JPWO2019222354A5 - - Google Patents

Claims (20)

Viral vectors containing one or more transgenes, each encoding an interfering ribonucleic acid (RNA) of at least 17 nucleotides in length, where each interfering RNA is an endogenous RNA transcript containing an extended repeat region. A viral vector comprising an annealing moiety, wherein said moiety of each interfering RNA annealies to a segment of said endogenous RNA transcript that does not overlap with said extended repeat region. The viral vector of claim 1, wherein the endogenous RNA transcript encodes a human dystrophy myotonica protein kinase (DMPK). The viral vector of claim 2, wherein the extended repeat region comprises 50 or more CUG trinucleotide repeats. The viral vector of claim 2 or 3 , wherein the vector further comprises a transgene encoding a human DMPK RNA transcript that does not anneal to the interfering RNA. The viral vector vector of claim 4 , wherein the human DMPK RNA transcript has less than 85% complementarity to the interfering RNA. The viral vector according to any one of claims 2 to 5 , wherein the endogenous RNA transcript comprises a moiety having at least 85% sequence identity to the nucleic acid sequence of SEQ ID NO: 1 or SEQ ID NO: 2. The part of each interfering RNA is
(a) Within any one of exons 1 to 15 of human DMPK ,
(b) Within any one of the human DMPK introns 1-14,
(c) Includes or contains an exon-intron boundary within the human DMPK
(d) Within the 5'UTR or 3'UTR of the human DMPK,
The viral vector according to any one of claims 2 to 6 , which has a nucleic acid sequence that is at least 85% complementary to the nucleic acid sequence of the segment. The viral vector according to any one of claims 2 to 6 , wherein the portion of each interfering RNA is annealed to a segment of an endogenous RNA transcript having the nucleic acid sequence of any one of SEQ ID NOs: 3 to 39. The viral vector according to any one of claims 2 to 8 , wherein the interfering RNA comprises a portion having at least 85% sequence identity to the nucleic acid sequence of any one of SEQ ID NOs: 3 to 161. The viral vector of claim 1, wherein the endogenous RNA transcript comprises a human chromosome 9 open reading frame 72 (C9ORF72) and an extended repeat region. The viral vector of claim 10 , wherein the extended repeat region comprises from about 25 to about 1600 GGGGCC hexanucleotide repeats. The viral vector according to claim 10 or 11 , wherein the endogenous RNA transcript comprises a moiety having at least 85% sequence identity to the nucleic acid sequence of SEQ ID NO: 163, 165, or 166. The viral vector according to any one of claims 10 to 12 , wherein each portion of the interfering RNA has a nucleic acid sequence that is at least 85% complementary to the nucleic acid sequence of the segment within human C9ORF72. The interfering RNA is short interfering RNA (siRNA), short hairpin RNA (shRNA), or microRNA (miRNA) , optionally the miRNA is U6 miRNA, and optionally the virus vector.
(a) Primary miRNA (pri-miRNA) transcripts encoding mature miRNAs, or
(b) pre-miRNA transcripts encoding mature miRNAs,
The viral vector according to any one of claims 1 to 13 , which comprises . The viral vector according to any one of claims 1 to 14 , wherein the interfering RNA is operably linked to a promoter that induces expression of the interfering RNA in muscle cells or neurons. The promoters are desmin promoter, phosphoglycerate kinase (PGK) promoter, muscle creatin kinase promoter, myosin light chain promoter, myosin heavy chain promoter, cardiac troponin C promoter, troponin I promoter, myoD gene family promoter, actin alpha promoter, actin. The viral vector of claim 15 , which is a beta promoter, an actin gamma promoter, or a promoter within Intron 1 of an ocular paired-like homeodomain 3 (PITX3). The virus vector is selected from the group consisting of adeno-associated virus (AAV), adenovirus, lentivirus, retrovirus, poxvirus, baculovirus, simple herpesvirus, vaccinia virus, and synthetic virus, and the AAV is optionally AAV1. , AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAVrh10, or AAVrh74 cellotype, the virus vector according to any one of claims 1 to 16 . 17. The viral vector of claim 17 , wherein the viral vector is pseudo-AAV, and optionally the pseudo-AAV is AAV2 / 8 or AAV2 / 9 . A nucleic acid encoding or containing an interfering RNA, wherein the interfering RNA comprises a moiety having at least 85% sequence identity to any one of the nucleic acid sequences of SEQ ID NOs: 3 to 161. A composition comprising the vector according to any one of claims 1 to 18 for use in a method for reducing or treating the occurrence of splice opacity in a human patient with myotonic dystrophy or myotonic lateral sclerosis . A composition comprising the step of administering to the patient a composition comprising a therapeutically effective amount of any one of the vectors of claims 1-18 , or the nucleic acid of claim 19 .