Charcot-Marie-Tooth disease is the most frequent form of the hereditary peripheral neuropathies, affecting one out of 2500 people. It is characterized by variable neuromuscular problems, often occurring prior to the age of 20 (20% of the patients have serious handicaps) (M. Shy et al, 2004).
CMT1A form, the most frequent disorder (50% of CMT patients) is a demyelinizing form of peripheral neuropathy. It was demonstrated in 1991 that a majority of patients showed a 1.5 Mb-duplication in the 17p11.2 region (Rayemekers et al, 1991, Lupski et al, 1991), encompassing a locus that encodes for the peripheral myelin protein PMP22 (Matsunami et al, Patel et al, Timmerman et al, Valentijn et al, 1992). It is well established that deregulation of functional PMP22 expression by either point mutation or gene duplication is responsible for CMT1A disease. But currently, no treatment can be offered to the patients.
We then thought to generate an animal model for a
better comprehension of the disease evolution and the evaluation of
potential therapeutic drugs.
Physiopathology issues. To create
an animal model mimicking the genomic duplication, we injected a YAC
containing the human PMP22 locus into fertilized murine oocytes. Two lines of mice, including the 7-copy C22 line,
presented a dominant autosomal CMT1A-like neuropathy. We showed the
pathology was linked to PMP22 over-expression in non-linear
correlation, suggesting that a slight correction of its expression
level might have a strong impact on the phenotype (Huxley et al, 1996,
1998). Our results strongly argued for a dysmyelination rather
than a demyelination process. We showed the nerve hypertrophy, often
noted in CMT1A patients, was due to the overproduction of collagen by
fibroblasts, suggesting that Schwann cells are not the only cells
affected by PMP22 over-expression (Robaglia et al, Brain 2002).
Regulation of PMP22 expression.
Since expression levels of PMP22 appeared to be critical for the rescue
of CMT1A phenotype, we further characterized the transcriptional
regulation of the 6-kb promoter. We showed the proximal 300 bp-region
was sufficient to drive the cell-specific expression of the gene. Two
additional CREB binding sites were required for the complete activation
of the full-length promoter in response to cAMP signalling (Sabéran et
We finally focused on the development of therapeutic procedures to
correct PMP22 over-expression. It was well known that ascorbic acid
(AA) was required for the co-culture of axons and Schwann cells, as
well as for the promotion of in vitro myelination (Carey et al, 1987;
Eldridge et al, 1987). There was also evidence of a link between
femoral neuropathies and AA deficiency. We then speculated that
ascorbic acid could be a good candidate to promote myelination in our
CMT1A model. Moreover, the drug had already received regulatory
approval for treatment, making possible further rapid phase II/III
clinical trials. Using the C22 mice, we found that high doses of
AA were able to correct the phenotype. PMP22 expression appeared to
decrease under treatment, probably by interfering with the
cAMP-dependent stimulation of the promoter (Passage et al, 2004). A
phase II/III clinical trial in three centres, Marseille (Timone), Paris
(Salpétrière) and Lyon began in the autumn of 2005 (coordination, Dr J.
Micaleff, Marseille) (see below).
Ascorbic acid action mechanism.
Other antioxidants (retinol or -tocopherol) appeared to be unable to
modulate PMP22 expression, indicating AA effect on myelination did not
rely on the oxidative stress response. We then demonstrated that AA but
not other antioxidants induced the reduction of the cAMP pool.
Importantly, the decrease was dose-dependent, and closely correlated
with AA-mediated suppression of PMP22 overexpression (Kaya et al,
2007). Recent data strongly indicate the drug behaves as a competitive
inhibitor of the adenylate cyclases (Kaya et al, 2008).
summary, we have largely contributed to the comprehension of CMT1A
disease, from physiopathology to medicine, making our team a group
leader in that field. This project is funded by French ANR program (350 000 Euros, up to the end of 2009).
the last years, our group has contributed to understand pathophysiology
of Charcot-Marie-Tooth disease. Therefore our project will be focused
on proposing therapeutical solutions. This will imply to put together
multidisciplinary approaches. The project will thus be supported by
molecular biologists, geneticists, phamacologists specialists of
galenic, and physiologists. To achieve this goal, we will develop the following sub projects.
-To better understand the pathophysiology of these disorders. -Confirm our hypothesis on the mode of action of AA. -Develop 2nd- and 3rd-generation drugs. -Construct a new murine model of CMT: CMT-X.
Ascorbic acid is a regulator of the intracellular cAMP concentration: old molecule, new functions? Kaya F, Belin S, Diamantidis G, Fontes M. FEBS Lett. 2008 Oct 29;582(25-26):3614-8. Epub 2008 Oct 1.
Ascorbic acid inhibits PMP22 expression by reducing cAMP levels. Kaya F, Belin S, Bourgeois P, Micaleff J, Blin O, Fontés M. Neuromuscul Disord. 2007 Mar;17(3):248-53. Epub 2007 Feb 15.
Ascorbic acid treatment corrects the phenotype of a mouse model of Charcot-Marie-Tooth disease.
Passage E, Norreel JC, Noack-Fraissignes P, Sanguedolce V, Pizant J,
Thirion X, Robaglia-Schlupp A, Pellissier JF, Fontés M. Nat Med. 2004
PMP22 overexpression causes dysmyelination in mice.
Robaglia-Schlupp A, Pizant J, Norreel JC, Passage E, Saberan-Djoneidi
D, Ansaldi JL, Vinay L, Figarella-Branger D, Levy N, Clarac F, Cau P,
Pellissier JF, Fontes M. Brain, 2002, 125, 2213-2221.
Molecular dissection of the Schwann cell specific promoter of the PMP22 gene. Sabéran-Djoneidi D, Sanguedolce V, Assouline Z, Lévy N, Passage E, Fontés M. Gene. 2000 May 2;248(1-2):223-31.
between varying levels of PMP22 expression and the degree of
demyelination and reduction in verve conduction velocity in trangenic
mice. C.Huxley, E .Passage, A.M.Robertson, B.Youl, S.Huston,
A.Manson, D.Saberan-Djoneidi, D.Figarella-Branger, J.F.Pellissier,
P.K.Thomas, M.Fontés. Hum.Mol.Genet, 1998, 7, 449-458.
Construction of a mouse model of the CMT1A disease by YAC injection in the murine oocyte. C.Huxley, E.Passage, A.Manson, G.Putzu, D.Figarella-Branger, J.F.Pellissier, M.Fontés. Human Mol.Genet. 1996, 5, 563-569.