The CMTA is committed to extending the STAR initiative to other kinds of CMT, such as CMT Type 2. Because CMT2 has diverse genetic causes, and likely multiple disease mechanisms, the CMTA sponsored a workshop in San Diego in November, 2010, and sought to obtain the advice and direction of the most knowledgeable experts in the world. The meeting was attended by more than 30 scientists, and, as a result of the dynamic exchange of ideas, the CMTA has formulated strategies for developing treatments for CMT Types 2A, 2C, and 2E.
CMT Type 2A
CMT2A is caused by dominant mutations in MFN2. It has been shown that MFN1 can compensate for MFN2 in cell models, and the over-expression of MFN1 can even compensate for the effects of dominant MFN2 mutants. Thus, the goal is to be able to use high-throughput screening (HTS) to identify small molecules that up-regulate the expression of MFN1 in human neuronal cell lines and determine whether they can rescue the effects of dominant MFN2 mutants in cells derived from CMT2A patients and in animal models of CMT2A.
Project 1, David Chan, MD, PhD, California Institute of Technology — To enable HTS for MFN1 up-regulators, the CMTA is funding a project by Dr. Chan to develop human neuronal cell lines with a stable expression vector comprised of the human MFN1 promoter and each of two reporter constructs: luciferase and beta-lactamase. SY5Y cells appear to be among the best candidates at this time, and a bacterial artificial chromosome (BAC) or fosmid containing the human MFN1 gene would be used to drive expression of luciferase and beta-lactamase.
Project 2, Stephan Zuchner, PhD, University of Miami — The CMTA is also funding a project by Dr. Zuchner to further refine the development of rodent models that will help to evaluate the CMT2A pathological process in vivo and provide a tool for pre-clinical testing. In addition to crossing mouse strains to obtain the best phenotypic model, crossing the MFN2 knock-in mouse into the existing MFN2 and MFN1 knock-out models could further solidfy, in vivo, the hypothesis that MFN1 expression rescues MFN2 mutants.
CMT Type 2C
Dominant mutations in the TRPV4 ion channel that result in increased channel conductance and presumably in increased calcium entry and toxicity have been reported as the cause of CMT2C. Since this is a well-studied family of channels, promising lead compounds already exist in the pharmaceutical industry. A promising result in favor of a gain-of-function mechanism and channel blocking therapeutic strategy is that mice completely lacking the TRPV4 gene have a relatively mild phenotype. A loss-of-function mechanism due to poor protein folding or trafficking may require alternative approaches such as chaperone-based strategies. To resolve this issue, animal models in which the human alleles (putative gain-of-function changes) are being introduced into the mouse gene are being generated. These animals could also serve in pre-clinical trials.
The CMTA is currently supporting research in development of immortalized pluri-potent stem (iPS) cells derived from patient skin biopsies to provide human screening models for therapeutic candidates. CMT2A neurons have already been derived from these iPS cells and CMT2E neurons are nearing completion.
CMT Type 2E
CMT2E is caused by dominant mutations in NEFL, which prevent the neurofilament light protein from assembling properly and result in the formation of aggregates or abnormal filaments. The goal is to find small molecules that can decrease aggregate formation and/or promote the formation of normal appearing neurofilament. Demonstration of these effects would be identified by rescuing the effects of dominant NEFL mutants in cells derived from CMT2E patients and in animal models of CMT2E.