTCCCCGTCTGTGCCTTCTCATCTGCCGGACCGTG TGCACTTCGCTTCACCTCTGCACGTCGCATGGAGACCACCGTGAACGCCCACCGG AACCTGCCCAAGGTCTTGCATAAGAGGACTCTTGGACTTTCAGCAATGTC

[0234] In some embodiments, the nucleic acid constructs of the disclosure include an HPRE having at least 50%, at least 60%, at least 70%, at least 80%. at least 85%. at least 90%. at least 95%. at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to the sequence of SEQ ID NOs: 97, 827, or 828.

[0235] The PRE can be located at any position relative to the other elements in the nucleic acid construct. In some embodiments, the PRE is located 3’ to the cargo of interest. In some embodiments, the PRE is located between the cargo of interest and the 3' LTR. In some embodiments, the PRE is located 3’ to the cargo of interest and the PRE is part of the 3’untranslated region (3’ UTR) of the transcript of the cargo of interest. In some embodiments, the PRE is comprised of alpha, beta, and / or gamma components from both WPRE and HPRE. As used herein, a PRE is not considered an exogenous promoter.

[0236] Potential PRE elements with sequence homology to the full WPRE or EIPRE sequences (SEQ ID NO:96 or 97 respectively) or to portions thereof (e.g., alpha and / or beta components for WPRE or HPRE, or alpha, beta, and / or gamma components for WPRE) can be functionally tested to confirm that the potential element can provide expression of one or more cargos of interest in the absence of an exogenous promoter. The functional testing can be as described for reporter gene expression as in Figures 1, 2, and 6. Potential PRE elements with sequence homology to WPRE or HPRE (full and portions thereof) are provided in Tables 1-7 below. The query sequence is indicated in the title of the Tables, i.e., the WPRE alpha, beta, gamma, or full-length sequence (SEQ ID NO:29, 30, 31, or 32, respectively) or the HPRE alpha, beta, or full-length sequence (SEQ ID NO:33 or alpha / beta portion thereof). The potential PRE sequences with homology to the query sequences are listed in the Tables under the column “Hit Sequence." The source of the Hit Sequence can be determined by searching the Genbank database of the National Center for Biotechnology Information via its accession number listed in the Tables. The potential PRE sequences may include IUPAC single letter codes to indicate that more than one type of nucleotide can be present at a position (B = C or G or T; D = A or G or T; H = A or C or T; K = G or T; M = A or C; N = A or C or G or T: R = A or G; S = C or G; V = A or C or G; W = A or T; Y = C or T). Thus, in some embodiments, the PRE element comprises a sequence listed in Tables 1-7.

[0237] TABLE 1: QUERY WITH WPRE-ALPHA SEQUENCE - POTENTIAL PRE SEQUENCES WITH HOMOLOGY TO WPRE-ALPHA

[0238] TABLE 2: QUERY WITH WPRE-BETA SEQUENCE - POTENTIAL PRE SEQUENCES WITH HOMOLOGY TO WPRE-BETAIll

[0239] TABLE 3: QUERY WITH WPRE-GAMMA SEQUENCE - POTENTIAL PRE SEQUENCES WITH HOMOLOGY TO WPRE-GAMMA

[0240] TABLE 4: QUERY WITH WPRE (FULL) SEQUENCE - POTENTIAL PRE SEQUENCES WITH HOMOLOGY TO WPRE (FULL)GTCCTTTCCATGGCTGCTCGCCTGTGTTGCCAACTGGATTCTGCGCGGGACGTCCTTCTGCTACGTCCCTTCGGCCCTCAATCCAGCGGACCTTCCTTCCCGCGGCCTGCTGCCGGTTCTGCGGCCTCTTCCGCGTCTTCGCCTTCGCCCTCAGACGAGTCGGATCTCCCTTTGGGCCGCCTCCCCGC SEQ ID NO:399GCGCATGCGTGGGACCTTTGTGTCTCCTCTGCCGATCCATACTGCGGAACTCCTCGCCGCCTGTTTTG CTCG CAG CAG GTCTG G GG CAAAACTCATCG G G ACTG ACAATTCTGTCGTGCTCTCCCG CAAGTATACATCGTTTCCATGGCTGCTAGGCTGTGCTGCCAACTGGATCCTGCGCGGGACGTCCTTTGTTTACGTCCCGTCG G CG CTG AATCCCG CG G ACG ACCCCTCCCGG G G CC-G CTTG G G G CTCTACCG C- —CCGCTTCTCCGCCT SEQ ID NO: 442GTCCCGTCGGCGCTGAATCCCGCGGACGACCCCTCCCGGGGCC-GCTTGGGGCTCTACCGC -CCGCTTCTCCGCCT SEQ ID NO: 449GCTCGCAGCAGGTCTGGGGCAAAACTCATCGGGACTGACAATTCTGTCGTGCTCTCCCGCAAGTAT ACATCCTTTCCATG G CTG CTAGG CTGTGCTG CCAACTG G ATCCTG CG CG G G ACGTCCTTTGTTTAC GTCCCGTCG G CG CTG AATCCCG CG G ACG ACCCCTCCCGG G G CC-G CTTG G G G CTCTACCG C- — CCG CTTCTCCG CCTGTTGTACCG ACCTTCCACG G G G CG CACCTCTCTTTACG CG G CCTCCCCG S EQID NO: 457GCTCGCAGCAGGTCTGGGGCAAAACTCATCGGGACTGACAATTCTGTCGTGCTCTCCCGCAAGTATACATCCTTTCCATG G CTG CTAGG CTGTGCTG CCAACTG G ATCCTG CG CG G G ACGTCCTTTGTTTACGTCCCGTCG G CG CTG AATCCCG CG G ACG ACCCCTCCCGG G G CC-G CTTG G G G CTCTACCG C- —CCGCTTCTCCGCCT SEQ ID NO: 464GCGCATGCGTGGGACCTTTGTGTCTCCTCTGCCGATCCATACTGCGGAACTCCTAGCCGCTTGTTTT GCTCGCAGCAGGTCTGGGGCGAACCTCATCGGGACTGACAATTCTGTCGTGCTCTCCCGCAAGTAT ACATCCTTTCCATG G CTG CTAGG CTGTGCTG CCAACTG G ATCCTG CG CG G G ACGTCCTTTGTTTAC GTCCCGTCG G CG CTG AATCCCG CG G ACG ACCCCTCCCGG G G CC-G CTTG G G G CTCTACCG C- —CCGCTTCTCCGCCT SEQ ID NO: 475TGCTGCCCCTTTCACGCAATGTGGATATCCTGCTTTAAAGCCTATATATGCATG-TATACAAG CAAAACAG G CTTTTAC I I I I I CG CCAACTT— ACAAG GCCTTTCTGC- GTCAACAGTATCTGAACCTTTACCCCGTTGCTCGGCAACG—G CCTG GTCTGTG CCAAGTGTTTG CTG ACG CAACCCCCACTGGTTG GG G CTTG G CCATAG G CCATCA G CG CATG CGTG G AACCTTTGTGTCTCCTCTGCCG ATCCATACTG CG G AACTCCTAG CCG CCTGTTTT G CTCG CAG CAG GTCTG G GG CAAAACTCATCG G G ACTG ACAATTCTGTCGTGCTCTCCCG CAAGTAT ACATCATTTCCATGGCTGCTCGGCTGTGCTGCCAACTGGATCCTGCGCGGGACGTCCTTTGTTTAC0 M 66957 GTCCCGTCGGCG CTG AATCCCG CG G ACG ACCCCTCCCGG G G CC-G CTTG G G G CTCTACCG C- —0 CCGCTTCTCCGCCT SEQ ID NO: 479GCTCGCAGCAGGTCTGGGGCAAAACTCATCGGGACTGACAATTCTGTCGTGCTCTCCCGCAAGTATACATCCTTTCCATG G CTG CTAGG CTGTGCTG CCAACTG G ATCCTG CG CG G G ACGTCCTTTGTTTACGTCCCGTCG G CG CTG AATCCCG CG G ACG ACCCCTCCCGG G G CC-G CTTG G G G CTCTACCG C- —CCGCTTCTCCGCCT SEQ ID NO: 487

[0241] TABLE 5: QUERY WITH HPRE-ALPHA SEQUENCE - POTENTIAL PRE SEQUENCES WITH HOMOLOGY TO HPRE-ALPHA

[0242] TABLE 6: QUERY WITH HPRE-BETA SEQUENCE - POTENTIAL PRE SEQUENCES WITH HOMOLOGY TO HPRE-BETA

[0243] TABLE 7: QUERY WITH HPRE (FULL) SEQUENCE - POTENTIAL PRE SEQUENCES WITH HOMOLOGY TO HPRE (FULL)Promoters

[0244] As demonstrated in the Examples provided herein, the compositions of the present disclosure can also include an exogenous promoter. As used herein, a promoter is referred to in its traditional sense as a sequence that allows a polymerase complex to initiate transcription of a target (e.g., cargo) gene. In standard retroviral vector production, a strong, exogenous, heterologous promoter is positioned upstream of the cargo of interest in order to drive expression of the cargo. For example, in many instances, exogenous promoters including, but not limited to, CMV, CMV IE, phosphoglycerate kinase- 1 (PGK), a simian virus 40 (SV40), EFla, SFFV, RSV, and Ubc are positioned upstream of a cargo of interest.

[0245] In some embodiments, recombinant integration deficient retroviral vector of the present disclosure includes one or more RNA polymerase II promoters. Exemplary RNA polymerase II promoters include, without limitation, CMV promoter, SFFV promoter, a PGK promoter, SV40 promoter, a CK6 promoter, a transthyretin promoter (TTR), a TK promoter, a tetracycline responsive promoter (TRE), an HBV promoter, an hAAT promoter, a LSP promoter, chimeric liver- specific promoters (LSPs), a E2F promoter, a EFla promoter, a telomerase (hTERT) promoter, and a cytomegalovirus enhancer / chicken beta- actin / Rabbit 0- globin promoter (CAG) promoter. In one embodiment, the promoter is an SFFV promoter.

[0246] In some embodiments, recombinant integration deficient retroviral vector of the present disclosure includes one or more RNA polymerase III promoters. Exemplar}' human RNA polymerase III promoters include, without limitation, human Hl, U6. 5S rRNA, 7SK and tRNA promoters.

[0247] In some embodiments, the compositions described herein contain both an RNA polymerase II and an RNA polymerase III promoter. In some embodiments, the compositions described herein contain one or more polymerase II and one or more polymerase III promoters. The choice and number of promoters used in the IDLV constructs of the present disclosure depend upon the number and ty pe of each cargo being expressed by the construct. For example, in some embodiments, a cargo of interest, such as a chimeric antigen receptor, can require an RNA polymerase II promoter, and the nucleic acid sequence encoding an RNA interference molecule or molecules that target one or more proteins involved in the epigenetic silencing of viral DNA require an RNA polymerase III promoter. In another embodiment, a cargo of interest such as a CRISPR protein, can require an RNA polymerase II promoter and another cargo of interest, such as an sgRNA, can require an RNA polymerase III promoter. Such a vector would also include the nucleic acid sequence encoding an RNA interference molecule or molecules that target one or more proteins involved in the epigenetic silencing of viral DNA under the control of another RNA polymerase III promoter.

[0248] In some embodiments, the compositions described herein include a modified promoter. For example, the promoters used in the composition described herein may be modified or altered to reduce or enhance their control characteristics. For example, as shown in the Examples herein, promoters can be modified to remove DNA binding sites for transcription factors involved in gene silencing. Sequences can be modified, for example by insertion, deletion, or replacement of template sequences in a PCR-based DNA modification approach. In some embodiments, a promoter contains changes in which one or more nucleotides of an original promoter is deleted, added, and / or substituted, preferably while substantially maintaining promoter function. For example, one or more base pairs may be deleted from the 5' or 3' end of a promoter to produce a "truncated" promoter. One or more base pairs can also be inserted, deleted, or substituted internally to a promoter. Modified promoters can be produced, for example, by7standard DNA mutagenesis techniques or by chemically synthesizing the modified promoter or a portion thereof. Those of skill in the art are familiar with the standard resource materials that describe specific conditions and procedures for the construction, manipulation, and isolation of macromolecules (e.g.. polynucleotide molecules, plasmids, etc.), as well as the generation of recombinant organisms and the screening and isolation of polynucleotide molecules.

[0249] In some embodiments, the modified promoter is a modified SFFV promoter.

[0250] In some embodiments, the SFFV promoter that is modified comprises the original promoter sequence ofGTAACGCCATTTTGCAAGGCATGGAAAAATACCAAACCAAGAATAGAGAAGTTC AGATCAAGGGCGGGTACATGAAAATAGCTAACGTTGGGCCAAACAGGATATCTG CGGTGAGCAGTTTCGGCCCCGGCCCGGGGCCAAGAACAGATGGTCACCGCAGTT TCGGCCCCGGCCCGAGGCCAAGAACAGATGGTCCCCAGATATGGCCCAACCCTC AGCAGTTTCTTAAGACCCATCAGATGTTTCCAGGCTCCCCCAAGGACCTGAAATG ACCCTGCGCCTTATTTGAATTAACCAATCAGCCTGCTTCTCGCTTCTGTTCGCGCG CTTCTGCTTCCCGAGCTCTATAAAAGAGCTCACAACCCCTCACTCGGCGCGCCAG TCCTCCGACAGACTGAGTCGCCCGGG (SEQ ID NO: 828)In some embodiments, the modified SFFV promoter comprises the sequence of GTAACGCCATTTTGCAAGGCATGGAAAAATACCAAACCAACTAACGTTGGGCCA AACAGGATATCTGCGGTGAGCAGTTTCGGCCCCGGCCCGGGGCCAAGAACAGAT GGTCACCGCAGTTTCGGCCCCGGCCCGAGGCCAAGAACAGATGGTCCCCAGATA TGGCCCAACCCTCAGCAGTTTCTTAAGACCCATCAGATGTTTCCAGGCTCCCCCA AGGACCTGAAATGACCCTGCGCCTTATTTGAATTAACCAATCAGCCTGCTTCTCG CTTCTGTTCGCGCGCTTCTGCTTCCCGAGCTCTATAAAAGAGCTCACAACCCCTC ACTCGGCGCGCCAGTCCTCCGACAGACTGAGTCGCCCGGG (SEQ ID NO: 829)

[0251] In some embodiments, the modified SFFV promoter comprises the sequence of GTAACGCCATTTTGCAAGGCATGGAAAAATACCAAACCAAGAATAGAGAAGTTC AGATCAAGGGCGGGTACATGAAAATATCGGCCCCGGCCCGGGGCCAAGAACAG ATGGTCACCGCAGTTTCGGCCCCGGCCCGAGGCCAAGAACAGATGGTCCCCAGA TATGGCCCAACCCTCAGCAGTTTCTTAAGACCCATCAGATGTTTCCAGGCTCCCC CAAGGACCTGAAATGACCCTGCGCCTTATTTGAATTAACCAATCAGCCTGCTTCT CGCTTCTGTTCGCGCGCTTCTGCTTCCCGAGCTCTATAAAAGAGCTCACAACCCC TCACTCGGCGCGCCAGTCCTCCGACAGACTGAGTCGCCCGGG (SEQ ID NO: 830)

[0252] In some embodiments, the modified SFFV promoter comprises the sequence ofGTAACGCCATTTTGCAAGGCATGGAAAAATACCAAACCAAGAATAGAGAAGTTC AGATCAAGGGCGGGTACATGAAAATAGCTAACGTTGGGCCAAACAGGATATCTG CGGTGAGCAGTTTTTCGGCCCCGGCCCGAGGCCAAGAACAGATGGTCCCCAGAT ATGGCCCAACCCTCAGCAGTTTCTTAAGACCCATCAGATGTTTCCAGGCTCCCCC AAGGACCTGAAATGACCCTGCGCCTTATTTGAATTAACCAATCAGCCTGCTTCTC GCTTCTGTTCGCGCGCTTCTGCTTCCCGAGCTCTATAAAAGAGCTCACAACCCCT CACTCGGCGCGCCAGTCCTCCGACAGACTGAGTCGCCCGGG (SEQ ID NO: 831)

[0253] In some embodiments, the modified SFFV promoter comprises the sequence ofGTAACGCCATTTTGCAAGGCATGGAAAAATACCAAACCAAGAATAGAGAAGTTC AGATCAAGGGCGGGTACATGAAAATAGCTAACGTTGGGCCAAACAGGATATCTG CGGTGAGCAGTTTCGGCCCCGGCCCGGGGCCAAGAACAGATGGTCACCGCAGAT ATGGCCCAACCCTCAGCAGTTTCTTAAGACCCATCAGATGTTTCCAGGCTCCCCC AAGGACCTGAAATGACCCTGCGCCTTATTTGAATTAACCAATCAGCCTGCTTCTC GCTTCTGTTCGCGCGCTTCTGCTTCCCGAGCTCTATAAAAGAGCTCACAACCCCT CACTCGGCGCGCCAGTCCTCCGACAGACTGAGTCGCCCGGG (SEQ ID NO: 832)

[0254] In some embodiments, the modified SFFV promoter comprises the sequence of GTAACGCCATTTTGCAAGGCATGGAAAAATACCAAACCAAGAATAGAGAAGTTC AGATCAAGGGCGGGTACATGAAAATAGCTAACGTTGGGCCAAACAGGATATCTG CGGTGAGCAGTTTCGGCCCCGGCCCGGGGCCAAGAACAGATGGTCACCGCAGTT TCGGCCCCGGCCCGAGGCCAAGAACAGATGGTCCCCAGATATGGCCCAACCCTC AGCAGTTTCTTAAGACCCATCAGATGTTTCCAGGCTCCCCCAAGGACCTGAAATG ACCCTGCGCTCGCGCGCTTCTGCTTCCCGAGCTCTATAAAAGAGCTCACAACCCC TCACTCGGCGCGCCAGTCCTCCGACAGACTGAGTCGCCCGGG (SEQ ID NO: 833)

[0255] In some embodiments, the modified SFFV promoter comprises the sequence of GTAACGCCATTTTGCAAGGCATGGAAAAATACCAAACCAAGAATAGAGAAGTTC AGATCAAGGGCGGGTACATGAAAATAGCTAACGTTGGGCCAAACAGGATATCTG CGGTGAGCAGTTATATGGCCCAACCCTCAGCAGTTTCTTAAGACCCATCAGATGT TTCCAGGCTCCCCCAAGGACCTGAAATGACCCTGCGCCTTATTTGAATTAACCAA TCAGCCTGCTTCTCGCTTCTGTTCGCGCGCTTCTGCTTCCCGAGCTCTATAAAAGA GCTCACAACCCCTCACTCGGCGCGCCAGTCCTCCGACAGACTGAGTCGCCCGGG (SEQ ID NO: 834)

[0256] In some embodiments, the modified SFFV promoter comprises a sequence having at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ ID NOs: 829-834.

[0257] In some embodiments, the compositions described herein contain no promoter at all. In some embodiments, the only promoter in the compositions described herein is a naturally occurring promoter in an LTR. In some embodiments, the compositions contain no promoter element between the end of the 5’ LTR and the cargo of interest. The person of ordinary skill knows how to determine the “end of the 5 ’ LTR” as retroviral LTR sequences are well characterized. For example, both 5’ and 3’ LTRs sometimes end with the dinucleotide “CA.” Accordingly, in some embodiments, the compositions described herein contain no promoterbetween the 5 ’ LTR dinucleotide and the cargo of interest. This includes both endogenous and exogenous promoters in between the end of the 5’ LTR and the cargo of interest as well as any promoter elements taken out of their natural sequence context (such as a portion of an LTR). Thus, in some embodiments, the only promoter sequence present in the compositions of the invention is the endogenous, naturally occurring promoter in an LTR, where the promoter is maintained in its natural sequence context of the LTR.

[0258] A person of ordinary skill in the art will understand that the LTRs themselves contain promoter sequences. However, in this context, such promoters represent endogenous promoters located within the LTR itself. In some embodiments, the recombinant integration defective retroviral vector of the present disclosure does not contain an exogenous LTR promoter.Cargos of Interest

[0259] As described above, the integration deficient retroviral vectors of the disclosure also include a nucleic acid sequence encoding one or more cargos of interest.

[0260] As used herein, a '‘cargo’’ includes one or more nucleic acid sequences of interest and refers to a nucleic acid that is transcribed. In some embodiments, where the cargo comprises two different nucleic acid sequences that are to be translated as separate protein molecules, the two different nucleic acid sequences are separated by an internal ribosomal entry site (IRES) sequence, or another connector sequence (e.g., a P2A sequence) as described below, such that the cargo is transcribed as a single molecule but translated into two different protein molecules, or translated as one polyprotein but subsequently proteolytically cleaved into two individual proteins.

[0261] In some embodiments, the cargo of interest is selected from a gene-editing nuclease, a gene-editing nickase, endonuclease deficient Cas, endonuclease deficient Cas effectors, endonuclease deficient Cas enzy mes, Cas nuclease fusion proteins, Cas nickase fusion proteins, endonuclease deficient Cas fusion proteins, engineered DNA binding protein, base editor, prime editor, epigenome editors, bridge editor, RNA interference (RNAi). RNA targeting system. CRISPR DNA binding protein, CR1SPR nickase, recombinase, programmable integrase, transposons, retrotransposons, transposase, chimeric antigen receptor (CAR), T cell receptors, HLA-independent T cell receptors, synNotch receptors, cytokine receptors, synthetic intramembrane proteolysis receptors, transcription factor, caspase, natural protease, programmable protease, reporter gene, and a selection marker.

[0262] Gene-editing nucleases are well known in the art. Any gene-editing nuclease as is known in the art can be encoded in the vectors of the present disclosure including, without limitation, zinc finger nucleases (ZFNs), Transcription Activator-Like Effector Nucleases (TALENs), nucleases of the CRISPR / Cas system.

[0263] In some embodiments, the one or more cargos of interest is a nuclease of the CRISPR / Cas sy stem. By w ay of example, CRISPR-based endonucleases include RNA- guided endonucleases that comprise at least one nuclease domain and at least one domain that interacts with a guide RNA. As used herein, a guide RNA can refer to an RNA required for the function and targeting of an RNA guided Cas protein. A guide RNA directs the CRISPR- based endonucleases to a targeted site in a nucleic acid at which site the CRISPR-based endonucleases cleaves at least one strand of the targeted nucleic acid sequence. As the guide RNA provides the speci ficity for the targeted cleavage, the CRISPR-based endonuclease is universal and can be used with different guide RNAs to cleave different target nucleic acid sequences. Thus, in some embodiments, the one or more cargos comprises a guide RNA. Guide RNAs include, without limitation, CRISPR RNA (crRNA), transactivating CRISPR RNA (tracrRNA), single guide RNA (sgRNA), and modified guide RNAs.

[0264] In some embodiments, the guide RNA is a crRNA. As is known in the art, crRNA is a guide RNA for a Cas protein. Some Cas enzymes naturally only have a crRNA and no tracrRNA. Examples include Casl2 and Casl3.

[0265] In some embodiments, the guide RNA is a tracrRNA. tracrRNA is a second RNA sometimes required in addition to the crRNA for RNA guided Cas proteins such as Cas9. tracrRNAs are described in the art, e.g., in Faure et al., “Comparative Genomics and Evolution of Trans -Activating RNAs in Class2 CRISPR-Cas Systems,” RNA Biol. 16(4):435-448 2019.

[0266] In some embodiments, the guide RNA is an sgRNA. sgRNAs are usually a chimeric fusion with an artificial linker between a crRNA and a tracrRNA. sgRNAs are described in, for example, Jinek et al., “A Programmable Dual-RNA-Guided DNA Endonuclease in Adaptive Bacterial Immunity." Science 337(6096):816-821 2012.

[0267] In some embodiments, the guide RNA is a modified guide RNA. Guide RNAs can be modified to include additional RNA sequences that act as aptamers to recruit additional proteins such as MCP resulting in a ribonucloprotein complex composed of a guide RNA, a Cas protein and another RNA binding protein. Both the Cas protein and the RNA binding protein can be fused to additional proteins resulting in non-natural fusion proteins. This is described, for example, in the SAM system (Konermann et al., “Genome-ScaleTranscriptional Activation by an Engineered CRISPR-Cas9 Complex,’" Nature 517(7536):583-588 2015.

[0268] CRISPR-based endonucleases are RNA-guided endonucleases derived from CRISPR / Cas systems. In an embodiment, a disclosed CRISPR-based endonuclease can be derived from a CRISPR / Cas type I, ty pe II, ty pe III, ty pe V, or ty pe VI system. Non-limiting examples of suitable CRISPR / Cas proteins include Cas3, Cas4, Cas5, Cas5e (or CasD), Cas6, Cas6e, Cas6f, Cas7, Cas8al, Cas8a2, Cas8b. Cas8c. Cas9. Casio, CaslOd. Cast 2 (including subtypes Cas 12a (formerly Cpfl), Cas l2b (previously C2C 1), and Casl2f (also known as Casl4)), Cas 13 (including subty pes Casl3a (aka C2c2), Casl3b, Casl3c, and Casl3d), CasF, CasG, CasH, Csyl, Csy2, Csy3, Csel (or CasA), Cse2 (or CasB), Cse3 (or CasE), Cse4 (or CasC), Cscl, Csc2, Csa5. Csn2, Csm2. Csm3, Csm4, Csm5, Csm6, Cmrl, Cmr3, Cmr4. Cmr5, Cmr6, Csbl, Csb2, Csb3, Csxl7, Csxl4, CsxlO, Csxl6, CsaX, Csx3, CszL Csxl5, Csfl, Csf2, Csf3, Csf4, Cul966, and orthologs thereof. All of these nuclease enzy mes can be rendered nickases or endonuclease deficient DNA binding proteins by mutation of one / both endonuclease domains. Further, all of these nuclease enzymes can be in the form of a fusion protein.

[0269] Endonuclease deficient Cas proteins are also contemplated for use herein. An exemplary, endonuclease deficient Cas includes dCas9 (Xu et al., “CRISPR / Cas Derivatives and Novel Gene Modulating Tools: Possibilities and In Vivo Applications,” Int J Mol Sci 21(0:3038 (2020).

[0270] Further contemplated for use herein are endonuclease deficient Cas effectors and endonuclease deficient Cas enzy mes. Examples include, without limitation, CRISPR activation (CRISPRa), CRISPR interference (CRISPRi), CRISPRoff,CRISPRon and CHARM In some embodiments, the one or more cargos comprises a CRISPRi.

[0271] In some embodiments, the integration deficient retroviral vector of the disclosure carries cargo comprising both a CRISPR endonuclease and a guide RNA (gRNA). In some of such embodiments, the minimal elements of an IDRV construct are: a 5’ LTR, a packaging signal, a coding sequence for a CRISPR endonuclease (including endonuclease deficient Cas proteins, Cas fusion proteins, or any wild-type or engineered Cas protein that associates with gRNA), a PRE, an RNA pol Ill-based promoter(s), and the coding sequence for a guide RNA. This construct can further minimally comprise a 3’ LTR downstream of the coding sequence for the guide RNA. In some embodiments, this construct contains an RNA polymerase II promoters 5’ of the CRISPR endonuclease. In some embodiments, the RNA polymerase II promoter is a modified SFFV promoter as described herein. In someembodiments, this construct contains no promoter between the end of the 5’ LTR and the beginning of the coding sequence for the CRISPR endonuclease; the only exogenous or ectopic promoter is the RNA pol Ill-based promoter(s) for guide RNA expression. In other such embodiments, the construct can also contain an RNA pol Ill-based promoter for expression of an RNA interference molecule as described herein, such as an shRNA targeting a protein involved epigenetic silencing.

[0272] As described above, engineered DNA binding proteins are also contemplated as cargo for use herein. Exemplary engineered DNA binding proteins include, without limitation, ZFPs, TALEs, and engineered Cas9 endonucleases. A ZFP, or zinc finger protein, is a protein, or a domain within a larger protein, that binds DNA in a sequence-specific manner through one or more zinc fingers, which are regions of amino acid sequence within the binding domain whose structure is stabilized through coordination of a zinc ion. Engineered DNA binding proteins can be fusion proteins to impart additional protein functions.

[0273] A "TALE DNA binding domain" or "TALE" is a polypeptide comprising one or more TALE repeat domains / units. The repeat domains are involved in binding of the TALE to its cognate target DNA sequence. A single "repeat unit" (also referred to as a "repeat") is typically 33-35 amino acids in length and exhibits at least some sequence homology with other TALE repeat sequences within a naturally occurring TALE protein. See, e.g., U.S. Patent Publication No. 20110301073, incorporated by reference herein in its entirety. Engineered TALE proteins can be fusion proteins to impart additional protein functions.

[0274] An engineered Cas9 endonuclease includes those that are engineered to have altered PAM specificity (Walton et al., “Unconstrained Genome Targeting with Near-PAMless Engineered CRISPR-Cas9 Variants,” Science 368(6488):290-296 (2020). Engineered Cas proteins can be fusion proteins to impart additional protein functions.

[0275] Zinc finger and TALE binding domains can be "engineered" to bind to a predetermined nucleotide sequence, for example via engineering (altering one or more amino acids) of the recognition helix region of a naturally occurring zinc finger. Similarly, TALEs can be "engineered" to bind to a predetermined nucleotide sequence, for example by engineering of the amino acids involved in DNA binding (the repeat variable diresidue or RVD region). Therefore, engineered DNA binding proteins (zinc fingers or TALEs) are proteins that are non-naturally occurring. Non- limiting examples of methods for engineering DNA-binding proteins are design and selection. A designed DNA binding protein is a protein not occurring in nature whose design / composition results principally from rational criteria.Rational criteria for design include application of substitution rules and computerized algorithms for processing information in a database storing information of existing ZFP and / or TALE designs and binding data. See, for example, U.S. Patents 6,140,081;6,453,242; and 6,534,261; see also WO 98 / 53058; WO 98 / 53059; WO 98 / 53060; WO 02 / 016536 and WO 03 / 016496 and U.S. Publication Nos. 201 10301073, 20110239315 and 20119145940.

[0276] Base editors can also be used as cargo of interest in the vectors of the disclosure. Base editors are described in Kantor et al.. “CRISPR-Cas9 DNA Base-Editing and Prime Editing,” International Journal of Molecular Sciences 21 :6240 (2020).

[0277] Prime editors for use as cargo in the vectors of the present disclosure include, without limitation, components of the twin prime editing system (Anzalone et al., “Programmable Deletion, Replacement, Integration and Inversion of Large DNA Sequences with Twin Prime Editing,” Nature Biotechnology 40:731-740 (2022). Prime editing is also described in Scholefield et al., “Prime Editing - an Update on the Field,” Gene Therapy 28:396-401 (2021) and Kantor et al.. “CRISPR-Cas9 DNA Base-Editing and Prime Editing,” International Journal of Molecular Sciences 21 :6240 (2020).

[0278] DNA polymerase editors can be used as cargo of interest in the vectors of the disclosure. DNA polymerase editors are described in Liu et al., “Targeted genome editing with a DNA-dependent DNA polymerase and exogenous DNA-containing templates” DOI: 10. 1038 / s41587-023-01947-w.

[0279] Bridge editors can be used as cargo of interest in the vectors of the disclosure. Bridge editors are described in Durrant et al., “Bridge RNAs direct modular and programmable recombination of target and donor DNA,” https : / / www. biorxi v . org / content / 10.1101 / 2024.01.24.577089 v 1.

[0280] Epigenome editors, including but not limited to CRISPRoff and CRISPRon, can be used as cargo of interest in the vectors of the disclosure. CRISPRoff and CRISPRon are described in Nunez et al., "Genome-wide programmable transcriptional memory by CRISPR- based epigenome editing,” Cell 184(9):2503-2519 (2021). Epigenome editors can also be composed of zinc fingers or TALEs as described in Cappelluti et al. “Durable and efficient gene silencing in vivo by hit-and-run epigenome editing” Nature 627. pages416-423 (2024). Epigenome editors can further be composed of CHARM as described in Neumann et al “Brainwide silencing of prion protein by AAV-mediated deliver}7of an engineered compact epigenetic editor” Science 2024 Jun 28;384(6703).

[0281] Also contemplated for use as cargo of interest is RNA interference (RNAi). RNAi is achieved by exposing the cellular messenger RNA (mRNA) molecules produced by the targetgene to double-stranded RNA (dsRNA) molecules that contain sequences complementary' to a short portion of the mRNA molecule. Inside the cell, the double-stranded RNA molecules are cleaved to produce short (21-23 nucleotides long) single and double-stranded fragments which can bind to the target mRNA molecules. Such binding leads to the cleavage or translation disruption of the target mRNA by nucleases, thus resulting in a reduction in the expression levels of the target gene. In some embodiments, the RNA interference molecule targets one or more proteins involved in the epigenetic silencing of viral DNA is a cargo as described herein.

[0282] RNA targeting systems for use as cargo in the vectors of the present disclosure include, without limitation, RNAse proficient Casl3 (e.g., Casl3d). RNAse deficient Casl3 (e.g., dCasl3d), CSM (Colognori et al., “Precise Transcript Targeting by CRISPR-Csm Complexes,” Nature Biotechnology 2023), and RADARs (Jiang et al., “Programmable Eukaryotic Protein Synthesis with RNA Sensors by Harnessing ADAR,” Nature Biotechnology 41:698-707).

[0283] CRISPR DNA binding proteins are also contemplated for use in the vectors of the present disclosure. These proteins include, without limitation, any' of the dCas enzymes (e.g., dCas9, dCasl2) or natural CRISPR DNA binding proteins without nuclease domains including without limit Casl2c. In some embodiments, the cargo comprises dCas9. CRISPR DNA binding proteins can be fusion proteins to impart additional protein function. Examples, without limitation include, fusions composed of CRISPR DNA binding proteins and recombinases, integrases, transposases, enzymes, protein scaffolds, retrotransposases, deaminases, reverse transcriptases, DNA polymerases.

[0284] Additional cargos for use in the vectors of the present disclosure include recombinases. Recombinases for use as cargo in the vectors of the present disclosure include, without limitation, large serine recombinases (Durrant, et al. Systematic discovery of recombinases for efficient integration of large DNA sequences into the human genome. Nat Biotechnol 41, 488-499 (2023) https: / / doi.org / 10.1038 / s41587-022-01494-w) and CRE.

[0285] Additional cargos for use in the vectors of the present disclosure include transcription factors. Transcription factors for use as cargo in the vectors of the present disclosure include, without limitation, reprogramming transcription factors such as Oct4, Sox2, Klf4 and c-Myc as described in Browder et al. “Zn vivo partial reprogramming alters age-associated molecular changes during physiological aging in mice” Nature Aging volume 2, pages 243-253 (2022).

[0286] Additional cargos for use in the vectors of the present disclosure include viral or bacterial proteins. Viral proteins for use as cargo in the vectors of the present disclosure include, without limitation, HSV-1 VP16, HSV-1 ICPO, HCMV IE1, EBV BNRF1, HBV HBx, HCMV pp71, KSHV RTA, EBV RTA, HIV-1 Vpr, and HIV / -2 / SIV Vpx.

[0287] Additional cargos for use in the vectors of the present disclosure include transposons, transposases, and retrotransposons. Retrotransposons for use as cargo in the vectors of the present disclosure include, without limitation, human LI retrotransposon as described in Wang et al. '‘CRTSPR-Enabled Autonomous Transposable Element (CREATE) for RNA-based gene editing and delivery” doi: https: / / doi.org / 10.1101 / 2024.01.29.577809.

[0288] Programmable integrases for use as cargo in the vectors of the present disclosure include, without limitation, Programmable Addition via Site-specific Targeting Element (PASTE) (Yamall et al., Drag-and-drop genome insertion of large sequences without doublestrand DNA cleavage using CRISPR-directed integrases,” Nature Biotechnology 41(4): SOO- 512 (2023).

[0289] Exemplary CRISPR nickases include, without limitation, CRISPR nickases are derived from CRISPR nucleases by inactivation of one of the nuclease domains. In specific embodiments, the CRISPR nickase can be derived from a type II CRISPR nuclease. For example, the type II CRISPR nuclease can be a Cas9 protein. Suitable Cas9 nucleases include Streptococcus pyogenes Cas9 (SpCas9), Francisella novicida Cas9 (FnCas9), Staphylococcus aureus (SaCas9). Streptococcus thermophilus Cas9 (StCas9), Streptococcus pasteurianus (SpaCas9), Campylobacter] ejum Cas9 (CjCas9), Neisseria meningitidis Cas9 (NmCas9), or Neisseria cinerea Cas9 (NcCas9). In other embodiments, the nickase can be derived from a type V CRISPR nuclease, such as a CpH nuclease. Suitable Cpfl nucleases include Francisella novicida CpH (FnCpfl), Acidaminococcus sp. Cpfl (AsCpH), or Lachnospiraceae bacterium ND2006 Cpfl (LbCpfl ). In yet another embodiment, the nickase can be derived from a type VI CRISPR nuclease, e.g., Leptotrichia wadei Casl3a (LwaCasl3a) or Leptotrichia shahii Casl3a (LshCasl3a).

[0290] The cargo included in the vectors of the present disclosure can also be nucleic acids encoding proteins designed to program cell function such as a transgene that encodes a chimeric antigen receptor (CAR) protein, a T cell receptor (TCR) protein, a synthetic Notch (SynNotch) receptor protein, a synthetic intramembrane proteolysis receptor (SNIPR) protein, or an HLA-independent T cell (HIT) receptor protein. In some embodiments, the transgene encodes the transgene encodes a chimeric antigen receptor (CAR) protein. In some embodiments, the transgene encodes a T cell receptor (TCR) protein. In some embodiments.the transgene encodes a synthetic Notch (SynNotch) receptor protein. In some embodiments, the transgene encodes a synthetic intramembrane proteolysis receptor (SNIPR) protein. In some embodiments, the transgene encodes an HLA-independent T cell (HIT) receptor protein. These include, without limitation, synnotch (Roybal et al., “Engineering T Cells with Customized Therapeutic Response Programs Using Synthetic Notch Receptors,” Cell 167(2):419-432 (2016); Morsut et al., “Engineering Customized Cell Sensing and Response Behaviors Using Synthetic Notch Receptors,” Cell 164(4):780-791 (2016)) and SNIPR CARs (Zhu et al., “Modular Design of Synthetic Receptors for Programmed Gene Regulation in Cell Therapies,” Cell 185(8): 1431-1443 (2022)). Generally, CARs, TCRs, SNIPRs, HITs, SynNotchs and their sequences are well known to one skilled in the art.

[0291] Exemplary programmable nucleases are also known in the art and include, without limitation, programmable Type V or Type VI CRISPR / Cas enzyme. In some aspects, the programmable Type V CRISPR / Cas enzyme is a programmable Casl2 nuclease. In some aspects, the programmable Casl2 nuclease is Casl2a, Casl2b, Casl2d, or Casl2e. In some aspects, the programmable Type V CRISPR / Cas enzyme is a programmable Cast 4 nuclease. In some aspects, the programmable Casl4 nuclease is Casl4a, Casl4b. Casl4c, Casl4d. Casl4e, Casl4f, Casl4g, or Casl4h. In some aspects, the programmable nuclease is a programmable Type VI CRISPR / Cas enzyme. In some aspects, the programmable Type VI CRISPR / Cas enzyme is a programmable Casl3 nuclease. In some aspects, the programmable Casl3 nuclease is Casl3a, Casl3b, Casl3c, Casl3d, or Casl3e.

[0292] Programmable proteases for use as cargo in the vectors of the present disclosure include, without limitation, CRASPASE (Hu et al., “Craspase is a CRISPR RNA-guided, RNA-activated Protease,” Science 377(6612): 1278-1285 (2022) and protease regulated CAR- T cell receptors (Labanieh et al, Enhanced Safety and Efficacy of Protease-regulated CAR-T Cell Receptors,” Cell 185: 1745-1763 (2022).

[0293] The cargo included in the vectors of the present disclosure can also be nucleic acids encoding proteases such as Caspases or viral proteases. Viral proteases are well known in the art and include, without limitation, HIV protease. Caspases are well know n in the art and include, without limitation, Caspase 1. 2, 3, 4. 5, 6, 7. 8, 9, 10, 11, 12, 13, and 14.

[0294] Examples of reporter genes which may be employed to identify transfected or transduced cell lines include alkaline phosphatase (AP), beta galactosidase (LacZ), beta glucoronidase (GUS), chloramphenicol acety ltransferase (CAT), green fluorescent protein (GFP), horseradish peroxidase (HRP), and luciferase (Luc). Possible antibiotic selectable markers include those that confer resistance to ampicillin, blasticidin, bleomycin.chloramphenicol, geneticin (G418), gentamycin, hygromycin, kanamycin, lincomycin, methotrexate, phosphinothricin, puromycin, and tetracyclin.

[0295] In some embodiments, the cargo includes a KRAB domain. A KRAB domain is a domain that is usually found in the N-terminal of several zinc finger protein based transcription factors. The KRAB domain can consist of 75-120 amino acids which repression may be accomplished by a module of about 45 amino acids. Hence, some embodiments of the invention may use KRAB domains or fragments thereof as repressor domains.

[0296] In some embodiments, the KRAB domain comprises the sequence of: RTLVTFKDVFVDFTREEWKLLDTAQQIVYRNVMLENYKNLVSLGYQLTKPDVILRL EKGEEP (SEQ ID NO: 796)

[0297] In some embodiments, the nucleic acid constructs of the present disclosure can include more than one cargo of interest. In some embodiments, the nucleic acid constructs disclosed herein can include at least one, at least two, at least three, at least four, at least five, or at least six cargos of interest. For example, in one embodiment, a first cargo can comprise an endonuclease deficient Cas effector (e.g.. a CRISPRi) and a second cargo comprises a guide RNA. In another embodiment, a first cargo can compnse Cas9 and a second cargo comprises a guide RNA. In some embodiments, the guide RNA is an sgRNA. In some embodiments, a third cargo comprises an shRNA or an siRNA. As described above, each cargo of interest can have its own RNA polymerase II or RNA polymerase III promoter. For example, the construct can include a Cas9 cargo expressed by an RNA polymerase II promoter, an sgRNA expressed by an RNA polymerase III promoter, and a TASOR shRNA expressed by another RNA polymerase III promoter.

[0298] In some embodiments, the cargo of interest is located immediately dow nstream of the 5' LTR.

[0299] In some embodiments, a polymerase II promoter is located in between the 5’ LTR and the cargo.

[0300] In some embodiments, the cargo of interest is located such that there is a PRE 3’ of the cargo.

[0301] In some embodiments, a polymerase II promoter is located in between the 5’ LTR and the cargo, and there is a PRE 3’ of the cargo.

[0302] In some embodiments, when the cargo comprises an sgRNA, shRNA, and / or an siRNA, one or more polymerase III promoters are located upstream of each cargo.

[0303] In some embodiments, each of the cargos of interest comprises a PRE sequence located 3’ to the cargo. In some embodiments, the sequence encoding the cargos of interestare operably linked to one another within a single open reading frame (e.g., in a polycistronic ORF).

[0304] In some embodiments, the sequence encoding the cargos of interest can be linked to one another directly or indirectly (e.g., via one or more connector sequences). For example, in some embodiments, the sequence encoding the cargos of interest can be directly linked to one another, e.g.. adjacently to one another. In some embodiments, at least two (e.g., 2, 3, 4, or 5) of the sequence encoding the cargos of interest are operably linked to one another by one or more connector sequences. In some embodiments, the length and amino acid composition of the connector sequences can be optimized to vary7the orientation, flexibility, and / or proximity of the polypeptides relative to one another to achieve a desired activity or property of the encoded protein. In some embodiments, a connector sequence of the plurality of connector sequences includes one or more coding sequences for autoproteolytic peptide sequences. Generally, any proteolytic cleavage site known in the art can be incorporated into the nucleic acid molecules of the disclosure and can be, for example, proteolytic cleavage sequences that are cleaved post-production by a protease. Further suitable proteolytic cleavage sites also include proteolytic cleavage sequences that can be cleaved following addition of an external protease. As used herein the term ‘'autoproteolytic peptide” refers to a “self-cleaving” peptide that possesses autoproteolytic activity7and is capable of cleaving itself from a larger polypeptide moiety7. First identified in the foot-and-mouth disease virus (FMDV), a member of the picomavirus group, several autoproteolytic peptides have been subsequently identified such as, for example, “2A like” peptides from equine rhinitis A virus (E2A), porcine teschovirus-1 (P2A) and Thosea asigna virus (T2A), and their activities in proteolytic cleavage have been shown in various ex vitro, in vitro, ex vivo, and in vivo eukaryotic systems. As such, the concept of autoproteolytic peptides is available to one of skill in the art with many naturally-occurring autoprotease systems have been identified. Well studied autoprotease systems are e.g., viral proteases, developmental proteins (e g., HetR, Hedgehog proteins), RumA autoprotease domain, UmuD, etc.). Non-limiting examples of autoproteolytic peptides suitable for the compositions and methods of the present disclosure include one or more autoproteolytic cleavage sequences from a calcium-dependent serine endoprotease (furin), a porcine teschovirus-1 2 A (P2A), a foot-and-mouth disease virus (FMDV) 2A (F2A), an Equine Rhinitis A Virus (ERAV) 2A (E2A), a Thosea asigna virus 2A (T2A), a cytoplasmic polyhedrosis virus 2A (BmCPV2A), a Flacherie Virus 2A (BmIFV2A), or a combination thereof.

[0305] In some embodiments, the sequence encoding the cargos of interest are operably linked to one another by a coding sequence for one or more internal ribosomal entry sites (IRES). An IRES or "internal ribosome entry site’’ is a sequence located between polycistronic genes that permits the production of the expression product originating from the second gene by internal initiation of the translation of the dicistronic mRNA. It promotes direct internal ribosome entry to the initiation codon, such as ATG, of a cistron (a protein encoding region), thereby leading to the cap-independent translation of the gene. See. e.g., Jackson et al., 1990. Trends Biochem Sci 15(12):477-83) and Jackson and Kaminski. 1995. RNA l(10):985-1000. In some embodiments, the IRES can be a viral IRES, a cellular IRES, or an artificial IRES. Examples of IRES generally employed by those of skill in the art include those described in U.S. Pat. No. 6.692,736. In some embodiments, the IRES is selected from a Kaposi’s sarcoma-associated herpesvirus (KSHV) IRES, a hepatitis virus IRES, a Pestivirus IRES, a Cripavirus IRES, a Rhopalosiphum padi virus IRES, a fibroblast growth factor IRES, a platelet-derived growth factor IRES, a vascular endothelial growth factor IRES, an insulin-like growth factor IRES, a picomavirus IRES, an encephalomyocarditis virus (EMCV) IRES. a Pim-1 IRES, a p53 IRES, an Apaf-1 IRES, a TDP2 IRES, an L-myc IRES, and a c-myc IRES., In some embodiments, the IRES is obtainable from EMCV.

[0306] One of skill in the art will appreciate that different configurations of sequence encoding the cargos of interest, the sequence encoding the autoproteolytic peptide, or an IRES can be employed as long as expression of sequence encoding the cargos of interest is adequately maintained. These sequences will typically be configured so that the polypeptide encoded by the gene of interest can be released from the protease and other sequences after cleavage by the autoprotease.

[0307] The term "operably linked”, as used herein, denotes a functional linkage betw een two or more sequences. For example, an operable linkage between a polynucleotide of interest and a regulatory sequence (for example, a promoter) is a functional link that allows for expression of the polynucleotide of interest, i.e., cargo of interest. In this sense, the term "operably linked” refers to the positioning of a regulatory region and a coding sequence to be transcribed so that the regulatory region is effective for regulating transcription or translation of the coding sequence of interest. In some embodiments disclosed herein, the term “operably linked” denotes a configuration in which a regulatory sequence is placed at an appropriate position relative to a sequence that encodes a polypeptide or functional RNA such that the control sequence directs or regulates the expression or cellular localization of the mRNAencoding the polypeptide, the polypeptide, and / or the functional RNA. Thus, a promoter is in operable linkage with a nucleic acid sequence if it can mediate transcription of the nucleic acid sequence. Operably linked elements may be contiguous or non-contiguous.

[0308] The basic techniques for operably linking two or more sequences of DNA together are familiar to one of ordinary7skill in the art, and such methods have been described in many books for standard molecular biological manipulation (see, for example, Maniatis et al., “Molecular Cloning: A Laboratory Manual” 2nd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.; and Gibson et al.. Nature Methods 6:343-45, 2009).Additional Components

[0309] The integration deficient retroviral vector described herein may also include transcriptional regulatory7control sequences operably linked to the retroviral genome to direct transcription of the genome in a production cell / packaging cell. Some vector genomes require additional sequences for efficient virus production. For example, particularly in the case of HIV, RRE sequences may be included. However, the requirement for RRE (and dependence on rev which is provided in trans) may be reduced or eliminated by codon optimization. Further details of this strategy can be found in WO 2001 / 79518.

[0310] Alternative sequences which perform the same function as the rev / RRE system are also known. For example, a functional analogue of the rev / RRE system is found in the Mason Pfizer monkey virus. This is known as the constitutive transport element (CTE) and comprises an RRE-type sequence in the genome which is believed to interact with a factor in the infected cell. The cellular factor can be thought of as a rev analogue. Thus, CTE may be used as an alternative to the rev / RRE system. Any other functional equivalents of the Rev protein which are known or become available may be relevant to the invention. For example, it is also known that the Rex protein of HTLV-I can functionally replace the Rev protein of HIV-1. Rev and RRE may7be absent or non-functional in the vector for use in the methods of the present disclosure; in the alternative rev and RRE, or functionally equivalent sy stem, may be present.

[0311] The retroviral vector disclosed herein can also include additional components such as, without limitation, splice donor sites (SD), splice acceptor sites (SA), and / or Rev responsive elements (RRE), Tat, and / or Rev.

[0312] A vector can also include other components as are known in the art, e.g. , any of various selection markers and / or reporter genes. Examples of reporter genes which may be employed to identify transfected or transduced cell lines include alkaline phosphatase (AP),beta galactosidase (LacZ), beta glucoronidase (GUS), chloramphenicol acetyltransferase (CAT), green fluorescent protein (GFP). horseradish peroxidase (HRP), and luciferase (Luc). Possible antibiotic selectable markers include those that confer resistance to ampicillin, blasticidin, bleomycin, chloramphenicol, geneticin (G418), gentamycin, hygromycin, kanamycin, lincomycin, methotrexate, phosphinothricin, puromycin, and tetracyclin.Exemplary IDLV Constructs

[0313] Exemplary constructs of the recombinant integration deficient retroviral vectors carrying multiple cargos of interest are provided in the Examples described herein. In some embodiments, the recombinant integration deficient retroviral vector includes, from N- terminus to C-terminus, a 5’ SIN LTR derived from HIV-1, a viral packaging signal, an SFFV promoter, a CRISPRi cargo, a WPRE sequence, and a 3’ SIN LTR derived from HIV-1 that includes a U6 promoter and an NP220 shRNA. Inclusion of an sgRNA cassette is also contemplated in this exemplary construct.

[0314] In some embodiments, the recombinant integration deficient retroviral vector comprises the nucleic acid sequence of SEQ ID NO: 797 as shown below.Bold text - HIV- 1 LTRItalic text - HIV-1 Psi (packaging signal)Capital text - HIV- 1 Rev responsive elementBold italic text - HIV-1 polypurine tract (PPT)Bold underline text - Spleen Focus Forming Virus (SFFV promoter) Underlined text - dCas9-KRAB-P2A-GFP (CRISPRi) Italic underlined text - WPREBold, italic, underlined text - human U6 promoter NP220 shRNA expression cassetteTggaagggctaattcactcccaaagaagacaagatatcctgatctgtggatctaccacacacaaggctacttccctgattag cagaactacacaccagggccaggggtcagatatccactgacctttggatggtgctacaagctagtaccagttgagccagata aggtagaagaggccaataaaggagagaacaccagcttgttacaccctgtgagcctgcatgggatggatgacccggagagag aagtgttagagtggaggttgacagccgcctagcattcatcacgtggcccgagagctgcatccggagtacttcaagaactgct gatatcgagctgctacaagggacttccgctggggacttccagggaggcgtggcctgggcgggactggggagtggcgagc cctcagatcctgcatataagcagctgcttttgcctgtactgggtctctctggtagaccagatctgagcctgggagctctctggc taactagggaacccactgcttaagcctcaataaagctgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgactctgg taactagagatccctcagacccttttagtcagtgtggaaaatctctagcagtggcgcccgaacagggacttgaaagcgaaaggg aaaccagaggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaagaggcgaggggcggcgactggtgagta cgccaaaaattttgactagcggaggctagaaggagagagatgggtgcgagagcgtcagtataagcgggggagaatagatcgc gatgggaaaaaattcggttaaggccagggggaaagaaaaaatataaattaaaacatatagtatgggcaagcagggagctagaacgat tcgcagttaatcctggcctgttagaaacatcagaaggctgtagacaaatactgggacagctacaaccatcccttcagacaggatcagaagaacttagalcattatataatacagtagcaaccclctattgtgtgcatcaaaggatagagataaaagacaccaaggaagctttagacaag atagaggaagagcaaaacaaaagtaagaccaccgcacagcaagcggccggccgctgatcttcagacctggaggaggagatatga gggacaattggagaagtgaatatataaatataaagtagtaaaaatgaaccataggagtagcaccCACCAAGGCAAAG AGAAGAGTGGTGCAGAGAGAAAAAAGAGCAGTGGGAATAGGAGCTTTGTTCCTT GGGTTCTTGGGAGCAGCAGGAAGCACTATGGGCGCAGCGTCAATGACGCTGACG GTACAGGCCAGACAATTATTGTCTGGTATAGTGCAGCAGCAGAACAATTTGCTG AGGGCTATTGAGGCGCAACAGCATCTGTTGCAACTCACAGTCTGGGGCATCAAG CAGCTCCAGGCAAGAATCCTGGCTGTGGAAAGATACCTAAAGGATCAACAGCTC CTGGGGATTTGGGGTTGCTCTGGAAAACTCATTTGCACCACTGCTGTGCCTTGgaat gctagttggagtaataaatctctggaacagatttggaatcacacgacctggatggagtgggacagagaaattaacaattacacaagctt aatacactccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaattatggaattagataaatgggcaagttgtgga attggtttaacataacaaattggctgtggtatataaaattattcataatgatagtaggaggctggtaggttaagaatagtttttgctgtactt ctatagtgaatagagttaggcagggatattcaccattatcgtttcagacccacctcccaaccccgaggggacccgacaggcccgaag gaatagaagaagaaggtggagagagagacagagacagatccatcgattagtgaacggatctcgacggtatcgccaaatggcagta XXcaXccacaatttaaaagaaaaggggggattggggggtacagtgcaggggaaagaatagtagacataatagcaacagacat acaaactaaagaatacaaaaacaaatacaaaaatcaaaatttc§,g^xxa!i\aca§,gg,aca§,cag,a§,a\cca^x^a!ic^a taagctgatatcgaattcgggaggtggtccctgcagtacgccaatgataacccccgccagaaaaatctagtagccttcccttttgtttt ccgtgccccaactcggcggatgactcggcccctccggaaacacccgaatcaacttctagtcaaatatgtcacgccgcaatgacc cacccctggcccgcgtctgtggaactgacccctggtgtacaggagagttcgctgctgaaagtggtcccaaaggggtactagttttaag ctcccaactccccctcccccagcgtctggaggattccacaccctcgcaccgcaggggcgaggaagtgggcggagtccggtttggc gccagccgctgaggctgccaagcagaaaagccaccgctgaggagactccggtcactgtcctcgccccgcctcccccttccctcccct tggggaccaccgggcgccacgccgcgaacggtaagtgccgcggtcgtcggcgcctccgccctccccctagggccccaattcccag cgggcgcggcgcgcggccccgcgcgcagctcccggctccctcccccttcggatgtggctgagctgtaggcgcggagtcctgcag ccccgataaaataaaagattttatttagtctccagaaaaaggggggaatgaaagaccccacctgtaggtttggcaagctagctgcagta acgccattttgcaaggcatggaaaaataccaaaccaagaatagagaagttcagatcaagggcgggtacatgaaaatagcta acgttgggccaaacaggatatctgcggtgagcagttcggccccggcccggggccaagaacagatggtcaccgcagttcgg ccccggcccgaggccaagaacagatggtccccagatatggcccaaccctcagcagtttctaagacccatcagatgttccag gctcccccaaggacctgaaatgaccctgcgccttatttgaattaaccaatcagcctgcttctcgcttctgtcgcgcgcttctgct cccgagctctataaaagagctcacaacccctcactcggcgcgccagtcctccgacagactgagtcgcccgggggggatctgga gctctcgagaattctcacgcgtctgcaggatatcaagctgcggtaccgcgggcccggccaccatggacaagaagtacagcateggc ctggccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaaggtgcccagcaagaaatcaaggtectgggcaa caccgaccggcacagcatcaagaagaacctgatcggcgccctgctgtcgacagcggagaaacagccgaggccacccggctgaa gagaaccgccagaagaagatacaccagacggaagaaccggatctgctatctgcaagagatctcagcaacgagatggccaaggtg gacgacagctctccacagactggaagagtcctcctggtggaagaggataagaagcacgagcggcaccccatctcggcaacatc gtggacgaggtggcctaccacgagaagtaccccaccatctaccacctgagaaagaaactggtggacagcaccgacaaggccgacc tgcggctgatctatctggccctggcccacatgatcaagtccggggccactcctgatcgagggcgacctgaaccccgacaacagcg acgtggacaagctgttcatccagclggtgcagacctacaaccagctgtlcgaggaaaaccccatcaacgccagcggcgtggacgcc aaggccatcctgtctgccagactgagcaagagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaagaatggcctgttcggcaacctgatgccctgagcctgggcctgacccccaactcaagagcaactcgacctggccgaggatgccaaactgcagct gagcaaggacacctacgacgacgacctggacaacctgctggcccagatcggcgaccagtacgccgacctgttctggccgccaag aacctgtccgacgccatcctgctgagcgacatcctgagagtgaacaccgagatcaccaaggcccccctgagcgcctctatgatcaag agatacgacgagcaccaccaggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgagaagtacaaagagattctc gaccagagcaagaacggctacgccggctacatcgatggcggagccagccaggaagagtctacaagtcatcaagcccatcctgga aaagatggacggcaccgaggaactgctcgtgaagctgaacagagaggacctgctgcggaagcagcggacctcgacaacggcag catcccccaccagatccacctgggagagctgcacgccatctgcggcggcaggaagatttacccatcctgaaggacaaccggga aaagatcgagaagatcctgacctccgcatcccctactacgtgggccctctggccaggggaaacagcagattcgcctggatgaccag aaagagcgaggaaaccatcaccccctggaactcgaggaagtggtggacaagggcgccagcgcccagagctcatcgagcggat gaccaacttcgataagaacctgcccaacgagaaggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtacaacgagct gaccaaagtgaaatacgtgaccgagggaatgagaaagcccgccttcctgagcggcgagcagaaaaaagccatcgtggacctgctg ttcaagaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaagaaaatcgagtgcttcgactccgtggaaatctcc ggcgtggaagatcggttcaacgcctccctgggcacataccacgatctgctgaaaattatcaaggacaaggacttcctggacaatgagg aaaacgaggacattctggaagatatcgtgctgaccctgacactgttgaggacagagagatgatcgaggaacggctgaaaacctatg cccacctgtcgacgacaaagtgatgaagcagctgaagcggcggagatacaccggctggggcaggctgagccggaagctgatcaa cggcatccgggacaagcagtccggcaagacaatcctggatttcctgaagtccgacggctcgccaacagaaactcatgcagctgat ccacgacgacagcctgaccttaaagaggacatccagaaagcccaggtgtccggccagggcgatagcctgcacgagcacatgcc aatctggccggcagccccgccataagaagggcatcctgcagacagtgaaggtggtggacgagctcgtgaaagtgatgggccggc acaagcccgagaacatcgtgatcgaaatggccagagagaaccagaccacccagaagggacagaagaacagccgcgagagaatg aagcggatcgaagagggcatcaaagagctgggcagccagatcctgaaagaacaccccgtggaaaacacccagctgcagaacgag aagctgtacctgtactacctgcagaatgggcgggatatgtacgtggaccaggaactggacatcaaccggctgtccgactacgatgtgg acgctatcgtgcctcagagctttctgaaggacgactccatcgataacaaagtgctgactcggagcgacaagaaccggggcaagagc gacaacgtgccctccgaagaggtcgtgaagaagatgaagaactactggcgccagctgctgaatgccaagctgattacccagaggaa gtcgacaatctgaccaaggccgagagaggcggcctgagcgaactggataaggccggctcatcaagagacagctggtggaaacc cggcagatcacaaagcacgtggcacagatcctggactcccggatgaacactaagtacgacgagaacgacaaactgatccgggaag tgaaagtgatcaccctgaagtccaagctggtgtccgatttccggaaggatttccagtttacaaagtgcgcgagatcaacaactaccacc acgcccacgacgcctacctgaacgccgtcgtgggaaccgccctgatcaaaaagtaccctaagctggaaagcgagttcgtgtacggc gactacaaggtgtacgacgtgcggaagatgatcgccaagagcgagcaggaaatcggcaaggctaccgccaagtacttcttctacag caacatcatgaacttttcaagaccgagattaccctggccaacggcgagatccggaagcggcctctgatcgagacaaacggcgaaac aggcgagatcgtgtgggataagggccgggacttgccaccgtgcggaaagtgctgtctatgccccaagtgaatatcgtgaaaaagac cgaggtgcagacaggcggcttcagcaaagagtctatcctgcccaagaggaacagcgacaagctgatcgccagaaagaaggactgg gaccctaagaagtacggcggctcgacagccccaccgtggcctatctgtgctggtggtggccaaagtggaaaagggcaagtccaa gaaactgaagagtgtgaaagagctgctggggatcaccatcatggaaagaagcagctcgagaagaatcccatcgacttctggaagc caagggctacaaagaagtgaaaaaggacctgatcatcaagctgcctaagtactccctgtcgagctggaaaacggccggaagagaat gctggcctctgccggcgaactgcagaagggaaacgaactggccctgccctccaaatatgtgaactcctgtacctggccagccactat gagaagctgaagggctcccccgaggataatgagcagaaacagctgttgtggaacagcacaaacactacctggacgagatcatcga gcagatcagcgagttctccaagagagtgalcctggccgacgctaatctggacaagglgctgagcgcctacaacaagcacagagaca agcctatcagagagcaggccgagaatatcatccacctgttaccctgaccaatctgggagcccctgccgcctcaagtactttgacaccaccatcgaccggaagaggtacaccagcaccaaagaggtgctggaceccaccctgatccaccagagcatcaccggcctgtaceaga cacggatcgacctgtctcagctgggaggcgacgcctatccctatgacgtgcccgattatgccagcctgggcagcggctcccccaaga aaaaacgcaaggtggaagatcctaagaaaaagcggaaagtggacggcattggtagtgggagcaacggcagcagcggatccggag ggccgagctctggcgcacccccaccaagtggagggtctcctgccgggtccccaacatctactgaagaaggcaccagcgaatccgc aacgcccgagtcaggccctggtacctccacagaaccatctgaaggtagtgcgcctggttccccagctggaagccctactccaccga agaaggcacgtcaaccgaaccaagtgaaggatctgcccctgggaccagcactgaaccatctgagatggatgctaagtcactaactgc ctggtcccggacactggtgaccttcaaggatgtatttgtggacttcaccagggaggagtggaagctgctggacactgctcagcagatc gtgtacagaaatgtgatgctggagaactataagaacctggtttccttgggttatcagcttactaagccagatgtgatcctccggttggaga agggagaagagcccagcggccgcgctactaacttcagcctgctgaagcaggctggagacgtggaggagaaccctggacctgtga gcaagggcgaggagctgttcaccggggtggtgcccatcctggtcgagctggacggcgacgtaaacggccacaagttcagcgtgtct ggcgagggcgagggcgatgccacctacggcaagctgaccctgaagttcatctgcaccaccggcaagctgcccgtgccctggccca ccctcgtgaccaccctgacctacggcgtgcagtgcttcagccgctaccccgaccacatgaagcagcacgacttcttcaagtccgccat gcccgaaggctacgtccaggagcgcaccatcttcttcaaggacgacggcaactacaagacccgcgccgaggtgaagttcgagggc gacaccctggtgaaccgcatcgagctgaagggcatcgacttcaaggaggacggcaacatcctggggcacaagctggagtacaact acaacagccacaacgtctatatcatggccgacaagcagaagaacggcatcaaggcgaacttcaagatccgccacaacatcgaggac ggcagcgtgcagctcgccgaccactaccagcagaacacccccatcggcgacggccccgtgctgctgcccgacaaccactacctga gcacccagtccgccctgagcaaagaccccaacgagaagcgcgatcacatggtcctgctggagttcgtgaccgccgccgggatcact ctcggcatggacgagctgtacaagtaataagcgatcgccctgcaggcatgcaagctgatatcaagcttatcgatoatcaaccZcfgg attacaaaatttgtgaaagattgactggtattcttaactatgttgctccttttacgctatgtggatacgctgctttaatgcctttgtatcatg ctattgcttcccgtatggctttcattttctcctccttgtataaatcctggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacg tggcgtggtgtgcactgtgtttgctgacgcaacccccactggttggggcattgccaccacctgtcagctcctttccgggactttcgcttt ccccctccctattgccacggcggaactcatcgccgcctgccttgcccgctgctggacaggggctcggctgttgggcactgacaattc cgtggtgttgtcggggaaatcatcgtcctttccttggctgctcgcctgtgttgccacctggattctgcgcgggacgtccttctgctacgtc ccttcggccctcaatccagcggaccttccttcccgcggcctgctgccggctctgcggcctcttccgcgtcttcgccttcgccctcagac gagfcggateteccfflgggccgcctecccgcatcgataccgtcgacctcgagggaattaattcgagctcggtacctttaagaccaatg acttacaaggcagctgtagatcttagccactttttaaaagaaaaggggggactggaagggctaatcactcccaacgaagtfaatfa agagggcctatttcccatgattccttcatatttgcatatacgatacaaggctgtagagagataatggaattaatttgactgtaaaca caaagatattagtacaaaatacgtgacgtagaaagtaataatttctgggtagtttgcagttttaaaattatgttttaaaatggactatc atatgcttaccgtaactgaaagtatttcgatttcttggctttatataicttgtggaaaggacgaaacaccggcgctacaaagagtgat tcaaactcgagttgaatcactcttgtagcgttttttgaatctcgacctcgagacaaatggcagtattctaat^acaagatetgct tttgcttgtactgggtctctctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgcttaagcctc aataaagcttgcctgagtgcttcaagtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagaccctttagt cagtgtggaaaatctctagca

[0315] In some embodiments, the recombinant integration deficient retroviral vector comprises the nucleic acid sequence of SEQ ID NO: 798 as shown below. Inclusion of an sgRNA cassette is also contemplated in this exemplary construct.Bold text - HIV-1 LTRItalic text - HIV-1 Psi (packaging signal)Capital text - HIV-1 Rev responsive elementBold italic text - HIV-1 polypurine tract (PPT)Bold underline text - Spleen Focus Forming Virus (SFFV promoter)Underlined text - dCas9-KRAB-P2A-GFP (CRISPRi)Italic underlined text - WPREBold, italic, underlined text- human U6 promoter NP220 shRNA expression cassetteTggaagggctaattcactcccaaagaagacaagatatcctgatctgtggatctaccacacacaaggctacttccctgattag cagaactacacaccagggccaggggtcagatatccactgacctttggatggtgctacaagctagtaccagttgagccagata aggtagaagaggccaataaaggagagaacaccagcttgttacaccctgtgagcctgcatgggatggatgacccggagagag aagtgtagagtggaggttgacagccgcctagcattcatcacgtggcccgagagctgcatccggagtacttcaagaactgct gatatcgagctgctacaagggacttccgctggggacttccagggaggcgtggcctgggcgggactggggagtggcgagc cctcagatcctgcatataagcagctgcttttgcctgtactgggtctctctggttagaccagatctgagcctgggagctctctggc taactagggaacccactgcttaagcctcaataaagctgcctgagtgctcaagtagtgtgtgcccgtctgttgtgtgactctgg taactagagatccctcagacccttttagtcagtgtggaaaatctctagcagtggcgcccgaacagggacttgaaagcgaaaggg aaaccagaggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaagaggcgaggggcggcgactggtgagta cgccaaaaattttgactagcggaggctagaaggagagagatgggtgcgagagcgtcagtataagcgggggagaattagatcgc gatgggaaaaaattcggttaaggccagggggaaagaaaaaatataaattaaaacatatagtatgggcaagcagggagctagaacgat tcgcagttaatcctggcctgttagaaacatcagaaggctgtagacaaatactgggacagctacaaccatcccttcagacaggatcagaa gaacttagatcattatataatacagtagcaaccctctattgtgtgcatcaaaggatagagataaaagacaccaaggaagctttagacaag atagaggaagagcaaaacaaaagtaagaccaccgcacagcaagcggccggccgctgatcttcagacctggaggaggagatatga gggacaattggagaagtgaattatataaatataaagtagtaaaaattgaaccattaggagtagcaccCACCAAGGCAAAG AGAAGAGTGGTGCAGAGAGAAAAAAGAGCAGTGGGAATAGGAGCTTTGTTCCTT GGGTTCTTGGGAGCAGCAGGAAGCACTATGGGCGCAGCGTCAATGACGCTGACG GTACAGGCCAGACAATTATTGTCTGGTATAGTGCAGCAGCAGAACAATTTGCTG AGGGCTATTGAGGCGCAACAGCATCTGTTGCAACTCACAGTCTGGGGCATCAAG CAGCTCCAGGCAAGAATCCTGGCTGTGGAAAGATACCTAAAGGATCAACAGCTC CTGGGGATTTGGGGTTGCTCTGGAAAACTCATTTGCACCACTGCTGTGCCTTGgaat gctagttggagtaataaatctctggaacagatttggaatcacacgacctggatggagtgggacagagaaattaacaattacacaagctt aatacactccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaattattggaattagataaatgggcaagtttgtgga attggtttaacataacaaattggctgtggtatataaaattattcataatgatagtaggaggcttggtaggtttaagaatagtttttgctgtacttt ctatagtgaatagagttaggcagggatattcaccattatcgtttcagacccacctcccaaccccgaggggacccgacaggcccgaag gaatagaagaagaaggtggagagagagacagagacagatccattcgattagtgaacggatctcgacggtatcgccaaatggcagta [{catccacaaMttaaaagaaaaggggggattggggggtacagtgcaggggaaagaatagtagacataatagcaacagacat acaaactaaagaatacaaaaacaaatacaaaaattcaaaattcgggtXattacagggacagcagagatccagttggatcga taagcttgatatcgaattcgggaggtggtccctgcagttacgccaatgataacccccgccagaaaaatcttagtagccttccctttttgtttt ccgtgccccaactcggcggattgactcggccccttccggaaacacccgaatcaacttctagtcaaattattgttcacgccgcaatgacccacccctggcccgcgtctgtggaactgacccctggtgtacaggagagtcgctgctgaaagtggtcccaaaggggtactagttttaag ctcccaactccccctcccccagcgtctggaggattccacaccctcgcaccgcaggggcgaggaagtgggcggagtccggtttggc gccagccgctgaggctgccaagcagaaaagccaccgctgaggagactccggtcactgtcctcgccccgcctcccccttccctcccct tggggaccaccgggcgccacgccgcgaacggtaagtgccgcggtcgtcggcgcctccgccctccccctagggccccaatcccag cgggcgcggcgcgcggccccgcgcgcagctcccggctccctccccctcggatgtggctgagctgtaggcgcggagtcctgcag ccccgataaaataaaagattttatttagtctccagaaaaaggggggaatgaaagaccccacctgtaggttggcaagctagctgcagta acgccattttgcaaggcatggaaaaataccaaaccaagaatagagaagttcagatcaagggcgggtacatgaaaatagcta acgttgggccaaacaggatatctgcggtgagcagtttcggccccggcccggggccaagaacagatggtcaccgcagttcgg ccccggcccgaggccaagaacagatggtccccagatatggcccaaccctcagcagtttcttaagacccatcagatgtttccag gctcccccaaggacctgaaatgaccctgcgccttatttgaattaaccaatcagcctgcttctcgcttctgtcgcgcgcttctgct cccgagctctataaaagagctcacaacccctcactcggcgcgccagtcctccgacagactgagtcgcccgggggggatctgga gctctcgagaattctcacgcgtctgcaggatatcaagctgcggtaccgcgggcccggccaccatggacaagaagtacagcatcggc ctggccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaaggtgcccagcaagaaattcaaggtgctgggcaa caccgaccggcacagcatcaagaagaacctgatcggcgccctgctgtcgacagcggagaaacagccgaggccacccggctgaa gagaaccgccagaagaagatacaccagacggaagaaccggatctgctatctgcaagagatctcagcaacgagatggccaaggtg gacgacagctctccacagactggaagagtcctcctggtggaagaggataagaagcacgagcggcaccccatctcggcaacatc gtggacgaggtggcctaccacgagaagtaccccaccatctaccacctgagaaagaaactggtggacagcaccgacaaggccgacc tgcggctgatctatctggccctggcccacatgatcaagtccggggccactcctgatcgagggcgacctgaaccccgacaacagcg acgtggacaagctgtcatccagctggtgcagacctacaaccagctgtcgaggaaaaccccatcaacgccagcggcgtggacgcc aaggccatcctgtctgccagactgagcaagagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaagaatggcc tgtcggcaacctgatgccctgagcctgggcctgacccccaactcaagagcaacttcgacctggccgaggatgccaaactgcagct gagcaaggacacctacgacgacgacctggacaacctgctggcccagatcggcgaccagtacgccgacctgttctggccgccaag aacctgtccgacgccatcctgctgagcgacatcctgagagtgaacaccgagatcaccaaggcccccctgagcgcctctatgatcaag agatacgacgagcaccaccaggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgagaagtacaaagagatttctc gaccagagcaagaacggctacgccggctacatcgatggcggagccagccaggaagagttctacaagtcatcaagcccatcctgga aaagatggacggcaccgaggaactgctcgtgaagctgaacagagaggacctgctgcggaagcagcggacctcgacaacggcag catcccccaccagatccacctgggagagctgcacgccatctgcggcggcaggaagatttttacccattcctgaaggacaaccggga aaagatcgagaagatcctgaccttccgcatcccctactacgtgggccctctggccaggggaaacagcagattcgcctggatgaccag aaagagcgaggaaaccatcaccccctggaacttcgaggaagtggtggacaagggcgccagcgcccagagctcatcgagcggat gaccaacttcgataagaacctgcccaacgagaaggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtacaacgagct gaccaaagtgaaatacgtgaccgagggaatgagaaagcccgccttcctgagcggcgagcagaaaaaagccatcgtggacctgctg tcaagaccaaccggaaagtgaccgtgaagcagctgaaagaggactactcaagaaaatcgagtgctcgactccgtggaaatctcc ggcgtggaagatcggtcaacgcctccctgggcacataccacgatctgctgaaaatatcaaggacaaggactcctggacaatgagg aaaacgaggacatctggaagatatcgtgctgaccctgacactgttgaggacagagagatgatcgaggaacggctgaaaacctatg cccacctgtcgacgacaaagtgatgaagcagctgaagcggcggagatacaccggctggggcaggctgagccggaagctgatcaa cggcatccgggacaagcagtccggcaagacaatcctggattcctgaagtccgacggctcgccaacagaaactcatgcagctgat ccacgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtccggccagggcgatagcctgcacgagcacattgcc aatctggccggcagccccgccattaagaagggcatcctgcagacagtgaaggtggtggacgagctcgtgaaagtgatgggccggcacaagcccgagaacatcgtgatcgaaatggccagagagaaccagaccacccagaagggacagaagaacagccgcgagagaatg aagcggatcgaagagggcatcaaagagctgggcagccagatcctgaaagaacaccccgtggaaaacacccagctgcagaacgag aagctgtacctgtactacctgcagaatgggcgggatatgtacgtggaccaggaactggacatcaaccggctgtccgactacgatgtgg acgctatcgtgcctcagagcttctgaaggacgactccatcgataacaaagtgctgactcggagcgacaagaaccggggcaagagc gacaacgtgccctccgaagaggtcgtgaagaagatgaagaactactggcgccagctgctgaatgccaagctgatacccagaggaa gtcgacaatctgaccaaggccgagagaggcggcctgagcgaactggataaggccggctcatcaagagacagctggtggaaacc cggcagatcacaaagcacgtggcacagatcctggactcccggatgaacactaagtacgacgagaacgacaaactgatccgggaag tgaaagtgatcaccctgaagtccaagctggtgtccgattccggaaggatttccagtttacaaagtgcgcgagatcaacaactaccacc acgcccacgacgcctacctgaacgccgtcgtgggaaccgccctgatcaaaaagtaccctaagctggaaagcgagtcgtgtacggc gactacaaggtgtacgacgtgcggaagatgatcgccaagagcgagcaggaaatcggcaaggctaccgccaagtactcttctacag caacatcatgaactttttcaagaccgagattaccctggccaacggcgagatccggaagcggcctctgatcgagacaaacggcgaaac aggcgagatcgtgtgggataagggccgggactttgccaccgtgcggaaagtgctgtctatgccccaagtgaatatcgtgaaaaagac cgaggtgcagacaggcggcttcagcaaagagtctatcctgcccaagaggaacagcgacaagctgatcgccagaaagaaggactgg gaccctaagaagtacggcggcttcgacagccccaccgtggcctattctgtgctggtggtggccaaagtggaaaagggcaagtccaa gaaactgaagagtgtgaaagagctgctggggatcaccatcatggaaagaagcagcttcgagaagaatcccatcgactttctggaagc caagggctacaaagaagtgaaaaaggacctgatcatcaagctgcctaagtactccctgtcgagctggaaaacggccggaagagaat gctggcctctgccggcgaactgcagaagggaaacgaactggccctgccctccaaatatgtgaactcctgtacctggccagccactat gagaagctgaagggctcccccgaggataatgagcagaaacagctgttgtggaacagcacaaacactacctggacgagatcatcga gcagatcagcgagtctccaagagagtgatcctggccgacgctaatctggacaaggtgctgagcgcctacaacaagcacagagaca agcctatcagagagcaggccgagaatatcatccacctgttaccctgaccaatctgggagcccctgccgcctcaagtacttgacacc accatcgaccgga; gaggtacaccagcaccaaagaggtgctggacgccaccctgatccaccagagcatcaccggcctgtacgaga cacggatcgacctgtctcagctgggaggcgacgcctatccctatgacgtgcccgatatgccagcctgggcagcggctcccccaaga aaaaacgcaaggtggaagatcctaagaaaaagcggaaagtggacggcatggtagtgggagcaacggcagcagcggatccggag ggccgagctctggcgcacccccaccaagtggagggtctcctgccgggtccccaacatctactgaagaaggcaccagcgaatccgc aacgcccgagtcaggccctggtacctccacagaaccatctgaaggtagtgcgcctggtccccagctggaagccctacttccaccga agaaggcacgtcaaccgaaccaagtgaaggatctgcccctgggaccagcactgaaccatctgagatggatgctaagtcactaactgc ctggtcccggacactggtgaccttcaaggatgtatttgtggacttcaccagggaggagtggaagctgctggacactgctcagcagatc gtgtacagaaatgtgatgctggagaactataagaacctggtttccttgggttatcagcttactaagccagatgtgatcctccggtggaga agggagaagagcccagcggccgcgctactaacttcagcctgctgaagcaggctggagacgtggaggagaaccctggacctgtga gcaagggcgaggagctgtcaccggggtggtgcccatcctggtcgagctggacggcgacgtaaacggccacaagtcagcgtgtct ggcgagggcgagggcgatgccacctacggcaagctgaccctgaagtcatctgcaccaccggcaagctgcccgtgccctggccca ccctcgtgaccaccctgacctacggcgtgcagtgctcagccgctaccccgaccacatgaagcagcacgactctcaagtccgccat gcccgaaggctacgtccaggagcgcaccatctctcaaggacgacggcaactacaagacccgcgccgaggtgaagtcgagggc gacaccctggtgaaccgcatcgagctgaagggcatcgactcaaggaggacggcaacatcctggggcacaagctggagtacaact acaacagccacaacgtctatatcatggccgacaagcagaagaacggcatcaaggcgaactcaagatccgccacaacatcgaggac ggcagcgtgcagctcgccgaccactaccagcagaacacccccatcggcgacggccccgtgctgctgcccgacaaccactacctga gcacccagtccgccctgagcaaagaccccaacgagaagcgcgatcacatggtcctgctggagltcglgaccgccgccgggatcact ctcggcatggacgagctgtacaagtaataagcgatcgccctgcaggcatgcaagcttgatatcaagcttatcgato7 / cflacc7c7,g,gattacaaaatttgtgaaagattgactggtattcttaactatgttgctccttttacgctatgtggatacgctgctttaatgcctttgtatcatg ctattgcttcccgtatggctttcattttctcctccttgtataaatcctggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacg tggcgtggtgtgcactgtgtttgctgacgcaacccccactggttggggcattgccaccacctgtcagctcctttccgggactttcgcttt ccccctccctattgccacggcggaactcatcgccgcctgccttgcccgctgctggacaggggctcggctgttgggcactgacaattc cgtggtgttgtcggggaaatcatcgtcctttccttggctgctcgcctgtgttgccacctggattctgcgcgggacgtccttctgctacgtc ccttcggccctcaatccagcggaccttccttcccgcggcctgctgccggctctgcggcctcttccgcgtcttcgccttcgccctcagac gagteggafercccttfgggccgccfecccgcatcgataccgtcgacctcgagggaattaattcgagctcggtacctttaagaccaatg acttacaaggcagctgtagatcttagccactttttaaaagaaaaggggggactggaagggctaatcactcccaacgaagrtaaffa agagggcctatttcccatgattccttcatatttgcatatacgatacaaggctgttagagagataattggaattaatttgactgtaaaca caaagutattagtacaaaatacgtgacgtagciaagtaataatttcttgggtagtttgcagttttaaaattatgttttaaaatggactatc atatgcttaccgtaacttgaaagtatttcgatttcttggctttiitiitiitcttgtggaaaggacgaaacaccggcctcaagtatcttagca agttctcgagaacttgctaagatacttgaggttttttgaattctcgacctcgagacaaatggcagtattcttaatta&&£wigatc\g£t ttgctgtactgggtctctctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgctaagcctc aataaagctgccttgagtgcttcaagtagtgtgtgcccgtctgtgtgtgactctggtaactagagatccctcagaccctttagt cagtgtggaaaatctctagca

[0316] In some embodiments, the recombinant integration deficient retroviral vector comprises the nucleic acid sequence of SEQ ID NO: 799 as shown below. Inclusion of an sgRNA cassette is also contemplated in this exemplary construct.Bold text - HIV- 1 LTRItalic text - HIV-1 Psi (packaging signal)Capital text - HIV-1 Rev responsive elementBold italic text - HIV-1 polypurine tract (PPT)Bold underline text - Spleen Focus Forming Virus (SFFV promoter)Underlined text - dCas9-KRAB-P2A-GFP (CRISPRi)Italic underlined text - WPREBold, italic, underlined text- human U6 promoter NP220 shRNA expression cassetteTggaagggctaattcactcccaaagaagacaagatatcctgatctgtggatctaccacacacaaggctacttccctgattag cagaactacacaccagggccaggggtcagatatccactgacctttggatggtgctacaagctagtaccagttgagccagata aggtagaagaggccaataaaggagagaacaccagcttgttacaccctgtgagcctgcatgggatggatgacccggagagag aagtgtagagtggaggttgacagccgcctagcatttcatcacgtggcccgagagctgcatccggagtacttcaagaactgct gatatcgagctgctacaagggacttccgctggggacttccagggaggcgtggcctgggcgggactggggagtggcgagc cctcagatcctgcatataagcagctgcttttgcctgtactgggtctctctggtagaccagatctgagcctgggagctctctggc taactagggaacccactgcttaagcctcaataaagcttgcctgagtgcttcaagtagtgtgtgcccgtctgttgtgtgactctgg taactagagatccctcagaccctttagtcagtgtggaaaatctctagcagtggcgcccgaacagggacttgaaagcgaaaggg aaaccagaggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaagaggcgaggggcggcgactggtgagtacgccaaaaaOgactagcggaggctagaoggGgagaga^gggrgcgogagcgfcagtattaagcgggggagaatagatcgc gatgggaaaaaattcggtaaggccagggggaaagaaaaaatataaataaaacatatagtatgggcaagcagggagctagaacgat tcgcagtaatcctggcctgtagaaacatcagaaggctgtagacaaatactgggacagctacaaccatcccttcagacaggatcagaa gaactagatcatatataatacagtagcaaccctctatgtgtgcatcaaaggatagagataaaagacaccaaggaagcttagacaag atagaggaagagcaaaacaaaagtaagaccaccgcacagcaagcggccggccgctgatctcagacctggaggaggagatatga gggacaattggagaagtgaattatataaatataaagtagtaaaaattgaaccattaggagtagcaccCACCAAGGCAAAG AGAAGAGTGGTGCAGAGAGAAAAAAGAGCAGTGGGAATAGGAGCTTTGTTCCTT GGGTTCTTGGGAGCAGCAGGAAGCACTATGGGCGCAGCGTCAATGACGCTGACG GTACAGGCCAGACAATTATTGTCTGGTATAGTGCAGCAGCAGAACAATTTGCTG AGGGCTATTGAGGCGCAACAGCATCTGTTGCAACTCACAGTCTGGGGCATCAAG CAGCTCCAGGCAAGAATCCTGGCTGTGGAAAGATACCTAAAGGATCAACAGCTC CTGGGGATTTGGGGTTGCTCTGGAAAACTCATTTGCACCACTGCTGTGCCTTGgaat gctagttggagtaataaatctctggaacagatttggaatcacacgacctggatggagtgggacagagaaattaacaattacacaagctt aatacactccttaatgaagaatcgcaaaaccagcaagaaaagaatgaacaagaatatggaattagataaatgggcaagttgtgga atggtttaacataacaaattggctgtggtatataaaatatcataatgatagtaggaggcttggtaggttaagaatagtttttgctgtactt ctatagtgaatagagttaggcagggatatcaccattatcgttcagacccacctcccaaccccgaggggacccgacaggcccgaag gaatagaagaagaaggtggagagagagacagagacagatccatcgattagtgaacggatctcgacggtatcgccaaatggcagta Ucatccacaattaaaagaaaaggggggatggggggtacagtgcaggggaaagaatagtagacataatagcaacagacat acaaactaaagaattacaaaaacaaattacaaaaattcaaaattttcgggUlaUacagggacagcagagaiccaglUggalcga taagctgatatcgaatcgggaggtggtccctgcagtacgccaatgataacccccgccagaaaaatctagtagccttccctttgtttt ccgtgccccaactcggcggattgactcggccccttccggaaacacccgaatcaacttctagtcaaattattgttcacgccgcaatgacc cacccctggcccgcgtctgtggaactgacccctggtgtacaggagagtcgctgctgaaagtggtcccaaaggggtactagttttaag ctcccaactccccctcccccagcgtctggaggatccacaccctcgcaccgcaggggcgaggaagtgggcggagtccggtttggc gccagccgctgaggctgccaagcagaaaagccaccgctgaggagactccggtcactgtcctcgccccgcctcccccttccctcccct tggggaccaccgggcgccacgccgcgaacggtaagtgccgcggtcgtcggcgcctccgccctccccctagggccccaattcccag cgggcgcggcgcgcggccccgcgcgcagctcccggctccctcccccttcggatgtggcttgagctgtaggcgcggagtcctgcag ccccgataaaataaaagatttatttagtctccagaaaaaggggggaatgaaagaccccacctgtaggtttggcaagctagctgcagta acgccatttgcaaggcatggaaaaataccaaaccaagaatagagaagttcagatcaagggcgggtacatgaaaatagcta acgttgggccaaacaggatatctgcggtgagcagttcggccccggcccggggccaagaacagatggtcaccgcagttcgg ccccggcccgaggccaagaacagatggtccccagatatggcccaaccctcagcagttcttaagacccatcagatgtttccag gctcccccaaggacctgaaatgaccctgcgccttatttgaattaaccaatcagcctgcttctcgcttctgtcgcgcgcttctgct cccgagctctataaaagagctcacaacccctcactcggcgcgccagtcctccgacagactgagtcgcccgggggggatctgga gctctcgagaatctcacgcgtctgcaggatatcaagctgcggtaccgcgggcccggccaccatggacaagaagtacagcatcggc ctggccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaaggtgcccagcaagaaatcaaggtgctgggcaa caccgaccggcacagcatcaagaagaacctgatcggcgccctgctgtcgacagcggagaaacagccgaggccacccggctgaa gagaaccgccagaagaagatacaccagacggaagaaccggatctgctatctgcaagagatctcagcaacgagatggccaaggtg gacgacagcUcttccacagactggaagagtccttcctggtggaagaggataagaagcacgagcggcaccccatcttcggcaacatc gtggacgaggtggcctaccacgagaagtaccccaccatctaccacctgagaaagaaactggtggacagcaccgacaaggccgacctgcggctgatctatctggccctggcccacatgatcaagtccggggccacttcctgatcgagggcgacctgaaccccgacaacagcg acgtggacaagctgtcatccagctggtgcagacctacaaccagctgtcgaggaaaaccccatcaacgccagcggcgtggacgcc aaggccatcctgtctgccagactgagcaagagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaagaatggcc tgtcggcaacctgatgccctgagcctgggcctgacccccaacttcaagagcaactcgacctggccgaggatgccaaactgcagct gagcaaggacacctacgacgacgacctggacaacctgctggcccagatcggcgaccagtacgccgacctgtttctggccgccaag aacctgtccgacgccatcctgctgagcgacatcctgagagtgaacaccgagatcaccaaggcccccctgagcgcctctatgatcaag agatacgacgagcaccaccaggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgagaagtacaaagagatttctc gaccagagcaagaacggctacgccggctacatcgatggcggagccagccaggaagagtctacaagttcatcaagcccatcctgga aaagatggacggcaccgaggaactgctcgtgaagctgaacagagaggacctgctgcggaagcagcggacctcgacaacggcag catcccccaccagatccacctgggagagctgcacgccatctgcggcggcaggaagattttacccattcctgaaggacaaccggga aaagatcgagaagatcctgaccttccgcatcccctactacgtgggccctctggccaggggaaacagcagattcgcctggatgaccag aaagagcgaggaaaccatcaccccctggaacttcgaggaagtggtggacaagggcgccagcgcccagagcttcatcgagcggat gaccaacttcgataagaacctgcccaacgagaaggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtacaacgagct gaccaaagtgaaatacgtgaccgagggaatgagaaagcccgccttcctgagcggcgagcagaaaaaagccatcgtggacctgctg ttcaagaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaagaaaatcgagtgcttcgactccgtggaaatctcc ggcgtggaagatcggtcaacgcctccctgggcacataccacgatctgctgaaaattatcaaggacaaggactcctggacaatgagg aaaacgaggacatctggaagatatcgtgctgaccctgacactgttgaggacagagagatgatcgaggaacggctgaaaacctatg cccacctgtcgacgacaaagtgatgaagcagctgaagcggcggagatacaccggctggggcaggctgagccggaagctgatcaa cggcatccgggacaagcagtccggcaagacaatcctggattcctgaagtccgacggctcgccaacagaaactcatgcagctgat ccacgacgacagcctgaccttaaagaggacatccagaaagcccaggtgtccggccagggcgatagcctgcacgagcacatgcc aatctggccggcagccccgccataagaagggcatcctgcagacagtgaaggtggtggacgagctcgtgaaagtgatgggccggc acaagcccgagaacatcgtgatcgaaatggccagagagaaccagaccacccagaagggacagaagaacagccgcgagagaatg aagcggatcgaagagggcatcaaagagctgggcagccagatcctgaaagaacaccccgtggaaaacacccagctgcagaacgag aagctgtacctgtactacctgcagaatgggcgggatatgtacgtggaccaggaactggacatcaaccggctgtccgactacgatgtgg acgctatcgtgcctcagagctttctgaaggacgactccatcgataacaaagtgctgactcggagcgacaagaaccggggcaagagc gacaacgtgccctccgaagaggtcgtgaagaagatgaagaactactggcgccagctgctgaatgccaagctgattacccagaggaa gtcgacaatctgaccaaggccgagagaggcggcctgagcgaactggataaggccggcttcatcaagagacagctggtggaaacc cggcagatcacaaagcacgtggcacagatcctggactcccggatgaacactaagtacgacgagaacgacaaactgatccgggaag tgaaagtgatcaccctgaagtccaagctggtgtccgattccggaaggatttccagtttacaaagtgcgcgagatcaacaactaccacc acgcccacgacgcctacctgaacgccgtcgtgggaaccgccctgatcaaaaagtaccctaagctggaaagcgagtcgtgtacggc gactacaaggtgtacgacgtgcggaagatgatcgccaagagcgagcaggaaatcggcaaggctaccgccaagtactctctacag caacatcatgaactttcaagaccgagattaccctggccaacggcgagatccggaagcggcctctgatcgagacaaacggcgaaac aggcgagatcgtgtgggataagggccgggacttgccaccgtgcggaaagtgctgtctatgccccaagtgaatatcgtgaaaaagac cgaggtgcagacaggcggctcagcaaagagtctatcctgcccaagaggaacagcgacaagctgatcgccagaaagaaggactgg gaccctaagaagtacggcggcttcgacagccccaccgtggcctatctgtgctggtggtggccaaagtggaaaagggcaagtccaa gaaactgaagagtgtgaaagagctgctggggatcaccatcatggaaagaagcagctcgagaagaatcccatcgacttctggaagc caagggctacaaagaagtgaaaaaggacctgatcatcaagctgcctaagtactccctgttcgagctggaaaacggccggaagagaat gctggcctctgccggcgaactgcagaagggaaacgaactggccctgccctccaaatatgtgaacttcctgtacctggccagccactatgagaagctgaagggctcccccgaggataatgagcagaaacagctgttetggaacagcacaaacactacctggacgagatcatcea gcagatcagcgagtctccaagagagtgatcctggccgacgctaatctggacaaggtgctgagcgcctacaacaagcacagagaca agcctatcagagagcaggccgagaatatcatccacctgttaccctgaccaatctgggagcccctgccgccttcaagtacttgacacc accatcgaccggaagaggtacaccagcaccaaagaggtgctggacgccaccctgatccaccagagcatcaccggcctgtacgaga cacggatcgacctgtctcagctgggaggcgacgcctatccctatgacgtgcccgatatgccagcctgggcagcggctcccccaaga aaaaacgcaaggtggaagatcctaagaaaaagcggaaagtggacggcatggtagtgggagcaacggcagcagcggatocggag ggccgagctctggcgcacccccaccaagtggagggtctcctgccgggtccccaacatctactgaagaaggcaccagcgaatccgc aacgcccgagtcaggccctggtacctccacagaaccatctgaaggtagtgcgcctggttccccagctggaagccctactccaccga agaaggcacgtcaaccgaaccaagtgaaggatctgcccctgggaccagcactgaaccatctgagatggatgctaagtcactaactgc ctggtcccggacactggtgaccttcaaggatgtattgtggacttcaccagggaggagtggaagctgctggacactgctcagcagatc gtgtacagaaatgtgatgctggagaactataagaacctggtttccttgggtatcagctactaagccagatgtgatcctccggtggaga agggagaagagcccagcggccgcgctactaactcagcctgctgaagcaggctggagacgtggaggagaaccctggacctgtga gcaagggcgaggagctgttcaccggggtggtgcccatcctggtcgagctggacggcgacgtaaacggccacaagttcagcgtgtct ggcgagggcgagggcgatgccacctacggcaagctgaccctgaagtcatctgcaccaccggcaagctgcccgtgccctggccca ccctcgtgaccaccctgacctacggcgtgcagtgctcagccgctaccccgaccacatgaagcagcacgactcttcaagtccgccat gcccgaaggctacgtccaggagcgcaccatctcttcaaggacgacggcaactacaagacccgcgccgaggtgaagtcgagggc gacaccctggtgaaccgcatcgagctgaagggcatcgactcaaggaggacggcaacatcctggggcacaagctggagtacaact acaacagccacaacgtctatatcatggccgacaagcagaagaacggcatcaaggcgaactcaagatccgccacaacatcgaggac ggcagcgtgcagctcgccgaccactaccagcagaacacccccatcggcgacggccccgtgctgctgcccgacaaccactacctga gcacccagtccgccctgagcaaagaccccaacgagaagcgcgatcacatggtcctgctggagtcgtgaccgccgccgggatcact ctcggcatggacgagctgtacaagtaataagcgatcgccctgcaggcatgcaagctgatatcaagcttatcgatoafcaacc?c'?gg attacaaaatttgtgaaagattgactgg.tattcttaactatg.ttgctccttttacgctatgtgg.atacgctgctttaatgcctttgtatcatg ctattgcttcccgtatggctttccittttctcctccttgtataaatcctggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacg tggcgtggtgtgcactgtgtttgctgacgcaacccccactggttggggcattgccaccacctgtcagctcctttccgggactttcgcttt ccccctccctattgccacggcggacictcatcgccgcctgccttgcccgctgctggacaggggctcggctgttgggcactgacaattc cgtggtgttgtcggggaaatcatcgtcctttccttggctgctcgcctgtgttgccacctggattctgcgcgggacgtccttctgctacgtc ccttcggccctcaatccagcggaccttccttcccgcggcctgctgccggctctgcggcctcttccgcgtcttcgccttcgccctcagac gagteggafcfcccfflgggccgccfccccgcatcgataccgtcgacctcgagggaattaattcgagctcggtacctttaagaccaatg actacaaggcagctgtagatcttagccacttttaaaagaaaaggggggactggaagggctaatcactcccaacgaagtfaatfa asassscctatttcccatsattccttcatatttecatatacgatacaassctsttasasasataattesaattaatttsactetaaaca caaasatatagtacaaaatacgtgacgtasaaagtaataattctsggtagttgcagttaaaatatgtttaaaatssactatc atatsctaccgtaactgaaagtattcsatttctsgcttatatatcttgtssaaassacsaaacaccsscctcaagtatctagca agttctcsasaactsctaasatactsasgttttsaattctcgacctcgasacaaatsscagtattctaattA.2L?LC2Laggdctgc ttttgcttgtactgggtctctctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgcttaagcct caataaagcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagacccttta gtcagtgtggaaaatctctagca

[0317] In some embodiments, the recombinant integration deficient retroviral vector includes, from N-terminus to C-terminus, a 5’ SIN LTR derived from HIV-1, a viral packaging signal, an SFFV promoter, a CRISPRi cargo, a WPRE sequence, and a 3’ SIN LTR derived from HIV-1 that includes a U6 promoter and a SETDB1 shRNA. Inclusion of an sgRNA cassette is also contemplated in this exemplary' construct.

[0318] In some embodiments, the recombinant integration deficient retroviral vector comprises the nucleic acid sequence of SEQ ID NO: 800 as shown below.Bold text - HIV-1 LTRItalic text - HIV-1 Psi (packaging signal)Capital text - HIV-1 Rev responsive elementBold italic text - HIV-1 polypurine tract (PPT)Bold underline text - Spleen Focus Forming Virus (SFFV promoter)Underlined text - dCas9-KRAB-P2A-GFP (CRISPRi)Italic underlined text - WPREBold, italic, underlined text- human U6 promoter SETDB 1 shRNA expression cassetteTggaagggctaattcactcccaaagaagacaagatatcctgatctgtggatctaccacacacaaggctacttccctgattag cagaactacacaccagggccaggggtcagatatccactgacctttggatggtgctacaagctagtaccagtgagccagata aggtagaagaggccaataaaggagagaacaccagcttgttacaccctgtgagcctgcatgggatggatgacccggagagag aagtgttagagtggaggttgacagccgcctagcattcatcacgtggcccgagagctgcatccggagtacttcaagaactgct gatatcgagctgctacaagggacttccgctggggacttccagggaggcgtggcctgggcgggactggggagtggcgagc cctcagatcctgcatataagcagctgctttttgcctgtactgggtctctctggttagaccagatctgagcctgggagctctctggc taactagggaacccactgcttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgactctgg taactagagatccctcagaccctttagtcagtgtggaaaatctctagcagtggcgcccgaacagggacttgaaagcgaaaggg aaaccagaggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaagaggcgaggggcggcgactggtgagta cgccaaaaattttgactagcggaggctagaaggagagagatgggtgcgagagcgtcagtataagcgggggagaatagatcgc gatgggaaaaaattcggttaaggccagggggaaagaaaaaatataaattaaaacatatagtatgggcaagcagggagctagaacgat tcgcagttaatcctggcctgttagaaacatcagaaggctgtagacaaatactgggacagctacaaccatcccttcagacaggatcagaa gaacttagatcattatataatacagtagcaaccctctattgtgtgcatcaaaggatagagataaaagacaccaaggaagctttagacaag atagaggaagagcaaaacaaaagtaagaccaccgcacagcaagcggccggccgctgatcttcagacctggaggaggagatatga gggacaattggagaagtgaattatataaatataaagtagtaaaaattgaaccattaggagtagcaccCACCAAGGCAAAG AGAAGAGTGGTGCAGAGAGAAAAAAGAGCAGTGGGAATAGGAGCTTTGTTCCTT GGGTTCTTGGGAGCAGCAGGAAGCACTATGGGCGCAGCGTCAATGACGCTGACG GTACAGGCCAGACAATTATTGTCTGGTATAGTGCAGCAGCAGAACAATTTGCTG AGGGCTATTGAGGCGCAACAGCATCTGTTGCAACTCACAGTCTGGGGCATCAAGCAGCTCCAGGCAAGAATCCTGGCTGTGGAAAGATACCTAAAGGATCAACAGCTC CTGGGGATTTGGGGTTGCTCTGGAAAACTCATTTGCACCACTGCTGTGCCTTggaat gctagttggagtaataaatctctggaacagattggaatcacacgacctggatggagtgggacagagaaataacaatacacaagct aatacactccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaattatggaattagataaatgggcaagtttgtgga attggtttaacataacaaattggctgtggtatataaaattattcataatgatagtaggaggcttggtaggttaagaatagtttttgctgtactt ctatagtgaatagagttaggcagggatattcaccattatcgtttcagacccacctcccaaccccgaggggacccgacaggcccgaag gaatagaagaagaaggtggagagagagacagagacagatccatcgattagtgaacggatctcgacggtatcgccaaatggcagta Mcatccacaatttaaaagaaaaggggggattggggggtacagtgcaggggaaagaatagtagacataatagcaacagacat acaaactaaagaatacaaaaacaaatacaaaaattcaaaattttcgggtttatacagggacagcagagatccagttggatcga taagcttgatatcgaattcgggaggtggtccctgcagttacgccaatgataacccccgccagaaaaatcttagtagccttccctttttgtttt ccgtgccccaactcggcggatgactcggcccctccggaaacacccgaatcaacttctagtcaaatatgttcacgccgcaatgacc cacccctggcccgcgtctgtggaactgacccctggtgtacaggagagttcgctgctgaaagtggtcccaaaggggtactagttttaag ctcccaactccccctcccccagcgtctggaggattccacaccctcgcaccgcaggggcgaggaagtgggcggagtccggtttggc gccagccgctgaggctgccaagcagaaaagccaccgctgaggagactccggtcactgtcctcgccccgcctcccccttccctcccct tggggaccaccgggcgccacgccgcgaacggtaagtgccgcggtcgtcggcgcctccgccctccccctagggccccaatcccag cgggcgcggcgcgcggccccgcgcgcagctcccggctccctcccccttcggatgtggctgagctgtaggcgcggagtcctgcag ccccgataaaataaaagattttattagtctccagaaaaaggggggaatgaaagaccccacctgtaggtttggcaagctagctgcagta acgccatttgcaaggcatggaaaaataccaaaccaagaatagagaagttcagatcaagggcgggtacatgaaaatagcta acgttgggccaaacaggatatctgcggtgagcagtttcggccccggcccggggccaagaacagatggtcaccgcagttcgg ccccggcccgaggccaagaacagatggtccccagatatggcccaaccctcagcagtttctaagacccatcagatgtttccag gctcccccaaggacctgaaatgaccctgcgccttatttgaattaaccaatcagcctgcttctcgcttctgtcgcgcgcttctgct cccgagctctataaaagagctcacaacccctcactcggcgcgccagtcctccgacagactgagtcgcccgggggggatctgga gctctcgagaattctcacgcgtctgcaggatatcaagctgcggtaccgcgggcccggccaccatggacaagaagtacagcatcggc ctggccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaaggtgcccagcaagaaatcaaggtgctgggcaa caccgaccggcacagcatcaagaagaacctgatcggcgccctgctgtcgacagcggagaaacagccgaggccacccggctgaa gagaaccgccagaagaagatacaccagacggaagaaccggatctgctatctgcaagagatcttcagcaacgagatggccaaggtg gacgacagctctccacagactggaagagtccttcctggtggaagaggataagaagcacgagcggcaccccatctcggcaacatc gtggacgaggtggcctaccacgagaagtaccccaccatctaccacctgagaaagaaactggtggacagcaccgacaaggccgacc tgcggctgatctatctggccctggcccacatgatcaagtccggggccactcctgatcgagggcgacctgaaccccgacaacagcg acgtggacaagctgttcatccagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaacgccagcggcgtggacgcc aaggccatcctgtctgccagactgagcaagagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaagaatggcc tgtcggcaacctgattgccctgagcctgggcctgacccccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagct gagcaaggacacctacgacgacgacctggacaacctgctggcccagatcggcgaccagtacgccgacctgtttctggccgccaag aacctgtccgacgccatcctgctgagcgacatcctgagagtgaacaccgagatcaccaaggcccccctgagcgcctctatgatcaagagatacgacgagcaccaccaggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgagaagtacaaagagatttctc gaccagagcaagaacggctacgccggctacatcgatggcggagccagccaggaagagtctacaagtcatcaagcccatcctgga aaagatggacggcaccgaggaactgctcgtgaagctgaacagagaggacctgctgcggaagcagcggacctcgacaacggcag catcccccaccagatccacctgggagagctgcacgccattctgcggcggcaggaagatttttacccattcctgaaggacaaccggga aaagatcgagaagatcctgaccttccgcatcccctactacgtgggccctctggccaggggaaacagcagattcgcctggatgaccag aaagagcgaggaaaccalcaccccctggaactlcgaggaagtgglggacaagggcgccagcgcccagagcttcatcgagcggat gaccaactcgataagaacctgcccaacgagaaggtgctgcccaagcacagcctgctgtacgagtactcaccgtgtacaacgagct gaccaaagtgaaatacgtgaccgagggaatgagaaagcccgcctcctgagcggcgagcagaaaaaagccatcgtggacctgctg ttcaagaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaagaaaatcgagtgcttcgactccgtggaaatctcc ggcgtggaagatcggtcaacgcctccctgggcacataccacgatctgctgaaaattatcaaggacaaggacttcctggacaatgagg aaaacgaggacatctggaagatatcgtgctgaccctgacactgttgaggacagagagatgatcgaggaacggctgaaaacctatg cccacctgtcgacgacaaagtgatgaagcagctgaagcggcggagatacaccggctggggcaggctgagccggaagctgatcaa cggcatccgggacaagcagtccggcaagacaatcctggatttcctgaagtccgacggcttcgccaacagaaacttcatgcagctgat ccacgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtccggccagggcgatagcctgcacgagcacatgcc aatctggccggcagccccgccataagaagggcatcctgcagacagtgaaggtggtggacgagctcgtgaaagtgatgggccggc acaagcccgagaacatcgtgatcgaaatggccagagagaaccagaccacccagaagggacagaagaacagccgcgagagaatg aagcggatcgaagagggcatcaaagagctgggcagccagatcctgaaagaacaccccgtggaaaacacccagctgcagaacgag aagctgtacctgtactacctgcagaatgggcgggatatgtacgtggaccaggaactggacatcaaccggctgtccgactacgatgtgg acgctatcgtgcctcagagctttctgaaggacgactccatcgataacaaagtgctgactcggagcgacaagaaccggggcaagagc gacaacgtgccctccgaagaggtcgtgaagaagatgaagaactactggcgccagctgctgaatgccaagctgatacccagaggaa gtcgacaatctgaccaaggccgagagaggcggcctgagcgaactggataaggccggctcatcaagagacagctggtggaaacc cggcagatcacaaagcacgtggcacagatcctggactcccggatgaacactaagtacgacgagaacgacaaactgatccgggaag tgaaagtgatcaccctgaagtccaagctggtgtccgattccggaaggatttccagttttacaaagtgcgcgagatcaacaactaccacc acgcccacgacgcctacctgaacgccgtcgtgggaaccgccctgatcaaaaagtaccctaagctggaaagcgagtcgtgtacggc gactacaaggtgtacgacgtgcggaagatgatcgccaagagcgagcaggaaatcggcaaggctaccgccaagtactctctacag caacatcatgaactttttcaagaccgagattaccctggccaacggcgagatccggaagcggcctctgatcgagacaaacggcgaaac aggcgagatcgtgtgggataagggccgggactttgccaccgtgcggaaagtgctgtctatgccccaagtgaatatcgtgaaaaagac cgaggtgcagacaggcggcttcagcaaagagtctatcctgcccaagaggaacagcgacaagctgatcgccagaaagaaggactgg gaccctaagaagtacggcggctcgacagccccaccgtggcctatctgtgctggtggtggccaaagtggaaaagggcaagtccaa gaaactgaagagtgtgaaagagctgctggggatcaccatcatggaaagaagcagcttcgagaagaatcccatcgactttctggaagc caagggctacaaagaagtgaaaaaggacctgatcatcaagctgcctaagtactccctgttcgagctggaaaacggccggaagagaat gctggcctctgccggcgaactgcagaagggaaacgaactggccctgccctccaaatatgtgaactcctgtacctggccagccactat gagaagctgaagggctcccccgaggataatgagcagaaacagctgttgtggaacagcacaaacactacctggacgagatcatcga gcagatcagcgagtctccaagagagtgatcctggccgacgctaatctggacaaggtgctgagcgcctacaacaagcacagagacaagcctatcagagagcaggccgagaatatcatccacctgttaccctgaccaatctgggagcccctgccgccttcaagtactttgacacc accatcgaccggaagaggtacaccagcaccaaagaggtgctggacgccaccctgatccaccagagcatcaccggcctgtacgaga cacggatcgacctgtctcagctgggaggcgacgcctatccctatgacgtgcccgatatgccagcctgggcagcggctcccccaaga aaaaacgcaaggtggaagatcctaagaaaaagcggaaagtggacggcattggtagtgggagcaacggcagcagcggatccggag ggccgagctctggcgcacccccaccaagtggagggtctcctgccgggtccccaacatctactgaagaaggcaccagcgaatccgc aacgcccgagtcaggccctgglacclccacagaaccatctgaaggtagtgcgcclggtlccccagctggaagccctacttccaccga agaaggcacgtcaaccgaaccaagtgaaggatctgcccctgggaccagcactgaaccatctgagatggatgctaagtcactaactgc ctggtcccggacactggtgacctcaaggatgtattgtggacttcaccagggaggagtggaagctgctggacactgctcagcagatc gtgtacagaaatgtgatgctggagaactataagaacctggtttccttgggtatcagctactaagccagatgtgatcctccggtggaga agggagaagagcccagcggccgcgctactaactcagcctgctgaagcaggctggagacgtggaggagaaccctggacctgtga gcaagggcgaggagctgtcaccggggtggtgcccatcctggtcgagctggacggcgacgtaaacggccacaagtcagcgtgtct ggcgagggcgagggcgatgccacctacggcaagctgaccctgaagtcatctgcaccaccggcaagctgcccgtgccctggccca ccctcgtgaccaccctgacctacggcgtgcagtgcttcagccgctaccccgaccacatgaagcagcacgactcttcaagtccgccat gcccgaaggctacgtccaggagcgcaccatctcttcaaggacgacggcaactacaagacccgcgccgaggtgaagttcgagggc gacaccctggtgaaccgcatcgagctgaagggcatcgacttcaaggaggacggcaacatcctggggcacaagctggagtacaact acaacagccacaacgtctatatcatggccgacaagcagaagaacggcatcaaggcgaactcaagatccgccacaacatcgaggac ggcagcgtgcagctcgccgaccactaccagcagaacacccccatcggcgacggccccgtgctgctgcccgacaaccactacctga gcacccagtccgccctgagcaaagaccccaacgagaagcgcgatcacatggtcctgctggagttcgtgaccgccgccgggatcact ctcggcatggacgagctgtacaagtaataagcgatcgccctgcaggcatgcaagcttgatatcaagctatcgataatcaacctctgga tacaaaattgtgaaagatgactggtatctaactatgtgctccttacgctatgtggatacgctgcttaatgccttgtatcatgctat gctcccgtatggcttcattctcctcctgtataaatcctggtgctgtctcttatgaggagtgtggcccgtgtcaggcaacgtggcgt ggtgtgcactgtgtttgctgacgcaacccccactggttggggcattgccaccacctgtcagctcctttccgggactttcgcttccccctc cctattgccacggcggaactcatcgccgcctgcctgcccgctgctggacaggggctcggctgttgggcactgacaatccgtggtgt gtcggggaaatcatcgtccttccttggctgctcgcctgtgtgccacctggatctgcgcgggacgtcctctgctacgtccctcggcc ctcaatccagcggacctcctcccgcggcctgctgccggctctgcggcctctccgcgtcttcgcctcgccctcagacgagtcggat ctccctttgggccgcctccccgcatcgataccgtcgacctcgagggaattaattcgagctcggtacctttaagaccaatgacttacaagg cagctgtagatctagccacttttaaaagaaaaggggggactggaagggctaattcactcccaacgaagtfaatfaagagggcc / aittcccatsattccttcataittscatatacsatacaassctstasasasataattssaaitaattsactstaaacacaaasatatt astacaaaatacstsacstasaaastaataattctssstasttscastttaaaatatsttaaaatssactaicatatsctacc QtaacttsaaaetattcsattcttQscttatatatcttgtssaaassacsaaacaccsssctcasatsataacttctstactcsagt acaeaaettatcatctsascttttttgaattctcsacctceasacaaatsscaetattcttaattsiasicsia2.sitct2Kttttt^.ctt^.tii ctgggtctctctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgctaagcctcaataaagctt gccttgagtgcttcaagtagtgtgtgcccgtctgtgtgtgactctggtaactagagatccctcagacccttttagtcagtgtgga aaatctctagca

[0319] In some embodiments, the recombinant integration deficient retroviral vector includes, from N-terminus to C-terminus, a 5’ SIN LTR derived from HIV-1, a viral packaging signal, an SFFV promoter, a CRISPRi cassette, a WPRE sequence, and a 3’ SIN LTR derived from HIV-1 that includes a U6 promoter and a TASOR shRNA. Inclusion of an sgRNA cassette is also contemplated in this exemplary construct.

[0320] In some embodiments, the recombinant integration deficient retroviral vector comprises the nucleic acid sequence of SEQ ID NO: 801 as shown below.Bold text - HIV-1 LTRItalic text - HIV-1 Psi (packaging signal)Capital text - HIV-1 Rev responsive elementBold italic text - HIV-1 polypurine tract (PPT)Bold underline text - Spleen Focus Forming Virus (SFFV promoter)Underlined text - dCas9-KRAB-P2A-GFP (CRISPRi)Italic underlined text - WPREBold, italic, underlined text - human U6 promoter TASOR shRNA expression cassetteTggaagggctaattcactcccaaagaagacaagatatccttgatctgtggatctaccacacacaaggctacttccctgattag cagaactacacaccagggccaggggtcagatatccactgacctttggatggtgctacaagctagtaccagttgagccagata aggtagaagaggccaataaaggagagaacaccagcttgttacaccctgtgagcctgcatgggatggatgacccggagagag aagtgttagagtggaggtttgacagccgcctagcatttcatcacgtggcccgagagctgcatccggagtacttcaagaactgct gatatcgagcttgctacaagggacttccgctggggacttccagggaggcgtggcctgggcgggactggggagtggcgagc cctcagatcctgcatataagcagctgctttttgcctgtactgggtctctctggttagaccagatctgagcctgggagctctctggc taactagggaacccactgcttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgactctgg taactagagatccctcagaccctttagtcagtgtggaaaatctctagcagtggcgcccgaacagggacttgaaagcgaaaggg aaaccagaggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaagaggcgaggggcggcgactggtgagta cgccaaaaattttgactagcggaggctagaaggagagagatgggtgcgagagcgtcagtattaagcgggggagaatagatcgc gatgggaaaaaattcggttaaggccagggggaaagaaaaaatataaattaaaacatatagtatgggcaagcagggagctagaacgat tcgcagttaatcctggcctgttagaaacatcagaaggctgtagacaaatactgggacagctacaaccatcccttcagacaggatcagaa gaacttagatcattatataatacagtagcaaccctctattgtgtgcatcaaaggatagagataaaagacaccaaggaagctttagacaag atagaggaagagcaaaacaaaagtaagaccaccgcacagcaagcggccggccgctgatcttcagacctggaggaggagatatga gggacaattggagaagtgaattatataaatataaagtagtaaaaattgaaccattaggagtagcaccCACCAAGGCAAAG AGAAGAGTGGTGCAGAGAGAAAAAAGAGCAGTGGGAATAGGAGCTTTGTTCCTT GGGTTCTTGGGAGCAGCAGGAAGCACTATGGGCGCAGCGTCAATGACGCTGACG GTACAGGCCAGACAATTATTGTCTGGTATAGTGCAGCAGCAGAACAATTTGCTG AGGGCTATTGAGGCGCAACAGCATCTGTTGCAACTCACAGTCTGGGGCATCAAG CAGCTCCAGGCAAGAATCCTGGCTGTGGAAAGATACCTAAAGGATCAACAGCTC CTGGGGATTTGGGGTTGCTCTGGAAAACTCATTTGCACCACTGCTGTGCCTTGgaatgctagttggagtaataaatctctggaacagattggaatcacacgacctggatggagtgggacagagaaataacaatacacaagct aatacactcctaatgaagaatcgcaaaaccagcaagaaaagaatgaacaagaatatggaattagataaatgggcaagttgtgga atggttaacataacaaatggctgtggtatataaaatatcataatgatagtaggaggctggtaggttaagaatagttttgctgtactt ctatagtgaatagagtaggcagggatatcaccatatcgttcagacccacctcccaaccccgaggggacccgacaggcccgaag gaatagaagaagaaggtggagagagagacagagacagatccatcgatagtgaacggatctcgacggtatcgccaaatggcagta Mcatccacwtttaaaagaaaaggggggattggggggtacagtgcaggggaaagaatagtagacaiaatagcaacagacat acaaactaaagaattacaaaaacaaatacaaaaattcaaaatttcgggtttaitacagggacagcagagatccagttggaicga taagcttgatatcgaattcgggaggtggtccctgcagttacgccaatgataacccccgccagaaaaatcttagtagccttccctttttgtttt ccgtgccccaactcggcggattgactcggccccttccggaaacacccgaatcaacttctagtcaaattattgttcacgccgcaatgacc cacccctggcccgcgtctgtggaactgacccctggtgtacaggagagttcgctgctgaaagtggtcccaaaggggtactagttttaag ctcccaactccccctcccccagcgtctggaggattccacaccctcgcaccgcaggggcgaggaagtgggcggagtccggtttggc gccagccgctgaggctgccaagcagaaaagccaccgctgaggagactccggtcactgtcctcgccccgcctcccccttccctcccct tggggaccaccgggcgccacgccgcgaacggtaagtgccgcggtcgtcggcgcctccgccctccccctagggccccaattcccag cgggcgcggcgcgcggccccgcgcgcagctcccggctccctcccccttcggatgtggctgagctgtaggcgcggagtcctgcag ccccgataaaataaaagattttatttagtctccagaaaaaggggggaatgaaagaccccacctgtaggtttggcaagctagctgcagta acgccatttgcaaggcatggaaaaataccaaaccaagaatagagaagttcagatcaagggcgggtacatgaaaatagcta acgttgggccaaacaggatatctgcggtgagcagttcggccccggcccggggccaagaacagatggtcaccgcagttcgg ccccggcccgaggccaagaacagatggtccccagatatggcccaaccctcagcagtttctaagacccatcagatgtttccag gctcccccaaggacctgaaatgaccctgcgccttatttgaattaaccaatcagcctgcttctcgcttctgtcgcgcgcttctgct cccgagctctataaaagagctcacaacccctcactcggcgcgccagtcctccgacagactgagtcgcccgggggggatctgga gctctcgagaattctcacgcgtctgcaggatatcaagcttgcggtaccgcgggcccggccaccatggacaagaagtacagcatcggc ctggccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaaggtgcccagcaagaaattcaaggtgctgggcaa caccgaccggcacagcatcaagaagaacctgatcggcgccctgctgttcgacagcggagaaacagccgaggccacccggctgaa gagaaccgccagaagaagatacaccagacggaagaaccggatctgctatctgcaagagatctcagcaacgagatggccaaggtg gacgacagctctccacagactggaagagtccttcctggtggaagaggataagaagcacgagcggcaccccatctcggcaacatc gtggacgaggtggcctaccacgagaagtaccccaccatctaccacctgagaaagaaactggtggacagcaccgacaaggccgacc tgcggctgatctatctggccctggcccacatgatcaagtccggggccacttcctgatcgagggcgacctgaaccccgacaacagcg acgtggacaagctgttcatccagctggtgcagacctacaaccagctgttcgaggaaaaccccatcaacgccagcggcgtggacgcc aaggccatcctgtctgccagactgagcaagagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaagaatggcc tgttcggcaacctgattgccctgagcctgggcctgacccccaacttcaagagcaacttcgacctggccgaggatgccaaactgcagct gagcaaggacacctacgacgacgacctggacaacctgctggcccagatcggcgaccagtacgccgacctgttctggccgccaag aacctgtccgacgccatcctgctgagcgacatcctgagagtgaacaccgagatcaccaaggcccccctgagcgcctctatgatcaag agatacgacgagcaccaccaggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgagaagtacaaagagattctc gaccagagcaagaacggctacgccggctacatcgatggcggagccagccaggaagagtctacaagtcatcaagcccatcctgga aaagatggacggcaccgaggaactgctcgtgaagctgaacagagaggacctgctgcggaagcagcggacctcgacaacggcag catcccccaccagatccacctgggagagctgcacgccatctgcggcggcaggaagattttacccatcctgaaggacaaccggga aaagatcgagaagalcctgaccttccgcatcccctaclacgtgggccctclggccaggggaaacagcagatlcgcclggatgaccag aaagagcgaggaaaccatcaccccctggaacttcgaggaagtggtggacaagggcgccagcgcccagagctcatcgagcggatgaccaactcgataagaacctgcccaacgagaaggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtacaacgagct gaccaaagtgaaatacgtgaccgagggaatgagaaagcccgcctcctgagcggcgagcagaaaaaagccatcgtggacctgctg ttcaagaccaaccggaaagtgaccgtgaagcagctgaaagaggactactcaagaaaatcgagtgctcgactccgtggaaatctcc ggcgtggaagatcggtcaacgcctccctgggcacataccacgatctgctgaaaatatcaaggacaaggactcctggacaatgagg aaaacgaggacattctggaagatatcgtgctgaccctgacactgtttgaggacagagagatgatcgaggaacggctgaaaacctatg cccacctgtcgacgacaaagtgatgaagcagctgaagcggcggagatacaccggctggggcaggctgagccggaagctgatcaa cggcatccgggacaagcagtccggcaagacaatcctggattcctgaagtccgacggctcgccaacagaaacttcatgcagctgat ccacgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtccggccagggcgatagcctgcacgagcacatgcc aatctggccggcagccccgccattaagaagggcatcctgcagacagtgaaggtggtggacgagctcgtgaaagtgatgggccggc acaagcccgagaacatcgtgatcgaaatggccagagagaaccagaccacccagaagggacagaagaacagccgcgagagaatg aagcggatcgaagagggcatcaaagagctgggcagccagatcctgaaagaacaccccgtggaaaacacccagctgcagaacgag aagctgtacctgtactacctgcagaatgggcgggatatgtacgtggaccaggaactggacatcaaccggctgtccgactacgatgtgg acgctatcgtgcctcagagctttctgaaggacgactccatcgataacaaagtgctgactcggagcgacaagaaccggggcaagagc gacaacgtgccctccgaagaggtcgtgaagaagatgaagaactactggcgccagctgctgaatgccaagctgattacccagaggaa gtcgacaatctgaccaaggccgagagaggcggcctgagcgaactggataaggccggctcatcaagagacagctggtggaaacc cggcagatcacaaagcacgtggcacagatcctggactcccggatgaacactaagtacgacgagaacgacaaactgatccgggaag tgaaagtgatcaccctgaagtccaagctggtgtccgattccggaaggatttccagttacaaagtgcgcgagatcaacaactaccacc acgcccacgacgcctacctgaacgccgtcgtgggaaccgccctgatcaaaaagtaccctaagctggaaagcgagtcgtgtacggc gactacaaggtgtacgacgtgcggaagatgatcgccaagagcgagcaggaaatcggcaaggctaccgccaagtactctctacag caacatcatgaacttttcaagaccgagataccctggccaacggcgagatccggaagcggcctctgatcgagacaaacggcgaaac aggcgagatcgtgtgggataagggccgggacttgccaccgtgcggaaagtgctgtctatgccccaagtgaatatcgtgaaaaagac cgaggtgcagacaggcggcttcagcaaagagtctatcctgcccaagaggaacagcgacaagctgatcgccagaaagaaggactgg gaccctaagaagtacggcggctcgacagccccaccgtggcctattctgtgctggtggtggccaaagtggaaaagggcaagtccaa gaaactgaagagtgtgaaagagctgctggggatcaccatcatggaaagaagcagcttcgagaagaatcccatcgactttctggaagc caagggctacaaagaagtgaaaaaggacctgatcatcaagctgcctaagtactccctgttcgagctggaaaacggccggaagagaat gctggcctctgccggcgaactgcagaagggaaacgaactggccctgccctccaaatatgtgaactcctgtacctggccagccactat gagaagctgaagggctcccccgaggataatgagcagaaacagctgtttgtggaacagcacaaacactacctggacgagatcatcga gcagatcagcgagtctccaagagagtgatcctggccgacgctaatctggacaaggtgctgagcgcctacaacaagcacagagaca agcctatcagagagcaggccgagaatatcatccacctgttaccctgaccaatctgggagcccctgccgccttcaagtactttgacacc accatcgaccggaagaggtacaccagcaccaaagaggtgctggacgccaccctgatccaccagagcatcaccggcctgtacgaga cacggatcgacctgtctcagctgggaggcgacgcctatccctatgacgtgcccgatatgccagcctgggcagcggctcccccaaga aaaaacgcaaggtggaagatcctaagaaaaagcggaaagtggacggcatggtagtgggagcaacggcagcagcggatccggag ggccgagctctggcgcacccccaccaagtggagggtctcctgccgggtccccaacatctactgaagaaggcaccagcgaatccgc aacgcccgagtcaggccctggtacctccacagaaccatctgaaggtagtgcgcctggtccccagctggaagccctactccaccga agaaggcacgtcaaccgaaccaagtgaaggatctgcccctgggaccagcactgaaccatctgagatggatgctaagtcactaactgc ctggtcccggacactggtgacctcaaggatgtattgtggactcaccagggaggagtggaagctgctggacactgctcagcagatc gtgtacagaaatgtgatgctggagaactataagaacctggtttccttgggttatcagcttactaagccagatgtgatcctccggttggaga agggagaagagcccagcggccgcgctactaactcagcctgctgaagcaggctggagacgtggaggagaaccctggacctgtgagcaagggcgaggagctgttcaccggggtggtgcccatcctggtcgagctggacggcgacgtaaacggccacaagttcagcgtgtct ggcgagggcgagggcgatgccacctacggcaagctgaccctgaagttcatctgcaccaccggcaagctgcccgtgccctggccca ccctcgtgaccaccctgacctacggcgtgcagtgcttcagccgctaccccgaccacatgaagcagcacgacttcttcaagtccgccat gcccgaaggctacgtccaggagcgcaccatcttcttcaaggacgacggcaactacaagacccgcgccgaggtgaagttcgagggc gacaccctggtgaaccgcatcgagctgaagggcatcgacttcaaggaggacggcaacatcctggggcacaagctggagtacaact acaacagccacaacgtctatatcatggccgacaagcagaagaacggcatcaaggcgaacttcaagatccgccacaacatcgaggac ggcagcgtgcagctcgccgaccactaccagcagaacacccccatcggcgacggccccgtgctgctgcccgacaaccactacctga gcacccagtccgccctgagcaaagaccccaacgagaagcgcgatcacatggtcctgctggagttcgtgaccgccgccgggatcact ctcggcatggacgagctgtacaagtaataagcgatcgccctgcaggcatgcaagcttgatatcaagcttatcgatoataMcctetgg attacaaaatttgtgaaagcittgactggtattcttaactatgttgctccttttacgctatgtggatacgctgctttaatgcctttgtatcatg ctattgcttcccgtatggctttcattttctcctccttgtataaatcctggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacg tggcgtggtgtgcactgtgtttgctgacgcaacccccactggttggggcattgccaccacctgtcagctcctttccgggactttcgcttt ccccctccctattgccacggcggaactcatcgccgcctgccttgcccgctgctggacaggggctcggctgttgggcactgcicaattc cgtggtgttgtcggggaaatcatcgtcctttccttggctgctcgcctgtgttgccacctggattctgcgcgggacgtccttctgctacgtc ccttcggccctcaatccagcggaccttccttcccgcggcctgctgccggctctgcggcctcttccgcgtcttcgccttcgccctcagac gagfcggQfcfcccfflgggccgcctecccgcatcgataccgtcgacctcgagggaataatcgagctcggtaccttaagaccaatg acttacaaggcagctgtagatcttagccactttttaaaagaaaaggggggactggaagggctaatcactcccaacgaagftoartfl asassscctattcccatsattccttcatattscatatacsatacaassctsttasasasaiaattssaattaattgactstaaaca caaasatattastacaaaatacstsacstasaaagtaataattctssstasttscasttaaaatatstttaaaatssactatc atatsctaccgtaactsaaastattcsattctsscttatatatctstsgaaassacsaaacaccssscassttcatcacaa gatactcgagtaictgtgaatgaaacctgctttttgaattctcgacctcgagacaaaiggcagtattcttaattasisigiiSLgsAct'S.^ ttttgcttgtactgggtctctctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgcttaagcctc aataaagctgcctgagtgctcaagtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagaccctttagt cagtgtggaaaatctctagca

[0321] In some embodiments, the recombinant integration deficient retroviral vector comprises the nucleic acid sequence of SEQ ID NO: 802 as shown below. Inclusion of an sgRNA cassette is also contemplated in this exemplary construct.Bold text - HIV-1 LTRItalic text - HIV-1 Psi (packaging signal)Capital text - HIV-1 Rev responsive elementBold italic text - HIV-1 polypurine tract (PPT)Bold underline text - Spleen Focus Forming Virus (SFFV promoter)Underlined text - dCas9-KRAB-P2A-GFP (CRISPRi) Italic underlined text - WPREBold, italic, underlined text- human U6 promoter TASOR shRNA expression cassetteTggaagggctaattcactcccaaagaagacaagatatcctgatctgtggatctaccacacacaaggctacttccctgatagcagaactacacaccagggccaggggtcagatatccactgacctttggatggtgctacaagctagtaccagttgagccagata aggtagaagaggccaataaaggagagaacaccagcttgttacaccctgtgagcctgcatgggatggatgacccggagagag aagtgttagagtggaggtttgacagccgcctagcatttcatcacgtggcccgagagctgcatccggagtacttcaagaactgct gatatcgagctgctacaagggacttccgctggggacttccagggaggcgtggcctgggcgggactggggagtggcgagc cctcagatcctgcatataagcagctgcttttgcctgtactgggtctctctggtagaccagatctgagcctgggagctctctggc taactagggaacccactgcttaagcctcaataaagcttgccttgagtgcttcaagtagtgtgtgcccgtctgttgtgtgactctgg taactagagatccctcagacccttttagtcagtgtggaaaatctctagcagtggcgcccgaacagggactgaaagcgaaaggg aaaccagaggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaagaggcgaggggcggcgactggtgagta cgccaaaaattttgactagcggaggctagaaggagagagatgggtgcgagagcgtcagtattaagcgggggagaatagatcgc gatgggaaaaaattcggttaaggccagggggaaagaaaaaatataaattaaaacatatagtatgggcaagcagggagctagaacgat tcgcagtaatcctggcctgtagaaacatcagaaggctgtagacaaatactgggacagctacaaccatcccttcagacaggatcagaa gaactagatcatatataatacagtagcaaccctctatgtgtgcatcaaaggatagagataaaagacaccaaggaagcttagacaag atagaggaagagcaaaacaaaagtaagaccaccgcacagcaagcggccggccgctgatcttcagacctggaggaggagatatga gggacaattggagaagtgaattatataaatataaagtagtaaaaattgaaccattaggagtagcaccCACCAAGGCAAAG AGAAGAGTGGTGCAGAGAGAAAAAAGAGCAGTGGGAATAGGAGCTTTGTTCCTT GGGTTCTTGGGAGCAGCAGGAAGCACTATGGGCGCAGCGTCAATGACGCTGACG GTACAGGCCAGACAATTATTGTCTGGTATAGTGCAGCAGCAGAACAATTTGCTG AGGGCTATTGAGGCGCAACAGCATCTGTTGCAACTCACAGTCTGGGGCATCAAG CAGCTCCAGGCAAGAATCCTGGCTGTGGAAAGATACCTAAAGGATCAACAGCTC CTGGGGATTTGGGGTTGCTCTGGAAAACTCATTTGCACCACTGCTGTGCCTTGgaat gctagttggagtaataaatctctggaacagattggaatcacacgacctggatggagtgggacagagaaataacaatacacaagctt aatacactccttaattgaagaatcgcaaaaccagcaagaaaagaatgaacaagaattatggaattagataaatgggcaagttgtgga attggtttaacataacaaattggctgtggtatataaaattattcataatgatagtaggaggctggtaggttaagaatagtttttgctgtactt ctatagtgaatagagtaggcagggatattcaccatatcgttcagacccacctcccaaccccgaggggacccgacaggcccgaag gaatagaagaagaaggtggagagagagacagagacagatccatcgattagtgaacggatctcgacggtatcgccaaatggcagta W.c,a ccacaattaaaagaaaaggggggatggggggtacagtgcaggggaaagaatagtagacataatagcaacagacat acaaactaaagaattacaaaaacaaattacaaaaattcaaaattttcgggtttattacagggacagcagagatccagttggatcga taagcttgatatcgaatcgggaggtggtccctgcagttacgccaatgataacccccgccagaaaaatctagtagcctcccttttgttt ccgtgccccaactcggcggatgactcggcccctccggaaacacccgaatcaacttctagtcaaatatgtcacgccgcaatgacc cacccctggcccgcgtctgtggaactgacccctggtgtacaggagagtcgctgctgaaagtggtcccaaaggggtactagttttaag ctcccaactccccctcccccagcgtctggaggattccacaccctcgcaccgcaggggcgaggaagtgggcggagtccggtttggc gccagccgctgaggctgccaagcagaaaagccaccgctgaggagactccggtcactgtcctcgccccgcctcccccttccctcccct tggggaccaccgggcgccacgccgcgaacggtaagtgccgcggtcgtcggcgcctccgccctccccctagggccccaatcccag cgggcgcggcgcgcggccccgcgcgcagctcccggctccctcccccttcggatgtggctgagctgtaggcgcggagtcctgcagccccgataaaataaaagattttatttagtctccagaaaaaggggggaatgaaagaccccacctgtaggtttggcaagctagctgcagta acgccattttgcaaggcatggaaaaataccaaaccaagaatagagaagttcagatcaagggcgggtacatgaaaatagcta acgttgggccaaacaggatatctgcggtgagcagttcggccccggcccggggccaagaacagatggtcaccgcagttcgg ccccggcccgaggccaagaacagatggtccccagatatggcccaaccctcagcagtttctaagacccatcagatgtttccag gctcccccaaggacctgaaatgaccctgcgccttatttgaattaaccaatcagcctgcttctcgcttctgtcgcgcgcttctgct cccgagctctataaaagagctcacaacccctcactcggcgcgccagtcctccgacagactgagtcgcccgggggggatctgga gctctcgagaattctcacgcgtctgcaggatatcaagctgcggtaccgcgggcccggccaccatggacaagaagtacagcatcggc ctggccatcggcaccaactctgtgggctgggccgtgatcaccgacgagtacaagetecccagcaagaaattcaaggtgctgggcaa caccgaccggcacagcatcaagaagaacctgatcggcgccctgctgttcgacagcggagaaacagccgaggccacccggctgaa gagaaccgccagaagaagatacaccagacggaagaaccggatctgctatctgcaagagatctcagcaacgagatggccaaggtg gacgacagctctccacagactggaagagtcctcctggtggaagaggataagaagcacgagcggcaccccatctcggcaacatc gtggacgaggtggcctaccacgagaagtaccccaccatctaccacctgagaaagaaactggtggacagcaccgacaaggccgacc tgcggctgatctatctggccctggcccacatgatcaagtccggggccacttcctgatcgagggcgacctgaaccccgacaacagcg acgtggacaagctgttcatccagctggtgcagacctacaaccagctgtcgaggaaaaccccatcaacgccagcggcgtggacgcc aaggccatcctgtctgccagactgagcaagagcagacggctggaaaatctgatcgcccagctgcccggcgagaagaagaatggcc tgtcggcaacctgatgccctgagcctgggcctgacccccaactcaagagcaactcgacctggccgaggatgccaaactgcagct gagcaaggacacctacgacgacgacctggacaacctgctggcccagatcggcgaccagtacgccgacctgttctggccgccaag aacctgtccgacgccatcctgctgagcgacatcctgagagtgaacaccgagatcaccaaggcccccctgagcgcctctatgatcaag agatacgacgagcaccaccaggacctgaccctgctgaaagctctcgtgcggcagcagctgcctgagaagtacaaagagatttctc gaccagagcaagaacggctacgccggctacatcgatggcggagccagccaggaagagtctacaagtcatcaagcccatcctgga aaagatggacggcaccgaggaactgctcgtgaagctgaacagagaggacctgctgcggaagcagcggacctcgacaacggcag catcccccaccagatccacctgggagagctgcacgccattctgcggcggcaggaagatttttacccattcctgaaggacaaccggga aaagatcgagaagatcctgacctccgcatcccctactacgtgggccctctggccaggggaaacagcagattcgcctggatgaccag aaagagcgaggaaaccatcaccccctggaactcgaggaagtggtggacaagggcgccagcgcccagagctcatcgagcggat gaccaactcgataagaacctgcccaacgagaaggtgctgcccaagcacagcctgctgtacgagtacttcaccgtgtacaacgagct gaccaaagtgaaatacgtgaccgagggaatgagaaagcccgccttcctgagcggcgagcagaaaaaagccatcgtggacctgctg ttcaagaccaaccggaaagtgaccgtgaagcagctgaaagaggactacttcaagaaaatcgagtgctcgactccgtggaaatctcc ggcgtggaagatcggttcaacgcctccctgggcacataccacgatctgctgaaaatatcaaggacaaggacttcctggacaatgagg aaaacgaggacatctggaagatatcgtgctgaccctgacactgttgaggacagagagatgatcgaggaacggctgaaaacctatg cccacctgtcgacgacaaagtgatgaagcagctgaagcggcggagatacaccggctggggcaggctgagccggaagctgatcaa cggcatccgggacaagcagtccggcaagacaatcctggatttcctgaagtccgacggctcgccaacagaaacttcatgcagctgat ccacgacgacagcctgacctttaaagaggacatccagaaagcccaggtgtccggccagggcgatagcctgcacgagcacatgcc aatctggccggcagccccgccattaagaagggcatcctgcagacagtgaaggtggtggacgagctcgtgaaagtgatgggccggc acaagcccgagaacatcgtgatcgaaatggccagagagaaccagaccacccagaagggacagaagaacagccgcgagagaatgaagcggatcgaagagggcatcaaagagctgggcagccagatcctgaaagaacaccccgtggaaaacacccagctgcagaacgag aagctgtacctgtactacctgcagaatgggcgggatatgtacgtggaccaggaactggacatcaaccggctgtccgactacgatgtgg acgctatcgtgcctcagagcttctgaaggacgactccatcgataacaaagtgctgactcggagcgacaagaaccggggcaagagc gacaacgtgccctccgaagaggtcgtgaagaagatgaagaactactggcgccagctgctgaatgccaagctgattacccagaggaa gtcgacaatctgaccaaggccgagagaggcggcctgagcgaactggataaggccggcttcatcaagagacagctggtggaaacc cggcagalcacaaagcacgtggcacagalcctggactcccggatgaacactaagtacgacgagaacgacaaactgatccgggaag tgaaagtgatcaccctgaagtccaagctggtgtccgattccggaaggatttccagttacaaagtgcgcgagatcaacaactaccacc acgcccacgacgcctacctgaacgccgtcgtgggaaccgccctgatcaaaaagtaccctaagctggaaagcgagtcgtgtacggc gactacaaggtgtacgacgtgcggaagatgatcgccaagagcgagcaggaaatcggcaaggctaccgccaagtactcttctacag caacatcatgaacttttcaagaccgagattaccctggccaacggcgagatccggaagcggcctctgatcgagacaaacggcgaaac aggcgagatcgtgtgggataagggccgggacttgccaccgtgcggaaagtgctgtctatgccccaagtgaatatcgtgaaaaagac cgaggtgcagacaggcggcttcagcaaagagtctatcctgcccaagaggaacagcgacaagctgatcgccagaaagaaggactgg gaccctaagaagtacggcggcttcgacagccccaccgtggcctattctgtgctggtggtggccaaagtggaaaagggcaagtccaa gaaactgaagagtgtgaaagagctgctggggatcaccatcatggaaagaagcagcttcgagaagaatcccatcgactttctggaagc caagggctacaaagaagtgaaaaaggacctgatcatcaagctgcctaagtactccctgtcgagctggaaaacggccggaagagaat gctggcctctgccggcgaactgcagaagggaaacgaactggccctgccctccaaatatgtgaactcctgtacctggccagccactat gagaagctgaagggctcccccgaggataatgagcagaaacagctgttgtggaacagcacaaacactacctggacgagatcatcga gcagatcagcgagtctccaagagagtgatcctggccgacgctaatctggacaaggtgctgagcgcctacaacaagcacagagaca agcctatcagagagcaggccgagaatatcatccacctgttaccctgaccaatctgggagcccctgccgcctcaagtactttgacacc accatcgaccggaagaggtacaccagcaccaaagaggtgctggacgccaccctgatccaccagagcatcaccggcctgtacgaga cacggatcgacctgtctcagctgggaggcgacgcctatccctatgacgtgcccgatatgccagcctgggcagcggctcccccaaga aaaaacgcaaggtggaagatcctaagaaaaagcggaaagtggacggcatggtagtgggagcaacggcagcagcggatccggag ggccgagctctggcgcacccccaccaagtggagggtctcctgccgggtccccaacatctactgaagaaggcaccagcgaatccgc aacgcccgagtcaggccctggtacctccacagaaccatctgaaggtagtgcgcctggtccccagctggaagccctactccaccga agaaggcacgtcaaccgaaccaagtgaaggatctgcccctgggaccagcactgaaccatctgagatggatgctaagtcactaactgc ctggtcccggacactggtgaccttcaaggatgtatttgtggacttcaccagggaggagtggaagctgctggacactgctcagcagatc gtgtacagaaatgtgatgctggagaactataagaacctggtttccttgggtatcagctactaagccagatgtgatcctccggtggaga agggagaagagcccagcggccgcgctactaactcagcctgctgaagcaggctggagacgtggaggagaaccctggacctgtga gcaagggcgaggagctgtcaccggggtggtgcccatcctggtcgagctggacggcgacgtaaacggccacaagtcagcgtgtct ggcgagggcgagggcgatgccacctacggcaagctgaccctgaagtcatctgcaccaccggcaagctgcccgtgccctggccca ccctcgtgaccaccctgacctacggcgtgcagtgcttcagccgctaccccgaccacatgaagcagcacgactcttcaagtccgccat gcccgaaggctacgtccaggagcgcaccatctcttcaaggacgacggcaactacaagacccgcgccgaggtgaagtcgagggc gacaccctggtgaaccgcatcgagctgaagggcatcgactcaaggaggacggcaacatcctggggcacaagctggagtacaact acaacagccacaacgtctatatcatggccgacaagcagaagaacggcatcaaggcgaactcaagatccgccacaacatcgaggacggcagcgtgcagctcgccgaccactaccagcagaacacccccatcggcgacggccccgtgctgctgcccgacaaccactacctga gcacccagtccgccctgagcaaagaccccaacgagaagcgcgatcacatggtcctgctggagttcgtgaccgccgccgggatcact ctcggcatggacgagctgtacaagtaataagcgatcgccctgcaggcatgcaagcttgatatcaagcttatcgatorztaTOccfcTgg attacaciaatttgtgaaagattgactggtattcttaactatgttgctccttttacgctatgtggcitacgctgctttaatgcctttgtatcatg ctattgcttcccgtatggctttccittttctcctccttgtataaatcctggttgctgtctctttatgaggagttgtggcccgttgtcaggcaacg Iggcgtggtgtgcaclglgltlgctgacgcaacccccaclggllggggcattgccaccacctgtcagclcctltccgggacltlcgcttt ccccctccctattgccacggcggaactcatcgccgcctgccttgcccgctgctggacaggggctcggctgttgggcactgacaattc cgtggtgttgtcggggaaatcatcgtcctttccttggctgctcgcctgtgttgccacctggattctgcgcgggacgtccttctgctacgtc ccttcggccctcaatccagcggaccttccttcccgcggcctgctgccggctctgcggcctcttccgcgtcttcgccttcgccctcagac gagteggarcYcccfflgggccgccfecccgcatcgataccgtcgacctcgagggaattaattcgagctcggtacctttaagaccaatg acttacaaggcagctgtagatcttagccactttttaaaagaaaaggggggactggaagggctaatcactcccaacgaag / taa / ta agagggcctatttcccatgattccttcatatttgcatatacgataciuiggctgttagagagataattggaattaatttgactgtaaaca caaagatattagtacaaaatacgtgacgtagaaagtaataatttctgggtagtttgcagttttaaaattatgttttaaaatggactatc atatgctaccgtaacttgaaagtatttcgatttcttggctttatatatcttgtggaaaggacgaaacaccggcaatggaaagtccact gtaaactcgagttacagtggactttccatgttttgaattctcgacctcgagacaaatggcagtattctaatnasLcasigsdctgct tttgcttgtactgggtctctctggttagaccagatctgagcctgggagctctctggctaactagggaacccactgcttaagcctc aataaagctgcctgagtgcttcaagtagtgtgtgcccgtctgttgtgtgactctggtaactagagatccctcagaccctttagt cagtgtggaaaatctctagca

[0322] In some embodiments, the recombinant integration deficient retroviral vector includes, from N-terminus to C-terminus, a 5’ SIN LTR derived from HIV-1, a viral packaging signal, an SFFV promoter, a CRISPRi cassette, a WPRE sequence, and a 3’ SIN LTR derived from HIV-1 that includes a U6 promoter and an MPP8 shRNA. Inclusion of an sgRNA cassette is also contemplated in this exemplary' construct.

[0323] In some embodiments, the recombinant integration deficient retroviral vector comprises the nucleic acid sequence of SEQ ID NO: 803 as shown below.Bold text - HIV-1 LTRItalic text - HIV-1 Psi (packaging signal)Capital text - HIV-1 Rev responsive elementBold italic text - HIV-1 polypurine tract (PPT)Bold underline text - Spleen Focus Forming Virus (SFFV promoter)Underlined text - dCas9-KRAB-P2A-GFP (CRISPRi)Italic underlined text - WPREBold, italic, underlined text- human U6 promoter MPP8 shRNA expression cassetteTggaagggctaattcactcccaaagaagacaagatatcctgatctgtggatctaccacacacaaggctacttccctgatagcagaactacacaccagggccaggggtcagatatccactgacctttggatggtgctacaagctagtaccagttgagccagata aggtagaagaggccaataaaggagagaacaccagcttgttacaccctgtgagcctgcatgggatggatgacccggagagag aagtgttagagtggaggtttgacagccgcctagcatttcatcacgtggcccgagagctgcatccggagtacttcaagaactgct gatatcgagcttgctacaagggacttccgctggggacttccagggaggcgtggcctgggcgggactggggagtggcgagc cctcagatcctgcatataagcagctgctttttgcctgtactgggtctctctggttagaccagatctgagcctgggagctctctggc taactagggaacccactgcttaagcctcaataaagcttgccttgagtgctcaagtagtgtgtgcccgtctgttgtgtgactctgg taactagagatccctcagacccttttagtcagtgtggaaaatctctagcagtggcgcccgaacagggacttgaaagcgaaaggg aaaccagaggagctctctcgacgcaggactcggcttgctgaagcgcgcacggcaagaggcgaggggcggcgactggtgagta cgccaaaaattttgactagcggaggctagaaggagagagatgggtgcgagagcgtcagtataagcgggggagaatagatcgc gatgggaaaaaattcggttaaggccagggggaaagaaaaaatataaattaaaacatatagtatgggcaagcagggagctagaacgat tcgcagttaatcctggcctgtagaaacatcagaaggctgtagacaaatactgggacagctacaaccatcccttcagacaggatcagaa gaacttagatcattatataatacagtagcaaccctctatgtgtgcatcaaaggatagagataaaagacaccaaggaagctttagacaag atagaggaagagcaaaacaaaagtaagaccaccgcacagcaagcggccggccgctgatcttcagacctggaggaggagatatga gggacaattggagaagtgaattatataaatataaagtagtaaaaatgaaccattaggagtagcaccCACCAAGGCAAAG AGAAGAGTGGTGCAGAGAGAAAAAAGAGCAGTGGGAATAGGAGCTTTGTTCCTT GGGTTCTTGGGAGCAGCAGGAAGCACTATGGGCGCAGCGTCAATGACGCTGACG GTACAGGCCAGACAATTATTGTCTGGTATAGTGCAGCAGCAGAACAATTTGCTG AGGGCTATTGAGGCGCAACAGC...

Claims

CLAIMSWHAT IS CLAIMED IS:

1. A recombinant integration deficient retroviral vector comprising a nucleic acid comprising: one or more virus-derived long terminal repeats (LTR); a viral packaging signal; a nucleic acid sequence encoding one or more cargos of interest; a post-transcriptional response element; and a nucleic acid sequence encoding an RNA interference molecule or molecules that targets one or more proteins involved in the epigenetic silencing of viral DNA.

2. The recombinant integration deficient retroviral vector according to claim 1, wherein the one or more proteins involved in the epigenetic silencing of viral DNA is part of the human silencing hub (HUSH) complex, the structural maintenance of chromosome (SMC) 5 / 6 complex, or chromatin modifiers activating transcription factor 7-interacting protein (Atf7ip) and / or its interacting partner SET domain bifurcated histone lysine methyltransferase 1 (Setdbl).

3. The recombinant integration deficient retroviral vector according to claim 1 or claim 2, wherein the one or more proteins involved in the epigenetic silencing of viral DNA is part of the HUSH complex.

4. The recombinant integration deficient retroviral vector according to claim 3, wherein the one or more proteins involved in the epigenetic silencing of viral DNA comprises NP220. MPP8. TASOR. PPHLN1, and / or M0RC2.

5. The recombinant integration deficient retroviral vector according to claim 4, wherein the one or more proteins involved in the epigenetic silencing of viral DNA comprises TASOR.

6. The recombinant integration deficient retroviral vector according to claim 5, wherein the nucleic acid sequence encoding an RNA interference molecule that targets TASOR comprises the nucleic acid sequence of SEQ ID NO: 22.

7. The recombinant integration deficient retroviral vector of any preceding claim, wherein the one or more proteins involved in the epigenetic silencing of viral DNA is part of the SMC5 / 6 complex.

8. The recombinant integration deficient retroviral vector according to claim 7, wherein the one or more proteins involved in the epigenetic silencing of viral DNA comprises SMC5. SMC6. NSMCE1, NSMCE2, NSMCE3, NSMCE4A, and / or SLF2.

9. The recombinant integration deficient retroviral vector of any preceding claim, wherein the one or more proteins involved in the epigenetic silencing of viral DNA is part of the chromatin modifiers activating transcription factor 7-interacting protein (Atf7ip) and / or its interacting partner SET domain bifurcated histone lysine methyltransferase 1 (Setdbl).

10. The recombinant integration deficient retroviral vector of any preceding claim, wherein the RNA interference molecule is selected from a short hairpin RNA (shRNA), a small interfering RNA (siRNA), a hairpin siRNA, a microRNA (miRNA). a precursor miRNA, or an miRNA-adapted shRNA.

11. The recombinant integration deficient retroviral vector of any preceding claim, wherein the RNA interference molecule is positioned within the 3’ LTR.

12. The recombinant integration deficient retroviral vector of any preceding claim, wherein the vector comprises an RNA polymerase III promoter.

13. The recombinant integration deficient retroviral vector of any preceding claim, wherein the vector comprises an RNA polymerase II promoter.

14. The recombinant integration deficient retroviral vector according to claim 13. wherein the RNA polymerase II promoter is an SFFV promoter.

15. The recombinant integration deficient retroviral vector according to claim 14, wherein the SFFV promoter is a modified SFFV promoter.

16. The recombinant integration deficient retroviral vector according to claim 15, wherein the modified SFFV promoter comprises the nucleic acid sequence of SEQ ID NOs: 829-834.

17. The recombinant integration deficient retroviral vector according to claim 1 , wherein the modified SFFV promoter comprises the nucleic acid sequence of SEQ ID NO: 831.

18. The recombinant integration deficient retroviral vector of any preceding claim, wherein the vector comprises an RNA polymerase II and an RNA polymerase III promoter.

19. The recombinant integration deficient retroviral vector of any preceding claim, wherein the vector comprises more than one RNA polymerase II and / or RNA polymerase III promoter.

20. The recombinant integration deficient retroviral vector of any preceding claim, wherein the one or more virus-derived LTRs are derived from murine leukemia virus (MLV), Bovine leukemia virus (BLV), Human T-lymphotropic vims 1 (HTLV-1), Human T- lympho tropic virus 2 (HTLV-2), Gibbon ape leukemia vims (GALV), Feline leukemia virus (FeLV), Porcine endogenous retrovimses (PERV), Feline Foamy Vims (FeFV). Bovine foamy virus (BFV), Mouse mammary tumor virus (MMTV), Jaagsiekte sheep retrovirus (JSRV), Mason-Pfizer monkey vims (MPMV), Avian sarcoma leukosis vims (ALV), Rous sarcoma virus (RSV), Human Endogenous Retrovirus-W (HERV-W), and Human endogenous retrovirus K (HERV-K), Equine infectious anemia virus (EIAV).

21. The recombinant integration deficient retroviral vector of any preceding claim, wherein the vector is derived from murine leukemia virus (MLV), Bovine leukemia vims (BLV). Human T-lymphotropic vims 1 (HTLV-1), Human T-lymphotropic vims 2 (HTLV- 2), Gibbon ape leukemia virus (GALV). Feline leukemia virus (FeLV). Porcine endogenous retrovimses (PERV), Feline Foamy Virus (FeFV), Bovine foamy vims (BFV), Mouse mammary7tumor virus (MMTV), Jaagsiekte sheep retrovirus (JSRV), Mason-Pfizer monkeyvirus (MPMV), Avian sarcoma leukosis virus (ALV), Rous sarcoma virus (RSV), Human Endogenous Retrovirus-W (HERV-W), and Human endogenous retrovirus K (HERV-K), Equine infectious anemia virus (EIAV).

22. The recombinant integration deficient retroviral vector of any one of claims 1 to 21, wherein the vector is a simple retroviral vector.

23. The recombinant integration deficient retroviral vector of any one of claims 1 to 21, wherein the vector is a complex retroviral vector.

24. The recombinant integration deficient retroviral vector of claim 22, wherein the one or more virus-derived LTRs are derived from MLV, GALV, FeLV, PERV, BFV, MMTV, JSRV, MPMV, ALV, or RSV.

25. The recombinant integration deficient retroviral vector of claim 24, wherein the one or more virus-derived LTRs are derived from HIV-1, F1V, HIV-2, S1V, HERV-W. HERV-K, BLV, HTLV-1, HTLV-2 or EIAV26. The recombinant integration deficient retroviral vector of any preceding claim, wherein the one or more virus-derived LTRs are self-inactivating (SIN) LTRs.

27. The recombinant integration deficient retroviral vector of any preceding claim, wherein the vector is derived from HIV.

28. The recombinant integration deficient retroviral vector of any preceding claim, wherein the vector further comprises a nucleic acid sequence encoding HIV Tat and / or Rev.

29. The recombinant integration deficient retroviral vector of any preceding claim, wherein the post-transcriptional response element is selected from the group consisting of Woodchuck Hepatitis Virus Posttranscriptional Regulatory Element (WPRE) and Hepatitis B Virus Posttranscriptional Regulatory' Element (HPRE).

30. The recombinant integration deficient retroviral vector of any preceding claim, wherein the post-transcriptional response element is WPRE.

31. The recombinant integration deficient retroviral vector of claim 30, wherein the WPRE comprises the sequence of SEQ ID NO: 835.

32. The recombinant integration deficient retroviral vector of any preceding claim, wherein the nucleic acid sequence encodes two or more cargos of interest.

33. The recombinant integration deficient retroviral vector of any preceding claim, wherein the nucleic acid sequence comprises coding sequences for two polypeptide cargos of interest and an IRES, protease cleavage or P2A sequence between the coding sequences.

34. The recombinant integration deficient retroviral vector of any preceding claim, wherein the vector further comprises a guide RNA (gRNA).

35. The recombinant integration deficient retroviral vector of any preceding claim, wherein the one or more cargos of interest are selected from the group consisting of geneediting nucleases, endonuclease deficient Cas, endonuclease deficient Cas effectors, endonuclease deficient Cas enzymes, engineered DNA binding proteins, base editors, prime editors, epigenome editors, bridge editors, RNA interference (RNAi), RNA targeting systems, CRISPR DNA binding proteins. CRISPR fusion proteins, CRISPR nickases, recombinases, integrases, programmable integrases, transposons, retrotransposons, DNA polymerases, reverse transcriptases, transposases, chimeric antigen receptors (CAR), T cell receptors, HLA-independent T cell receptors. synNotch receptors, cytokine receptors, synthetic intramembrane proteolysis receptors, transcription factors, caspases, natural proteases, programmable proteases, reporter genes, and selection markers.

36. The recombinant integration deficient retroviral vector according to claim 35, wherein the gene-editing nuclease is selected from the group consisting of zinc finger nucleases (ZFNs), Transcription Activator-Like Effector Nucleases (TALENs), and nucleases of the CRISPR / Cas system.

37. The recombinant integration deficient retroviral vector according to claim 36, wherein the gene-editing nuclease is Cas9.

38. A cell comprising the recombinant integration deficient retroviral vector according to any preceding claim.

39. A composition comprising the recombinant integration deficient retroviral vector of any one of claims 1-37 or the cell of claim 38.

40. A recombinant integration deficient retroviral particle comprising a nucleic acid comprising: one or more virus-derived long terminal repeats (LTR); a viral packaging signal; a nucleic acid sequence encoding one or more cargos of interest; a post-transcriptional response element; and a nucleic acid sequence encoding an RNA interference molecule or molecules that targets one or more proteins involved in the epigenetic silencing of viral DNA.

41. The recombinant integration deficient retroviral particle according to claim 40, wherein the one or more proteins involved in the epigenetic silencing of viral DNA is part of the human silencing hub (HUSH) complex, the structural maintenance of chromosome (SMC) 5 / 6 complex, or chromatin modifiers activating transcription factor 7-interacting protein (Atf7ip) and / or its interacting partner SET domain bifurcated histone lysine methyltransferase 1 (Setdbl).

42. The recombinant integration deficient retroviral particle according to claim 40 or claim 41 , wherein the one or more proteins involved in the epigenetic silencing of viral DNA is part of the HUSH complex.

43. The recombinant integration deficient retroviral particle according to claim 42, wherein the one or more proteins involved in the epigenetic silencing of viral DNA comprises NP220, MPP8, TASOR, PPHLN1, and / or MORC2.

44. The recombinant integration deficient retroviral particle according to claim 43, wherein the one or more proteins involved in the epigenetic silencing of viral DNA comprises TASOR45. The recombinant integration deficient retroviral particle according to claim 44, wherein the RNA interference molecule that targets TASOR comprises the sequence of SEQ ID NO: 22.

46. The recombinant integration deficient retroviral particle according to any one of claims 40-45, wherein the one or more proteins involved in the epigenetic silencing of viral DNA is part of the SMC5 / 6 complex.

47. The recombinant integration deficient retroviral particle according to claim 46, wherein the one or more proteins involved in the epigenetic silencing of viral DNA comprises SMC5. SMC6. NSMCE1, NSMCE2, NSMCE3, NSMCE4A, and / or SLF2.

48. The recombinant integration deficient retroviral particle according to any one of claims 40-41, wherein the one or more proteins involved in the epigenetic silencing of viral DNA is part of the chromatin modifiers activating transcription factor 7-interacting protein (Atf7ip) and / or its interacting partner SET domain bifurcated histone lysine methyltransferase 1 (Setdbl).

49. The recombinant integration deficient retroviral particle according to any one of claims 40-48, wherein the RNA interference molecule is selected from a short hairpin RNA (shRNA), a small interfering RNA (siRNA), a hairpin siRNA, a microRNA (miRNA), a precursor miRNA, or an miRNA-adapted shRNA.

50. The recombinant integration deficient retroviral particle according to any one of claims 40-49, wherein the RNA interference molecule is positioned within the 3?LTR.

51. The recombinant integration deficient retroviral particle according to any one of claims 40-50, wherein the vector comprises an RNA polymerase III promoter.

52. The recombinant integration deficient retroviral particle according to any one of claims 40-51, wherein the vector comprises an RNA polymerase II promoter.

53. The recombinant integration deficient retroviral particle according to claim 52, wherein the RNA polymerase II promoter is an SFFV promoter.

54. The recombinant integration deficient retroviral particle according to claim 53, wherein the SFFV promoter is a modified SFFV promoter.

55. The recombinant integration deficient retroviral particle according to claim 54, wherein the modified SFFV promoter comprises the nucleic acid sequence of SEQ ID NOs: 829-834.

56. The recombinant integration deficient retroviral particle according to claim 55, wherein the modified SFFV promoter comprises the nucleic acid sequence of SEQ ID NO: 831.

57. The recombinant integration deficient retroviral particle according to any one of claims 40-56, wherein the vector comprises an RNA polymerase II and an RNA polymerase III promoter.

58. The recombinant integration deficient retroviral particle according to any one of claims 40-57, wherein the vector comprises more than one RNA polymerase II and / or RNA polymerase III promoter.

59. The recombinant integration deficient retroviral particle according to any one of claims 40-58, wherein the one or more virus-derived LTRs are derived from murine leukemia virus (MLV), Bovine leukemia virus (BLV), Human T-lympho tropic virus 1 (HTLV-1). Human T-lymphotropic virus 2 (HTLV-2). Gibbon ape leukemia virus (GALV), Feline leukemia virus (FeLV), Porcine endogenous retroviruses (PERV), Feline Foamy Virus (FeFV), Bovine foamy virus (BFV), Mouse mammary' tumor virus (MMTV), Jaagsiekte sheep retrovirus (JSRV). Mason-Pfizer monkey virus (MPMV), Avian sarcoma leukosis virus (ALV). Rous sarcoma virus (RSV). Human Endogenous Retrovirus-W (HERV-W), and Human endogenous retrovirus K (HERV-K), Equine infectious anemia virus (El AV).

60. The recombinant integration deficient retroviral particle according to any one of claims 40-59, wherein the vector is derived from murine leukemia virus (MLV), Bovine leukemia virus (BLV). Human T-lymphotropic virus 1 (HTLV-1), Human T-lymphotropic virus 2 (HTLV-2), Gibbon ape leukemia virus (GALV), Feline leukemia virus (FeLV), Porcine endogenous retroviruses (PERV), Feline Foamy Virus (FeFV), Bovine foamy virus(BFV), Mouse mammary' tumor virus (MMTV). Jaagsiekte sheep retrovirus (JSRV), Mason- Pfizer monkey virus (MPMV), Avian sarcoma leukosis virus (ALV), Rous sarcoma vims (RSV), Human Endogenous Retrovirus-W (HERV-W), and Human endogenous retrovirus K (HERV-K), Equine infectious anemia virus (EIAV).

61. The recombinant integration deficient retroviral particle according to any one of claims 40-60, wherein the vector is a simple retroviral vector.

62. The recombinant integration deficient retroviral particle according to any one of claims 40-60, wherein the vector is a complex retroviral vector.

63. The recombinant integration deficient retroviral particle according to claim 61, wherein the one or more virus-derived LTRs are derived from MLV, GALV, FeLV, PERV, BFV, MMTV, JSRV, MPMV, ALV, or RSV.

64. The recombinant integration deficient retroviral particle according to claim 62, wherein the one or more virus-derived LTRs are derived from HIV-L FIV, EIAV, HIV-2, SIV, HERV-W, HERV-K, BLV, HTLV-1, HTLV-2 or EIAV.

65. The recombinant integration deficient retroviral particle according to any one of claims 40-64, wherein the one or more virus-derived LTRs are self-inactivating (SIN) LTRs.

66. The recombinant integration deficient retroviral particle according to any one of claims 40-65, wherein the vector is derived from HIV.

67. The recombinant integration deficient retroviral particle according to any one of claims 40-66, wherein the vector further comprises a nucleic acid sequence encoding HIV Tat and / or Rev.

68. The recombinant integration deficient retroviral particle according to any one of claims 40-67, wherein the post-transcriptional response element is selected from the group consisting of Woodchuck Hepatitis Vims Posttranscriptional Regulatory’ Element (WPRE) and Hepatitis B Vims Posttranscriptional Regulatory' Element (HPRE).

69. The recombinant integration deficient retroviral particle according to any one of claims 40-68, wherein the nucleic acid sequence encodes two or more cargos of interest.

70. The recombinant integration deficient retroviral particle according to any one of claims 40-69. wherein the nucleic acid sequence comprises coding sequences for two polypeptide cargos of interest and an IRES or P2A sequence between the coding sequences.

71. The recombinant integration deficient retroviral particle according to any one of claims 40-70. further comprising a guide RNA (gRNA).

72. The recombinant integration deficient retroviral particle according to any one of claims 40-71, wherein the one or more cargos of interest are selected from the group consisting of gene-editing nucleases, endonuclease deficient Cas, endonuclease deficient Cas effectors, endonuclease deficient Cas enzymes, engineered DNA binding proteins, base editors, epigenome editors, prime editors. RNA interference (RNAi), RNA targeting systems, CRISPR DNA binding proteins, CRISPR fusion proteins, CRISPR nickases, recombinases, integrases, programmable integrases, transposons, retrotransposons, DNA polymerases, reverse transcriptases, transposases, chimeric antigen receptors (CAR), T cell receptors, HLA-independent T cell receptors. synNotch receptors, cytokine receptors, synthetic intramembrane proteolysis receptors, transcription factors, caspases, natural proteases, programmable proteases, reporter genes, and selection markers.

73. The recombinant integration deficient retroviral particle according to claim 72, wherein the gene-editing nuclease is selected from the group consisting of zinc finger nucleases (ZFNs), Transcription Activator-Like Effector Nucleases (TALENs), and nucleases of the CRISPR / Cas system.

74. The recombinant integration deficient retroviral particle according to claim 73, wherein the gene-editing nuclease is Cas9.

75. A cell comprising the recombinant integration deficient retroviral particle according to any one of claims 40-74.

76. A composition comprising the recombinant integration deficient retroviral particle of any one of claims 40-74 or the cell of claim 75.

77. A nucleic acid construct encoding a modified Gag protein comprising a Vpx interaction motif.

78. The nucleic acid construct according to claim 77, wherein the Vpx interaction motif is an STV Vpx interaction motif.

79. The nucleic acid construct according to claims 77 or 78, wherein the Vpx interaction motif is located in the C-terminal tail of the Gag protein.

80. The nucleic acid construct according to claims 77 to 79, wherein the modified Gag protein is a modified HIV-1 Gag protein.

81. The nucleic acid construct according to any one of claims 77 to 80, wherein the Vpx interaction motif comprises the sequence of SEQ ID NO: 839.

82. The nucleic acid construct according to any one of claims 77 to 81, wherein the modified Gag protein comprises the nucleic acid sequence of SEQ ID NO: 841.

83. The nucleic acid construct according to any one of claims 77 to 82, wherein the nucleic acid construct comprises the sequence of SEQ ID NO: 842.

84. A method of reducing the silencing of an integration deficient retroviral vector, the method comprising: administering the recombinant integration deficient retroviral vector of any one of claims 1-37 or the recombinant integration deficient retroviral particle of any one of claims 40-74 to a cell of interest under conditions effective to reduce the silencing of the vector cargo within said cell.

85. A system comprising:(i) a recombinant integration deficient retroviral vector comprising a nucleic acid comprising:one or more virus-derived long terminal repeats (LTR); a viral packaging signal; a nucleic acid sequence encoding one or more cargos of interest; a post-transcriptional response element; and(ii) one or more viral proteins capable of inhibiting the silencing of viral DNA.

86. The system of claim 85, wherein the one or more viral proteins is a heterologous viral protein.

87. The system of claim 85 or claim 86, wherein the one or more viral proteins capable of inhibiting the silencing of viral DNA is selected from Vpr. Vpx. a Vpr-Vpx fusion protein, VP16, ICPO, IE1 , BNRF1 , Hbx, pp71, and RTA.

88. The system of any one of claims 85 to 87, wherein the one or more viral proteins is Vpr.

89. The system of claim 88, wherein the Vpr comprises a mutation at amino acid position 67.

90. The system of claim 89, wherein the mutation is a substitution, deletion, or insertion.

91. The system of claim 90, wherein the substitution is a conservative substitution.

92. The system of claim 91, wherein the substitution is L67E.

93. The system of any one of claims 85 to 87, wherein the one or more viral proteins is Vpx.

94. The system of claim 93, further comprising a nucleic acid construct encoding a modified Gag protein comprising a Vpx interaction motif.

95. The system according to claim 94, wherein the Vpx interaction motif is an SIV Vpx interaction motif.

96. The system according to claims 94 or 95, wherein the Vpx interaction motif is located in the C -terminal tail of the Gag protein.

97. The system according to claims 94 to 96, wherein the modified Gag protein is a modified HIV-1 Gag protein.

98. The system according to any one of claims 94 to 97, wherein the Vpx interaction motif comprises the sequence of SEQ ID NO: 838.

99. The system according to any one of claims 94 to 98, wherein the modified Gag protein comprises the nucleic acid sequence of SEQ ID NO: 841.

100. The system according to any one of claims 94 to 99. wherein the nucleic acid construct comprises the sequence of SEQ ID NO: 842.

101. The system of any one of claims 85 to 87, wherein the one or more viral proteins comprises Vpr and Vpx.

102. The system of claim 101, further comprising a nucleic acid construct encoding a modified Gag protein comprising a Vpx interaction motif.

103. The system of any one of claims 85 to 102, wherein (i) further comprises a nucleic acid sequence encoding an RNA interference molecule or molecules that targets one or more proteins involved in the epigenetic silencing of viral DNA.

104. The system of claim 103. wherein the one or more proteins involved in the epigenetic silencing of viral DNA is part of the human silencing hub (HUSH) complex, the structural maintenance of chromosome (SMC) 5 / 6 complex, or the chromatin modifiers activating transcription factor 7-interacting protein (Atf7ip) and / or its interacting partner SET domain bifurcated histone lysine methyltransferase 1 (Setdbl).

105. The system of claim 104, wherein the one or more proteins involved in the epigenetic silencing of viral DNA is part of the HUSH complex.

106. The system of claim 105. wherein the one or more proteins involved in the epigenetic silencing of viral DNA comprises NP220, MPP8, TASOR, PPHLN1, and / or MORC2.

107. The system of claim 106, wherein the one or more proteins involved in the epigenetic silencing of viral DNA comprises TASOR.

108. The system of claim 107, wherein RNA interference molecule that targets TASOR comprises the sequence of SEQ ID NO: 22.

109. The system of any one of claims 103 to 104, wherein the one or more proteins involved in the epigenetic silencing of viral DNA is part of the SMC5 / 6 complex.

110. The system of claim 109. wherein the one or more proteins involved in the epigenetic silencing of viral DNA comprises SMC5, SMC6, NSMCE1, NSMCE2, NSMCE3, NSMCE4A, and / or SLF2.

111. The system of any one of claims 103 to 110, wherein the one or more proteins involved in the epigenetic silencing of viral DNA is part of the chromatin modifiers activating transcription factor 7-interacting protein (Atf7ip) and / or its interacting partner SET domain bifurcated histone lysine methyltransferase 1 (Setdbl).

112. The system of any one of claims 103 to 111. wherein the RNA interference molecule is selected from a short hairpin RNA (shRNA), a small interfering RNA (siRNA), a hairpin siRNA, a microRNA (miRNA), a precursor miRNA, or an miRNA-adapted shRNA.

113. The system of any one of claims 103 to 112. wherein the RNA interference molecule is positioned within the 3’ LTR.

114. The system of any one of claims 103 to 113, wherein the vector comprises an RNA polymerase III promoter.

115. The system of any one of claims 103 to 114, wherein the vector comprises an RNA polymerase II promoter.

116. The system according to claim 115, wherein the RNA polymerase II promoter is an SFFV promoter.

117. The system according to claim 116, wherein the SFFV promoter is a modified SFFV promoter.

118. The system according to claim 117, wherein the modified SFFV promoter comprises the nucleic acid sequence of SEQ ID NOs: 829-834.

119. The system according to claim 118, wherein the modified SFFV promoter comprises the nucleic acid sequence of SEQ ID NO: 831.

120. The system of any one of claims 103 to 119, wherein the vector comprises an RNA polymerase II and an RNA polymerase III promoter.

121. The system of any one of claims 103 to 120, wherein the vector comprises more than one RNA polymerase II and / or RNA polymerase III promoter.

122. The system of any one of claims 103 to 121, wherein the one or more virus- derived LTRs are derived from murine leukemia virus (MLV), Bovine leukemia virus (BLV), Human T-lymphotropic virus 1 (HTLV-1), Human T-lymphotropic vims 2 (HTLV- 2), Gibbon ape leukemia virus (GALV), Feline leukemia virus (FeLV), Porcine endogenous retroviruses (PERV). Feline Foamy Virus (FeFV), Bovine foamy vims (BFV), Mouse mammary tumor virus (MMTV), Jaagsiekte sheep retrovims (JSRV). Mason-Pfizer monkey virus (MPMV), Avian sarcoma leukosis virus (ALV), Rous sarcoma vims (RSV), Human Endogenous Retrovims-W (HERV-W), and Human endogenous retrovims K (HERV-K), Equine infectious anemia vims (EIAV).

123. The system of any one of claims 103 to 122, wherein the vector is derived from murine leukemia virus (MLV), Bovine leukemia vims (BLV), Human T-lymphotropic virus 1 (HTLV-1), Human T-lymphotropic virus 2 (HTLV-2), Gibbon ape leukemia vims (GALV), Feline leukemia vims (FeLV), Porcine endogenous retrovimses (PERV), Feline Foamy Vims (FeFV), Bovine foamy vims (BFV). Mouse mammary tumor virus (MMTV), Jaagsiekte sheep retrovirus (JSRV), Mason-Pfizer monkey vims (MPMV), Avian sarcoma leukosis virus (ALV), Rous sarcoma vims (RSV), Human Endogenous Retrovims-W (HERV-W), and Human endogenous retrovims K (HERV-K), Equine infectious anemia virus (EIAV).

124. The system of any one of claims 103 to 123, wherein the vector is a simple retroviral vector.

125. The system of any one of claims 103 to 123, wherein the vector is a complex retroviral vector.

126. The system of claim 124, wherein the one or more virus-derived LTRs are derived from MLV, GALV, FeLV, PERV, BFV, MMTV, JSRV, MPMV, ALV, or RSV.

127. The system of claim 125. wherein the one or more virus-derived LTRs are derived from HIV-1 , FIV, EIAV, HIV-2, SIV, HERV-W, HERV-K, BLV, HTLV-1 , HTLV-2 or EIAV.

128. The system of any one of claims 103 to 127, wherein the one or more virus- derived LTRs are self-inactivating (SIN) LTRs.

129. The system of any one of claims 103 to 128, wherein the vector is derived from HIV.

130. The system of any one of claims 103 to 129, wherein the vector further comprises a nucleic acid sequence encoding HIV Tat and / or Rev.

131. The system of any one of claims 103 to 130, wherein the post-transcriptional response element is selected from the group consisting of Woodchuck Hepatitis Virus Posttranscriptional Regulatory Element (WPRE) and Hepatitis B Virus Posttranscriptional Regulatory Element (HPRE).

132. The system according to claim 131, wherein the post-transcriptional response element is WPRE.

133. The system according to claim 132, wherein the WPRE comprises the nucleic acid sequence of SEQ ID NO: 835.

134. The system of any one of claims 103 to 133, wherein the nucleic acid sequence encodes two or more cargos of interest.

135. The system of any one of claims 103 to 134, wherein the nucleic acid sequence comprises coding sequences for two polypeptide cargos of interest and an IRES or P2A sequence between the coding sequences.

136. The system of any one of claims 103 to 135, further comprising a guide RNA (gRNA).

137. The system of any one of claims 103 to 136, wherein the one or more cargos of interest are selected from the group consisting of gene-editing nucleases, endonuclease deficient Cas, endonuclease deficient Cas effectors, endonuclease deficient Cas enzymes, engineered DNA binding proteins, base editors, epigenome editors, prime editors, bridge editors, RNA interference (RNAi), RNA targeting systems, CRISPR DNA binding proteins, CRISPR fusion proteins, CRISPR nickases, recombinases, integrases, programmable integrases, transposons, retrotransposons, DNA polymerases, reverse transcriptases, transposases, chimeric antigen receptors (CAR), T cell receptors, HLA-independent T cell receptors, synNotch receptors, cytokine receptors, synthetic intramembrane proteolysis receptors, transcription factors, caspases, natural proteases, programmable proteases, reporter genes, and selection markers.

138. The system according to claim 137, wherein the gene-editing nuclease is selected from the group consisting of zinc finger nucleases (ZFNs), Transcription Activator- Like Effector Nucleases (TALENs), and nucleases of the CRISPR / Cas system.

139. The system according to claim 138, wherein the gene-editing nuclease is Cas9.

140. A cell comprising the system according to any one of claims 103 to 139.

141. A recombinant integration deficient retroviral particle comprising a nucleic acid comprising:(i) a recombinant integration deficient retroviral vector comprising a nucleic acid comprising: one or more virus-derived long terminal repeats (LTR); a viral packaging signal; a nucleic acid sequence encoding one or more cargos of interest;a post-transcriptional response element; and(ii) one or more viral proteins capable of inhibiting the silencing of viral DNA.

142. A method of reducing the silencing of an integration deficient retroviral vector, the method comprising: administering the system of any one of claims 85-139 to a cell of interest under conditions effective to reduce the silencing of the vector cargo within said cell.

143. A method of increasing the efficiency of the deliver}' of one or more cargos via an integration deficient retroviral vector, the method comprising: administering the system of any one of claims 85-139 to a cell of interest.

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