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Development of genetic quality tests for good manufacturing practice-compliant induced pluripotent stem cells and their derivatives

ABSTRACT: Purpose : We investigated RNA-seq-based broad-range genetic quality tests on GMP-compliant human leucocyte antigen (HLA)-homozygous hiPSCs and their derivatives under postdistribution conditions to investigate whether sequencing data could provide a basis for future quality control. Methods : Transcriptome profiling of the differentiated cell lines in three lineages, including neuronal cells, cardiomyocytes and hepatocytes, at early and late passages, respectively, were carried out using RNA sequencing. Results : We performed SNV, indel, CNV, ekaryotyping, HLA genotyping and transcriptomic analyses in which differentiated cells were compared with hiPSCs using RNA-seq data from undifferentiated and differentiated cultures returned from multiple centres. In particular, we evaluated whether prolonged culture conditions have the potential to give rise to genomic and phenotypic changes in the CMC3 hiPSC line, a candidate line for manufacturing cell therapies. Conclusion : RNA-seq analysis appears to offer an informative genetic quality testing approach for cell types and allows the early screening of candidate hiPSC seed stocks for clinical use by facilitating safety and potential risk evaluation

ORGANISM(S): Homo sapiens

PROVIDER: GSE148517 | GEO | 2020/04/13

REPOSITORIES: GEO

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Project description:Cell-based models of many neurological and psychiatric diseases, established by reprogramming patient somatic cells into human induced pluripotent stem cells (hiPSCs), have now been reported. While numerous reports have demonstrated that neuronal cells differentiated from hiPSCs are electrophysiologically active mature neurons, the âageâ of these cells relative to cells in the human brain remains unresolved. Comparisons of gene expression profiles of hiPSC-derived neural progenitor cells (NPCs) and neurons to the Allen BrainSpan Atlas indicate that hiPSC neural cells most resemble first trimester neural tissue. Consequently, we posit that hiPSC-derived neural cells may most accurately be used to model the early developmental defects that contribute to disease predisposition rather than the late features of the disease. Though the characteristic symptoms of schizophrenia SZ generally appear late in adolescence, it is now thought to be a neurodevelopmental condition, often predated by a prodromal period that can appear in early childhood. Postmortem studies of SZ brain tissue typically describe defects in mature neurons, such as reduced neuronal size and spine density in the prefrontal cortex and hippocampus, but abnormalities of neuronal organization, particularly in the cortex, have also been reported. We postulated that defects in cortical organization in SZ might result from abnormal migration of neural cells. To test this hypothesis, we directly reprogrammed fibroblasts from SZ patients into hiPSCs and subsequently differentiated these disorder-specific hiPSCs into NPCs. SZ hiPSC differentiated into forebrain NPCs have altered expression of a number of cellular adhesion genes and WNT signaling. Methods: We compared global transcription of forebrain NPCs from six control and four SZ patients by RNAseq. Results: Multi-dimensional scaling (MDS) resolved most SZ and control hiPSC NPC samples; 848 genes were significantly differentially expressed (FDR<0.01) Conclusions: The WNT signaling pathway was enriched 2-fold (fisher exact test p-value = 0.031). 1-2 independent differentiations (biological replicates) for each of four control and four schizophrenia patients were analyzed; samples were generated in parallel to neuron RNAseq data.

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Development of genetic quality tests for good manufacturing practice-compliant induced pluripotent stem cells and their derivatives

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