Epigenetic gene regulation for the treatment of neurological diseases and pain

Abstract

Described herein are compositions and methods for modulating gene expression without permanently editing the genome. The compositions and methods may be useful for treating neurological diseases and pain.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of U.S. Provisional Application No. 63 / 144,408, filed February 1, 2021, the entirety of which is incorporated herein by reference.

[0002] U.S. Government Support This invention was made with Government support under 1R43CA239940 awarded by the Department of Health and Human Services. The Government has certain rights in this invention. [Background technology]

[0003] Chronic pain affects 19%-50% of the world's population, with over 100 million people in the United States alone (Institute of Medicine (US) Committee on Advancing Pain Research, Care, 2011). Despite their side effects and limited effectiveness, opioids have become the preferred treatment for chronic pain among both private and VA prescribers in recent years. However, opioids are highly addictive, and more than 130 Americans die from overdoses every day. Thus, opioid overdose represents a threat with significant public health implications. Although chronic pain is more common than cancer, diabetes, and cardiovascular disease combined, drug development for chronic pain has not undergone the remarkable advances seen in these other therapeutic areas. Despite decades of research, broad-acting, long-acting, non-addictive, and effective treatments for chronic pain remain elusive. Summary of the Invention [Means for solving the problem]

[0004] Epigenetic modulation of gene(s) associated with pain can provide a non-permanent, long-term therapy for pain. In one aspect, epigenetic modulation includes delivery of a nuclease null Cas protein with one or more guide RNAs and repression and / or activation domains to repress and / or activate the target gene(s). Similarly, zinc finger proteins (ZFPs) can be delivered with repressor and / or activation domains to repress and / or activate the target gene(s).

[0005] In one aspect, provided herein is a nucleic acid comprising a sequence encoding a nucleic acid binding domain, a sequence encoding an epigenetic regulator that regulates transcription of one or more target molecules, and a promoter comprising a Nav1.7 promoter, a promoter having a sequence at least about 90% identical to a sequence in Table 5, or a promoter of a gene selected from Tables 1-3. In some embodiments, the one or more target molecules comprise a nucleic acid encoding an ion channel and / or are associated with an ion channel. In some embodiments, the one or more target molecules comprise SCN9A, SCN10A, or SCN11A, or a combination thereof. In some embodiments, the one or more target molecules are associated with Dravet syndrome, epilepsy syndrome, familial hemiplegic migraine, Otahara syndrome, West syndrome, Lennox-Gastaut syndrome, sodium channel myotonia, autism, long QT syndrome, Brugada syndrome, or progressive cardiac conduction disease (also known as LeNegre's disease), or a combination thereof. In some embodiments, the one or more target molecules comprise one or more genes in Table 1. In some embodiments, the one or more target molecules are associated with pain. In some embodiments, the one or more target molecules comprise one or more genes in Table 2 or Table 3. In some embodiments, the nucleic acid binding domain comprises a zinc finger protein. In some embodiments, the nucleic acid binding domain comprises a clustered regularly interspaced short palindromic repeats associated protein (dCas) that does not have nuclease activity. In some embodiments, the epigenetic regulator comprises a domain with transcriptional repression activity (repressor domain). In some embodiments, the repressor domain comprises a ZIM3 repressor domain or a Krueppel-associated box (KRAB) domain (recruiting histone methyltransferases and deacetylases). In some embodiments, the epigenetic regulator comprises a domain with transcriptional activation activity (activator domain).In some embodiments, the epigenetic regulator is ZIM3, KRAB (also referred to as KOX), SID, MBD2, MBD3, HP1a, DNMT family (including DNMT1, DNMT3A, DNMT3B, DNMT3L, DNMT2A), Sin3a, Rb, MeCP2, ROM2, AtHD2A, LSD1, SUV39H1, G9a (EHMT2), ZIM3, ZNF554, ZNF264, ZNF324, ZNF354A, ZNF189, ZNF543, ZNP82, ZNF669, ZNF582, KOX1-MeCP2, ZNF30, ZNF680, ZNF331, ZNF33A, ZNF 528, ZNF320, ZNF350, ZNF175, ZNF214, ZNF184, ZNF8, ZNF596, KOX1, ZNF37A, ZNF394, ZNF610, ZNF273, ZNF34, ZNF250, ZNF98, ZNF675, ZNF213, NLuc, ZFP28-2, ZNF224, ZNF257, VP64, Rta, P16, P65, p300, TET1 catalytic domain, TDG, Ldb1 self-association domain, SAM activator (VP64, p65, HSF1), VPR (VP64, p65, Rta), CD, or SunTag, or variants or combinations thereof. In some embodiments, the promoter comprises a Nav1.7 promoter. In some embodiments, the promoter comprises a sequence in Table 5, optionally at least about 90% identical to promoter 1 or promoter 2. In some embodiments, the promoter is a promoter of a gene selected from Tables 1-3. In some embodiments, the nucleic acid is delivered to the subject via a delivery vehicle. In some embodiments, the delivery vehicle is a viral delivery vehicle (e.g., a retroviral vector, a lentiviral vector, or an adenoviral vector), a liposome, a nanoparticle, or an exosome. In some embodiments, the viral delivery vehicle is a recombinant adeno-associated virus (AAV). In some embodiments, the AAV is AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAVhu68, AAVrh.10, AAVrh74, or AAVDJ, or a combination thereof.In some embodiments, the AAV is AAV9. In some embodiments, the AAV has a recombinant capsid. In some embodiments, the recombinant capsid includes a targeting moiety or a sequence encoding the same. In some embodiments, the delivery vehicle includes or is linked to a targeting moiety. In some embodiments, the targeting moiety targets a cell that includes a target molecule in a subject. In some embodiments, the targeting moiety binds to a peptide product of the target molecule. In some embodiments, the target molecule is present on a target cell. In some embodiments, the target cell is associated with a disease or condition of the subject. In some embodiments, the targeting moiety includes a peptide. In some embodiments, the peptide includes JNJ63955, m3-Huwentoxin-IV, Phlotoxin1 (PhlTx1), Protoxin-II (ProTx-II), or Ceratotoxin-1 (CcoTx1), or variants thereof. In some embodiments, the peptide includes a sequence that is at least about 90% identical to a peptide in Table 7.

[0006] In one aspect, provided herein is a composition comprising: a) a nucleic acid comprising a sequence encoding a nucleic acid binding domain and a sequence encoding an epigenetic regulator that regulates the transcription of one or more target molecules; and b) a delivery vehicle for delivering the nucleic acid, the delivery vehicle comprising JNJ63955, m3-Huwentoxin-IV, Phlotoxin1 (PhlTx1), Protoxin-II (ProTx-II), or Ceratotoxin-1 (CcoTx1), or a variant thereof, or a target peptide comprising a sequence at least about 90% identical to a peptide in Table 7. In some embodiments, the delivery vehicle is a viral delivery vehicle (e.g., a retroviral vector, a lentiviral vector, or an adenoviral vector), a liposome, a nanoparticle, or an exosome. In some embodiments, the viral delivery vehicle is a recombinant adeno-associated virus (AAV). In some embodiments, the AAV is AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAVhu68, AAVrh.10, AAVrh74, or AAVDJ, or a combination thereof. In some embodiments, the AAV is AAV9. In some embodiments, the AAV has a recombinant capsid. In some embodiments, the recombinant capsid includes a targeting moiety or includes a sequence encoding the same. In some embodiments, the delivery vehicle includes or is linked to a targeting moiety. In some embodiments, the targeting moiety targets a cell that includes a target molecule in a subject. In some embodiments, the targeting moiety binds to a peptide product of the target molecule. In some embodiments, the target molecule is present on a target cell. In some embodiments, the target cell is associated with a disease or condition of the subject. In some embodiments, the targeting moiety includes a peptide. In some embodiments, the peptide comprises JNJ63955, m3-Huwentoxin-IV, Phlotoxin1 (PhlTx1), Protoxin-II (ProTx-II), or Ceratotoxin-1 (CcoTx1), or a variant thereof.In some embodiments, the peptide comprises a sequence that is at least about 90% identical to a peptide in Table 7.

[0007] In one aspect, provided herein is a method for modulating expression of one or more target molecules associated with a disease or condition in a subject, the method comprising administering to the subject a nucleic acid sequence encoding a nucleic acid binding domain and an epigenetic regulator that modulates transcription of the one or more target molecules. In some embodiments, the modulation of transcription of the one or more target molecules is transient. In some embodiments, the subject's genome is not edited. In some embodiments, the disease or condition comprises pain. In some embodiments, the pain comprises neuropathic pain, inflammatory pain (e.g., rheumatoid arthritis), visceral pain, migraine, erythromycin pain, fibromyalgia pain, idiopathic pain, or somatic pain, or a combination thereof. In some embodiments, the disease or condition comprises a channelopathy. In some embodiments, the channelopathy comprises Dravet syndrome, epilepsy syndrome, familial hemiplegic migraine, Otahara syndrome, West syndrome, Lennox-Gastaut syndrome, sodium channelotonopathy, autism, long QT syndrome, Brugada syndrome, or progressive cardiac conduction disease (also called Lenegre's disease), or a combination thereof. In some embodiments, the disease or condition comprises a neurological disease. In some embodiments, the neurological disease is dementia. In some embodiments, the disease or condition comprises Alzheimer's disease. In some embodiments, the disease or condition comprises Parkinson's disease. In some embodiments, the disease or condition comprises Huntington's disease. In some embodiments, the disease or condition comprises schizophrenia. In some embodiments, the disease or condition comprises amyotrophic lateral sclerosis (ALS). In some embodiments, the disease or condition comprises multiple sclerosis. In some embodiments, the disease or condition comprises a disease of the central nervous system. In some embodiments, the disease or condition is an infectious disease. In some embodiments, the disease or condition comprises β-thalassemia, fragile X syndrome, centronuclear myopathy, prion disease, Angelman syndrome, Lafora disease, or Alexander disease.

[0008] In some embodiments, the one or more target molecules comprise DNA. In some embodiments, the one or more target molecules comprise RNA. In some embodiments, the one or more target molecules comprise a coding region of a gene. In some embodiments, the one or more target molecules comprise DNA complementary to a non-coding RNA. In some embodiments, the non-coding RNA is associated with neuropathic pain. In some embodiments, the neuropathic pain comprises spinal nerve ligation injury, nerve branch ligation injury, chronic constriction injury, or diabetic neuropathy, or a combination thereof. In some embodiments, the non-coding RNA comprises SCN9A natural antisense transcript (NAT), Kcna2 antisense RNA, H19, Gm21781, MRAK009713, uc.48+, NONRATT021972, BC168687, Speer7-ps, Uc007pbc.1, XLOC_041439, Mlxipl, Rn50_X_0739.1, CCAT1, rno circ0004058, rno_circRNA_007512, or Egr2 antisense RNA, or a combination thereof. In some embodiments, the one or more target molecules comprise a nucleic acid encoding a channel. In some embodiments, the one or more target molecules comprise a nucleic acid associated with a channel. In some embodiments, the channel is an ion channel. In some embodiments, the ion channel is a sodium channel. In some embodiments, the ion channel is a potassium channel. In some embodiments, the ion channel is a calcium channel. In some embodiments, the ion channel is a chloride channel. In some embodiments, the one or more target molecules are associated with Dravet syndrome, epilepsy syndrome, familial hemiplegic migraine, Otahara syndrome, West syndrome, Lennox-Gastaut syndrome, sodium channel myotonia, autism, long QT syndrome, Brugada syndrome, or progressive cardiac conduction disease (also called Lenegre's disease), or a combination thereof. In some embodiments, the one or more target molecules comprise one or more genes in Table 1. In some embodiments, the one or more target molecules comprise SCN9A, SCN10A, or SCN11A, or a combination thereof.In some embodiments, the one or more target molecules comprise a native antisense transcript of SCN9A. In some embodiments, the one or more target molecules are associated with pain. In some embodiments, the pain comprises neuropathic pain, inflammatory pain (e.g., rheumatoid arthritis), visceral pain, migraine, erythromycin pain, fibromyalgia pain, idiopathic pain, or somatic pain, or a combination thereof. In some embodiments, the one or more target molecules comprise one or more genes in Table 2. In some embodiments, the one or more target molecules are associated with a neurological disease. In some embodiments, the neurological disease comprises dementia. In some embodiments, the one or more target molecules are associated with Alzheimer's disease. In some embodiments, the one or more target molecules comprise one or more genes in Table 3. In some embodiments, the one or more target molecules are associated with Parkinson's disease. In some embodiments, the one or more target molecules comprise one or more genes selected from the group comprising SNCA, GBA, and LRRK2. In some embodiments, the one or more target molecules are associated with Huntington's disease. In some embodiments, the one or more target molecules are associated with schizophrenia. In some embodiments, the one or more target molecules include GPR52. In some embodiments, the one or more target molecules are associated with amyotrophic lateral sclerosis (ALS). In some embodiments, the one or more target molecules include one or more genes selected from the group including SOD1, ataxin-2, TDP43, FUS, C9ORF72, and SCA2. In some embodiments, the one or more target molecules are associated with multiple sclerosis. In some embodiments, the one or more target molecules are associated with a disease of the central nervous system. In some embodiments, the one or more target molecules include one or more genes selected from the group including BFD1, FUS, C9orf72, brain-derived neurotrophic factor, nerve growth factor, neurotrophin, BCL11A, FMR1, DNM2, PrP, UBE3A, GYS1, STING, and GFAP.

[0009] In some embodiments, the nucleic acid binding domain binds to at least one of the one or more target molecules. In some embodiments, the nucleic acid binding domain comprises a clustered regularly interspaced short palindromic repeats associated protein (dCas) that does not have nuclease activity. In some embodiments, the dCas comprises a mutant Cas protein. In some embodiments, the dCas is a truncated Cas protein. In some embodiments, the dCas is a mutant or truncated Cas protein of Table 4. In some embodiments, the dCas comprises a sequence that is at least 90% identical to a dCas of Table 4. In some embodiments, the dCas comprises dCas9. In some embodiments, the dCas9 is a truncated or mutant Cas9 protein. In some embodiments, the dCas comprises dCas12. In some embodiments, the dCas12 is a truncated or mutant Cas12 protein. In some embodiments, the dCas comprises a dCas9 from Streptococcus pyogenes, Staphylococcus aureus, Campylobacter jejuni, S. thermophilus, S. pneumoniae, Neisseria meningitidis, Corynebacter diphtheriae, Eubacterium ventriosum, Streptococcus pasteurianus, Lactobacillus farciminis, Sphaerochaeta globus, Azospirillum B510, Gluconacetobacter diazotrophicus, Neisseria cinerea, Roseburia intestinalis, Parvibaculum lavamentivorans, Nitratifractor salsuginis DSM 16511, Campylobacter lari CF89-12, or Streptococcus thermophilus LMD-9. In some embodiments, the dCas has a REC2 domain deletion. In some embodiments, the dCas has a REC3 domain deletion.In some embodiments, the dCas has an HNH deletion. In some embodiments, the dCas has a nuclease (NUC) lobe deletion. In some embodiments, the nucleic acid binding domain comprises a zinc finger protein. In some embodiments, the nucleic acid binding domain comprises a meganuclease. In some embodiments, the nucleic acid binding domain comprises a transcription activator-like effector nuclease (TALEN).

[0010] In some embodiments, the epigenetic regulator comprises a transcriptional regulatory domain. In some embodiments, the epigenetic regulator comprises a domain with transcriptional repression activity (repressor domain). In some embodiments, the repressor domain comprises a Krueppel-associated box (KRAB) domain (recruiting histone methyltransferases and deacetylases). In some embodiments, the epigenetic regulator comprises a domain with transcriptional activation activity (activator domain). In some embodiments, the epigenetic regulator comprises KRAB (also called KOX), SID, MBD2, MBD3, HP1a, DNMT family (including DNMT1, DNMT3A, DNMT3B, DNMT3L, DNMT2A), Sin3a, Rb, MeCP2, ROM2, AtHD2A, LSD1, SUV39H1, or G9a (EHMT2), or variants or combinations thereof. In some embodiments, the epigenetic regulator comprises ZIM3, ZNF554, ZNF264, ZNF324, ZNF354A, ZNF189, ZNF543, ZNP82, ZNF669, ZNF582, KOX1-MeCP2, ZNF30, ZNF680, ZNF331, ZNF33A, ZNF528, ZNF320, ZNF350, ZNF175, ZNF214, ZNF184, ZNF8, ZNF596, KOX1, ZNF37A, ZNF394, ZNF610, ZNF273, ZNF34, ZNF250, ZNF98, ZNF675, ZNF213, NLuc, ZFP28-2, ZNF224, or ZNF257, or a variant or combination thereof. In some embodiments, the epigenetic regulator comprises VP64, Rta, P16, P65, p300, TET1 catalytic domain, TDG, Ldb1 self-association domain, SAM activator (VP64, p65, HSF1), VPR (VP64, p65, Rta), CD, or SunTag, or a variant or combination thereof.In some embodiments, the epigenetic regulator comprises a domain that recruits a transcriptional activator, a histone acetyltransferase, a DNA demethylase, a domain that recruits enhancer-associated endogenous Ldb1, a domain that recruits histone methyltransferases and deacetylases, a domain that recruits histone deacetylases, a histone demethylase, a histone methyltransferase, a DNA methyltransferase, an acetylation domain, or a deacetylation domain, or a combination thereof. In some embodiments, the epigenetic regulator comprises VP64 (recruiting transcription activators), p65 (recruiting transcription activators), p300 catalytic domain (histone acetyltransferase), TET1 catalytic domain (DNA demethylase), TDG (DNA demethylase), Ldb1 self-association domain (recruiting enhancer-associated endogenous Ldb1), SAM activator (VP64, p65, HSF1) (recruits transcription activators), VPR (VP64, p65, Rta) (recruits transcription activators), Sin3a (recruiting histone deacetylase), LSD1 (histone demethylase), SUV39H1 (histone methyltransferase), G9a (EHMT2) (histone methyltransferase), DNMT3a (DNA methyltransferase), or DNMT3a-DNMT3L (DNA methyltransferase), or a combination thereof. In some embodiments, the epigenetic regulator is linked to the nucleic acid binding domain at the N-terminus or C-terminus of the nucleic acid binding domain via a linker. In some embodiments, the linker is a flexible linker.In some embodiments the linker is selected from the group consisting of (GGS)n, (GGGS)n, (GGGGS)n, (G)n, (EAAAK)n, A(EAAAK)nALEA(EAAAK)nA, PAPAP, AEAAAKEAAAKA, (Ala-Pro)x, LE, GlySer-polyPro(Glyc)-polyPro(Glyc)-polyPro(Glyc), GlySer-polyPro-polyPro(Glyc)-polyPro, GlySer-polyP ro-GlySer(Glyc)-polyPro, GlySer-polyPro-polyPro-polyPro, GlySer-polyPro-β2m-polyPro, GlySer-polyPro-β2m-GlyS er, polyPro-β2m-GlySer-β2m-GlySer, GlySer-polyPro-β2m-GlySer-β2m-polyPro, GlySer-polyPro-Ub-GlySer, GlySer-pol and (G4S)n, wherein n is independently selected from 1 to 10 and x is 10 to 3. 4, polyPro is the proline-rich hinge sequence from IgA1, polyPro(Glyc) is the proline-rich hinge sequence from IgA1 with an embedded potential N-linked glycosylation site (Asn-Ser-Ser), β2m is β2-microglubulin, Ub is ubiquitin, ZAG is Zn-α2-glycoprotein, and cTPRX is the consensus tetratricopeptide repeat sequence with a repeat number of X. In some embodiments, the nucleic acid binding domain and the epigenetic regulator are linked by a disulfide bond.

[0011] In some embodiments, the nucleic acid sequence comprises a nuclear localization sequence (NLS). In some embodiments, the nucleic acid sequence comprises a promoter. In some embodiments, the promoter comprises a sequence that is at least about 90% identical to a sequence in Table 5. In some embodiments, the promoter comprises a nuclear localization sequence. In some embodiments, the NLS drives nuclear import of the nucleic acid. In some embodiments, the promoter drives expression of a nucleic acid binding domain. In some embodiments, the promoter drives expression of an epigenetic regulator. In some embodiments, the promoter is a promoter naturally associated with the target molecule. In some embodiments, the promoter is a promoter of a gene from Tables 1-3. In some embodiments, the promoter is an SCN9A promoter or an SCN10A promoter. In some embodiments, the promoter is a pan-neuronal gene promoter. In some embodiments, the promoter is a promoter naturally associated with a gene associated with pain. In some embodiments, the promoter is a promoter of microtubule-associated protein 2 (MAP-2), a promoter of neuron-specific enolase (NSE), a promoter of choline acetyltransferase (ChAT), a promoter of protein gene product 9.5 (PGP9.5) (also called ubiquitin C-terminal hydrolase 1 (UCHL-1)), a gene promoter of human synapsin 1 (hSYN1), a promoter of the NeuN gene (Fox-3, Rbfox3, or hexaribonucleotide-binding protein-3), a promoter of α-calcium / calmodulin-dependent protein kinase II [CaMKIIα], a promoter of the Rheb gene (ras homolog enriched in brain), a TRKA promoter (tyrosine kinase A), or jET. In some embodiments, the promoter is a promoter naturally associated with a gene associated with a channel.In some embodiments, the promoter is a promoter naturally associated with Dravet syndrome, epilepsy syndrome, familial hemiplegic migraine, Ohtahara syndrome, West syndrome, Lennox-Gastaut syndrome, sodium channel myotonia, autism, long QT syndrome, Brugada syndrome, or progressive cardiac conduction disease (also known as Lenegre's disease), or a combination thereof. In some embodiments, the promoter is a promoter naturally associated with inflammatory pain, visceral pain, migraine, erythromycin pain, fibromyalgia pain, idiopathic pain, or somatic pain, or a combination thereof. In some embodiments, the promoter is a promoter naturally associated with a neurological disease. In some embodiments, the promoter is a promoter naturally associated with dementia. In some embodiments, the promoter is a promoter naturally associated with Alzheimer's disease. In some embodiments, the promoter is a promoter naturally associated with Parkinson's disease. In some embodiments, the promoter is a promoter naturally associated with ALS. In some embodiments, the promoter is a promoter naturally associated with multiple sclerosis. In some embodiments, the promoter is a promoter naturally associated with a gene associated with a central nervous system disease. In some embodiments, the promoter is a promoter of a gene selected from SNCA, GBA, and LRRK2, SOD1, ataxin-2, SCA2, BFD1, FUS, C9orf72, brain-derived neurotrophic factor, nerve growth factor, neurotrophin, BCL11A, FMR1, DNM2, PrP, UBE3A, GYS1, and GFAP. In some embodiments, the promoter is a pol II promoter (e.g., Thy1 and H1xb9), a small latency associated promoter (e.g., from the herpesvirus pseudorabies virus), a cytomegalovirus promoter, SV40, elongation factor 1-alpha (EF1a) promoter, a cytomegalovirus enhancer / chicken-beta actin (CAG) promoter, or a herpes simplex virus (HSV) promoter.In some embodiments, the promoter is controlled by a small molecule. In some embodiments, the promoter is a tetracycline-responsive promoter, a glucocorticoid-responsive promoter, a RU-486-responsive promoter, a peroxide-inducible promoter, or a tamoxifen-inducible promoter. In some embodiments, expression of the epigenetic regulator and / or the transcriptional regulatory domain occurs upon natural or physiological induction of the promoter. In some embodiments, the promoter is induced upon a pathology in the subject. In some embodiments, the pathology is injury and / or inflammation. In some embodiments, the promoter is a galanin promoter or an NF-κB promoter. In some embodiments, the nucleic acid sequence comprises two or more promoters. In some embodiments, the nucleic acid sequence comprises a tandem promoter.

[0012] In some embodiments, the nucleic acid sequence comprises an enhancer. In some embodiments, the nucleic acid sequence comprises an intron. In some embodiments, the nucleic acid comprises an inverted terminal repeat (ITR). In some embodiments, the nucleic acid comprises a terminator sequence.

[0013] In some embodiments, the method comprises administering one or more guide RNA sequences to the subject. In some embodiments, the one or more guide RNA sequences are selected from the sequences in Table 6. In some embodiments, the one or more guide RNA sequences bind to one or more of the target molecules. In some embodiments, the one or more target molecules are 2, 3, 4, or 5 target molecules. In some embodiments, the one or more guide RNA sequences are 2, 3, 4, or 5 guide RNA sequences. In some embodiments, the method comprises administering at least two guide RNA sequences, and at least two guide RNA sequences are different. In some embodiments, the at least one or more guide RNA sequences target the promoter described herein, creating a regulatory feedback loop to control expression levels.

[0014] In some embodiments, the nucleic acid is delivered as a naked (or unmodified) nucleic acid. In some embodiments, the nucleic acid is delivered in a complex with a cationic molecule. In some embodiments, the nucleic acid is delivered to the subject via a vehicle. In some embodiments, the vehicle is a viral delivery vehicle (e.g., a retroviral vector, a lentiviral vector, or an adenoviral vector), a liposome, a nanoparticle, or an exosome. In some embodiments, the vehicle is a viral delivery vehicle. In some embodiments, the viral delivery vehicle is a retroviral vector, a lentiviral vector, or an adenoviral vector. In some embodiments, the viral delivery vehicle is a recombinant adeno-associated virus (AAV). In some embodiments, the AAV is AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAVhu68, AAVrh.10, AAVrh74, or AAVDJ, or a combination thereof. In some embodiments, the AAV is AAV9. In some embodiments, the AAV has a recombinant capsid. In some embodiments, the recombinant capsid encodes a targeting moiety. In some embodiments, the nucleic acid comprises a sequence encoding the targeting moiety. In some embodiments, the vehicle is a liposome, lipid nanoparticle, nanocapsule, or exosome. In some embodiments, the vehicle comprises or is linked to a targeting moiety. In some embodiments, the targeting moiety targets a cell comprising a target molecule in the subject. In some embodiments, the targeting moiety binds to a peptide product of the target molecule. In some embodiments, the target molecule is present on a target cell. In some embodiments, the target cell is associated with a disease or condition of the subject. In some embodiments, the targeting moiety comprises a peptide. In some embodiments, the peptide comprises JNJ63955, m3-Huwentoxin-IV, Phlotoxin1, Protoxin-II (ProTx-II), Huwentoxin IV (HwTx-IV), or Ceratotoxin-1 (CcoTx1), or variants thereof.In some embodiments, the peptide comprises a sequence that is at least about 90% identical to a peptide in Table 7.

[0015] In some embodiments, the method further comprises administering to the subject an additional therapeutic agent.

[0016] In some embodiments, the nucleic acid is administered via lumbar intrathecal puncture, epidural, intravenous, transdermal, intranasal, oral, mucosal, intracisternal, or intraganglionic administration.

[0017] In another aspect, a nucleic acid sequence is further provided that encodes a nucleic acid binding domain and an epigenetic regulator that regulates the transcription of one or more target molecules. In some embodiments, the regulation of the transcription of one or more target molecules is transient. In some embodiments, the subject's genome is not edited. In some embodiments, the one or more target molecules comprise DNA. In some embodiments, the one or more target molecules comprise RNA. In some embodiments, the one or more target molecules comprise a coding region of a gene. In some embodiments, the one or more target molecules comprise DNA complementary to a non-coding RNA. In some embodiments, the non-coding RNA is associated with neuropathic pain. In some embodiments, the neuropathic pain comprises spinal nerve ligation injury, nerve branch ligation injury, chronic constriction injury, or diabetic neuropathy, or a combination thereof. In some embodiments, the non-coding RNA comprises SCN9A natural antisense transcript (NAT), Kcna2 antisense RNA, H19, Gm21781, MRAK009713, uc.48+, NONRATT021972, BC168687, Speer7-ps, Uc007pbc.1, XLOC_041439, Mlxipl, Rn50_X_0739.1, CCAT1, rno circ0004058, rno_circRNA_007512, or Egr2 antisense RNA, or a combination thereof. In some embodiments, the one or more target molecules comprise a nucleic acid encoding a channel. In some embodiments, the one or more target molecules comprise a nucleic acid associated with a channel. In some embodiments, the channel is an ion channel. In some embodiments, the ion channel is a sodium channel. In some embodiments, the ion channel is a potassium channel. In some embodiments, the ion channel is a calcium channel. In some embodiments, the ion channel is a chloride channel.In some embodiments, the one or more target molecules are associated with Dravet syndrome, epilepsy syndrome, familial hemiplegic migraine, Otahara syndrome, West syndrome, Lennox-Gastaut syndrome, sodium channel myotonia, autism, long QT syndrome, Brugada syndrome, or progressive cardiac conduction disease (also called Lenegre's disease), or a combination thereof. In some embodiments, the one or more target molecules comprise one or more genes in Table 1. In some embodiments, the one or more target molecules comprise SCN9A, SCN10A, or SCN11A, or a combination thereof. In some embodiments, the one or more target molecules comprise a native antisense transcript of SCN9A. In some embodiments, the one or more target molecules are associated with pain. In some embodiments, the pain comprises neuropathic pain, inflammatory pain, visceral pain, migraine, erythromycin pain, fibromyalgia pain, idiopathic pain, or somatic pain, or a combination thereof. In some embodiments, the one or more target molecules comprise one or more genes in Table 2. In some embodiments, the one or more target molecules are associated with a neurological disease. In some embodiments, the neurological disease comprises dementia. In some embodiments, the one or more target molecules are associated with Alzheimer's disease. In some embodiments, the one or more target molecules comprise one or more genes in Table 3. In some embodiments, the one or more target molecules are associated with Parkinson's disease. In some embodiments, the one or more target molecules comprise one or more genes selected from the group comprising SNCA, GBA, and LRRK2. In some embodiments, the one or more target molecules are associated with Huntington's disease. In some embodiments, the one or more target molecules are associated with schizophrenia. In some embodiments, the one or more target molecules comprise GPR52. In some embodiments, the one or more target molecules are associated with amyotrophic lateral sclerosis (ALS). In some embodiments, the one or more target molecules comprise one or more genes selected from the group comprising SOD1, ataxin-2, TDP43, FUS, C9ORF72, and SCA2. In some embodiments, the one or more target molecules are associated with multiple sclerosis.In some embodiments, the one or more target molecules are associated with a disease of the central nervous system. In some embodiments, the one or more target molecules include one or more genes selected from the group including BFD1, FUS, C9orf72, brain-derived neurotrophic factor, nerve growth factor, neurotrophin, BCL11A, FMR1, DNM2, PrP, UBE3A, GYS1, STING, and GFAP.

[0018] In some embodiments, the nucleic acid binding domain binds to at least one of the one or more target molecules. In some embodiments, the nucleic acid binding domain comprises a clustered regularly interspaced short palindromic repeats associated protein (dCas) that does not have nuclease activity. In some embodiments, the dCas comprises a mutant Cas protein. In some embodiments, the dCas is a truncated Cas protein. In some embodiments, the dCas is a mutant or truncated Cas protein of Table 4. In some embodiments, the dCas comprises a sequence that is at least 90% identical to a dCas of Table 4. In some embodiments, the dCas comprises dCas9. In some embodiments, the dCas9 is a truncated or mutant Cas9 protein. In some embodiments, the dCas comprises dCas12. In some embodiments, the dCas12 is a truncated or mutant Cas12 protein. In some embodiments, the dCas comprises a dCas9 from Streptococcus pyogenes, Staphylococcus aureus, Campylobacter jejuni, S. thermophilus, S. pneumoniae, Neisseria meningitidis, Corynebacter diphtheriae, Eubacterium ventriosum, Streptococcus pasteurianus, Lactobacillus farciminis, Sphaerochaeta globus, Azospirillum B510, Gluconacetobacter diazotrophicus, Neisseria cinerea, Roseburia intestinalis, Parvibaculum lavamentivorans, Nitratifractor salsuginis DSM 16511, Campylobacter lari CF89-12, or Streptococcus thermophilus LMD-9. In some embodiments, the dCas has a REC2 domain deletion. In some embodiments, the dCas has a REC3 domain deletion.In some embodiments, the dCas has an HNH deletion. In some embodiments, the dCas has a nuclease (NUC) lobe deletion. In some embodiments, the nucleic acid binding domain comprises a zinc finger protein. In some embodiments, the nucleic acid binding domain comprises a meganuclease. In some embodiments, the nucleic acid binding domain comprises a transcription activator-like effector nuclease (TALEN).

[0019] In some embodiments, the epigenetic regulator comprises a transcriptional regulatory domain. In some embodiments, the epigenetic regulator comprises a domain with transcriptional repression activity (repressor domain). In some embodiments, the repressor domain comprises a Krueppel-associated box (KRAB) domain (recruiting histone methyltransferases and deacetylases). In some embodiments, the epigenetic regulator comprises a domain with transcriptional activation activity (activator domain). In some embodiments, the epigenetic regulator comprises KRAB (also called KOX), SID, MBD2, MBD3, HP1a, DNMT family (including DNMT1, DNMT3A, DNMT3B, DNMT3L, DNMT2A), Sin3a, Rb, MeCP2, ROM2, AtHD2A, LSD1, SUV39H1, or G9a (EHMT2), or variants or combinations thereof. In some embodiments, the epigenetic regulator comprises ZIM3, ZNF554, ZNF264, ZNF324, ZNF354A, ZNF189, ZNF543, ZNP82, ZNF669, ZNF582, KOX1-MeCP2, ZNF30, ZNF680, ZNF331, ZNF33A, ZNF528, ZNF320, ZNF350, ZNF175, ZNF214, ZNF184, ZNF8, ZNF596, KOX1, ZNF37A, ZNF394, ZNF610, ZNF273, ZNF34, ZNF250, ZNF98, ZNF675, ZNF213, NLuc, ZFP28-2, ZNF224, or ZNF257, or a variant or combination thereof. In some embodiments, the epigenetic regulator comprises VP64, Rta, P16, P65, p300, TET1 catalytic domain, TDG, Ldb1 self-association domain, SAM activator (VP64, p65, HSF1), VPR (VP64, p65, Rta), CD, or SunTag, or a variant or combination thereof.In some embodiments, the epigenetic regulator comprises a domain that recruits a transcriptional activator, a histone acetyltransferase, a DNA demethylase, a domain that recruits enhancer-associated endogenous Ldb1, a domain that recruits histone methyltransferases and deacetylases, a domain that recruits histone deacetylases, a histone demethylase, a histone methyltransferase, a DNA methyltransferase, an acetylation domain, or a deacetylation domain, or a combination thereof. In some embodiments, the epigenetic regulator comprises VP64 (recruiting transcription activators), p65 (recruiting transcription activators), p300 catalytic domain (histone acetyltransferase), TET1 catalytic domain (DNA demethylase), TDG (DNA demethylase), Ldb1 self-association domain (recruiting enhancer-associated endogenous Ldb1), SAM activator (VP64, p65, HSF1) (recruits transcription activators), VPR (VP64, p65, Rta) (recruits transcription activators), Sin3a (recruiting histone deacetylase), LSD1 (histone demethylase), SUV39H1 (histone methyltransferase), G9a (EHMT2) (histone methyltransferase), DNMT3a (DNA methyltransferase), or DNMT3a-DNMT3L (DNA methyltransferase), or a combination thereof. In some embodiments, the epigenetic regulator is linked to the nucleic acid binding domain at the N-terminus or C-terminus of the nucleic acid binding domain via a linker. In some embodiments, the linker is a flexible linker.In some embodiments the linker is selected from the group consisting of (GGS)n, (GGGS)n, (GGGGS)n, (G)n, (EAAAK)n, A(EAAAK)nALEA(EAAAK)nA, PAPAP, AEAAAKEAAAKA, (Ala-Pro)x, LE, GlySer-polyPro(Glyc)-polyPro(Glyc)-polyPro(Glyc), GlySer-polyPro-polyPro(Glyc)-polyPro, GlySer-polyP ro-GlySer(Glyc)-polyPro, GlySer-polyPro-polyPro-polyPro, GlySer-polyPro-β2m-polyPro, GlySer-polyPro-β2m-GlyS er, polyPro-β2m-GlySer-β2m-GlySer, GlySer-polyPro-β2m-GlySer-β2m-polyPro, GlySer-polyPro-Ub-GlySer, GlySer-pol and (G4S)n, wherein n is independently selected from 1 to 10 and x is 10 to 3. 4, polyPro is the proline-rich hinge sequence from IgA1, polyPro(Glyc) is the proline-rich hinge sequence from IgA1 with an embedded potential N-linked glycosylation site (Asn-Ser-Ser), β2m is β2-microglubulin, Ub is ubiquitin, ZAG is Zn-α2-glycoprotein, and cTPRX is the consensus tetratricopeptide repeat sequence with a repeat number of X. In some embodiments, the nucleic acid binding domain and the epigenetic regulator are linked by a disulfide bond.

[0020] In some embodiments, the nucleic acid sequence comprises a nuclear localization sequence (NLS). In some embodiments, the nucleic acid sequence comprises a promoter. In some embodiments, the promoter comprises a sequence that is at least about 90% identical to a sequence in Table 5. In some embodiments, the promoter comprises a nuclear localization sequence. In some embodiments, the NLS drives nuclear import of the nucleic acid. In some embodiments, the promoter drives expression of a nucleic acid binding domain. In some embodiments, the promoter drives expression of an epigenetic regulator. In some embodiments, the promoter is a promoter naturally associated with the target molecule. In some embodiments, the promoter is a promoter of a gene from Tables 1-3. In some embodiments, the promoter is an SCN9A promoter or an SCN10A promoter. In some embodiments, the promoter is a pan-neuronal gene promoter. In some embodiments, the promoter is a promoter naturally associated with a gene associated with pain. In some embodiments, the promoter is a promoter of microtubule-associated protein 2 (MAP-2), a promoter of neuron-specific enolase (NSE), a promoter of choline acetyltransferase (ChAT), a promoter of protein gene product 9.5 (PGP9.5) (also called ubiquitin C-terminal hydrolase 1 (UCHL-1)), a gene promoter of human synapsin 1 (hSYN1), a promoter of the NeuN gene (Fox-3, Rbfox3, or hexaribonucleotide-binding protein-3), a promoter of α-calcium / calmodulin-dependent protein kinase II [CaMKIIα], a promoter of the Rheb gene (ras homolog enriched in brain), a TRKA promoter (tyrosine kinase A), or jET. In some embodiments, the promoter is a promoter naturally associated with a gene associated with a channel.In some embodiments, the promoter is a promoter naturally associated with Dravet syndrome, epilepsy syndrome, familial hemiplegic migraine, Ohtahara syndrome, West syndrome, Lennox-Gastaut syndrome, sodium channel myotonia, autism, long QT syndrome, Brugada syndrome, or progressive cardiac conduction disease (also known as Lenegre's disease), or a combination thereof. In some embodiments, the promoter is a promoter naturally associated with inflammatory pain, visceral pain, migraine, erythromycin pain, fibromyalgia pain, idiopathic pain, or somatic pain, or a combination thereof. In some embodiments, the promoter is a promoter naturally associated with a neurological disease. In some embodiments, the promoter is a promoter naturally associated with dementia. In some embodiments, the promoter is a promoter naturally associated with Alzheimer's disease. In some embodiments, the promoter is a promoter naturally associated with Parkinson's disease. In some embodiments, the promoter is a promoter naturally associated with ALS. In some embodiments, the promoter is a promoter naturally associated with multiple sclerosis. In some embodiments, the promoter is a promoter naturally associated with a gene associated with a central nervous system disease. In some embodiments, the promoter is a promoter of a gene selected from SNCA, GBA, and LRRK2, SOD1, ataxin-2, SCA2, BFD1, FUS, C9orf72, brain-derived neurotrophic factor, nerve growth factor, neurotrophin, BCL11A, FMR1, DNM2, PrP, UBE3A, GYS1, and GFAP. In some embodiments, the promoter is a pol II promoter (e.g., Thy1 and H1xb9), a small latency associated promoter (e.g., from the herpesvirus pseudorabies virus), a cytomegalovirus promoter, SV40, elongation factor 1-alpha (EF1a) promoter, a cytomegalovirus enhancer / chicken-beta actin (CAG) promoter, or a herpes simplex virus (HSV) promoter.In some embodiments, the promoter is controlled by a small molecule. In some embodiments, the promoter is a tetracycline-responsive promoter, a glucocorticoid-responsive promoter, a RU-486-responsive promoter, a peroxide-inducible promoter, or a tamoxifen-inducible promoter. In some embodiments, expression of the epigenetic regulator and / or the transcriptional regulatory domain occurs upon natural or physiological induction of the promoter. In some embodiments, the promoter is induced upon a pathology in the subject. In some embodiments, the pathology is injury and / or inflammation. In some embodiments, the promoter is a galanin promoter or an NF-κB promoter. In some embodiments, the nucleic acid sequence comprises two or more promoters. In some embodiments, the nucleic acid sequence comprises a tandem promoter.

[0021] In some embodiments, the nucleic acid sequence comprises an enhancer. In some embodiments, the nucleic acid sequence comprises an intron. In some embodiments, the nucleic acid comprises an inverted terminal repeat (ITR). In some embodiments, the nucleic acid comprises a terminator sequence.

[0022] In some embodiments, the nucleic acid is administered with one or more guide RNA sequences. In some embodiments, the one or more guide RNA sequences are selected from the sequences in Table 6. In some embodiments, the one or more guide RNA sequences bind to one or more of the target molecules. In some embodiments, the one or more target molecules are 2, 3, 4, or 5 target molecules. In some embodiments, the one or more guide RNA sequences are 2, 3, 4, or 5 guide RNA sequences. In some embodiments, at least two guide RNA sequences are administered, and at least two guide RNA sequences are different. In some embodiments, at least one or more guide RNA sequences target the promoter of claims 221-252, creating a regulatory feedback loop to control expression levels.

[0023] In some embodiments, the nucleic acid is delivered as a naked (or unmodified) nucleic acid. In some embodiments, the nucleic acid is delivered in a complex with a cationic molecule. In some embodiments, the nucleic acid is delivered to the subject via a vehicle. In some embodiments, the vehicle is a viral delivery vehicle (e.g., a retroviral vector, a lentiviral vector, or an adenoviral vector), a liposome, a nanoparticle, or an exosome. In some embodiments, the vehicle is a viral delivery vehicle. In some embodiments, the viral delivery vehicle is a retroviral vector, a lentiviral vector, or an adenoviral vector. In some embodiments, the viral delivery vehicle is a recombinant adeno-associated virus (AAV). In some embodiments, the AAV is AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAVhu68, AAVrh.10, AAVrh74, or AAVDJ, or a combination thereof. In some embodiments, the AAV is AAV9. In some embodiments, the AAV has a recombinant capsid. In some embodiments, the recombinant capsid encodes a targeting moiety. In some embodiments, the nucleic acid comprises a sequence encoding the targeting moiety. In some embodiments, the vehicle is a liposome, lipid nanoparticle, nanocapsule, or exosome. In some embodiments, the vehicle comprises or is linked to a targeting moiety. In some embodiments, the targeting moiety targets a cell comprising a target molecule in the subject. In some embodiments, the targeting moiety binds to a peptide product of the target molecule. In some embodiments, the target molecule is present on a target cell. In some embodiments, the target cell is associated with a disease or condition of the subject. In some embodiments, the targeting moiety comprises a peptide. In some embodiments, the peptide comprises JNJ63955, m3-Huwentoxin-IV, Phlotoxin1, Protoxin-II (ProTx-II), or Ceratotoxin-1 (CcoTx1), Huwentoxin IV, or variants thereof.In some embodiments, the peptide comprises a sequence that is at least about 90% identical to a peptide in Table 7.

[0024] Additionally provided are combinations comprising a nucleic acid sequence and an additional therapeutic agent.

[0025] Exemplary embodiments are illustrated in the referenced figures. It is intended that the embodiments and figures disclosed herein be considered illustrative and not restrictive. [Brief description of the drawings]

[0026] [Figure 1] 1 is a schematic diagram of an exemplary nucleic acid composition described herein. [Diagram 2] FIG. 2 is a schematic diagram of another exemplary nucleic acid composition described herein. [Diagram 3] 1 shows expression of mCherry driven by different NaV1.7 promoters in the high NaV1.7 expressing human cell line HuH7. [Figure 4] 1 shows suppression of NaV1.7 gene expression following transfection of HuH7 cells with a nucleic acid composition encoding a ZFP and KRAB targeting NaV1.7. [Diagram 5] FIG. 1 is a schematic diagram of an exemplary nucleic acid composition described herein for the inhibition of a single target molecule. [Figure 6] FIG. 1 is a schematic diagram of an exemplary nucleic acid composition described herein for the inhibition of two target molecules. [Figure 7] AB show improvement in reversal of allodynia in male mice with single and dual targeted gene suppression. [Figure 8] AB show improvement in reversal of allodynia in female mice with single and dual targeted gene suppression. [Figure 9A] 1 shows expression of mCherry driven by different chemical conjugations of NaV1.7-binding peptides bound to the surface of AAV9 on different cell lines. [Figure 9B]1 shows expression of mCherry driven by different chemical conjugations of NaV1.7-binding peptides bound to the surface of AAV9 on different cell lines. [Figure 9C] 1 shows expression of mCherry driven by different chemical conjugations of NaV1.7-binding peptides bound to the surface of AAV9 on different cell lines. [Figure 10] 1 shows expression of mCherry driven by expression of the NaV1.7 binding peptide JNJ63955918-Indel and a short 6 bp linker on the AAV9 capsid. [Figure 11A] Expression of mCherry from different NaV1.7-specific promoters on different cell lines is shown. [Figure 11B] Expression of mCherry from different NaV1.7-specific promoters on different cell lines is shown. [Figure 11C] Expression of mCherry from different NaV1.7-specific promoters on different cell lines is shown. [Figure 11D] Expression of mCherry from different NaV1.7-specific promoters on different cell lines is shown. [Figure 12] FIG. 1 shows expression of mCherry from a NaV1.7-specific promoter in NaV1.7-expressing neurons of mouse dorsal root ganglia (DRG). DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0027] Described herein are compositions and methods for regulating gene expression via nucleic acid binding proteins that do not permanently edit the genome. The compositions and methods may be used to treat diseases or conditions, such as neurological diseases and / or pain. The compositions include nucleic acid sequences encoding nuclease-inactivated CRISPR-Cas systems (DeadCas for CRISPRi or CRISPRa) or zinc finger proteins, along with epigenetic regulators for gene repression, activation, and epigenetic editing. The methods include treating a subject with a disease or condition associated with a gene or non-coding RNA by introducing the compositions into a target cell and regulating expression of the gene or non-coding RNA in the cell. In an exemplary embodiment, the methods provide a route for targeted nucleic acid delivery using specific promoters and adeno-associated virus (AAV) modifications to enhance viral tropism.

[0028] Nucleic acid composition In one aspect, provided herein is a nucleic acid composition comprising a nucleic acid sequence encoding a DNA binding domain and an epigenetic regulator. Non-limiting examples of such DNA binding domains and epigenetic regulators are described herein. In exemplary embodiments, the nucleic acid binding domain and the epigenetic regulator are connected via a linker to form an epigenetic repressor or activator of gene expression. In some cases, the composition further comprises one or more of a guide RNA, a promoter, an enhancer, an intron, a nuclear localization signal, an ITR, a terminator sequence, and / or a delivery vehicle.

[0029] Also provided herein is a nucleic acid composition comprising an adeno-associated virus modified with a sequence encoding a peptide specific to the protein product of a target molecule. Such a composition can be useful for targeting nucleic acid to cells expressing the target molecule for targeted therapeutic approaches. For example, the peptide specifically binds to the protein product of the target molecule. Non-limiting examples of specific binding include peptides that bind to the protein product of the target molecule with high affinity, for example, nanomolar range affinity.

[0030] Figure 1 provides a schematic diagram of an exemplary nucleic acid composition described herein. The composition of Figure 1 includes an inverted terminal repeat (ITR), a promoter, a nuclear localization sequence, a sequence encoding a nucleic acid binding protein (exemplified as a zinc finger protein), a sequence encoding an epigenetic regulator (exemplified as a KRAB domain), and a terminator sequence. The promoter may be a cell-specific promoter (e.g., Na V 1.7, Na V 1.8, TRPV1, TRPA1 expressing cells, or neuron specific). The DNA sequence may be packaged into a single-stranded adeno-associated virus or a self-complementary adeno-associated virus (e.g., AAV9).

[0031] Nucleic Acid Binding Proteins Provided herein is a nucleic acid binding protein that is used to regulate gene expression without permanently editing genome.In certain embodiments, the nucleic acid binding domain binds to target molecule.In certain embodiments, the nucleic acid binding domain binds to DNA.In certain embodiments, the nucleic acid binding domain binds to RNA or the DNA complementary to RNA.

[0032] CRISPR / Cas An exemplary nucleic acid binding domain is a clustered regularly interspaced short palindromic repeats associated protein (known as dCas or CRISPRi) that does not have nuclease activity. The Cas molecules described herein do not have nuclease activity and therefore do not edit genomes. Non-limiting examples of dCas are provided in Table 4. In some cases, the dCas has a sequence that is at least about 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to the sequence in Table 4.

[0033] In some cases, dCas comprises a mutant Cas protein. In some cases, dCas is a truncated Cas protein. In some cases, dCas truncations are utilized to repress or activate genes. These truncations include, but are not limited to, REC2 domain deletion, REC3 domain deletion, HNH deletion, and deletion of the nuclease (NUC) lobe domain, or any combination of the aforementioned domains.

[0034] In some cases, the dCas comprises dCas9. In some cases, the dCas9 is a truncated or mutated Cas9 protein. In some cases, the dCas comprises dCas12. In some cases, the dCas12 is a truncated or mutated Cas12 protein.

[0035] In some cases, dCas has been linked to Streptococcus pyogenes, Staphylococcus aureus, Campylobacter jejuni, S.thermophilus, S.pneumoniae, Neisseria meningitidis, Corynebacter diphtheriae, Eubacterium ventriosum, Streptococcus pasteurianus, Lactobacillus farciminis, Sphaerochaeta globus, Azospirillum B510, Gluconacetobacter diazotrophicus, Neisseria cinerea, Roseburia intestinalis, Parvibaculum lavamentivorans, Nitratifractor salsuginis DSM 16511, Campylobacter lari CF89-12, or dCas9 from Streptococcus thermophilus LMD-9.

[0036] Zinc Finger Proteins Zinc finger proteins (ZFPs) contain a DNA-binding domain composed of a Cys2His2 zinc finger. ZFPs constitute the largest individual family of transcriptional regulators encoded by the genomes of higher organisms.

[0037] In some embodiments, a nucleic acid composition comprises a sequence encoding a ZFP. The ZFP can comprise a naturally occurring or modified sequence.

[0038] Non-limiting examples of ZFPs include the molecules in Table 8.

[0039] Additional Nucleic Acid Binding Domains Further non-limiting examples of nucleic acid binding domains include meganucleases and transcription activator-like effector nucleases (TALENs).

[0040] Guide RNA In the compositions and methods described herein using the CRISPR / Cas system, the nucleic acid comprises and / or is administered with one or more guide RNAs that bind to one or more target molecules. In some cases, the one or more guide RNAs are two, three, four, or five guide RNA sequences.

[0041] Non-limiting example guide RNAs are provided in Table 6. In some cases, the guide RNA comprises a sequence that is at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to a sequence in Table 6.

[0042] In some embodiments, the guide RNA sequence may target the promoter used to drive expression of the final construct, creating a regulatory feedback loop to control expression levels of the therapeutic agent.

[0043] Epigenetic regulators Provided herein are epigenetic regulators that regulate the expression of target molecules. In certain embodiments, the regulators activate expression. In certain embodiments, the regulators repress expression. The epigenetic regulators may be transcriptional regulatory domains with transcriptional repression activity (repressor domain) and / or transcriptional activation activity (activator domain). In some cases, the repressor domain comprises ZIM3. In some cases, the repressor domain comprises a Krueppel-associated box (KRAB) domain (recruitment of histone methyltransferases and deacetylases). Non-limiting examples of repressor domains are provided in Table 9 (e.g., ZIM3). Thus, provided herein are compositions comprising or encoding a repressor domain of Table 9, or a sequence at least about 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to a sequence of Table 9. The repressor domain may be a variant or combination of the repressor domains of Table 9. In some embodiments, the repressor domain comprises a sequence at least about 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to ZIM3, e.g., as shown in Table 9.

[0044] In certain embodiments, the epigenetic regulator is VP64 (recruits transcriptional activators), p65 (recruits transcriptional activators), p300 catalytic domain (histone acetyltransferase), TET1 catalytic domain (DNA demethylase), TDG (DNA demethylase), Ldb1 self-association domain (recruits enhancer-associated endogenous Ldb1), SAM activator (VP64, p65, HSF1) (recruits transcriptional activators), VPR (VP64, p65, Rta ) (recruits transcriptional activators), Sin3a (recruits histone deacetylases), LSD1 (histone demethylase), SUV39H1 (histone methyltransferase), G9a (EHMT2) (histone methyltransferase), DNMT3a (DNA methyltransferase), or DNMT3a-DNMT3L (DNA methyltransferases), p16, p300, CD, SunTag, or variants or combinations thereof.

[0045] In certain embodiments, the epigenetic regulator comprises KRAB (also known as KOX), SID, MBD2, MBD3, HP1a, the DNMT family (including DNMT1, DNMT3A, DNMT3B, DNMT3L, DNMT2A), Sin3a, Rb, MeCP2 (methyl-CpG binding protein 2), ROM2, AtHD2A, LSD1, SUV39H1, or G9a (EHMT2), or a variant or combination thereof. Variants of the KRAB domain include ZIM3, ZNF554, ZNF264, ZNF324, ZNF354A, ZNF189, ZNF543, ZNP82, ZNF669, ZNF582, KOX1-MeCP2, ZNF30, ZNF680, ZNF331, ZNF33A, ZNF528, ZNF320, ZNF350, ZNF175, ZNF214, ZNF184, ZNF8, ZNF596, KOX1, ZNF37A, ZNF394, ZNF610, ZNF273, ZNF34, ZNF250, ZNF98, ZNF675, ZNF213, NLuc, ZFP28-2, ZNF224, or ZNF257, or variants or combinations thereof.

[0046] In certain embodiments, the epigenetic regulator comprises a domain that recruits a transcriptional activator, a histone acetyltransferase, a DNA demethylase, a domain that recruits enhancer-associated endogenous Ldb1, a domain that recruits histone methyltransferases and deacetylases, a domain that recruits histone deacetylases, a histone demethylase, a histone methyltransferase, a DNA methyltransferase, an acetylation domain, or a deacetylation domain, or a combination thereof.

[0047] Simultaneous activation and inhibition In some embodiments, two or more genes are activated and repressed, and / or one gene is activated and another gene is repressed. A non-limiting example of a system for simultaneous activation and repression of two genes is shown in the schematic diagram of FIG. 2. In this diagram, the system includes a first sequence including the amino terminus of dCas9 (1-573), a guide RNA of SCN9A, and a KRAB repressor domain, and a second sequence including the carboxy terminus of dCas9 (574-1398), a guide RNA of PenK, and a VP64 activation domain. This system can be utilized to simultaneously repress SCN9A and activate PenK. The simultaneous activation and repression is via gRNA-M2M recruiting MCP-VP64 and gRNA-Com recruiting Com-KRAB. This system can be utilized with other activation and / or repressor domains, as well as gRNAs, for simultaneous targeting of different target molecules. While certain exemplary components are shown, the diagram should not be construed as limiting. For example, although the CMV promoter is shown, other promoters may alternatively be used, e.g., Na V It will be appreciated that the 1.7 promoter may be used.

[0048] Linker In certain embodiments, the epigenetic regulator is linked to the nucleic acid binding domain. The epigenetic regulator may be located at the N-terminus or C-terminus of the nucleic acid binding domain. The domains may be linked via a peptide linker. The domains may be linked via a disulfide bond.

[0049] In some embodiments, the epigenetic regulator is linked to the nucleic acid binding domain via a peptide linker. Non-limiting examples of peptide linkers include (GGS)n, (GGGS)n, (GGGGS)n, (G)n, (EAAAK)n, A(EAAAK)nALEA(EAAAK)nA, PAPAP, AEAAAKEAAAKA, (Ala-Pro)x, LE, GlySer-polyPro(Glyc)-polyPro(Glyc)-polyPro(Glyc), GlySer-polyPro-polyPro(Glyc)-polyPro, GlySer-pol yPro-GlySer(Glyc)-polyPro, GlySer-polyPro-polyPro-polyPro, GlySer-polyPro-β2m-polyPro, GlySer-polyPro-β2m-Gl ySer, polyPro-β2m-GlySer-β2m-GlySer, GlySer-polyPro-β2m-GlySer-β2m-polyPro, GlySer-polyPro-Ub-GlySer, GlySer-p and (G4S)n, wherein n is independently selected from 1 to 10 and x is an integer from 10 to 3. 4, polyPro is the proline-rich hinge sequence from IgA1, polyPro(Glyc) is the proline-rich hinge sequence from IgA1 with an embedded potential N-linked glycosylation site (Asn-Ser-Ser), β2m is β2-microglubulin, Ub is ubiquitin, ZAG is Zn-α2-glycoprotein, and cTPRX is the consensus tetratricopeptide repeat sequence with a repeat number of X.

[0050] promoter Further provided herein is a promoter that drives the expression of the nucleic acid binding domain and / or epigenetic regulator herein. In some embodiments, the nucleic acid composition comprises one or more promoters. In one example, the nucleic acid composition comprises a tandem promoter.

[0051] In some embodiments, the promoter is specific for the target molecule, so that the nucleic acid composition is specific to or expressed only in cells that express the target for increased therapeutic selectivity. As a non-limiting example, the target molecule is SCN9A.

[0052] In some embodiments, the promoter further increases the specificity of AAV tropism to cells expressing the target molecule. As a non-limiting example, the target molecule is SCN9A. Downregulating or upregulating the target molecule only in target molecule-expressing cells can reduce off-target effects and general toxicity. This is important to prevent the expression of effectors (e.g., dCas or ZFP) in immune cells such as glial cells, microglia, macrophages, astrocytes, etc., which can mediate immune responses to the gene therapy proposed herein.

[0053] In some embodiments, the promoter is SCN9A (Na V 1.7) and is specific for Na V 1.7 or other gene products. In some embodiments, the promoter is specific for any one of the genes in Tables 1-3.

[0054] In some embodiments, some promoters can be induced with small molecules or other means. These inducible expression promoters include tetracycline-responsive promoters, glucocorticoid-responsive promoters, RU-486-responsive promoters, peroxide-inducible promoters, and tamoxifen-inducible promoters.

[0055] In addition, there are promoters that can be induced when pathology such as injury or inflammation occurs. Injury-induced promoters include the galanin promoter, which is specific to nociceptive afferent neurons. Inflammation-induced promoter NF-κB can also be used for pain associated with inflammation.

[0056] Tandem promoters and promoter combinations can also be used to prevent immune responses and generate more cell type specific expression.

[0057] In some embodiments, expression of the epigenetic regulator and / or the transcriptional regulatory domain occurs upon natural or physiological induction of the promoter.

[0058] In some embodiments, pan-neuronal gene promoters are used to regulate gene expression in neurological diseases and suppress pain-related genes. Non-limiting exemplary promoters include the promoter of microtubule-associated protein 2 (MAP-2), the promoter of neuron-specific enolase (NSE), the promoter of choline acetyltransferase (ChAT), the promoter of protein gene product 9.5 (PGP9.5) (also called ubiquitin C-terminal hydrolase 1 (UCHL-1)), the gene promoter of human synapsin 1 (hSYN1), the promoter of the NeuN gene (Fox-3, Rbfox3, or hexaribonucleotide-binding protein-3), the promoter of α-calcium / calmodulin-dependent protein kinase II [CaMKIIα], the promoter of the Rheb gene (ras homolog enriched in brain), and the TRKA promoter (tyrosine kinase A). In some embodiments, the promoter is neuron-specific, such as pol II promoters, including Thy1 and H1xb9. The small latency-associated promoter from the herpesvirus pseudorabies virus can also be used for pan-neuronal expression of effectors (dCas9 or ZFPs) fused to a repressor domain.

[0059] Additional promoters include cytomegalovirus (CMV), SV40, elongation factor 1-alpha (EF1a) promoter, cytomegalovirus enhancer / chicken beta-actin (CAG) promoter, jET promoter, and herpes simplex virus (HSV).

[0060] In some cases, the promoter comprises a sequence that is at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to a sequence in Table 5.

[0061] In some embodiments, the promoter is an SCN9A promoter or an SCN10A promoter.

[0062] In some embodiments, the promoter is naturally associated with a gene associated with a channelopathy, Dravet syndrome, epilepsy syndrome, familial hemiplegic migraine, Ohtahara syndrome, West syndrome, Lennox-Gastaut syndrome, sodium channel myotonia, autism, long QT syndrome, Brugada syndrome, or progressive cardiac conduction disease (also known as Lenegre's disease), pain (e.g., inflammatory pain, visceral pain, migraine, red pulp pain, fibromyalgia, idiopathic pain, somatic pain), neurological disease, dementia, Alzheimer's disease, Parkinson's disease, ALS, multiple sclerosis, a disease of the central nervous system, or a combination thereof.

[0063] In some embodiments, the promoter is a promoter of a gene selected from SNCA, GBA, LRRK2, SOD1, ataxin-2, SCA2, BFD1, FUS, C9orf72, brain-derived neurotrophic factor, nerve growth factor, neurotrophin, BCL11A, FMR1, DNM2, PrP, UBE3A, GYS1, and GFAP.

[0064] In some embodiments, the promoter is a pol II promoter (e.g., Thy1 and H1xb9), a mini-latency associated promoter (e.g., from the herpesvirus pseudorabies virus), a cytomegalovirus promoter, SV40, an elongation factor 1-alpha (EF1a) promoter, a cytomegalovirus enhancer / chicken beta-actin (CAG) promoter, or a herpes simplex virus (HSV) promoter.

[0065] nucleic acid delivery In some embodiments, the nucleic acid compositions herein are delivered as naked or unmodified nucleic acids, hi other embodiments, the nucleic acid compositions are delivered complexed with cationic molecules.

[0066] In some embodiments, the nucleic acid is delivered to the subject via a vehicle, which may be a liposome, a lipid nanoparticle, a nanocapsule, or an exosome.

[0067] In some embodiments, the nucleic acid is delivered via a viral vehicle. Non-limiting viral vehicles include, but are not limited to, retroviral vectors, lentiviral vectors, adenoviral vectors, adeno-associated viral vectors (e.g., AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAVhu68, AAVrh.10, AAVrh74, AAVDJ), and the like. In some cases, the vehicle is a recombinant adeno-associated virus (AAV). In some cases, the AAV is AAV9. AAV9 is a recombinant adeno-associated virus (AAV). V It can be selected for its highest tropism for dorsal root ganglion (DRG) neurons, where pain-related proteins such as 1.7 are highly expressed. Furthermore, AAV9 has been shown to be safe.

[0068] Any delivery vehicle (viral vector or non-viral) can have improved tropism for cells expressing the protein product of the target molecule. For example, the vehicle can include a peptide that binds to the protein product of the target molecule. As a non-limiting example, the peptide can be a peptide that binds to the protein product of the target molecule. V 1.7 Binding of nucleic acids to Na V 1.7 Targeting to expressing cells.

[0069] targeting part In some embodiments, the nucleic acid compositions herein, along with a nucleic acid sequence encoding a peptide targeting moiety, are delivered via a viral vehicle, such as an AAV capsid as described herein.

[0070] In some embodiments, the nucleic acid compositions herein are delivered via a non-viral delivery vehicle, such as, but not limited to, a liposome, lipid nanoparticle, nanocapsule, or exosome. In some such embodiments, the vehicle may include and / or be linked to a targeting moiety, such as a small molecule or peptide targeting moiety.

[0071] In some embodiments, the targeting moiety comprises a peptide targeting moiety that binds to a protein product of a targeting molecule to target the nucleic acid to a particular cell. In some cases, the targeting molecule is present on a target cell. In some cases, the target cell is associated with a disease or condition of interest.

[0072] Non-limiting examples of peptide targeting moieties for use in viral and non-viral delivery methods include JNJ63955, m3-Huwentoxin-IV, Phlotoxin1 (PhlTx1), Protoxin-II (ProTx-II), Ceratotoxin-1 (CcoTx1), Huwentoxin-IV (HwTx-IV), μ-TRTX-Pn3a, Jz-Tx-V, GsAFI, Tp1a (Protoxin-III), GpTx-1, HpTx1, Hm1a, and variants and combinations thereof. The peptides may be derived from one or more of the following organisms: Thrixopelma prurient, Pamphobeteus nigricolor, Chilobrachys jingzhao, Grammostola spatulata, Phlogiellus genus, Thrixopelma pruriens, Grammostola porteri, Selenocosmia huwena, Ceratogyrus cornuatus, Heteropoda venatoria, and / or Heteroscodra maculate.

[0073] In some cases, the peptide comprises a sequence that is at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to a peptide in Table 7. The peptides in Table 7 include Na V 1.7 and therefore Na V 1.7 may be useful for providing AAV tropism to expressing cells. In some cases, the peptide comprises ProTx-II, ProTx-III, ProTx-II variant JNJ63955, HwTx-IV variant m3-HwTx-IV, CcoTx1 variant 2670, PhlTx1 variant D7A-PhlTx1, or JxTx-V variant AM-0422.

[0074] AAV vectors Recombinant adeno-associated virus (AAV) is one of the most commonly used vehicles for in vivo gene delivery. To enter a host cell, the virus 1) binds to a receptor and glycan co-receptor on the cell surface, 2) is endocytosed, 3) progresses through an endosomal compartment, 4) escapes the endosome, and 5) is transported to the nucleus. Once inside the nucleus, the virus sheds its coat and its single-stranded genome is converted into a double-stranded genome that the host cell can now use as a template for gene expression. The AAV receptor (AAVR, KIAA0319L) has been reported to be essential for AAV to enter cells, although some recombinant AAVs can enter cells independently of AAVR. Glycosylation is also known to play a role in viral transduction efficiency. Altering the capsid composition of AAV alters its ability to enter cells. There are at least four major techniques currently being used to modify and improve the tropism of AAVs: rational engineering, directed evolution, evolutionary phylogenetic analysis, and chemical conjugation.

[0075] One method to modify viral tropism is to generate a large library of peptides for addition to the AAV surface and then characterize the resulting variants to determine their tropism. This method is labor intensive and requires extensive screening efforts since the library of screened peptides is generated more or less randomly, so success is based on a numbers game. In a similar technique called directed viral evolution, organs are collected from a first round of viral infection and screened to select viral variants with the desired tropism (e.g., brain-specific) for subsequent rounds of infection and even more specific tropisms. However, some of these screening methods cannot be translated between species.

[0076] Instead of using random library screening, described herein is a rational design method to improve AAV tropism.Rational design strategies for AAV capsid engineering include peptide domain insertion and chemical biology approaches.Peptides known to specifically interact with cells of interest can be added.By using binding peptides that bind to proteins that code for target molecules, the AAV described herein enhances tropism for target molecule-expressing cells, generating a more targeted strategy.

[0077] In some embodiments, the peptide for increasing tropism is Na V Binds to 1.7 Na V 1.7 Peptides that increase tropism include: JNJ63955, m3-Huwentoxin-IV, Phlotoxin1 (PhlTx1), Protoxin-II (ProTx-II), Ceratotoxin-1 (CcoTx1), Huwentoxin-IV (HwTx-IV), and those described elsewhere herein, e.g., a peptide in Table 7, or a sequence that is at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to a peptide in Table 7.

[0078] target molecule In certain embodiments, the compositions described herein modulate the expression of one or more target molecules associated with a disease or condition in a subject. In some cases, the expression of the target molecule is activated. In some cases, the expression of the target molecule is inhibited.

[0079] One or more target molecules may comprise DNA or RNA. In some embodiments, the target molecule comprises DNA. In some embodiments, the target molecule comprises a coding region of a gene. In some embodiments, the target molecule comprises DNA complementary to a non-coding RNA. The non-coding RNA may be associated with a disease or condition (e.g., pain) described herein. For example, the non-coding RNA is associated with neuropathic pain, such as spinal nerve ligation injury, nerve branch ligation injury, chronic constriction injury, or diabetic neuropathy, or a combination thereof.

[0080] In some embodiments, the non-coding RNA comprises SCN9A natural antisense transcript (NAT), Kcna2 antisense RNA, H19, Gm21781, MRAK009713, uc.48+, NONRATT021972, BC168687, Speer7-ps, Uc007pbc.1, XLOC_041439, Mlxipl, Rn50_X_0739.1, CCAT1, rno circ0004058, rno_circRNA_007512, or Egr2 antisense RNA, or a combination thereof.

[0081] In some embodiments, the one or more target molecules comprise a nucleic acid associated with a disease or condition described herein. Non-limiting examples of nucleic acids associated with diseases and conditions described herein are provided in Tables 1-3. In some embodiments, the one or more target molecules comprise a nucleic acid associated with pain. Pain includes neuropathic pain, inflammatory pain, visceral pain, migraine, erythromycin pain, fibromyalgia pain, idiopathic pain, and somatic pain.

[0082] In some embodiments, the one or more target molecules comprise a nucleic acid associated with a channelopathy. In some cases, the one or more target molecules comprise a nucleic acid encoding a channel. The channel can be an ion channel, e.g., a sodium channel, a potassium channel, a calcium channel, and / or a chloride channel.

[0083] In some embodiments, the one or more target molecules comprise a nucleic acid associated with a neurological disease. In some embodiments, the one or more target molecules comprise a nucleic acid associated with dementia. In some embodiments, the one or more target molecules comprise a nucleic acid associated with Alzheimer's disease. In some embodiments, the one or more target molecules comprise a nucleic acid associated with Parkinson's disease. In some embodiments, the one or more target molecules comprise a nucleic acid associated with Huntington's disease. In some embodiments, the one or more target molecules comprise a nucleic acid associated with schizophrenia. In some embodiments, the one or more target molecules comprise a nucleic acid associated with amyotrophic lateral sclerosis (ALS). In some embodiments, the one or more target molecules comprise a nucleic acid associated with multiple sclerosis. In some embodiments, the one or more target molecules comprise a nucleic acid associated with a central nervous system disease. In some embodiments, the one or more target molecules comprise a nucleic acid associated with Dravet syndrome, epilepsy syndrome, familial hemiplegic migraine, Ohtahara syndrome, West syndrome, Lennox-Gastaut syndrome, sodium channel dystonia, autism, long QT syndrome, Brugada syndrome, or progressive cardiac conduction disease (also known as LeNegret disease), or a combination thereof.

[0084] In some embodiments, the one or more target molecules comprise one or more genes in Tables 1-3. In some embodiments, the one or more target molecules comprise SCN9A, SCN10A, or SCN11A, or a combination thereof. In some embodiments, the one or more target molecules comprise SNCA, GBA, or LRRK2, or a combination thereof. In some embodiments, the one or more target molecules comprise GPR52. In some embodiments, the one or more target molecules comprise SOD1, ataxin-2, or SCA2, or a combination thereof. In some embodiments, the one or more target molecules comprise one or more genes selected from the group comprising BFD1, FUS, C9orf72, brain-derived neurotrophic factor, nerve growth factor, neurotrophin, BCL11A, FMR1, DNM2, PrP, UBE3A, GYS1, and GFAP.

[0085] Genes involved in pain The human genome encodes genes that may confer protection against unwanted pain. Genetic studies have demonstrated that the human voltage-gated sodium channel (Na V 1.7 (SCN9A)) with a rare genetic disorder that results in insensitivity to pain without other neurodevelopmental changes. This sodium channel therefore represents an attractive target for developing chronic pain therapies. However, Na V Efforts to develop selective small molecule inhibitors have been hampered by the high sequence identity between subtypes, and in fact, Na V Many small molecule drugs targeting 1.7 have failed due to lack of specificity. Antibodies face a similar situation, as there is a trade-off between selectivity and potency due to the antibody binding to a specific (open or closed) conformation of the channel. In fact, even commercially available antibodies targeting the human channel are insufficient for Western blot. Interfering RNA (RNAi) has also been shown to inhibit Na V 1.7. However, as an exogenous system, RNAi competes with endogenous mechanisms such as microRNA or RISC complex function. Thus, RNAi can compete with and impair basic homeostatic mechanisms of RNA synthesis and degradation. In addition, due to high RNA turnover, RNAi methods have a lower pharmacokinetic prospect and require higher doses. The main cause of these shortcomings is the lack of Na+ delivery systems based on these methods. V1.7 None of the targeted therapies have yet been successful in reaching the final stage of clinical trials. In contrast, disclosed herein is the use of nucleic acid binding domains (e.g., also known as nuclease-inactivated "dead" CRISPR-Cas (dCas)-CRISPR interference or CRISPRi and zinc finger proteins) and epigenetic regulators (e.g., KRAB repressors) to suppress the transcription of SCN9A and other genes associated with pain. Since permanent ablation of pain is undesirable, there is no permanent genome editing using such methods. Instead, these epigenetic engineering methods are directed to Na V 1.7 Allows for transient regulation of gene expression. Furthermore, this approach may have less risk of off-target effects than other approaches. Rather than pharmacologically targeting proteins or RNA, this approach targets Na at the DNA level. V 1.7, which may provide longer-lasting results than methods that target proteins or RNA. This approach allows the design of highly specific, long-term, and reversible treatments for pain. The duration of treatment is important, since many pain conditions resulting from chronic inflammation and nerve damage are typically enduring conditions that require continuous remedication. This genetic approach provides continuous, controllable regulation of abnormal pain processing. Furthermore, it represents a new paradigm in pain management, since the disclosed approach can be easily designed to target several genes, providing a synergistic way to target single or multiple sodium channels for more potent pain relief.

[0086] In addition to SCN9A, Table 2 provides other genes involved in pain. For genes that are upregulated, the methods herein can suppress expression, and for genes that are downregulated, the methods herein can activate expression. In some cases, the methods both suppress and activate expression of one or more genes.

[0087] Genes involved in channelopathies In certain embodiments, the target molecule comprises a gene involved in a channelopathy. The gene may be associated with or may encode a channel, such as a sodium channel, a potassium channel, a calcium channel, and / or a chloride channel. Non-limiting examples of such genes are provided in Table 1.

[0088] Genes involved in neurological diseases or conditions In certain embodiments, the target molecule comprises a gene involved in a neurological disease or condition of the central nervous system. Non-limiting exemplary diseases and conditions include Alzheimer's disease, Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis (ALS), and multiple sclerosis.

[0089] Alzheimer's disease (AD) is a progressive disease that causes brain cells to wear out (degenerate) and die. Alzheimer's is the most common cause of dementia, with a continuing decline in thinking, behavior, and social skills preventing a person from functioning independently. Activating and / or suppressing genes may slow the progression or prevent Alzheimer's disease.

[0090] Studies have shown that the binding of B-amyloid to LilrB2 (e.g., Uniprot reference number Q8N423) is one of the first steps leading to Alzheimer's disease. Thus, inhibition of LilrB2, which affects B-amyloid binding, can prevent the onset of Alzheimer's disease. Dl is associated with Uniprot reference number P21728. D2 is associated with Uniprot reference number 14416. Thus, compositions and methods for inhibiting the expression of LilrB2 are provided herein.

[0091] MS4A4A and TREM2 operate in microglia, the immune cells of the brain. They affect the risk of Alzheimer's disease by altering the levels of TREM2, a protein that helps microglial cells remove excess amounts of Alzheimer's disease proteins amyloid and tau from the brain. Thus, activation of TREM2 in cerebrospinal fluid (CSF) may help prevent and / or slow the progression of Alzheimer's disease. Thus, compositions and methods are provided herein that activate the expression of TREM2.

[0092] Apolipoprotein E4 (apoE4), the most common genetic risk factor for Alzheimer's disease, is expressed in more than half of Alzheimer's disease patients and may therefore be an important therapeutic target. Of the three polymorphisms of apoE, namely apoE2, apoE3, and apoE4, carriers of apoE4 are more likely to develop Alzheimer's disease. Blocking apoE4 effects helps 40-60% of AD patients who carry apoE4, but if all apoE forms are in fact toxic, another approach is to block all apoE effects. In addition, several studies have revealed that apoE4 is also a risk factor for other diseases, including cerebral amyloid angiopathy, dementia with Lewy bodies, tauopathy, cerebrovascular disease, multiple sclerosis, and vascular dementia. Thus, provided herein are compositions and methods for inhibiting APOE4 expression.

[0093] VEGF (vascular endothelial growth factor), originally described as an important angiogenic factor, has been shown to play a key role in neurogenesis and neuroprotection, affecting neuroplasticity and repair. Animal model studies have revealed that the brain levels of VEGF and its receptor (VEGFR-2) are reduced in the hippocampus of apoE4 mice. Thus, upregulation of hippocampal VEGF levels can reverse the accumulation of Aβ and hyperphosphorylated tau in hippocampal neurons. Thus, provided herein are compositions and methods for upregulating VEGF expression.

[0094] Similarly, upregulation of ABCA1 (ATP-binding cassette transporter ABCA1) reverses apoE4-driven cognitive and brain pathology, making ABCA1 a target molecule for upregulation and treatment of Alzheimer's disease.

[0095] Transactive response DNA-binding protein 43 (TDP-43), an RNA-binding protein that functions in axon skipping, has recently been shown to be deposited in AD brains. TDP-43 is present in the brains of 65-80% of AD patients and has been shown to be associated with progressive hippocampal atrophy. Thus, in some embodiments, the target molecule is TDP-43, and the compositions described herein inhibit the expression of TDP-43.

[0096] Target molecules also include genes with known mutations that cause early-onset Alzheimer's disease, such as presenilin 1 (PSEN1) and presenilin 2 (PSEN2), and thus activation of these genes may be another potential avenue for the treatment of Alzheimer's disease.

[0097] Additional non-limiting target molecules involved in neurological diseases, such as Alzheimer's disease, are shown in Table 3. Such genes may be inhibited or upregulated using the compositions and methods described herein.

[0098] Additional target molecules include α-synuclein, microtubule-associated protein tau, APP, and huntingtin (SNCA, MAPT, APP, and HTT, respectively). Precise transcriptional regulation is achieved via CRISPRa of neurodegenerative disease-related genes in human iPSC-derived neurons. TSS2-2 sgRNA and dCas9-VPR transcriptional activator mediated activation of α-synuclein in neurons derived from normal α-synuclein levels (NAS) iPSCs, normal control patients, and iPSCs derived from patients with Parkinson's disease caused by α-synuclein triplication (AST). Eight-fold activation of endogenous SNCA expression was achieved in NAS iPSC-derived neurons, and dCas9-KRAB inhibition reduced α-synuclein mRNA levels by 40% in AST iPSC-derived neurons. In addition, targeting genes close to the TSS region significantly reduced off-target effects and reduced the negative side effects of using SpCas9. Overall, these findings suggest the potential to harness the regulatable CRISPRa / CRISPRi platform for multiplex transcriptional repression of molecular pathological signatures in vivo in the mammalian brain and to address neurodegeneration in familial and sporadic disease states.

[0099] In some embodiments, the target molecule comprises one or more of alpha-synuclein (SNCA), glucocerebrosidase (GBA), and / or leucine-rich repeat kinase (LRRK2). In some cases, SNCA degradation is prevented by inhibiting the tyrosine kinase c-ABL. In some cases, expression of SNCA is inhibited. The target molecule may be inhibited for the treatment of Parkinson's disease or another condition of the nervous system.

[0100] In some embodiments, the target molecule comprises HTT, which encodes the Huntington protein. Inhibition of mutant HTT can be performed to treat Huntington's disease.

[0101] In some embodiments, the target molecule comprises GPR52. Modulation of GPR52 may be performed to treat Huntington's disease and / or schizophrenia. GPR52 upregulation can be used for schizophrenia, cognitive impairment, psychosis, brain malformation and hyperactivity. Inhibition of GPR52 can be used for the treatment of Huntington's disease, since GPR52 is associated with the abnormal expression of huntingtin observed in patients with this disorder.

[0102] In some embodiments, the target molecule comprises at least one of superoxide dismutase 1 (SOD1), ataxin-2, and / or transactive response DNA binding protein 43 (TDP-43). Modulation of the target molecule can be performed to treat ALS, such as inhibition of SOD1, ataxin-2, or TDP43. Modulation of the target molecule may also be performed to treat frontotemporal dementia (FTD) or other neurological diseases.

[0103] In some embodiments, the target molecule comprises BFD1, a transcription factor protein that encodes bradycardia formation defective 1. BFD1 can drive expression of genes required for bradycardia formation in Toxoplasma Gondii, and thus its inhibition can prevent chronicity of this and other infections using similar chronicity pathways.

[0104] Mutations in the C9orf72 gene are the most common genetic cause of two neurodegenerative diseases. Mouse and human studies suggest the role of loss of C9orf72 protein in some neurodegenerative disorders, as well as reduced C9orf72 levels, and there is more inflammation mediated by STING protein in immune cells and brain cells. Thus, in some embodiments, the target molecule comprises C9orf72, and the compositions herein upregulate C9orf72 expression. In some embodiments, the target molecule comprises STING gene.

[0105] Additional genes that may be upregulated to treat neurological disorders include neurotrophic factors such as brain-derived neurotrophic factor, nerve growth factor, and neurotrophins. The DDC gene, which encodes AADC, may be upregulated to treat Parkinson's disease.

[0106] Genes that may be downregulated to treat the following diseases include B-thalassemia (BCL11A), Fragile X syndrome (FMR1), centronuclear myopathy (DNM2), prion disease (PrP), Angelman syndrome (UBE3A), Lafora disease (GYS1), and Alexander disease (GFAP).

[0107] Treatment method Various embodiments provide methods of treating a disease or condition in a subject with a composition described herein. In some embodiments, the composition comprises a nucleic acid described herein encoding a nucleic acid binding domain and an epigenetic regulator. The composition can be delivered via AAV or a non-viral vehicle. Non-limiting exemplary compositions include those generally depicted in Figures 1-2.

[0108] In an exemplary embodiment, the disease or condition comprises pain. Pain includes neuropathic, inflammatory, visceral, migraine, red pulp pain, fibromyalgia, idiopathic and somatic pain. Inflammatory pain includes rheumatoid arthritis pain. The disease or condition includes Na V 1.7 or other genes involved in pain.

[0109] In some embodiments, the disease or condition comprises a channelopathy, including Dravet syndrome, epilepsy syndrome, familial hemiplegic migraine, Otahara syndrome, West syndrome, Lennox-Gastaut syndrome, sodium channelotonopathy, autism, long QT syndrome, Brugada syndrome, progressive cardiac conduction disease (also called Lenegre's disease), and the like. Exemplary diseases and conditions are shown in Table 1.

[0110] In some embodiments, the disease or condition comprises a neurological disease or condition, including dementia, Alzheimer's disease, Parkinson's disease, Huntington's disease, schizophrenia, amyotrophic lateral sclerosis (ALS), and multiple sclerosis. The disease or condition may comprise a disease of the central nervous system.

[0111] In some embodiments, the disease or condition comprises an inflammatory disease or condition. As a non-limiting example, the inflammatory disease or condition is rheumatoid arthritis.

[0112] In some embodiments, the disease or condition comprises an infectious disease.

[0113] In some embodiments, the disease or condition comprises β-thalassemia, fragile X syndrome, centronuclear myopathy, prion disease, Angelman syndrome, Lafora disease, or Alexander disease, or a combination thereof.

[0114] In some embodiments, the method of treatment comprises administering a nucleic acid composition described herein and one or more additional active agents, e.g., the additional active agents may be used to complementarily treat a disease or condition.

[0115] In some embodiments, the subject refers to any animal, including, but not limited to, humans, non-human primates, rodents, and livestock and game animals. Primates include chimpanzees, cynomolgus monkeys, spider monkeys, and macaques, such as rhesus monkeys. Rodents include mice, rats, woodchucks, ferrets, rabbits, and hamsters. Livestock and game animals include cattle, horses, pigs, deer, bison, buffalo, feline species, such as domestic cats, canine species, such as dogs, foxes, wolves, avian species, such as chickens, emus, ostriches, and fish, such as trout, catfish, and salmon. In certain embodiments, the subject is a human.

[0116] In various embodiments, the subject may be a subject suffering from, or previously diagnosed with, or identified as suffering from, a disease or condition requiring treatment. In various embodiments, a subject suffering from, or previously diagnosed with, or identified as suffering from a disease or condition may or may not have been treated for the condition. In other embodiments, the subject may also be a subject not previously diagnosed with a disease or condition, and the therapeutic agent is used for prevention (prophylactic).

[0117] A "subject" in need of treatment for a particular condition may be a subject who has the condition, has been diagnosed with the condition, or is at risk for developing the condition. In some embodiments, the subject is a "patient" who has been diagnosed with a disease or condition described herein. In some embodiments, the subject is a military personnel for the prevention or relief of pain and / or physical suffering.

[0118] In some embodiments, the term "therapeutically effective amount" refers to an amount of a composition (e.g., a nucleic acid) effective to "treat" a disease or disorder in a subject or mammal. In some cases, a therapeutically effective amount of a composition reduces the severity of symptoms of a disease or condition. In some cases, a therapeutically effective amount of a composition prevents the onset of a disease or condition.

[0119] In some embodiments, the term "treat" or "treating" as used herein refers to both therapeutic treatment and prophylactic or preventative measures (e.g., disease progression), the purpose of which is to prevent or slow (alleviate) the targeted pathological condition. In some aspects provided herein, subjects in need of treatment include those already suffering from a disease or condition, as well as those susceptible to developing a disease or condition.

[0120] Pharmaceutical Compositions, Administration and Dosage In various embodiments, the compositions herein are formulated for delivery via any route of administration. "Route of administration" can refer to any route of administration known in the art, including but not limited to intrathecal, epidural, intravenous, transdermal, intranasal, oral, mucosal, or other delivery methods, and / or via single or multiple doses. Examples of routes of administration of nucleic acids described herein include lumbar intrathecal puncture, intracisternal administration, and intraganglionic administration.

[0121] In an exemplary embodiment, the composition is delivered to the spinal intrathecal space using any suitable delivery method. V Because the role played by 1.7 is in nociceptive afferents, whose cell bodies are located in the respective segmental dorsal root ganglion (DRG) neurons, Na V 1.7. Thus, delivery to the spinal intrathecal space may be particularly useful for targeting Na V Compositions targeting 1.7 can be efficiently delivered to DRG neurons, which can minimize the possibility of off-target biodistribution and reduce the viral load required for transduction.

[0122] It is understood that the actual dosage may vary depending on the route of administration, the delivery system used (e.g., AAV or liposomes, etc.), the target cell, organ, or tissue, the subject, and the degree of effect sought. The size and weight of the tissue, organ, and / or patient may also affect dosing. The dose may further include additional agents, including, but not limited to, carriers. Non-limiting examples of suitable carriers are known in the art, such as water, saline, ethanol, glycerol, lactose, sucrose, dextran, agar, pectin, vegetable-derived oils, phosphate-buffered saline, and / or diluents.

[0123] The pharmaceutical composition may also contain any pharma- ceutically acceptable carrier. A "pharma-ceutically acceptable carrier" refers to a pharma-ceutically acceptable material, composition, or vehicle involved in carrying or transporting the composition from one tissue, organ, or part of the body to another tissue, organ, or part of the body. For example, the carrier may be a liquid or solid filler, diluent, excipient, solvent, or encapsulating material, or a combination thereof. Each component of the carrier must be "pharma-ceutically acceptable" in that it must be compatible with the other components of the formulation. It must also be suitable for use in contact with any tissue or organ with which it may come into contact. That is, it must not be associated with a risk of toxicity, irritation, allergic reaction, immunogenicity, or other complications that unduly outweigh its therapeutic benefits.

[0124] In various embodiments, a pharmaceutical composition is provided that includes a therapeutically effective amount of a nucleic acid as described herein together with a pharma- ceutically acceptable excipient. By "pharmaceutically acceptable excipient" is meant an excipient that is generally safe, non-toxic, and useful in the preparation of a desired pharmaceutical composition, including excipients that are acceptable for veterinary use as well as for human pharmaceutical use. The active ingredient can be mixed with an excipient that is pharma- ceutically acceptable and compatible with the active ingredient, in an amount suitable for use in the therapeutic methods described herein. Such excipients can be solid, liquid, semi-solid, or, in the case of an aerosol composition, gaseous. Suitable excipients are, for example, starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glycerol monostearate, talc, sodium chloride, dried skim milk, water, saline, dextrose, polyethylene glycol, glycerol, ethanol, mannitol, polysorbate, and the like, and combinations thereof. In addition, if necessary, the composition may contain auxiliary substances such as wetting or emulsifying agents, pH buffering agents, etc., which enhance or maintain the effectiveness of the active ingredient or increase the stability of the pharmaceutical. In addition, if necessary, the composition may contain auxiliary substances to modify the density of the pharmaceutical. The therapeutic compositions described herein may contain pharmaceutically acceptable salts. Pharmaceutically acceptable salts include acid addition salts formed with inorganic acids, such as hydrochloric acid or phosphoric acid, organic acids, such as acetic acid, tartaric acid or mandelic acid, inorganic bases, such as sodium, potassium, ammonium, calcium or ferric hydroxide, and organic bases, such as isopropylamine, trimethylamine, 2-ethylaminoethanol, histidine, procaine, etc. Liquid compositions may contain liquid phases in addition to and to the exclusion of water, such as water, vegetable oils, such as glycerin, cottonseed oil, and water-oil emulsions. Physiologically acceptable carriers are well known in the art.

[0125] The pharmaceutical composition may be delivered in a therapeutically effective amount. The exact therapeutically effective amount is the amount of the composition that will produce the most effective results in terms of efficacy of treatment in a given subject. This amount will vary depending on a variety of factors, including, but not limited to, the characteristics of the nucleic acid (including, but not limited to, activity, pharmacokinetics, pharmacodynamics, and bioavailability), the physiological state of the subject (including, but not limited to, age, sex, type and stage of disease, general physical condition, responsiveness to a given dose, and type of drug), the nature of the pharmacologic acceptable carrier in the formulation, and the route of administration. If the intrathecal route is recommended, the pharmaceutical agent may be diluted ex vivo with the subject's cerebrospinal fluid prior to administration to achieve an isobaric solution.

[0126] kit Further provided is a kit for carrying out the method described herein. The kit is a collection of components, including at least one of the compositions described herein. Thus, in some embodiments, the kit includes a nucleic acid encoding a nucleic acid binding domain and an epigenetic regulator. The nucleic acid may be combined or complexed with another component, such as a vehicle, for delivery, or may be unmodified for direct delivery. In some cases, the nucleic acid is complexed with a cationic molecule. In some cases, the nucleic acid is configured to be delivered via a viral delivery vehicle, such as an AAV capsid protein. For certain kits in which the nucleic acid binding domain includes a dCas protein, the kit includes one or more guide RNA sequences.

[0127] The kit may include instructions for use of the components. Optionally, the kit also includes other useful components, such as diluents, buffers, pharma- ceutically acceptable carriers, syringes, catheters, applicators, pipettes or measuring tools, dressings, or other useful paraphernalia that will be readily recognized by one of skill in the art.

[0128] The materials or components assembled in the kit can be provided to the practitioner stored in any convenient and suitable manner that maintains their operability and usefulness. For example, the components can be in dissolved, dehydrated, or lyophilized form, and they can be provided at room temperature, refrigerated temperature, or frozen temperature. The components are typically included in a suitable packaging material(s). As used herein, the phrase "packaging material" refers to one or more physical structures used to contain the contents of the kit, such as the compositions of the present invention. The packaging material is constructed by well-known methods to provide a preferably sterile, contaminant-free environment. The packaging materials used in the kit are those that are commonly utilized in gene expression assays and administration of therapeutics. As used herein, the term "package" refers to a suitable solid matrix or material, such as glass, plastic, paper, foil, etc., capable of holding the individual kit components. Thus, for example, the package can be a glass vial or a pre-filled syringe used herein to contain the appropriate amount of the composition containing the nucleic acid. The packaging material generally has an exterior label that indicates the contents and / or purpose of the kit and / or its components.

[0129] Numbered embodiments 1. A method for modulating expression of one or more target molecules associated with a disease or condition in a subject, the method comprising administering to the subject a nucleic acid sequence encoding a nucleic acid binding domain and an epigenetic regulator that modulates transcription of the one or more target molecules. 2. The method of embodiment 1, wherein said modulation of transcription of said one or more target molecules is transient. 3. The method of embodiment 1, wherein the subject's genome is not edited. 4. The method of any one of embodiments 1-3, wherein the disease or condition comprises pain. 5. The method of embodiment 4, wherein said pain comprises neuropathic pain, inflammatory pain, visceral pain, migraine, erythromycin pain, fibromyalgia pain, idiopathic pain, or somatic pain, or a combination thereof. 6. The method of any one of embodiments 1-3, wherein the disease or condition comprises a channelopathy. 7. The method of embodiment 6, wherein the channelopathy comprises Dravet syndrome, epilepsy syndrome, familial hemiplegic migraine, Ohtahara syndrome, West syndrome, Lennox-Gastaut syndrome, sodium channelotomyopathy, autism, long QT syndrome, Brugada syndrome, or progressive cardiac conduction disease (also known as Lenegret's disease), or a combination thereof. 8. The method of any one of embodiments 1-3, wherein the disease or condition comprises a neurological disease. 9. The method of embodiment 8, wherein the neurological disease is dementia. 10. The method of any one of embodiments 1-3, wherein the disease or condition comprises Alzheimer's disease. 11. The method of any one of embodiments 1-3, wherein the disease or condition comprises Parkinson's disease. 12. The method of any one of embodiments 1-3, wherein the disease or condition comprises Huntington's disease. 13. The method of any one of embodiments 1-3, wherein the disease or condition comprises schizophrenia. 14. The method of any one of embodiments 1-3, wherein the disease or condition comprises amyotrophic lateral sclerosis (ALS). 15. The method of any one of embodiments 1-3, wherein the disease or condition comprises multiple sclerosis. 16. The method of any one of embodiments 1-3, wherein the disease or condition comprises a disease of the central nervous system. 17. The method of any one of embodiments 1-3, wherein the disease or condition comprises an infectious disease. 18. The method of any one of embodiments 1-3, wherein the disease or condition comprises β-thalassemia, fragile X syndrome, centronuclear myopathy, prion disease, Angelman syndrome, Lafora disease, or Alexander disease. 19. The method of any one of the preceding claims, wherein the one or more target molecules comprise DNA. 20. The method of any one of the preceding claims, wherein the one or more target molecules comprise RNA. 21. The method of any one of embodiments 1 to 20, wherein the one or more target molecules comprise a coding region of a gene. 22. The method of any one of embodiments 1 to 21, wherein the one or more target molecules comprise DNA complementary to a non-coding RNA. 23. The method of embodiment 22, wherein the non-coding RNA is associated with neuropathic pain. 24. The method of embodiment 23, wherein the neuropathic pain comprises spinal nerve ligation injury, nerve branch ligation injury, chronic constriction injury, or diabetic neuropathy, or a combination thereof. 25. The method of any one of embodiments 22 to 24, wherein the non-coding RNA comprises SCN9A natural antisense transcript (NAT), Kcna2 antisense RNA, H19, Gm21781, MRAK009713, uc.48+, NONRATT021972, BC168687, Speer7-ps, Uc007pbc.1, XLOC_041439, Mlxipl, Rn50_X_0739.1, CCAT1, rno circ0004058, rno_circRNA_007512, or Egr2 antisense RNA, or a combination thereof. 26. The method of any one of embodiments 1-21, wherein the one or more target molecules comprise a nucleic acid encoding a channel. 27. The method of any one of embodiments 1 to 21, wherein the one or more target molecules comprise a nucleic acid associated with a channel. 28. The method of embodiment 26 or embodiment 27, wherein the channel is an ion channel. 29. The method of embodiment 28, wherein the ion channel is a sodium channel. 30. The method of embodiment 28, wherein the ion channel is a potassium channel. 31. The method of embodiment 28, wherein the ion channel is a calcium channel. 32. The method of embodiment 28, wherein the ion channel is a chloride channel. 33. The method of any one of embodiments 1 to 32, wherein the one or more target molecules are associated with Dravet syndrome, epilepsy syndrome, familial hemiplegic migraine, Ohtahara syndrome, West syndrome, Lennox-Gastaut syndrome, sodium channel myotonia, autism, long QT syndrome, Brugada syndrome, or progressive cardiac conduction disease (also known as Lenegret's disease), or a combination thereof. 34. The method of any one of embodiments 1-33, wherein the one or more target molecules comprise one or more genes in Table 1. 35. The method of any one of embodiments 1-34, wherein the one or more target molecules include SCN9A, SCN10A, or SCN11A, or a combination thereof. 36. The method of any one of embodiments 1 to 35, wherein the one or more target molecules comprises a native antisense transcript of SCN9A. 37. The method of any one of embodiments 1-36, wherein the one or more target molecules are associated with pain. 38. The method of embodiment 37, wherein the pain comprises neuropathic pain, inflammatory pain, visceral pain, migraine, erythromycin pain, fibromyalgia pain, idiopathic pain, or somatic pain, or a combination thereof. 39. The method of any one of embodiments 1-38, wherein the one or more target molecules comprise one or more genes in Table 2. 40. The method of any one of embodiments 1-39, wherein the one or more target molecules are associated with a neurological disease. 41. The method of embodiment 40, wherein the neurological disease comprises dementia. 42. The method of any one of embodiments 1-41, wherein the one or more target molecules are associated with Alzheimer's disease. 43. The method of any one of embodiments 1-42, wherein the one or more target molecules comprise one or more genes in Table 3. 44. The method of any one of embodiments 1-43, wherein the one or more target molecules are associated with Parkinson's disease. 45. The method of any one of embodiments 1 to 44, wherein the one or more target molecules comprise one or more genes selected from the group comprising SNCA, GBA, and LRRK2. 46. ​​The method of any one of embodiments 1-45, wherein the one or more target molecules are associated with Huntington's disease. 47. The method of any one of embodiments 1 to 46, wherein the one or more target molecules are associated with schizophrenia. 48. The method of any one of embodiments 1 to 47, wherein the one or more target molecules comprises GPR52. 49. The method of any one of embodiments 1-48, wherein the one or more target molecules are associated with amyotrophic lateral sclerosis (ALS). 50. The method of any one of embodiments 1 to 49, wherein the one or more target molecules comprise one or more genes selected from the group comprising SOD1, ataxin-2, TDP43, FUS, C9ORF72 and SCA2. 51. The method of any one of embodiments 1-50, wherein the one or more target molecules are associated with multiple sclerosis. 52. The method of any one of the preceding embodiments, wherein the one or more target molecules are associated with a disease of the central nervous system. 53. The method of any one of embodiments 1 to 52, wherein the one or more target molecules comprise one or more genes selected from the group including BFD1, FUS, C9orf72, brain-derived neurotrophic factor, nerve growth factor, neurotrophins, BCL11A, FMR1, DNM2, PrP, UBE3A, GYS1, STING, and GFAP. 54. The method of any one of embodiments 1 to 53, wherein the nucleic acid binding domain binds to at least one of one or more target molecules. 55. The method of any one of embodiments 1 to 53, wherein the nucleic acid binding domain comprises a clustered regularly interspaced short palindromic repeats associated protein (dCas) that has no nuclease activity. 56. The method of embodiment 55, wherein the dCas comprises a mutant Cas protein. 57. The method of embodiment 55, wherein the dCas is a truncated Cas protein. 58. The method of embodiment 55, wherein the dCas is a mutated or truncated Cas protein of Table 4. 59. The method of embodiment 55, wherein the dCas comprises a sequence at least 90% identical to a dCas in Table 4. 60. The method of embodiment 55, wherein the dCas comprises dCas9. 61. The method of embodiment 60, wherein the dCas9 is a truncated or mutated Cas9 protein. 62. The method of embodiment 55, wherein the dCas comprises dCas12. 63. The method of embodiment 62, wherein the dCas12 is a truncated or mutated Cas12 protein. 64. The dCas is Streptococcus pyogenes, Staphylococcus aureus, Campylobacter jejuni, S.thermophilus, S.pneumoniae, Neisseria meningitidis, Corynebacter diphtheriae, Eubacterium ventriosum, Streptococcus pasteurianus, Lactobacillus farciminis, Sphaerochaeta globus, Azospirillum 61. The method of embodiment 60, comprising dCas9 derived from B510, Gluconacetobacter diazotrophicus, Neisseria cinerea, Roseburia intestinalis, Parvibaculum lavamentivorans, Nitratifractor salsuginis DSM 16511, Campylobacter lari CF89-12, or Streptococcus thermophilus LMD-9. 65. The method of embodiment 55, wherein the dCas has a REC2 domain deletion. 66. The method of embodiment 55, wherein the dCas has a REC3 domain deletion. 67. The method of embodiment 55, wherein the dCas has an HNH deletion. 68. The method of embodiment 55, wherein the dCas has a nuclease (NUC) lobe deletion. 69. The method of any one of embodiments 1 to 53, wherein the nucleic acid binding domain comprises a zinc finger protein. 70. The method of any one of embodiments 1 to 53, wherein the nucleic acid binding domain comprises a meganuclease. 71. The method of any one of embodiments 1 to 53, wherein the nucleic acid binding domain comprises a transcription activator-like effector nuclease (TALEN). 72. The method of any one of embodiments 1 to 71, wherein the epigenetic regulatory factor comprises a transcriptional regulatory domain. 73. The method of embodiment 72, wherein the epigenetic regulator comprises a domain having transcriptional repression activity (repressor domain). 74. The method of embodiment 73, wherein the repressor domain comprises a Krueppel-associated box (KRAB) domain (recruiting histone methyltransferases and deacetylases). 75. The method of embodiment 72, wherein the epigenetic regulator comprises a domain having transcriptional activation activity (activation domain). 76. The method of any one of embodiments 1 to 75, wherein the epigenetic regulator comprises KRAB (also known as KOX), SID, MBD2, MBD3, HP1a, the DNMT family (including DNMT1, DNMT3A, DNMT3B, DNMT3L, DNMT2A), Sin3a, Rb, MeCP2, ROM2, AtHD2A, LSD1, SUV39H1, or G9a (EHMT2), or a variant or combination thereof. 77. The method of any one of embodiments 1-76, wherein the epigenetic regulator comprises ZIM3, ZNF554, ZNF264, ZNF324, ZNF354A, ZNF189, ZNF543, ZNP82, ZNF669, ZNF582, KOX1-MeCP2, ZNF30, ZNF680, ZNF331, ZNF33A, ZNF528, ZNF320, ZNF350, ZNF175, ZNF214, ZNF184, ZNF8, ZNF596, KOX1, ZNF37A, ZNF394, ZNF610, ZNF273, ZNF34, ZNF250, ZNF98, ZNF675, ZNF213, NLuc, ZFP28-2, ZNF224, or ZNF257, or a variant or combination thereof. 78. The method of any one of embodiments 1 to 77, wherein the epigenetic regulator comprises VP64, Rta, P16, P65, p300, TET1 catalytic domain, TDG, Ldb1 self-association domain, SAM activator (VP64, p65, HSF1), VPR (VP64, p65, Rta), CD, or SunTag, or a variant or combination thereof. 79. The method of any one of embodiments 1 to 78, wherein the epigenetic regulator comprises a domain that recruits a transcriptional activator, a histone acetyltransferase, a DNA demethylase, a domain that recruits enhancer-associated endogenous Ldb1, a domain that recruits histone methyltransferases and deacetylases, a domain that recruits histone deacetylases, a histone demethylase, a histone methyltransferase, a DNA methyltransferase, an acetylation domain, or a deacetylation domain, or a combination thereof. 80. The epigenetic regulatory factor is VP64 (recruiting transcription activators), p65 (recruiting transcription activators), p300 catalytic domain (histone acetyltransferase), TET1 catalytic domain (DNA demethylase), TDG (DNA demethylase), Ldb1 self-association domain (recruits enhancer-associated endogenous Ldb1), SAM activator (VP64, p65, HSF1) (recruits transcription activators), VPR (VP64, p65, Rt The method of any one of embodiments 1-79, comprising: a) (recruiting transcriptional activators), Sin3a (recruiting histone deacetylases), LSD1 (histone demethylase), SUV39H1 (histone methyltransferase), G9a (EHMT2) (histone methyltransferase), DNMT3a (DNA methyltransferase), or DNMT3a-DNMT3L (DNA methyltransferases), or a combination thereof. 81. The method of any one of embodiments 1 to 80, wherein the epigenetic regulator is linked to the nucleic acid binding domain via a linker at the N-terminus or C-terminus of the nucleic acid binding domain. 82. The method of embodiment 81, wherein the linker is a flexible linker. 83. The linker is selected from the group consisting of (GGS)n, (GGGS)n, (GGGGS)n, (G)n, (EAAAK)n, A(EAAAK)nALEA(EAAAK)nA, PAPAP, AEAAAKEAAAKA, (Ala-Pro)x, LE, GlySer-polyPro(Glyc)-polyPro(Glyc)-polyPro(Glyc), GlySer-polyPro-polyPro(Glyc)-polyPro, GlySer-polyPro-GlySer(G lyc)-polyPro, GlySer-polyPro-polyPro-polyPro, GlySer-polyPro-β2m-polyPro, GlySer-polyPro-β2m-GlySer, polyPro-β2 m-GlySer-β2m-GlySer, GlySer-polyPro-β2m-GlySer-β2m-polyPro, GlySer-polyPro-Ub-GlySer, GlySer-polyPro-ZAG-polyP ro, GlySer-GlySer-ZAG-GlySer-ZAG-polyPro, GlySer(Glyc)-GlySer(Glyc)-polyPro, (G4S)3-cTPR3-(G4S)3, (G4S)3-cTPR6-(G4S)3, (G4S)3-cTPR9-(G4S)3, (G4S)3-cTPR12-(G4S)3, or (G4S)n, where n is independently selected from 1 to 10, x is 10 to 34, and polyPro is IgA1 83. The method of embodiment 81 or embodiment 82, wherein: polyPro(Glyc) is the proline-rich hinge sequence from IgA1 with an embedded potential N-linked glycosylation site (Asn-Ser-Ser); β2m is β2-microglubulin; Ub is ubiquitin; ZAG is Zn-α2-glycoprotein; and cTPRX is a consensus tetratricopeptide repeat sequence having a repeat number of X. 84. The method of any one of embodiments 1 to 80, wherein the nucleic acid binding domain and the epigenetic regulatory factor are linked by a disulfide bond. 85. The method of any one of embodiments 1 to 84, wherein the nucleic acid sequence comprises a nuclear localization sequence (NLS). 86. The method of any one of embodiments 1 to 85, wherein the nucleic acid sequence comprises a promoter. 87. The method of embodiment 86, wherein the promoter comprises a sequence that is at least about 90% identical to a sequence in Table 5. 88. The method of embodiment 86 or embodiment 87, wherein the promoter comprises a nuclear localization sequence. 89. The method of embodiment 85 or embodiment 88, wherein the NLS drives nuclear import of a nucleic acid. 90. The method of any one of embodiments 86 to 89, wherein the promoter drives expression of a nucleic acid binding domain. 91. The method of any one of embodiments 86 to 90, wherein the promoter drives expression of an epigenetic regulatory factor. 92. The method of any one of embodiments 86 to 91, wherein the promoter is the promoter naturally associated with the target molecule. 93. The method of any one of embodiments 86-92, wherein the promoter is a promoter of a gene in Tables 1-3. 94. The method of any one of embodiments 86 to 92, wherein the promoter is the SCN9A promoter or the SCN10A promoter. 95. The method of any one of embodiments 86 to 92, wherein the promoter is a whole neuronal gene promoter. 96. The method of any one of embodiments 86-92, wherein the promoter is a promoter naturally associated with a gene associated with pain. 97. The method according to any one of embodiments 86 to 92, wherein the promoter is a promoter of microtubule-associated protein 2 (MAP-2), a promoter of neuron-specific enolase (NSE), a promoter of choline acetyltransferase (ChAT), a promoter of protein gene product 9.5 (PGP9.5) (also called ubiquitin C-terminal hydrolase 1 (UCHL-1)), a promoter of the gene for human synapsin 1 (hSYN1), a promoter of the NeuN gene (Fox-3, Rbfox3, or hexaribonucleotide-binding protein-3), a promoter of α-calcium / calmodulin-dependent protein kinase II [CaMKIIα], a promoter of the Rheb gene (ras homolog enriched in brain), a promoter of TRKA (tyrosine kinase A), or jET. 98. The method of any one of embodiments 86-92, wherein the promoter is a promoter naturally associated with a gene associated with the channel. 99. The method of any one of embodiments 86 to 92, wherein the promoter is a promoter naturally associated with Dravet syndrome, epilepsy syndrome, familial hemiplegic migraine, Otahara syndrome, West syndrome, Lennox-Gastaut syndrome, sodium channel myotonia, autism, long QT syndrome, Brugada syndrome, or progressive cardiac conduction disease (also known as LeNegret's disease), or a combination thereof. 100. The method of any one of embodiments 86-92, wherein the promoter is a promoter naturally associated with inflammatory pain, visceral pain, migraine, erythromycin pain, fibromyalgia pain, idiopathic pain, or somatic pain, or a combination thereof. 101. The method of any one of embodiments 86-92, wherein the promoter is a promoter naturally associated with a neurological disease. 102. The method of any one of embodiments 86-92, wherein the promoter is a promoter naturally associated with dementia. 103. The method of any one of embodiments 86-92, wherein the promoter is a promoter naturally associated with Alzheimer's disease. 104. The method of any one of embodiments 86-92, wherein the promoter is a promoter naturally associated with Parkinson's disease. 105. The method of any one of embodiments 86-92, wherein the promoter is a promoter naturally associated with ALS. 106. The method of any one of embodiments 86-92, wherein the promoter is a promoter naturally associated with multiple sclerosis. 107. The method of any one of embodiments 86 to 92, wherein the promoter is a promoter naturally associated with a gene associated with a disease of the central nervous system. 108. The method of any one of embodiments 86 to 92, wherein the promoter is a promoter of a gene selected from SNCA, GBA, and LRRK2, SOD1, ataxin-2, SCA2, BFD1, FUS, C9orf72, brain-derived neurotrophic factor, nerve growth factor, neurotrophin, BCL11A, FMR1, DNM2, PrP, UBE3A, GYS1, and GFAP. 109. The method of any one of embodiments 86 to 92, wherein the promoter is a pol II promoter (e.g., Thy1 and H1xb9), a small latency-associated promoter (e.g., from the herpesvirus pseudorabies virus), a cytomegalovirus promoter, SV40, an elongation factor 1-alpha (EF1a) promoter, a cytomegalovirus enhancer / chicken beta-actin (CAG) promoter, or a herpes simplex virus (HSV) promoter. 110. The method of any one of embodiments 86 to 92, wherein the promoter is controlled by a small molecule. 111. The method of embodiment 110, wherein the promoter is a tetracycline-responsive promoter, a glucocorticoid-responsive promoter, a RU-486-responsive promoter, a peroxide-inducible promoter, or a tamoxifen-inducible promoter. 112. The method according to any one of embodiments 86 to 109, wherein expression of the epigenetic regulatory factor and / or transcriptional regulatory domain occurs upon natural or physiological induction of the promoter. 113. The method of embodiment 112, wherein the promoter is induced when a pathology is caused in a subject. 114. The method of embodiment 113, wherein the pathology is injury and / or infection. 115. The method according to embodiment 113 or embodiment 114, wherein the promoter is a galanin promoter or an NF-κB promoter. 116. The method of any one of embodiments 1 to 115, wherein the nucleic acid sequence comprises two or more promoters. 117. The method of any one of embodiments 1 to 116, wherein the nucleic acid sequence comprises a tandem promoter. 118. The method of any one of embodiments 1 to 117, wherein the nucleic acid sequence comprises an enhancer. 119. The method of any one of embodiments 1 to 118, wherein the nucleic acid sequence comprises an intron. 120. The method of any one of embodiments 1 to 119, wherein the nucleic acid comprises an inverted terminal repeat (ITR). 121. The method of any one of embodiments 1 to 120, wherein the nucleic acid comprises a terminator sequence. 122. The method of any one of embodiments 1 to 121, comprising administering one or more guide RNA sequences to the subject. 123. The method of embodiment 122, wherein the one or more guide RNA sequences are selected from the sequences in Table 6. 124. The method of embodiment 122 or embodiment 123, wherein the one or more guide RNA sequences bind to one or more target molecules. 125. The method of embodiment 124, wherein the one or more target molecules are two, three, four, or five target molecules. 126. The method of any one of embodiments 122-125, wherein the one or more guide RNA sequences are two, three, four, or five guide RNA sequences. 127. The method of embodiment 126, comprising at least two guide RNA sequences, wherein said at least two guide RNA sequences are different. 128. The method according to any one of embodiments 122 to 127, wherein the at least one or more guide RNA sequences target a promoter according to embodiments 86 to 115, creating a regulatory feedback loop to control expression levels. 129. The method of any one of embodiments 1 to 128, wherein the nucleic acid is delivered as a naked (or unmodified) nucleic acid. 130. The method of any one of embodiments 1 to 128, wherein the nucleic acid is delivered complexed with a cationic molecule. 131. The method of any one of embodiments 1 to 128, wherein the nucleic acid is delivered to the subject via a vehicle (e.g., a viral delivery vehicle (e.g., a retroviral vector, a lentiviral vector, or an adenoviral vector), a liposome, a nanoparticle, or an exosome). 132. The method of embodiment 131, wherein the vehicle is a viral delivery vehicle. 133. The method of embodiment 132, wherein the viral delivery vehicle is a retroviral vector, a lentiviral vector, or an adenoviral vector. 134. The method of embodiment 133, wherein the viral delivery vehicle is a recombinant adeno-associated virus (AAV). 135. The method of embodiment 134, wherein the AAV is AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAVhu68, AAVrh.10, AAVrh74 or AAVDJ, or a combination thereof. 136. The method of embodiment 135, wherein the AAV is AAV9. 137. The method of any one of embodiments 134 to 136, wherein the AAV has a recombinant capsid. 138. The method of embodiment 137, wherein the recombinant capsid encodes a targeting moiety. 139. The method of any one of embodiments 1 to 137, wherein the nucleic acid comprises a sequence encoding a targeting moiety. 140. The method of embodiment 131, wherein the vehicle is a liposome, lipid nanoparticle, nanocapsule, or exosome. 141. The method of embodiment 140, wherein the vehicle comprises or is linked to a targeting moiety. 142. The method of any one of embodiments 138-141, wherein the targeting moiety targets cells containing the target molecule in the subject. 143. The method of any one of embodiments 138 to 142, wherein the targeting moiety binds to a peptide product of the target molecule. 144. The method of embodiment 142, wherein the target molecule is present on a target cell. 145. The method of embodiment 143, wherein the target cells are associated with a disease or condition of the subject. 146. The method of any one of embodiments 138 to 145, wherein the targeting moiety comprises a peptide. 147. The method of embodiment 146, wherein the peptide comprises JNJ63955, m3-Huwentoxin-IV, Phlotoxin1 (PhlTx1), Protoxin-II (ProTx-II), or Ceratotoxin-1 (CcoTx1), or a variant thereof. 148. The method of embodiment 146, wherein the peptide comprises a sequence at least about 90% identical to a peptide in Table 7. 149. The method of any one of embodiments 1-147, further comprising administering an additional therapeutic agent to the subject. 150. The method of any one of embodiments 1-149, wherein the nucleic acid is administered via lumbar intrathecal puncture, epidural, intravenous, transdermal, intranasal, oral, mucosal, intracisternal, or intraganglionic administration. 151. A nucleic acid sequence encoding a nucleic acid binding domain and an epigenetic regulator that regulates transcription of one or more target molecules. 152. The nucleic acid sequence according to embodiment 151, wherein the regulation of transcription of the one or more target molecules is transient. 153. The nucleic acid sequence of embodiment 151, wherein the subject's genome is not edited. 154. The nucleic acid sequence according to any one of embodiments 151 to 153, wherein the one or more target molecules comprise DNA. 155. The nucleic acid sequence according to any one of embodiments 151 to 154, wherein the one or more target molecules comprises RNA. 156. The nucleic acid sequence according to any one of embodiments 151 to 155, wherein the one or more target molecules comprise a coding region of a gene. 157. The nucleic acid sequence according to any one of embodiments 151 to 156, wherein the one or more target molecules comprises DNA complementary to a non-coding RNA. 158. The nucleic acid sequence of embodiment 157, wherein the non-coding RNA is associated with neuropathic pain. 159. The nucleic acid sequence of embodiment 158, wherein the neuropathic pain comprises spinal nerve ligation injury, nerve branch ligation injury, chronic constriction injury, or diabetic neuropathy, or a combination thereof. 160. The nucleic acid sequence of any one of embodiments 157 to 159, wherein the non-coding RNA comprises SCN9A natural antisense transcript (NAT), Kcna2 antisense RNA, H19, Gm21781, MRAK009713, uc.48+, NONRATT021972, BC168687, Speer7-ps, Uc007pbc.1, XLOC_041439, Mlxipl, Rn50_X_0739.1, CCAT1, rno circ0004058, rno_circRNA_007512, or Egr2 antisense RNA, or a combination thereof. 161. The nucleic acid sequence according to any one of embodiments 151 to 160, wherein the one or more target molecules comprises a nucleic acid encoding a channel. 162. The nucleic acid sequence according to any one of embodiments 151 to 160, wherein the one or more target molecules comprises a nucleic acid associated with a channel. 163. The nucleic acid sequence according to embodiment 161 or embodiment 162, wherein the channel is an ion channel. 164. The nucleic acid sequence of embodiment 163, wherein the ion channel is a sodium channel. 165. The nucleic acid sequence according to embodiment 163, wherein the ion channel is a potassium channel. 166. The nucleic acid sequence according to embodiment 163, wherein the ion channel is a calcium channel. 167. The nucleic acid sequence of embodiment 163, wherein the ion channel is a chloride channel. 168. The nucleic acid sequence of any one of embodiments 151 to 167, wherein the one or more target molecules are associated with Dravet syndrome, epilepsy syndrome, familial hemiplegic migraine, Otahara syndrome, West syndrome, Lennox-Gastaut syndrome, sodium channel myotonia, autism, long QT syndrome, Brugada syndrome, or progressive cardiac conduction disease (also known as Lenegret disease), or a combination thereof. 169. The nucleic acid sequence according to any one of embodiments 151 to 168, wherein the one or more target molecules comprise one or more genes of Table 1. 170. The nucleic acid sequence of any one of embodiments 151 to 169, wherein the one or more target molecules comprises SCN9A, SCN10A, or SCN11A, or a combination thereof. 171. The nucleic acid sequence according to any one of embodiments 151 to 170, wherein the one or more target molecules comprises a naturally occurring antisense transcript of SCN9A. 172. The nucleic acid sequence according to any one of embodiments 151 to 171, wherein the one or more target molecules are associated with pain. 173. The nucleic acid sequence of embodiment 172, wherein the pain comprises neuropathic pain, inflammatory pain, visceral pain, migraine, erythromycin pain, fibromyalgia pain, idiopathic pain, or somatic pain, or a combination thereof. 174. The nucleic acid sequence of any one of embodiments 151 to 173, wherein the one or more target molecules comprise one or more genes of Table 2. 175. The nucleic acid sequence according to any one of embodiments 151 to 174, wherein the one or more target molecules are associated with a neurological disease. 176. The nucleic acid sequence of embodiment 175, wherein the neurological disease comprises dementia. 177. The nucleic acid sequence according to any one of embodiments 151 to 176, wherein the one or more target molecules are associated with Alzheimer's disease. 178. The nucleic acid sequence of any one of embodiments 151 to 177, wherein the one or more target molecules comprise one or more genes of Table 3. 179. The nucleic acid sequence according to any one of embodiments 151 to 178, wherein the one or more target molecules are associated with Parkinson's disease. 180. The nucleic acid sequence according to any one of embodiments 151 to 179, wherein the one or more target molecules comprise one or more genes selected from the group comprising SNCA, GBA, and LRRK2. 181. The nucleic acid sequence according to any one of embodiments 151 to 180, wherein the one or more target molecules are associated with Huntington's disease. 182. The nucleic acid sequence according to any one of embodiments 151 to 181, wherein the one or more target molecules are associated with schizophrenia. 183. The nucleic acid sequence according to any one of embodiments 151 to 182, wherein the one or more target molecules comprises GPR52. 184. The nucleic acid sequence of any one of embodiments 151 to 183, wherein the one or more target molecules are associated with amyotrophic lateral sclerosis (ALS). 185. The nucleic acid sequence according to any one of embodiments 151 to 184, wherein the one or more target molecules comprise one or more genes selected from the group comprising SOD1, ataxin-2, TDP43, C9ORF72, FUS and SCA2. 186. The nucleic acid sequence according to any one of embodiments 151 to 185, wherein the one or more target molecules are associated with multiple sclerosis. 187. The nucleic acid sequence according to any one of embodiments 151 to 186, wherein the one or more target molecules are associated with a disease of the central nervous system. 188. The nucleic acid sequence of any one of embodiments 151 to 187, wherein the one or more target molecules comprise one or more genes selected from the group comprising BFD1, C9orf72, FUS, SOD1, TPD43, brain-derived neurotrophic factor, nerve growth factor, neurotrophins, BCL11A, FMR1, DNM2, PrP, UBE3A, GYS1, STING, and GFAP. 189. The nucleic acid sequence according to any one of embodiments 151 to 188, wherein the nucleic acid binding domain binds to at least one of the one or more target molecules. 190. The nucleic acid sequence of any one of embodiments 151 to 189, wherein the nucleic acid binding domain comprises a clustered regularly interspaced short palindromic repeats associated protein (dCas) that has no nuclease activity. 191. The nucleic acid sequence of embodiment 190, wherein the dCas comprises a mutated Cas protein. 192. The nucleic acid sequence of embodiment 190, wherein the dCas is a truncated Cas protein. 193. The nucleic acid sequence of embodiment 190, wherein the dCas is a mutated or truncated Cas protein of Table 4. 194. The nucleic acid sequence of embodiment 190, wherein the dCas comprises a sequence at least 90% identical to a dCas in Table 4. 195. The nucleic acid sequence of embodiment 190, wherein the dCas comprises dCas9. 196. The nucleic acid sequence of embodiment 195, wherein the dCas9 is a truncated or mutated Cas9 protein. 197. The nucleic acid sequence of embodiment 195, wherein the dCas comprises dCas12. 198. The nucleic acid sequence of embodiment 197, wherein the dCas12 is a truncated or mutated Cas12 protein. 199. The dCas is Streptococcus pyogenes, Staphylococcus aureus, Campylobacter jejuni, S.thermophilus, S.pneumoniae, Neisseria meningitidis, Corynebacter diphtheriae, Eubacterium ventriosum, Streptococcus pasteurianus, Lactobacillus farciminis, Sphaerochaeta globus, Azospirillum 191. The nucleic acid sequence of embodiment 190, comprising dCas9 from B510, Gluconacetobacter diazotrophicus, Neisseria cinerea, Roseburia intestinalis, Parvibaculum lavamentivorans, Nitratifractor salsuginis DSM 16511, Campylobacter lari CF89-12, or Streptococcus thermophilus LMD-9. 200. The nucleic acid sequence of embodiment 190, wherein the dCas has a REC2 domain deletion. 201. The nucleic acid sequence of embodiment 190, wherein the dCas has a REC3 domain deletion. 202. The nucleic acid sequence of embodiment 190, wherein the dCas has an HNH deletion. 203. The nucleic acid sequence of embodiment 190, wherein the dCas has a nuclease (NUC) lobe deletion. 204. The nucleic acid sequence according to any one of embodiments 151 to 189, wherein the nucleic acid binding domain comprises a zinc finger protein. 205. The nucleic acid sequence according to any one of embodiments 151 to 189, wherein the nucleic acid binding domain comprises a meganuclease. 206. The nucleic acid sequence of any one of embodiments 151-189, wherein the nucleic acid binding domain comprises a Transcription Activator-Like Effector Nuclease (TALEN). 207. The nucleic acid sequence according to any one of embodiments 151 to 206, wherein the epigenetic regulator comprises a transcriptional regulator domain. 208. The nucleic acid sequence according to embodiment 207, wherein the epigenetic regulator comprises a domain having transcriptional repression activity (repressor domain). 209. The nucleic acid sequence of embodiment 208, wherein the repressor domain comprises a Krueppel-associated box (KRAB) domain (recruitment of histone methyltransferases and deacetylases). 210. The nucleic acid sequence according to embodiment 207, wherein the epigenetic regulator comprises a domain having transcriptional activation activity (activator domain). 211. The nucleic acid sequence according to any one of embodiments 151 to 210, wherein the epigenetic regulator comprises KRAB (also called KOX), SID, MBD2, MBD3, HP1a, DNMT family (including DNMT1, DNMT3A, DNMT3B, DNMT3L, DNMT2A), Sin3a, Rb, MeCP2, ROM2, AtHD2A, LSD1, SUV39H1, or G9a (EHMT2), or a variant or combination thereof. 212. The epigenetic regulator is ZIM3, ZNF554, ZNF264, ZNF324, ZNF354A, ZNF189, ZNF543, ZNP82, ZNF669, ZNF582, KOX1-MeCP2, ZNF30, ZNF680, ZNF331, ZNF33A, ZNF528, ZNF320, ZNF350, ZNF175, ZNF214, ZNF 184, ZNF8, ZNF596, KOX1, ZNF37A, ZNF394, ZNF610, ZNF273, ZNF34, ZNF250, ZNF98, ZNF675, ZNF213, Nluc, ZFP28-2, ZNF224, or ZNF257, or a variant or combination thereof. 213. The nucleic acid sequence of any one of embodiments 151 to 212, wherein the epigenetic regulator comprises VP64, Rta, P16, P65, p300, TET1 catalytic domain, TDG, Ldb1 self-association domain, SAM activator (VP64, p65, HSF1), VPR (VP64, p65, Rta), CD, or SunTag, or a variant or combination thereof. 214. The nucleic acid sequence of any one of embodiments 151 to 213, wherein the epigenetic regulator comprises a domain that recruits a transcriptional activator, a histone acetyltransferase, a DNA demethylase, a domain that recruits enhancer-associated endogenous Ldb1, a domain that recruits histone methyltransferases and deacetylases, a domain that recruits histone deacetylases, a histone demethylase, a histone methyltransferase, a DNA methyltransferase, an acetylation domain, or a deacetylation domain, or a combination thereof. 215. The epigenetic regulator is VP64 (recruiting transcription activators), p65 (recruiting transcription activators), p300 catalytic domain (histone acetyltransferase), TET1 catalytic domain (DNA demethylase), TDG (DNA demethylase), Ldb1 self-association domain (recruits enhancer-associated endogenous Ldb1), SAM activator (VP64, p65, HSF1) (recruits transcription activators), VPR (VP64, p65, Rta) 215. The nucleic acid sequence of any one of embodiments 151-214, comprising Sin3a (recruits transcriptional activators), Sin3a (recruits histone deacetylases), LSD1 (histone demethylase), SUV39H1 (histone methyltransferase), G9a (EHMT2) (histone methyltransferase), DNMT3a (DNA methyltransferase), or DNMT3a-DNMT3L (DNA methyltransferases), or a combination thereof. 216. The nucleic acid sequence according to any one of embodiments 151 to 215, wherein the epigenetic regulator is linked to the nucleic acid binding domain at the N-terminus or C-terminus of the nucleic acid binding domain via a linker. 217. The nucleic acid sequence according to embodiment 216, wherein the linker is a flexible linker. 218. The linker is selected from the group consisting of (GGS)n, (GGGS)n, (GGGGS)n, (G)n, (EAAAK)n, A(EAAAK)nALEA(EAAAK)nA, PAPAP, AEAAAKEAAAKA, (Ala-Pro)x, LE, GlySer-polyPro(Glyc)-polyPro(Glyc)-polyPro(Glyc), GlySer-polyPro-polyPro(Glyc)-polyPro, GlySer-polyPro-GlySer(G lyc)-polyPro, GlySer-polyPro-polyPro-polyPro, GlySer-polyPro-β2m-polyPro, GlySer-polyPro-β2m-GlySer, polyPro-β2m -GlySer-β2m-GlySer, GlySer-polyPro-β2m-GlySer-β2m-polyPro, GlySer-polyPro-Ub-GlySer, GlySer-polyPro-ZAG-polyPro , GlySer-GlySer-ZAG-GlySer-ZAG-polyPro, GlySer(Glyc)-GlySer(Glyc)-polyPro, (G4S)3-cTPR3-(G4S)3, (G4S)3-cTPR6-(G4S)3, (G4S)3-cTPR9-(G4S)3, (G4S)3-cTPR12-(G4S)3, or (G4S)n, where n is independently selected from 1 to 10, x is 10 to 34, and polyPro is a polypeptide from IgA1. 218. The nucleic acid sequence of embodiment 216 or embodiment 217, wherein: polyPro(Glyc) is a proline-rich hinge sequence from IgA1 with an embedded potential N-linked glycosylation site (Asn-Ser-Ser); β2m is β2-microglubulin; Ub is ubiquitin; ZAG is Zn-α2-glycoprotein; and cTPRX is a consensus tetratricopeptide repeat sequence having a repeat number of X. 219. The nucleic acid sequence according to any one of embodiments 151 to 218, wherein the nucleic acid binding domain and the epigenetic regulator are linked by a disulfide bond. 220. The nucleic acid sequence of any one of embodiments 151 to 219, wherein the nucleic acid sequence comprises a nuclear localization sequence (NLS). 221. The nucleic acid sequence according to any one of embodiments 151 to 220, wherein the nucleic acid sequence comprises a promoter. 222. The nucleic acid sequence of embodiment 221, wherein the promoter comprises a sequence that is at least about 90% identical to a sequence in Table 5. 223. The nucleic acid sequence of embodiment 221 or embodiment 222, wherein the promoter comprises a nuclear localization sequence. 224. The nucleic acid sequence of embodiment 220 or embodiment 223, wherein the NLS drives nuclear import of the nucleic acid. 225. The nucleic acid sequence according to any one of embodiments 221 to 224, wherein the promoter drives expression of the nucleic acid binding domain. 226. The nucleic acid sequence according to any one of embodiments 221 to 225, wherein the promoter drives expression of an epigenetic regulatory factor. 227. The nucleic acid sequence according to any one of embodiments 221 to 226, wherein the promoter is the promoter naturally associated with the target molecule. 228. The nucleic acid sequence according to any one of embodiments 221-227, wherein the promoter is a promoter of a gene of Tables 1-3. 229. The nucleic acid sequence according to any one of embodiments 221 to 227, wherein the promoter is the SCN9A promoter or the SCN10A promoter. 230. The nucleic acid sequence according to any one of embodiments 221 to 227, wherein the promoter is a global neurogenic promoter. 231. The nucleic acid sequence according to any one of embodiments 221 to 227, wherein the promoter is a promoter naturally associated with a gene associated with pain. 232. The nucleic acid sequence according to any one of embodiments 221 to 227, wherein the promoter is a promoter of microtubule-associated protein 2 (MAP-2), a promoter of neuron-specific enolase (NSE), a promoter of choline acetyltransferase (ChAT), a promoter of protein gene product 9.5 (PGP9.5) (also called ubiquitin C-terminal hydrolase 1 (UCHL-1)), a promoter of the gene for human synapsin 1 (hSYN1), a promoter of the NeuN gene (Fox-3, Rbfox3, or hexaribonucleotide-binding protein-3), a promoter of α-calcium / calmodulin-dependent protein kinase II [CaMKIIα], a promoter of the Rheb gene (ras homolog enriched in brain), a TRKA promoter (tyrosine kinase A), or jET. 233. The nucleic acid sequence according to any one of embodiments 221-227, wherein the promoter is a promoter naturally associated with a gene associated with the channel. 234. The nucleic acid sequence of any one of embodiments 221 to 227, wherein the promoter is a promoter naturally associated with Dravet syndrome, epilepsy syndrome, familial hemiplegic migraine, Otahara syndrome, West syndrome, Lennox-Gastaut syndrome, sodium channel myotonia, autism, long QT syndrome, Brugada syndrome, or progressive cardiac conduction disease (also known as Lenegret's disease), or a combination thereof. 235. The nucleic acid sequence of any one of embodiments 221 to 227, wherein the promoter is a promoter naturally associated with inflammatory pain, visceral pain, migraine, erythema pain, fibromyalgia, idiopathic pain, or somatic pain, or a combination thereof. 236. The nucleic acid sequence according to any one of embodiments 221 to 227, wherein the promoter is a promoter naturally associated with a neurological disease. 237. The nucleic acid sequence according to any one of embodiments 221 to 227, wherein the promoter is a promoter naturally associated with dementia. 238. The nucleic acid sequence according to any one of embodiments 221-227, wherein the promoter is a promoter naturally associated with Alzheimer's disease. 239. The nucleic acid sequence according to any one of embodiments 221 to 227, wherein the promoter is a promoter naturally associated with Parkinson's disease. 240. The nucleic acid sequence according to any one of embodiments 221 to 227, wherein the promoter is a promoter naturally associated with ALS. 241. The nucleic acid sequence according to any one of embodiments 221 to 227, wherein the promoter is a promoter naturally associated with multiple sclerosis. 242. The nucleic acid sequence according to any one of embodiments 221 to 227, wherein the promoter is a promoter naturally associated with a gene associated with a disease of the central nervous system. 243. The nucleic acid sequence according to any one of embodiments 221 to 227, wherein the promoter is a promoter of a gene selected from SNCA, GBA, and LRRK2, SOD1, ataxin-2, SCA2, BFD1, FUS, TDP43, C9orf72, brain-derived neurotrophic factor, nerve growth factor, neurotrophins, BCL11A, FMR1, DNM2, PrP, UBE3A, GYS1, and GFAP. 244. The nucleic acid sequence according to any one of embodiments 221 to 227, wherein the promoter is a pol II promoter (e.g., Thy1 and H1xb9), a mini-latency associated promoter (e.g., from the herpesvirus pseudorabies virus), a cytomegalovirus promoter, SV40, an elongation factor 1-alpha (EF1a) promoter, a cytomegalovirus enhancer / chicken beta-actin (CAG) promoter, or a herpes simplex virus (HSV) promoter. 245. The nucleic acid sequence according to any one of embodiments 221 to 227, wherein the promoter is controlled by a small molecule. 246. The nucleic acid sequence according to embodiment 245, wherein the promoter is a tetracycline-responsive promoter, a glucocorticoid-responsive promoter, a RU-486-responsive promoter, a peroxide-inducible promoter, or a tamoxifen-inducible promoter. 247. The nucleic acid sequence according to any one of embodiments 221 to 244, wherein expression of the epigenetic regulatory factor and / or the transcriptional regulatory domain occurs upon natural or physiological induction of the promoter. 248. The nucleic acid sequence of embodiment 247, wherein the promoter is induced when a pathology is caused in a subject. 249. The nucleic acid sequence according to embodiment 248, wherein the pathology is injury and / or infection. 250. The nucleic acid sequence according to embodiment 248 or embodiment 249, wherein the promoter is a galanin promoter or an NF-κB promoter. 251. The nucleic acid sequence according to any one of embodiments 151 to 250, wherein the nucleic acid sequence comprises two or more promoters. 252. The nucleic acid sequence according to any one of embodiments 151 to 251, wherein the nucleic acid sequence comprises a tandem promoter. 253. The nucleic acid sequence according to any one of embodiments 151 to 252, wherein the nucleic acid sequence comprises an enhancer. 254. The nucleic acid sequence of any one of embodiments 151 to 253, wherein the nucleic acid sequence comprises an intron. 255. The nucleic acid sequence of any one of embodiments 151 to 254, wherein the nucleic acid comprises an inverted terminal repeat (ITR). 256. The nucleic acid sequence according to any one of embodiments 151 to 255, wherein the nucleic acid comprises a terminator sequence. 257. The nucleic acid sequence according to any one of embodiments 151 to 256, comprising administering to said subject one or more guide RNA sequences. 258. The nucleic acid sequence of embodiment 257, wherein the one or more guide RNA sequences are selected from the sequences in Table 6. 259. The nucleic acid sequence of embodiment 257 or embodiment 258, wherein the one or more guide RNA sequences bind to one or more of the target molecules. 260. The nucleic acid sequence of embodiment 259, wherein the one or more target molecules are two, three, four, or five target molecules. 261. The nucleic acid sequence of any one of embodiments 257 to 260, wherein the one or more guide RNA sequences are 2, 3, 4, or 5 guide RNA sequences. 262. The nucleic acid sequence of embodiment 261, comprising at least two guide RNA sequences, wherein said at least two guide RNA sequences are different. 263. The nucleic acid sequence according to any one of embodiments 257 to 262, wherein the at least one or more guide RNA sequences create a regulatory feedback loop to control the expression level and target a promoter according to embodiments 221 to 252. 264. The nucleic acid sequence according to any one of embodiments 151 to 263, wherein the nucleic acid is delivered as a naked (or unmodified) nucleic acid. 265. The nucleic acid sequence according to any one of embodiments 151 to 263, wherein the nucleic acid is delivered complexed with a cationic molecule. 266. The nucleic acid sequence of any one of embodiments 151 to 263, wherein the nucleic acid is delivered to the subject via a vehicle (e.g., a viral delivery vehicle (e.g., a retroviral vector, a lentiviral vector, or an adenoviral vector), a liposome, a nanoparticle, or an exosome). 267. The nucleic acid sequence according to embodiment 266, wherein the vehicle is a viral delivery vehicle. 268. The nucleic acid sequence of embodiment 267, wherein the viral delivery vehicle is a retroviral vector, a lentiviral vector, or an adenoviral vector. 269. The nucleic acid sequence of embodiment 268, wherein the viral delivery vehicle is a recombinant adeno-associated virus (AAV). 270. The nucleic acid sequence of embodiment 269, wherein the AAV is AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAVhu68, AAVrh.10, AAVrh74 or AAVDJ, or a combination thereof. 271. The nucleic acid sequence of embodiment 269, wherein the AAV is AAV9. 272. The nucleic acid sequence of any one of embodiments 269 to 271, wherein the AAV has a recombinant capsid. 273. The nucleic acid sequence of embodiment 272, wherein the recombinant capsid encodes a targeting moiety. 274. The nucleic acid sequence according to any one of embodiments 151 to 273, wherein the nucleic acid comprises a sequence encoding a targeting moiety. 275. The nucleic acid sequence according to embodiment 266, wherein the vehicle is a liposome, lipid nanoparticle, nanocapsule, or exosome. 276. The nucleic acid sequence according to embodiment 275, wherein the vehicle comprises or is linked to a targeting moiety. 277. The nucleic acid sequence of any one of embodiments 273 to 276, wherein the targeting moiety targets cells containing the target molecule in a subject. 278. The nucleic acid sequence according to any one of embodiments 273 to 277, wherein the targeting moiety binds to a peptide product of the target molecule. 279. The nucleic acid sequence of embodiment 278, wherein the target molecule is present on a target cell. 280. The nucleic acid sequence of embodiment 279, wherein the target cells are associated with a disease or condition of the subject. 281. The nucleic acid sequence according to any one of embodiments 273 to 280, wherein the targeting moiety comprises a peptide. 282. The nucleic acid sequence of embodiment 281, wherein the peptide comprises JNJ63955, m3-Huwentoxin-IV, Phlotoxin1 (PhlTx1), Protoxin-II (ProTx-II), or Ceratotoxin-1 (CcoTx1), or a variant thereof. 283. The nucleic acid sequence of embodiment 281, wherein the peptide comprises a sequence at least about 90% identical to a peptide of Table 7. 284. A combination comprising the nucleic acid sequence of any one of embodiments 151 to 283 and an additional therapeutic agent.

[0130] Specific Definitions Percent sequence identity (%) with respect to a reference polypeptide or polynucleotide sequence is defined as the percentage of amino acid or nucleotide residues in a candidate sequence that are identical to the amino acid or nucleotide residues in the reference polypeptide or polynucleotide sequence, without considering any conservative substitutions as part of sequence identity, after aligning sequences and introducing gaps as necessary to achieve maximum sequence identity percentage. Alignment for determining percent amino acid sequence identity can be achieved in various known ways, for example, using publicly available computer software such as BLAST, BLAST-2, ALIGN, or Megalign (DNASTAR) software. Appropriate parameters for aligning sequences can be determined, including the algorithm required to achieve maximum alignment over the entire length of the sequences to be compared. However, for the purposes herein, percent amino acid or polynucleotide sequence identity values ​​are generated using sequence comparison computer program ALIGN-2. The ALIGN-2 sequence comparison computer program was written by Genentech, Inc., and the source code, along with user documentation, has been filed with the US Copyright Office, Washington DC, 20559, and is registered under US Copyright Registration No. TXU510087. The ALIGN-2 program is publicly available from Genentech, Inc., South San Francisco, Calif., or can be compiled from the source code. The ALIGN-2 program should be compiled for use on UNIX operating systems, including Digital UNIX V4.0D. All sequence comparison parameters are set by the ALIGN-2 program and are not altered.

[0131] When ALIGN-2 is used for amino acid or polynucleotide sequence comparison, the amino acid or polynucleotide sequence identity percentage of a given sequence A to a given sequence B (alternatively, it can be expressed as a given sequence A having or containing a certain sequence identity percentage to a given sequence B) is calculated as follows: 100 x fraction X / Y, where X is the number of amino acid residues scored as perfect matches by the sequence alignment program ALIGN-2 in the alignment of A and B in the program, and Y is the total number of amino acid residues in B. It is understood that if the length of amino acid sequence A is not equal to the length of amino acid sequence B, the amino acid sequence identity percentage of A to B is not equal to the amino acid sequence identity percentage of B to A. Unless specifically indicated otherwise, all amino acid sequence identity percentage values ​​used herein are obtained using the ALIGN-2 computer program as described in the immediately preceding paragraph.

[0132] In some embodiments, the term "about" means within 10% of the stated amount. For example, a peptide that contains about 80% identity to a reference peptide may contain 72%-88% identity to the reference peptide sequence. EXAMPLES

[0133] The following examples are illustrative of the embodiments described herein and should not be construed as limiting the scope of the present disclosure. To the extent that specific materials are mentioned, they are for illustrative purposes only and are not intended to be limiting. Those skilled in the art can develop equivalent means or reactants without departing from the scope of the present disclosure and without the exercise of the inventive capacity.

[0134] Example 1: Na V 1.7 Preparation of Compositions Containing Binding Peptides Peptide binders of sodium channels are known in the art. For example, venomous animals such as spiders produce molecules in their venom that bind to and inhibit sodium channels in their prey. In particular, spider venom peptides such as Protoxin-II (ProTx-II), Huwentoxin-IV (HwTx-IV), m3-Huwentoxin-IV (variant m3-HwTX-IV), Ceratotoxin-1 (CcoTx1), and Phlotoxin1 (PhlTx1) bind to and inhibit sodium channels in their prey. V Combined into 1.7.

[0135] In this experiment, each of the following peptides are conjugated to the surface of different AAV9 packaging mCherry reporter constructs in Table 7: ProTx-II, ProTx-III, ProTx-II variant JNJ63955, HwTx-IV, variant m3-HwTx-IV, CcoTx1 variant 2670, PhlTx1 variant D7A-PhlTx1, and JzTx-V variant AM-0422. V 1.7 Binding ligands have been established, and Na V The peptides were chosen to represent a diverse selection of peptides with different sequences and differential interactions with the 1.7 channel. Peptide binding to AAV is achieved by both a C-terminal NHS modification of the peptide and an N-terminal benzoyl NH modification that binds to nine lysine residues on the AAV9 surface. Cross-linking the peptides at the N- and C-termini indicates that at least one of them is a Na V 1.7 increases the likelihood of maintaining the peptide conformation for binding.

[0136] Example 2: Na V 1.7 Preparation of compositions encoding binding peptides Since attaching peptides to AAV9 is not an economically viable option in terms of scaled-up production for therapeutic applications, sequences encoding the peptides of Example 1 are integrated into the viral genome and expressed on viral protein loops. The AAV viral capsid is composed of three proteins: VP1, VP2, and VP3. The N-terminus of the VP2 capsid protein accepts large peptide insertions as well as proteins, and therefore accepts insertion of target peptides with VP2 retargeting vector tropism. Another potential site for inserting these target peptides is found in VP3.

[0137] In the first experiment, the sequences encoding the peptides of Example 1 are cloned in frame within -GS-flanked linkers for expression in different loops of the AAV9 capsid. In this example, five insertion positions are included: 453, 447, 588, 589, and downstream of the N-terminal methionine of the VP2 start codon. Transduction efficiency is evaluated using the Na V 1.7 As determined by mCherry reporter expression in the expressing cell lines HuH7 and Neuro2a.

[0138] Quality assessment tests are performed to ensure that the surface modification does not adversely affect the packaging and capsid structure. This assessment focuses on viral titer, since yield is usually the trait most affected after capsid modification. Titer is measured by RT-qPCR. The ratio of viral proteins VP1, VP2, and VP3 is analyzed by protein gel.

[0139] Example 3: In vivo biodistribution and immunogenicity Na as described in Example 2 V 1.7 Unmodified AAV9-mCherry and AAV9 modified with a sequence encoding a binding peptide were incubated with a low viral titer load of 1 10 11 Inject IT into male C57BL / 6 mice (n=6 / group) with vg / mouse. Lower titers transduce larger numbers of neurons while higher titers transduce larger numbers of neurons. V1.7 results in better differentiation of transduction efficacy in expressing cells. Organs (brain, spinal cord, dorsal root ganglion, eye, lung, heart, liver, spleen, and gonads) and blood are collected for biodistribution and immunological analysis. Importantly, transduction of sensory neurons in the DRG results in better differentiation of Na V As this is important to ensure the validity of future studies focused on the inhibition of 1.7, a subset of neurons in the DRG will be transduced via RNAscope with mCherry and Na V 1.7 are analyzed simultaneously. To define peptidergic and non-peptidergic nociceptor subpopulations, probes for calcitonin gene-related peptide (CGRP) and P2X purinergic ion channel type 3 receptor (P2X3R) are also used. Viral genomes are also quantified via RT-qPCR. Viral tropism, as indicated by mCherry expression, is determined from images captured by confocal microscopy and RT-qPCR.

[0140] Example 4: Na V 1.7 Design of promoter region To determine specificity, different Na V 1.7 Promoters (#1, #2, and #7) were V Investigate the expression of mCherry in Nav1.7-expressing cell lines and, as a negative control, in non-Nav1.7-expressing cell lines.

[0141] Minimal Human Na V 1.7 Design of the promoter region: The promoters tested vary in length between 700 bp and 1.2 kbp. Promoters of various lengths will be tested to allow their packaging into a single AAV. In addition, Na V 1.7 Gene enhancers are tested even if they are not part of the promoter region (https: / / epd.epfl.ch / index.php). These new sequences are cloned in the mCherry expression vector, CMV-free, as previously performed.

[0142] Human Na V 1.7 In vitro testing of promoters: All cloned sequences will be tested in human cell lines and mCherry expression will be compared to vectors carrying the CMV promoter. V 1.7 Expression lines HuH7 and IMR-90 are utilized. The negative control is non-Na V 1.7 expressing cell line MCF-7. RT-qPCR, FACS sorting, and imaging analysis are performed to determine the expression (activity and intensity) levels of the transgenes using these promoters.

[0143] Human Na V 1.7 iPCS test of promoters: specific promoters (Na V mCherry expression (high expression in MCF-7 expressing cells and no expression in MCF-7 negative control) is tested in nociceptor-like iPCS cells. RT-qPCR, FACS sorting, and imaging analysis are performed to determine the expression (activity and intensity) levels of mCherry using these promoters.

[0144] In Figure 3, seven NaCl molecules drive the expression of mCherry. V 1.7 Promoter, high Na V 1.7 was transfected into the expressing human cell line HuH7. The three promoters had high transgene expression.

[0145] Example 5: Human Na in mice to determine biodistribution V 1.7 Pilot Testing of Promoters This study was carried out in mice to evaluate the efficacy of the human Na V 1.7 To determine the activity and biodistribution of the promoter in mice. V There is low sequence homology between the promoter region of 1.7 and humans. However, our preliminary assays indicate that promoters #1, #2 and #7 are also homologous to Neuro2a (mouse Na VIn addition, there are other examples of promoters with low homology between mouse and human that can have similar expression characteristics in rodents, such as the human synapsin 1 (hSYN1) promoter. Thus, human Na V The 1.7 promoter will be tested in mice and compared to the CMV and hSyn1 promoters for activity and biodistribution. Importantly, this will confirm expression of the transgene (mCherry) in the nociceptors. This pilot study will help identify promoters for dose ranging studies.

[0146] C57BL / 6 male mice, 1.10 12 AAV9-hNa in vg V Inject IT (n=6 / group) with 1.7promoter-mCherry, AAV9-CMV-mCherry, AAV9-hSyn1-mCherry, or saline. Biodistribution is determined by protein immunostaining, qPCR and RNAscope. After 3 weeks, n=3 mice / group are perfused and n=3 mice / group are euthanized and tissues are placed in RNA at harvest to preserve RNA. Organs (brain, spinal cord, dorsal root ganglion, eye, lung, heart, liver, kidney, skeletal muscle, spleen, and gonads) and blood are harvested. Immunostaining for mCherry (ab167453) and subsequent confocal analysis are performed. Additionally, mCherry mRNA is quantified via RT-qPCR. Importantly, transduction of sensory neurons in the DRG is critical to ensure efficacy, so a subset of neurons in the DRG are transduced via RNAscope to express mCherry and Na. V 1.7 are analyzed simultaneously. Probes for calcitonin gene-related peptide (CGRP) and P2X purinergic ion channel type 3 receptor (P2X3R) are also used to define peptidergic and non-peptidergic nociceptor subpopulations. ELISA and ELISPOT analyses are performed to determine whether any immune response to the transgene is detected and whether it varies between promoters.

[0147] Example 6: ZFP-KRAB repressor-based Na V 1.7 Gene silencing HuH7 cells were transfected with nucleic acids encoding one of the ZFPs 1-11 (see Table 8 for ZFP sequences) and the KRAB repressor domain using standard Lipofectamine transfection. V The Na+ levels after transfection of ZFP-KRAB were compared to the mCherry control 3 days after transfection using qPCR. V A fold expression change of 1.7 is shown.

[0148] Example 7: Na+ expression using ZFP-KRAB repressors V 1.7 and Na V Suppression of 1.8 Single target (Na V 1.7 or Na V 1.8) Composition and dual targeting (Na V 1.7 and Na V 1.8) Compositions were designed as shown in the schematic diagrams in Figures 5-6. Mice were treated with paclitaxel to induce allodynia and then treated with single or dual targeting compositions to identify improvements in reversing allodynia. For male mice with mechanical allodynia, Na V When suppressing 1.7, Na V Compared to 1.8, the relative threshold is increased by about 50%, and the targeting Na V The relative threshold for the dual target increased by approximately 13.4% compared to 1.7. For female mice with mechanical allodynia, as shown in Figures 8A-8B, the Na V When suppressing 1.7, Na V Compared to 1.8, the relative threshold is increased by about 20%, and the targeting Na V The relative threshold for the dual target increased by approximately 22% compared to 1.7. These results suggest that the Na V 1.7 and Na V The double inhibition of 1.8 is Na V1.7 inhibition alone was more effective than that of 1.7 alone. Furthermore, no significant changes were seen in grip strength or rotarod testing, and no neuronal loss, axonopathy, or apoptosis was observed.

[0149] Example 8: Na+ to the surface of AAV9 V 1.7 Chemical conjugation of binding peptides and in vitro transduction Na V 1.7 Binding peptides (Protoxin-II, Phlotoxin-I, Huwentoxin-IV, m3-Huwentoxin-IV, see Table 7) were conjugated to the surface of AAV9 CMV mCherry by chemical cross-linking. The virus was then transfected with Na V 9A-9C show that AAV9 expression on the surface of Na+ was significantly increased in three cell lines expressing AAV9. Cells were incubated with the virus for 72 h (MOI Neuro-2A: 50,000; HuH-7: 100,000; SH-SY5Y: 100,000). Naïve cells did not receive any virus. qRT-PCR was used to measure expression of the transgene reporter, mCherry. Expression of mCherry was quantified relative to the housekeeping gene, glyceraldehyde 3-phosphate dehydrogenase (GAPDH). The fold expression change of mCherry was calculated relative to the expression of mCherry under unmodified AAV9 (WT) conditions. Error bars indicate SD. Figures 9A-9C show that Na+ was increased in three cell lines expressing AAV9. V 1.7 Chemical conjugation of binding peptides V We show that capsid modifications increase transduction in Neuro-2A, HuH-7, and SH-SY5Y cell lines expressing 1.7. Several capsid modifications increased transduction by approximately 2- to 6-fold compared to WT.

[0150] Example 9: Na in the AAV9 capsid V 1.7 Expression of binding peptides V 1.7 Increasing viral transduction in conjugated cell lines Peptide JNJ63955918-Indel and a short 6 bp linker were expressed in the viral protein VR-III region of the AAV9 capsid. Viruses were generated using WT capsids or mature capsids containing peptide JNJ63955918 (SEQ ID NO: 97). An mCherry reporter was packaged into all viruses. Viruses were incubated with cells for 72 hours at an MOI of 10,000. Naïve cells did not receive any virus. Cells were analyzed for mCherry expression by qPCR. Expression of mCherry was quantified relative to the housekeeping gene, glyceraldehyde 3-phosphate dehydrogenase (GAPDH). The fold expression change of mCherry was calculated relative to the expression of mCherry under unmodified AAV9 (WT) conditions. Error bars indicate SD. Figure 10 shows that expression of JNJ63955918-Indel on the surface of AAV9 significantly increased the expression of Na V 1.7 expressing cell line HuH-7, thus increasing transduction by 2-fold.

[0151] Example 10: Na V 1.7 and transgene expression in cell lines expressing Na V 1.7 Use of Promoters The cell lines are transfected with a plasmid carrying a promoter driving mCherry reporter expression. Naïve is the non-transfected state. The cytomegalovirus (CMV) promoter is an industry standard universal promoter. Promoter 3, Promoter 1, Promoter 2, and Promoter 7 (see Table 5) are Na V The three cell lines tested were HuH-7, SH-SY5Y, and Neuro-2A. After 72 h, cells are imaged under a fluorescent microscope. Red fluorescence intensity was measured and averaged from three non-overlapping images using the Histogram tool in ImageJ. Error bars indicate SD. Figure 11A shows the effect of different Na+-specific promoters tested in HuH-7 and Neuro-2A cell lines. V1.7 Images from a fluorescent microscope showing mCherry expression driven by a specific promoter. V 1.7 Specific promoters V 1.7 expressing cell lines.

[0152] Example 11: Mouse dorsal root ganglion (DRG) Na V 1.7 Expressing Na in Neurons V 1.7 Specific promoters and transgene expression C57BL / 6 mice were V 1.7 1 x 10 packaged with a specific promoter, mCherry reporter driven by promoter 1 12 gc AAV9 virus was intrathecally injected. Four weeks after injection, mice were sacrificed and lumbar DRGs were harvested, fixed, and cryoprotected. DRGs were sectioned (10 μm) and Na V The cells were labeled using RNAScope Multiplex Fluorescent (Advanced Cell Diagnostics, Inc) reagent with probes targeting promoter 1.7, mCherry, and WPRE (a marker for viral transgenes). Z-stacks were captured from each sample using a Leica DMi8 fluorescent microscope with a 20x objective. Maximum intensity projection images were created from each Z-stack using ImageJ. Hoechst (blue) was used for nuclear localization. Figure 12 shows the promoter 1 Na V 1.7 Specific promoters are used to target Na V The results show that Na+ drives mCherry expression (red) in the 1.7-expressing neuronal population (green). V Colocalization of 1.7 (asterisk), mCherry (plus sign), and WPRE (dagger) indicates that the virus V 1.7 expression and show that the reporter is expressed in these cells (arrows). Each punctate red dot in the mCherry channel represents a single mCherry mRNA molecule, and many punctate dots are observed in some cells, indicating that Na VFigure 1 shows that the 1.7-specific promoter can drive relatively high expression of a transgene in neurons of the mouse DRG. Scale bar indicates 50 μm. [Table 1] [Table 2-1] [Table 2-2] [Table 2-3] [Table 2-4] [Table 2-5] [Table 2-6] [Table 3-1] [Table 3-2] [Table 4-1] [Table 4-2] [Table 4-3] [Table 4-4] [Table 4-5] [Table 4-6] [Table 4-7]

Table 4-8

Table 4-9

Table 4-10

Table 4-11

Table 4-12

Table 4-13

Table 4-14

Table 4-15

Table 4-16

Table 4-17

Table 4-18

Table 4-19

Table 4-20

Table 4-21

Table 4-22

Table 4-23

Table 4-24

Table 4-25

Table 4-26

Table 4-27

Table 4-28

Table 4-29

Table 4-30

Table 4-31

Table 4-32

Table 4-33

Table 4-34

Table 4-35

Table 4-36

Table 4-37

Table 4-38

Table 4-39

Table 4-40

Table 4-41

Table 4-42

Table 4-43

Table 5-1

Table 5-2

Table 5-3

Table 5-4

Table 5-5

Table 6-1

Table 6-2

Table 7-1

Table 7-2

Table 8-1

Table 8-2

Table 8-3

Table 9-1

Table 9-2

Claims

1. A nucleic acid comprising a sequence encoding a nucleic acid binding domain, a sequence encoding an epigenetic regulator that regulates transcription of one or more target molecules, and a promoter comprising the Navl.7 promoter, a promoter having a sequence at least about 90% identical to a sequence in Table 5, or a promoter of a gene selected from Tables 1-3.

2. The nucleic acid of claim 1 , wherein the one or more target molecules comprises a nucleic acid encoding an ion channel and / or is associated with an ion channel.

3. 3. The nucleic acid of claim 1 or claim 2, wherein the one or more target molecules include SCN9A, SCN10A, or SCN11A, or a combination thereof.

4. 3. The nucleic acid of claim 1 or 2, wherein the one or more target molecules are associated with Dravet syndrome, epilepsy syndrome, familial hemiplegic migraine, Ohtahara syndrome, West syndrome, Lennox-Gastaut syndrome, sodium channel myotonia, autism, long QT syndrome, Brugada syndrome, or progressive cardiac conduction disease (also known as Renegre's disease), erythromelalgia, diabetic neuropathy, small fiber neuropathy, rheumatoid arthritis, trigeminal neuralgia, or a combination thereof.

5. 3. The nucleic acid of claim 1 or claim 2, wherein the one or more target molecules comprise one or more genes in Table 1.

6. The nucleic acid of claim 1 or claim 2, wherein the one or more target molecules are associated with pain.

7. 3. The nucleic acid of claim 1 or claim 2, wherein the one or more target molecules comprise one or more genes of Table 2 or Table 3.

8. The nucleic acid of any one of claims 1 to 7, wherein the nucleic acid binding domain comprises a zinc finger protein.

9. 8. The nucleic acid of any one of claims 1 to 7, wherein the nucleic acid binding domain comprises a clustered regularly interspaced short palindromic repeats associated protein (dCas) that has no nuclease activity.

10. The nucleic acid according to any one of claims 1 to 9, wherein the epigenetic regulatory factor comprises a domain having transcriptional repression activity (repressor domain).

11. 11. The nucleic acid of claim 10, wherein the repressor domain comprises a ZIM3 repressor domain or a Krueppel-associated box (KRAB) domain (recruiting histone methyltransferases and deacetylases).

12. The nucleic acid according to any one of claims 1 to 11, wherein the epigenetic regulatory factor comprises a domain having transcription activation activity (activator domain).

13. The epigenetic regulators include ZIM3, KRAB (also called KOX), SID, MBD2, MBD3, HP1a, DNMT family (including DNMT1, DNMT3A, DNMT3B, DNMT3L, DNMT2A), Sin3a, Rb, MeCP2, ROM2, AtHD2A, LSD1, SUV 39H1, G9a (EHMT2), ZIM3, ZNF554, ZNF264, ZNF324, ZNF354A, ZNF189, ZNF543, ZNP82, ZNF669, ZNF582, KOX1-MeCP2, ZNF30, ZNF680, ZNF331, ZNF33A, ZNF528, ZNF320, ZNF3 50, ZNF175, ZNF214, ZNF184, ZNF8, ZNF596, KOX1, ZNF37A, ZNF394, ZNF610, ZNF273, ZNF34, ZNF250, ZNF98, ZNF675, ZNF213, NLuc, ZFP28-2, ZNF224, ZNF257, VP64, Rta, P16, P65, p300, TET1 catalytic domain, TDG, Ldbl self-association domain, SAM activator (VP64, p65, HSF1), VPR (VP64, p65, Rta), CD, or SunTag, or a variant or combination thereof.

14. The nucleic acid according to any one of claims 1 to 13, wherein the promoter comprises the Nav1.7 promoter.

15. 14. The nucleic acid of any one of claims 1 to 13, wherein the promoter comprises a sequence at least about 90% identical to a sequence of Table 5, optionally promoter 1 or promoter 2.

16. The nucleic acid sequence according to any one of claims 1 to 13, wherein the promoter is a promoter of a gene selected from Tables 1 to 3.

17. The nucleic acid according to any one of claims 1 to 16, wherein the nucleic acid is delivered to the subject via a delivery vehicle.

18. 1. A composition comprising: a) a nucleic acid comprising a sequence encoding a nucleic acid binding domain and a sequence encoding an epigenetic regulator that regulates transcription of one or more target molecules; and b) a delivery vehicle for delivering said nucleic acid, said delivery vehicle comprising a targeting peptide comprising a sequence at least about 90% identical to JNJ63955, m3-Huwentoxin-IV, Phlotoxin 1 (PhlTx1), Protoxin-II (ProTx-II), or Ceratotoxin-1 (CcoTx1), or a variant thereof, or a peptide in Table 7.

19. 19. The nucleic acid of claim 17 or the composition of claim 18, wherein the delivery vehicle is a viral delivery vehicle (e.g., a retroviral vector, a lentiviral vector, or an adenoviral vector), a liposome, a nanoparticle, or an exosome.

20. 20. The nucleic acid of claim 19, wherein the viral delivery vehicle is a recombinant adeno-associated virus (AAV).

21. 21. The nucleic acid or composition of claim 20, wherein the AAV is AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAVhu68, AAVrh.10, AAVrh74, or AAVDJ, or a combination thereof.

22. 22. The nucleic acid or composition of claim 21 , wherein the AAV is AAV9.

23. The nucleic acid or composition of any one of claims 20 to 22, wherein the AAV has a recombinant capsid.

24. 24. The nucleic acid or composition of claim 23, wherein the recombinant capsid comprises a sequence that includes or encodes a targeting moiety.

25. The nucleic acid of claim 17 or 19-23, or the composition of any one of claims 18-23, wherein the delivery vehicle comprises or is linked to a targeting moiety.

26. 26. The nucleic acid or composition of claim 25, wherein the targeting moiety targets cells containing the target molecule in the subject.

27. The nucleic acid or composition of any one of claims 24 to 26, wherein the targeting moiety binds to a peptide product of the target molecule.

28. 28. The nucleic acid or composition of claim 27, wherein the target molecule is present on a target cell.

29. 29. The nucleic acid or composition of claim 28, wherein the target cell is associated with the disease or condition of the subject.

30. The nucleic acid or composition of any one of claims 24 to 29, wherein the targeting moiety comprises a peptide.

31. 31. The nucleic acid or composition of claim 30, wherein the peptide comprises JNJ63955, m3-Huwentoxin-IV, Phlotoxin 1 (PhlTx1), Protoxin-II (ProTx-II), or Ceratotoxin-1 (CcoTx1), or a variant thereof.

32. 31. The nucleic acid or composition of claim 30, wherein the peptide comprises a sequence that is at least about 90% identical to a peptide of Table 7.

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