Genetic correction and analysis of induced pluripotent stem cells from a patient with gyrate atrophy.

Gene-corrected patient-specific induced pluripotent stem (iPS) cells offer a unique approach to gene therapy. Here, we begin to assess whether the mutational load acquired during gene correction of iPS cells is compatible with use in the treatment of genetic causes of retinal degenerative disease. We isolated iPS cells free of transgene sequences from a patient with gyrate atrophy caused by a point mutation in the gene encoding ornithine-δ-aminotransferase (OAT) and used homologous recombination to correct the genetic defect. Cytogenetic analysis, array comparative genomic hybridization (aCGH), and exome sequencing were performed to assess the genomic integrity of an iPS cell line after three sequential clonal events: initial reprogramming, gene targeting, and subsequent removal of a selection cassette. No abnormalities were detected after standard G-band metaphase analysis. However, aCGH and exome sequencing identified two deletions, one amplification, and nine mutations in protein coding regions in the initial iPS cell clone. Except for the targeted correction of the single nucleotide in the OAT locus and a single synonymous base-pair change, no additional mutations or copy number variation were identified in iPS cells after the two subsequent clonal events. These findings confirm that iPS cells themselves may carry a significant mutational load at initial isolation, but that the clonal events and prolonged cultured required for correction of a genetic defect can be accomplished without a substantial increase in mutational burden.

Clonal chromosome abnormalities in a plexiform cellular schwannoma.

Cellular schwannomas are uncommon tumors of Schwann cells that can rarely have a plexiform architecture. Cellular schwannomas can be confused with low-grade malignant peripheral nerve sheath tumors (MPNST) but have been noted to have a benign clinical course. There are few published cytogenetic analyses of cellular schwannomas and, to our knowledge, there are no reports of the plexiform variant of cellular schwannoma to date. Cellular schwannomas are reported to have cytogenetic changes similar to those seen in benign schwannomas with near-diploid karyotypes having simple numerical changes often involving chromosomes 22, 7, and the sex chromosomes. MPNST are markedly different, with extensive genetic heterogeneity and complex karyotypes. We report clonal numerical changes in a cellular schwannoma with plexiform architecture: 47,XY,+17 and 48,XY,+17,+18. These findings add to the karyotypic spectrum of peripheral nerve sheath tumors.