Targeted PMP22 TATA-box editing by CRISPR/Cas9 reduces demyelinating neuropathy of Charcot-Marie-Tooth disease type 1A in mice

Abstract

Charcot-Marie-Tooth 1A (CMT1A) is the most common inherited neuropathy without a known therapy, which is caused by a 1.4 Mb duplication on human chromosome 17, which includes the gene encoding the peripheral myelin protein of 22 kDa (PMP22). Overexpressed PMP22 protein from its gene duplication is thought to cause demyelination and subsequently axonal degeneration in the peripheral nervous system (PNS). Here, we targeted TATA-box of human PMP22 promoter to normalize overexpressed PMP22 level in C22 mice, a mouse model of CMT1A harboring multiple copies of human PMP22. Direct local intraneural delivery of CRISPR/Cas9 designed to target TATA-box of PMP22 before the onset of disease, downregulates gene expression of PMP22 and preserves both myelin and axons. Notably, the same approach was effective in partial rescue of demyelination even after the onset of disease. Collectively, our data present a proof-of-concept that CRISPR/Cas9-mediated targeting of TATA-box can be utilized to treat CMT1A.

Squalenoyl siRNA PMP22 nanoparticles are effective in treating mouse models of Charcot-Marie-Tooth disease type 1 A

“…route instead of intraperitoneal injection. In addition, our data are consistent with two recent studies showing that inhibiting PMP22 expression can ameliorate the neuropathological symptoms caused by PMP22 overexpression 50 , 51 . In contrast with our study, where siRNA was injected via i.v.…”

Exploiting Sphingo- and Glycerophospholipid Impairment to Select Effective Drugs and Biomarkers for CMT1A

“…As for other hereditary neuromuscular diseases in which gene therapy proved to be effective, the negative regulation of PMP22 expression represents the ideal biological endpoint and therapeutic goal (12)(13)(14). To date, in contrast to preclinical studies, clinical trials in CMT1A patients aimed to down-regulate PMP22 showed negative results (7,(15)(16)(17). Furthermore, gene therapy, in addition to the obvious technical difficulties, seems to be still poorly acceptable for CMT1A patients in which life expectancy and quality of life are not dramatically affected (18,19).…”

Gene Therapy Options as New Treatment for Inherited Peripheral Neuropathy

“…Reducing the protein expression of PMP22 can also be achieved by modulating the transcriptional activity of a gene. Recently, two independent research groups demonstrated the feasibility of PMP22 reduction by disrupting either the promoter or enhancer of PMP22 with gene editing technology [81,82]. The clustered regularly interspaced short palindromic repeats (CRISPR) and related Cas genes are now emerging as essential tools for gene editing [83,84].…”

“…The in vivo efficacy of gene editing was evaluated by another group. Lee et al [82] targeted the TATA-box promoter of PMP22 to reduce the transcription. After intraneural delivery of CRISPR/ Cas9 protein targeting the TATA-box promoter of PMP22, the expression level of PMP22 in the sciatic nerve was effectively reduced in C22 mice.…”

“…The CRISPR/Cas9 delivery also ameliorated demyelination, muscle atrophy, and defects in the locomotor function. By duplicating the experiment in accordance with the administration time-points (before onset and after onset), Lee et al [82] also validated the efficacy of CRISPR/Cas9-mediated gene editing on reversing the neuropathic phenotype even after the onset; a crucial clinical benefit of treatment in regards to human patients. Additionally, the safety of gene editing in vivo was analyzed for future clinical application.…”