Molecular Neurobiology

, Volume 56, Issue 9, pp 6460–6471 | Cite as

Disease Modeling and Therapeutic Strategies in CMT2A: State of the Art

  • Kordelia Barbullushi
  • Elena Abati
  • Federica Rizzo
  • Nereo Bresolin
  • Giacomo P. Comi
  • Stefania CortiEmail author


Mitofusin 2 (MFN2) is a protein of the mitochondrial outer membrane that belongs to a family of highly conserved dynamin-related GTPases. It is implicated in several intracellular pathways; however, its main role is the regulation of mitochondrial dynamics, in particular mitochondrial fusion. Mutations in MFN2 are associated with Charcot–Marie–Tooth disease type 2A (CMT2A), a neurological disorder characterized by a wide spectrum of clinical features, primarily a motor sensory neuropathy. The cellular and molecular mechanisms by which MFN2 mutations lead to neuronal degeneration are largely unknown, and there is currently no cure for patients. Here, we present the most recent in vitro and in vivo models of CMT2A and the more promising therapeutic approaches under development. These models and therapies may represent relevant tools for the study and recovery of defective mitochondrial dynamics that seem to play a significant role in the pathogenesis of other more common neurodegenerative diseases.


Mitofusin2 Charcot–Marie–Tooth disease type 2 Hereditary neuropathies Mitochondrial diseases Molecular therapy Gene therapy Mitofusin agonists 



Charcot–Marie–Tooth disease


Motor nerve conduction velocity


Motor neurons


Sensory neurons


Induced pluripotent stem cells


Antisense oligonucleotide


RNA interference


Clustered regularly interspersed short palindromic repeats


Caspase 9





We thank Associazione Progetto Mitofusina 2 Onlus and Associazione Amici del Centro Dino Ferrari for their support.


This study is supported by grant “Ricerca Corrente 2019: Analisi dei pathway molecolari condivisi coinvolti nella patogenesi delle malattie neurodegenerative mediante modelli in vitro basati su cellule staminali” from the Italian Ministry of Health.

Compliance with Ethical Standards

Conflict of Interest

The authors declare that they have no conflict of interest.


  1. 1.
    Züchner S, Mersiyanova IV, Muglia M, Bissar-Tadmouri N, Rochelle J, Dadali EL, Zappia M, Nelis E et al (2004) Mutations in the mitochondrial GTPase mitofusin 2 cause Charcot-Marie-Tooth neuropathy type 2A. Nat Genet 36(5):449–451CrossRefGoogle Scholar
  2. 2.
    Stuppia G, Rizzo F, Riboldi G, Del Bo R, Nizzardo M, Simone C et al (2015) MFN2-related neuropathies: clinical features, molecular pathogenesis and therapeutic perspectives. J Neurol Sci 356(1–2):7–18. Available from: CrossRefPubMedGoogle Scholar
  3. 3.
    Braathen GJ, Sand JC, Lobato A, Høyer H, Russell MB (2011) Genetic epidemiology of Charcot-Marie-Tooth in the general population. Eur J Neurol 18(1):39–48CrossRefGoogle Scholar
  4. 4.
    Barreto LCLS, Oliveira FS, Nunes PS, De França Costa IMP, Garcez CA, Goes GM et al (2016) Epidemiologic study of Charcot-Marie-Tooth disease: a systematic review. Neuroepidemiology 46(3):157–165CrossRefGoogle Scholar
  5. 5.
    Bergamin G, Boaretto F, Briani C, Pegoraro E, Cacciavillani M, Martinuzzi A, Muglia M, Vettori A et al (2014) Mutation analysis of MFN2, GJB1, MPZ and PMP22 in Italian patients with axonal Charcot-Marie-Tooth disease. NeuroMolecular Med 16(3):540–550CrossRefGoogle Scholar
  6. 6.
    Verhoeven K, Claeys KG, Züchner S, Schröder JM, Weis J, Ceuterick C et al (2006) MFN2 mutation distribution and genotype/phenotype correlation in Charcot-Marie-Tooth type 2. Brain 129(8):2093–2102CrossRefGoogle Scholar
  7. 7.
    Feely SME, Laura M, Siskind CE, Sottile S, Davis M, Gibbons VS, Reilly MM, Shy ME (2011) MFN2 mutations cause severe phenotypes in most patients with CMT2A. Neurology 76(20):1690–1696CrossRefGoogle Scholar
  8. 8.
    Piscosquito G, Saveri P, Magri S, Ciano C, Di Bella D, Milani M et al (2015) Mutational mechanisms in MFN2 -related neuropathy: compound heterozygosity for recessive and semidominant mutations. J Peripher Nerv Syst 20(4):380–386CrossRefGoogle Scholar
  9. 9.
    Nicholson GA, Magdelaine C, Zhu D, Grew S, Ryan MM, Sturtz F, Vallat JM, Ouvrier RA (2008) Severe early-onset axonal neuropathy with homozygous and compound heterozygous MFN2 mutations. Neurology 70:1678–1681CrossRefGoogle Scholar
  10. 10.
    Polke JM, Laura M, Pareyson D, Taroni F, Milani M, Bergamin G, Gibbons VS, Houlden H et al (2011) Recessive axonal Charcot-Marie-Tooth disease due to compound heterozygous mitofusin 2 mutations. Neurology 77:168–173CrossRefGoogle Scholar
  11. 11.
    Tan CA, Rabideau M, Blevins A, Westbrook MJ, Ekstein T, Nykamp K, Deucher A, Harper A et al (2016) Autosomal recessive MFN2-related Charcot-Marie-Tooth disease with diaphragmatic weakness: case report and literature review. Am J Med Genet 170(6):1580–1584CrossRefGoogle Scholar
  12. 12.
    Zhu D, Kennerson ML, Walizada G, Vance JM, Nicholson GA (2005) Charcot-Marie-Tooth with pyramidal signs is genetically heterogeneous: families with and without MFN2 mutations. Neurology 65:496–497CrossRefGoogle Scholar
  13. 13.
    Züchner S, De Jonghe P, Jordanova A, Claeys KG, Guergueltcheva V, Cherninkova S et al (2006) Axonal neuropathy with optic atrophy is caused by mutations in mitofusin 2. Ann Neurol 59(2):276–281CrossRefGoogle Scholar
  14. 14.
    Filadi R, Pendin D, Pizzo P (2018) Mitofusin 2: from functions to disease. Cell Death Dis 9(3). Available from:
  15. 15.
    Chandhok G, Lazarou M, Neumann B (2017) Structure, function, and regulation of mitofusin-2 in health and disease. Biol Rev 61Google Scholar
  16. 16.
    El Fissi N, Rojo M, Aouane A, Karatas E, Poliacikova G, David C, Royet J, Rival T (2018) Mitofusin gain and loss of function drive pathogenesis in Drosophila models of CMT2A neuropathy. EMBO Rep 19:e45241. Available from:
  17. 17.
    Bombelli F, Stojkovic T, Dubourg O, Echaniz-Laguna A, Tardieu S, Larcher K, Amati-Bonneau P, Latour P et al (2014) Charcot-Marie-Tooth disease type 2A from typical to rare phenotypic and genotypic features. JAMA Neurol 71:1036–1042CrossRefGoogle Scholar
  18. 18.
    Lawson VH, Graham BV, Flanigan KM (2005) Clinical and electrophysiologic features of CMT2A with mutations in the mitofusin 2 gene. Neurology 65(2):197–204CrossRefGoogle Scholar
  19. 19.
    Takahashi K, Yamanaka S (2006) Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 126(4):663–676CrossRefGoogle Scholar
  20. 20.
    Saporta MA, Dang V, Volfson D, Zou B, Xie XS, Adebola A et al (2015) Axonal Charcot-Marie-Tooth disease patient-derived motor neurons demonstrate disease-specific phenotypes including abnormal electrophysiological properties. Exp Neurol 263:190–199CrossRefGoogle Scholar
  21. 21.
    Rizzo F, Ronchi D, Salani S, Nizzardo M, Fortunato F, Bordoni A, Stuppia G, del Bo R et al (2016) Selective mitochondrial depletion, apoptosis resistance, and increased mitophagy in human Charcot-Marie-Tooth 2A motor neurons. Hum Mol Genet 25(19):4266–4281CrossRefGoogle Scholar
  22. 22.
    Cartoni R, Arnaud E, Médard JJ, Poirot O, Courvoisier DS, Chrast R, Martinou JC (2010) Expression of mitofusin 2R94Q in a transgenic mouse leads to Charcot-Marie-Tooth neuropathy type 2A. Brain. 133(5):1460–1469CrossRefGoogle Scholar
  23. 23.
    Bannerman P, Burns T, Xu J, Miers L, Pleasure D (2016) Mice hemizygous for a pathogenic mitofusin- 2 allele exhibit hind limb/foot gait deficits and phenotypic perturbations in nerve and muscle. PLoS One 11(12):17–22CrossRefGoogle Scholar
  24. 24.
    Chen H, Detmer SA, Ewald AJ, Griffin EE, Fraser SE, Chan DC (2003) Mitofusins Mfn1 and Mfn2 coordinately regulate mitochondrial fusion and are essential for embryonic development. J Cell Biol 160(2):189–200CrossRefGoogle Scholar
  25. 25.
    Misko A, Jiang S, Wegorzewska I, Milbrandt J, Baloh RH (2010) Mitofusin 2 is necessary for transport of axonal mitochondria and interacts with the Miro/Milton complex. J Neurosci 30(12):4232–4240. Available from: CrossRefPubMedPubMedCentralGoogle Scholar
  26. 26.
    Misko AL, Sasaki Y, Tuck E, Milbrandt J, Baloh RH (2012) Mitofusin2 mutations disrupt axonal mitochondrial positioning and promote axon degeneration. J Neurosci 32(12):4145–4155. Available from: CrossRefPubMedPubMedCentralGoogle Scholar
  27. 27.
    Bhandari P, Song M, Chen Y, Burelle Y, Dorn GW II (2014) Mitochondrial contagion induced by parkin deficiency in drosophila hearts and its containment by suppressing mitofusin. Circulation Research 114(2):257–265. Available from:
  28. 28.
    Vettori A, Bergamin G, Moro E, Vazza G, Polo G, Tiso N, Argenton F, Mostacciuolo ML (2011) Developmental defects and neuromuscular alterations due to mitofusin 2 gene (MFN2) silencing in zebrafish: a new model for Charcot-Marie-Tooth type 2A neuropathy. Neuromuscul Disord 21(1):58–67. Available from: CrossRefPubMedGoogle Scholar
  29. 29.
    Chapman AL, Bennett EJ, Ramesh TM, De Vos KJ, Grierson AJ (2013) Axonal transport defects in a mitofusin 2 loss of function model of Charcot-Marie-Tooth disease in zebrafish. PLoS One 8(6)Google Scholar
  30. 30.
    Pareyson D, Saveri P, Pisciotta C (2017) New developments in Charcot–Marie–Tooth neuropathy and related diseases. Curr Opin Neurol 30:471–480CrossRefGoogle Scholar
  31. 31.
    Eldridge CF, Bunge MB, Bunge RP (1989) Differentiation of axon-related Schwann cells in vitro: II. Control of myelin formation by basal lamina. J Neurosci 9(2):625–638CrossRefGoogle Scholar
  32. 32.
    Gess B, Baets J, De Jonghe P, Reilly MM, Pareyson D, Young P (2015) Ascorbic acid for the treatment of Charcot-Marie-Tooth disease ( review ). Cochrane Database Syst Rev (12)Google Scholar
  33. 33.
    Sereda MW, Meyer Zu Hörste G, Suter U, Uzma N, Nave KA (2003) Therapeutic administration of progesterone antagonist in a model of Charcot-Marie-Tooth disease (CMT-1A). Nat Med 9(12):1533–1537CrossRefGoogle Scholar
  34. 34.
    Chahbouni M, Lòpez MS, Molina-Carballo A, Haro D, Muñoz-Hoyos A, Fernàndez-Ortis M, Guerra-Librero A, Acuna-Castroviejo D (2017) Melatonin treatment reduces oxidative damage and normalizes plasma pro-inflammatory cytokines in patients suffering from Charcot-Marie-Tooth neuropathy: a pilot study in three children. Molecules 22(1728):1-14. Available from:
  35. 35.
    Smith CA, Chetlin RD, Gutmann L, Yeater RA, Alway SE (2006) Effects of exercise and creatine on myosin heavy chain isoform composition in patients with Charcot-Marie-Tooth disease. Muscle Nerve 34(5):586–594CrossRefGoogle Scholar
  36. 36.
    Franco A, Kitsis RN, Fleischer JA, Gavathiotis E, Kornfeld OS, Gong G, Biris N, Benz A et al (2016) Correcting mitochondrial fusion by manipulating mitofusin conformations. Nature 540(7631):74–79. Available from: CrossRefPubMedPubMedCentralGoogle Scholar
  37. 37.
    Rocha AG, Franco A, Krezel AM, Rumsey JM, Alberti JM, Knight WC et al (2018) MFN2 agonists reverse mitochondrial defects in preclinical models of Charcot-Marie-Tooth disease type 2A. Science (80- ) 360(6386):336–341CrossRefGoogle Scholar
  38. 38.
    Mendell JR, Al-Zaidy S, Shell R, Arnold WD, Rodino-Klapac LR, Prior TW et al (2017) Single-dose gene-replacement therapy for spinal muscular atrophy. N Engl J Med 377(18):1713–1722. Available from: CrossRefPubMedGoogle Scholar
  39. 39.
    Xu GX, Zhou H, Zhou S, Yu Y, Wu R, Xu Z (2005) An RNAi strategy for treatment of amyotrophic lateral sclerosis caused by mutant Cu,Zn superoxide dismutase. J Neurochem 92(2):362–367CrossRefGoogle Scholar
  40. 40.
    Rinaldi C, Wood MJA (2017) Antisense oligonucleotides: the next frontier for treatment of neurological disorders. Nat Rev Neurol. Available from:
  41. 41.
    Herrmann DN (2008) Experimental therapeutics in hereditary neuropathies: the past, the present, and the future. Neurotherapeutics. 5(4):507–515CrossRefGoogle Scholar
  42. 42.
    Hoy SM (2017) Nusinersen: first global approval. Drugs. 77(4):473–479CrossRefGoogle Scholar
  43. 43.
    Voelker R (2016) First DMD drug gains approval. JAMA 316(17):1756Google Scholar
  44. 44.
    Deng Y, Wang CC, Choy KW, Du Q, Chen J, Wang Q et al (2014) Therapeutic potentials of gene silencing by RNA interference: principles, challenges, and new strategies. Gene 538(2):217–227CrossRefGoogle Scholar
  45. 45.
    Aagaard L, Rossi JJ (2007) RNAi therapeutics: principles, prospects and challenges. Adv Drug Deliv Rev 59(2–3):75–86CrossRefGoogle Scholar
  46. 46.
    Lam JKW, Chow MYT, Zhang Y, Leung SWS (2015) siRNA versus miRNA as therapeutics for gene silencing. Mol Ther - Nucleic Acids 4(9):1–20Google Scholar
  47. 47.
    Cong L, Ran FA, Cox D, Lin S, Barretto R, Habib N, Hsu PD, Wu X, Jiang W, Marraffini LA, Zhang F (2013) Multiplex genome engineering using CRISPR/Cas systems. Science 339(6121):819-823. Available from:
  48. 48.
    Zhan T, Rindtorff N, Betge J, Ebert MP, Boutros M (2018) CRISPR/Cas9 for cancer research and therapy. Semin Cancer Biol. Available from:
  49. 49.
    Zhang XH, Tee LY, Wang XG, Huang QS, Yang SH (2015) Off-target effects in CRISPR/Cas9-mediated genome engineering. Mol Ther - Nucleic Acids 4(11):e264. Available from:
  50. 50.
    Liao HK, Hatanaka F, Araoka T, Reddy P, Wu MZ, Sui Y et al (2017) In vivo target gene activation via CRISPR/Cas9-mediated trans-epigenetic modulation. Cell 171(7):1495–1507.e15CrossRefGoogle Scholar
  51. 51.
    Baylis F, McLeod M (2017) First-in-human phase 1 CRISPR gene editing cancer trials: are we ready? Curr Gene Ther 17:309–319. Available from:
  52. 52.
    Joshi CR, Labhasetwar V, Ghorpade A (2017) Destination brain: The past, present, and future of therapeutic gene delivery. J NeuroImmune Pharmacol 12(1):51–83CrossRefGoogle Scholar
  53. 53.
    DeVos SL, Miller T (2013) Direct intraventricular delivery of drugs to the rodent central nervous system. J Vis Exp.75:e50326. Available from:
  54. 54.
    Weinberg MS, Samulski RJ, McCown TJ (2013) Adeno-associated virus (AAV) gene therapy for neurological disease. Neuropharmacology 69:82–88. Available from: CrossRefPubMedGoogle Scholar
  55. 55.
    Klein RL, Dayton RD, Leidenheimer NJ, Jansen K, Golde TE, Zweig RM (2006) Efficient neuronal gene transfer with AAV8 leads to neurotoxic levels of tau or green fluorescent proteins. Mol Ther 13(3):517–527. Available from: CrossRefPubMedGoogle Scholar
  56. 56.
    Markakis EA, Vives KP, Bober J, Leichtle S, Leranth C, Beecham J, Elsworth JD, Roth RH et al (2010) Comparative transduction efficiency of AAV vector serotypes 1-6 in the substantia nigra and striatum of the primate brain. Mol Ther 18(3):588–593. Available from: CrossRefPubMedGoogle Scholar
  57. 57.
    Hadaczek P, Forsayeth J, Mirek H, Munson K, Bringas J, Pivirotto P, McBride JL, Davidson BL, Bankiewicz KS (2009) Transduction of nonhuman primate brain with adeno-associated virus serotype 1: vector trafficking and immune response. Hum Gene Ther 20:225–237Google Scholar
  58. 58.
    Foust KD, Nurre E, Montgomery CL, Hernandez A, Chan CM, Kaspar BK (2009) Intravascular AAV9 preferentially targets neonatal neurons and adult astrocytes. Nat Biotechnol 27(1):59–65CrossRefGoogle Scholar
  59. 59.
    Foust KD, Salazar DL, Likhite S, Ferraiuolo L, Ditsworth D, Ilieva H, Meyer K, Schmelzer L, Braun L, Cleveland DW, Kaspar BK (2013) Therapeutic AAV9-mediated suppression of mutant SOD1 slows disease progression and extends survival in models of inherited ALS. Molecular Therapy 21(12):2148–2159. Available from:
  60. 60.
    Meyer K, Ferraiuolo L, Schmelzer L, Braun L, Mcgovern V, Likhite S et al (2015) Improving single injection CSF delivery of AAV9-mediated gene therapy for SMA: a dose–response study in mice and nonhuman primates. Mol Ther 23(3):477–487. Available from: CrossRefPubMedGoogle Scholar
  61. 61.
    Miyanohara A, Kamizato K, Juhas S, Juhasova J, Navarro M, Marsala S et al (2016) Potent spinal parenchymal AAV9-mediated gene delivery by subpial injection in adult rats and pigs. Mol Ther - Methods Clin Dev 3:16046. Available from: CrossRefPubMedPubMedCentralGoogle Scholar
  62. 62.
    Ameri H (2018) Prospect of retinal gene therapy following commercialization of voretigene neparvovec-rzyl for retinal dystrophy mediated by RPE65 mutation. J Curr Ophthalmol 30(1):1–2. Available from: CrossRefPubMedPubMedCentralGoogle Scholar
  63. 63.
    Nizzardo M, Simone C, Rizzo F, Salani S, Dametti S, Rinchetti P et al (2015) Gene therapy rescues disease phenotype in a spinal muscular atrophy with respiratory distress type 1 (SMARD1) mouse model. Sci Adv 1(2):1–10CrossRefGoogle Scholar
  64. 64.
    Johnson-Kerner BL, Roth L, Greene JP, Wichterle H, Sproule DM (2014) Giant axonal neuropathy: An updated perspective on its pathology and pathogenesis. Muscle Nerve 50(4):467–476CrossRefGoogle Scholar
  65. 65.
    Sahenk Z, Nagaraja HN, McCracken BS, King WM, Freimer ML, Cedarbaum JM et al (2005) NT-3 promotes nerve regeneration and sensory improvement in CMT1A mouse models and in patients. Neurology. 65(5):681–689CrossRefGoogle Scholar
  66. 66.
    Flax JD, Aurora S, Yang C, Simonin C, Wills AM, Billinghurst LL, Jendoubi M, Sidman RL et al (1998) Engraftable human neural stem cells respond to development cues, replace neurons, and express foreign genes. Nat Biotechnol 16(11):1033–1039CrossRefGoogle Scholar
  67. 67.
    Corti S, Locatelli F, Papadimitriou D, Donadoni C, Del Bo R, Crimi M et al (2006) Transplanted ALDHhiSSCloneural stem cells generate motor neurons and delay disease progression of nmd mice, an animal model of SMARD1. Hum Mol Genet 15(2):167–187CrossRefGoogle Scholar
  68. 68.
    Nizzardo M, Bucchia M, Ramirez A, Trombetta E, Bresolin N, Comi GP et al (2015) iPSC-derived LewisX+CXCR4+β1-integrin+ neural stem cells improve the amyotrophic lateral sclerosis phenotype by preserving motor neurons and muscle innervation in human and rodent models. Hum Mol Genet 25(15):3152–3163CrossRefGoogle Scholar
  69. 69.
    Yang YM, Gupta SK, Kim KJ, Powers BE, Cerqueira A, Wainger BJ, Ngo HD, Rosowski KA et al (2013) A small molecule screen in stem-cell-derived motor neurons identifies a kinase inhibitor as a candidate therapeutic for ALS. Cell Stem Cell 12(6):713–726. Available from: CrossRefPubMedPubMedCentralGoogle Scholar
  70. 70.
    Simone C, Nizzardo M, Rizzo F, Ruggieri M, Riboldi G, Salani S, Bucchia M, Bresolin N et al (2014) IPSC-derived neural stem cells act via kinase inhibition to exert neuroprotective effects in spinal muscular atrophy with respiratory distress type 1. Stem Cell Reports 3(2):297–311. Available from: CrossRefPubMedPubMedCentralGoogle Scholar
  71. 71.
    Leal A, Ichim TE, Marleau AM, Lara F, Kaushal S, Riordan NH (2008) Immune effects of mesenchymal stem cells: Implications for Charcot-Marie-Tooth disease. Cell Immunol 253(1–2):11–15CrossRefGoogle Scholar
  72. 72.
    Kimbrel EA, Lanza R (2015) Current status of pluripotent stem cells: moving the first therapies to the clinic. Nat Rev Drug Discov 14(10):681–692. Available from: CrossRefPubMedGoogle Scholar
  73. 73.
    Shi Y, Inoue H, Wu JC, Yamanaka S (2017) Induced pluripotent stem cell technology: a decade of progress. Nat Rev Drug Discov 16(2):115–130. Available from: CrossRefPubMedGoogle Scholar
  74. 74.
    Murphy SM, Herrmann DN, McDermott MP, Scherer SS, Shy ME, Reilly MM et al (2011) Reliability of the CMT neuropathy score (second version) in Charcot-Marie-Tooth disease. J Peripher Nerv Syst 16(3):191–198CrossRefGoogle Scholar
  75. 75.
    Cornett KMD, Menezes MP, Shy RR, Moroni I, Pagliano E, Pareyson D, Estilow T, Yum SW et al (2017) Natural history of Charcot-Marie-Tooth disease during childhood. Ann Neurol 82(3):353–359CrossRefGoogle Scholar
  76. 76.
    Fledrich R, Mannil M, Leha A, Ehbrecht C, Solari A, Pelayo-Negro AL, Berciano J, Schlotter-Weigel B et al (2017) Biomarkers predict outcome in Charcot-Marie-Tooth disease 1A. J Neurol Neurosurg Psychiatry 88(11):941–952CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Kordelia Barbullushi
    • 1
  • Elena Abati
    • 1
  • Federica Rizzo
    • 1
  • Nereo Bresolin
    • 1
    • 2
  • Giacomo P. Comi
    • 1
    • 2
  • Stefania Corti
    • 1
    • 2
    Email author
  1. 1.Dino Ferrari Centre, Neuroscience Section, Department of Pathophysiology and Transplantation (DEPT)University of MilanMilanItaly
  2. 2.Foundation IRCCS Ca’ Granda Ospedale Maggiore Policlinico, Neurology UnitMilanItaly

Personalised recommendations