Characterization of molecular mechanisms underlying the axonal Charcot-Marie-Tooth neuropathy caused by MORC2 mutations.

Mutations in MORC2 lead to an axonal form of Charcot-Marie-Tooth (CMT) neuropathy type 2Z. To date, 31 families have been described with mutations in MORC2, indicating that this gene is frequently involved in axonal CMT cases. While the genetic data clearly establish the causative role of MORC2 in CMT2Z, the impact of its mutations on neuronal biology and their phenotypic consequences in patients remains to be clarified. We show that the full-length form of MORC2 is highly expressed in both embryonic and adult human neural tissues and that Morc2 expression is dynamically regulated in both the developing and the maturing murine nervous system. To determine the effect of the most common MORC2 mutations, p.S87L and p.R252W, we used several in vitro cell culture paradigms. Both mutations induced transcriptional changes in patient-derived fibroblasts and when expressed in rodent sensory neurons. These changes were more pronounced and accompanied by abnormal axonal morphology, in neurons expressing the MORC2 p.S87L mutation, which is associated with a more severe clinical phenotype. These data provide insight into the neuronal specificity of the mutated MORC2-mediated phenotype and highlight the importance of neuronal cell models to study the pathophysiology of CMT2Z.

MORC2 mutations cause axonal Charcot-Marie-Tooth disease with pyramidal signs.

OBJECTIVE: To use linkage analysis and whole exome sequencing to identify the genetic mutation in a multigenerational Australian family with Charcot-Marie-Tooth disease type 2 (CMT2) and pyramidal signs. METHODS: Genome-wide linkage analysis was performed to map the locus. Whole exome sequencing was undertaken on selected individuals (3 affected, 1 normal), and segregation analysis and mutation screening were carried out using high-resolution melt analysis. The GEM.app database was queried to identify additional families with mutations. RESULTS: Significant linkage (2-point LOD score ≥ +3) and haplotype analysis mapped a new locus for CMT2 and pyramidal signs to a 6.6Mb interval on chromosome 22q12.1-q12.3. Whole exome sequencing identified a novel mutation (p.R252W) in the microrchidia CW-type zinc finger 2 (MORC2) gene mapping within the linkage region. The mutation fully segregated with the disease phenotype in the family. Screening additional families and querying unsolved CMT2 exomes, we identified the p.R252W mutation in 2 unrelated early onset CMT2 families and a second mutation p.E236G in 2 unrelated CMT2 families. Both the mutations occurred at highly conserved amino acid residues and were absent in the normal population. INTERPRETATION: We have identified a new locus in which MORC2 mutations are the likely pathogenic cause of CMT2 and pyramidal signs in these families. MORC2 encodes the human CW-type zinc finger 2 protein, which is a chromatin modifier involved in the regulation of DNA repair as well as gene transcription.

X-linked Charcot-Marie-Tooth disease type 6 (CMTX6) patients with a p.R158H mutation in the pyruvate dehydrogenase kinase isoenzyme 3 gene.

Charcot-Marie-Tooth disease (CMT) is the most common inherited peripheral neuropathy. Mutations in the pyruvate dehydrogenase kinase isoenzyme 3 (PDK3) gene have been found to cause X-linked dominant CMT type 6 (CMTX6). This study identified the p.R158H PDK3 mutation after screening 67 probable X-linked CMT families. The mutation fully segregated with the phenotype, and genotyping the family indicated the mutation arose on a different haplotype compared with the original Australian CMTX6 family. Results of bisulphite sequencing suggest that methylated deamination of a CpG dinucleotide may cause the recurrent p.R158H mutation. The frequency of the p.R158H PDK3 mutation in Koreans is very rare. Magnetic resonance imaging revealed fatty infiltration involving distal muscles in the lower extremities. In addition, fatty infiltrations were predominantly observed in the soleus muscles, with a lesser extent in tibialis anterior muscles. This differs from demyelinating CMT1A patients and is similar to axonal CMT2A patients. The clinical, neuroimaging, and electrophysiological findings from a second CMTX6 family with the p.R158H PDK3 mutation were similar to the axonal neuropathy reported in the Australian family.

A case study of a long-term glioblastoma survivor with unmethylated MGMT and hypermutated genotype.

Effective treatments that extend survival of malignant brain tumor glioblastoma (GBM) have not changed in more than a decade; however, there exists a minority patient group (<5%) whose survival is longer than 3 yr. We herein present a case report of a long-term surviving 51-yr-old female diagnosed with a MGMT unmethylated GBM. The patient was progression-free for 23 mo. Fresh primary and recurrent tumor samples were collected and processed for patient-derived model development. Whole-genome sequencing (WGS) was performed concurrently with additional standard of care diagnostics. WGS revealed a hypermutated genotype in the germline tissue and in both the primary and recurrent tumor samples. Specific to the matched tumors, an average of 30 cancer driver genes were mutated. Noteworthy was the identification of a nonsynonymous mutation in the POLE gene. As a possible instigator of the hypermutational genotype observed in the tumors, we identified nonsynonymous germline mutations within the mismatch repair genes, MLH1 and PMS2 Mutations within these genes are often indicative of the pan-cancer phenotype known as Lynch syndrome; however, their pathogenicity remains unreported. We performed a drug screen of 165 compounds, which identified one compound, YM155, an experimental survivin inhibitor, that showed effectivity to the patient-derived cell lines of both tumors. Treatment selection based on a patient's genome to individualize treatment for GBM patients could potentially be useful in the clinic. This is a promising avenue for further translational research, with larger databases and integrated platforms to increase the efficiency of analyzing and interpreting the individual genomic data of GBM.

Whole genome and biomarker analysis of patients with recurrent glioblastoma on bevacizumab: A subset analysis of the CABARET trial.

The CABARET trial (ACTRN12610000915055) reported no difference in overall survival (OS) between patients with recurrent glioblastoma (GBM) randomized to either bevacizumab monotherapy or bevacizumab plus carboplatin. However, a subset of patients showed durable responses and prolonged survival, with recorded survival times of over 30 months in five of 122 patients (4%). Patient selection for bevacizumab therapy would be enhanced if a predictive biomarker of response or survival could be identified; this biomarker sub-study attempted to identify novel biomarkers. Patients who opted to participate in this sub-study and who had adequate biospecimens for analysis (n = 54) were retrospectively evaluated for the expression of a series of tumor proteins. Immunohistochemistry (IHC) was used to measure the expression of 19 proteins previously implicated in cancer treatment response to bevacizumab. MGMT promoter methylation was also assessed. Tumor DNA from five patients with outlying survival duration ('poor' and 'exceptional' survivors) was subjected to whole genome sequencing (WGS). No single protein expression level, including VEGF-A, predicted OS in the cohort. WGS of poor and exceptional survivors identified a gain in Chromosome 19 that was exclusive to the exceptional survivors. Validation of this finding requires examination of a larger independent cohort.