PROJECT SUMMARY:
Charcot-Marie-Tooth (CMT) disease is the most frequent inherited neuropathy affecting the peripheral nervous
system and is characterized by a group of genetically and clinically heterogeneous disorders leading to
progressive weakness and atrophy in distal muscles, sensory loss, hyporeflexia and skeletal deformity. CMT
type 1A (CMT1A) is the most prevalent form, affecting 1 in 10,000 people, and is associated with a 1.4-Mbp
duplication in the chromosome 17p11.2 region, which contains the peripheral myelin protein 22 (PMP22) gene.
PMP22 is essential for the structure, development and maintenance of peripheral nerve myelin. PMP22
overexpression prompts cycles of demyelination-remyelination resulting in dysfunction in Schwann cells. Due to
the association of PMP22 gene dosage with neuropathic phenotypes, therapeutic strategies are primarily
focused on repressing PMP22 overexpression. RNA therapeutics, like antisense oligonucleotides (ASO) and
siRNA, are attractive because they target messenger RNA and thus can modulate the expression of protein
targets inaccessible to other therapeutic modalities. Challenges identifying safe and effective ways to deliver
RNA therapeutics into cells outside the liver have limited the clinical deployment of this promising therapeutic
class. For instance, a recent study used an ASO to decrease PMP22 mRNA in affected nerves, improving
phenotypes in rat and mouse models of CMT1A. However, very high drug doses (multiple 100 mg/kg doses)
were required to see a beneficial effect. At such high doses, the risk of toxicities related to ASO treatment, such
as thrombocytopenia and renal dysfunction, preclude further development. DTx Pharma has identified a fatty
acid motif that when covalently coupled to siRNA/ASO results in efficient delivery to multiple cells and tissues,
including sciatic nerve (relevant for CMT), resulting in potent repression of target gene mRNA expression.
Herein, we propose to explore whether DTx technology can be applied to PMP22-targeting siRNAs to correct its
overexpression in Schwann cells in a mouse model of CMT1A, providing strong proof of concept for designing
future therapeutic efficacy studies. We will explore this in 2 aims. Aim 1 will screen a library (~36 in addition to
what we’ve screened to date) of siRNA candidates targeting PMP22 in vitro using both primary human Schwann
cells and HEK293 cells engineered to express human PMP22 to identify potent and non-toxic siRNAs that will
be conjugated to the DTx motif and further validated in vitro. In Aim 2, the 10 most active hits from aim 1 will be
dosed in two parallel studies via intravenous or intrathecal administration in C3-PMP22 mice, a model of CMT1A,
to assess dosing route, safety and target engagement in the sciatic nerve and Schwann cells. The more
successful dosing route will be utilized in follow-on studies to explore dose range and duration of action for
suppressing PMP22 expression to wildtype levels. Data from these studies will help us understand if DTx PMP22
siRNA is a viable approach for treatment of CMT1A and provide the foundation for a phase 2 SBIR grant focused
on efficacy trials in rodents, non-GLP toxicity studies and validation in higher species.
