Cytogenetic resource enables mechanistic resolution of changing trends in human pluripotent stem cell aberrations linked to feeder-free culture

Abstract

Since the first derivation of human pluripotent stem cells (hPSCs), the number of culture conditions has steadily increased, making hPSC culture more facile. Nonetheless, there remains the persistent issue of culture-acquired genetic changes, hampering the reproducibility of hPSC research and jeopardising their clinical use. Here, we utilised comprehensive karyotyping datasets from over 20,000 hPSC cultures sampled under different conditions to ascertain association of genetic changes with specific culture regimens. We found condition-dependent patterns of aberrations, with higher prevalence of chromosome 1q gains in recent years, associated with increased use of contemporary, feeder-free cultures. Mechanistically, we show the context-dependent selection of 1q variants is mainly driven by MDM4, a gene amplified in many cancers, located on chromosome 1q. To facilitate reproducibility of hPSC research and their safe clinical utility, we provide a unique hPSC karyotype resource for informing the risk assessment of genetic aberrations and developing strategies to suppress their occurrence.

Aberrant Wnt activation in recurrent genetically variant human pluripotent stem cells impairs cardiomyocyte differentiation and phenotype

“…Specifically, we noted an increase in prevalence of chromosome 1q gains in recent years, associated with increased use of feeder-free cultures 15 . Our work 15 and that of others 16 , showed that MDM4 is a likely driver gene for the selective advantage of variant 1q cells, allowing variants to rapidly take over cultures through faster proliferation and reduced apoptosis. Given the prevalence of chromosome 1q variants in hPSC culture in recent years, a question arises whether the gains of chromosome 1q affect downstream application of hPSCs, such as their use for generating differentiated cells?…”

“…Although the recurrent nature of genetic aberrations in hPSCs is well established, our recent large-scale analysis of karyotyping datasets from over 23,000 hPSC cultures demonstrated that trends of karyotypic abnormalities are culture condition-dependent 15 . Specifically, we noted an increase in prevalence of chromosome 1q gains in recent years, associated with increased use of feeder-free cultures 15 .…”

“…Although the recurrent nature of genetic aberrations in hPSCs is well established, our recent large-scale analysis of karyotyping datasets from over 23,000 hPSC cultures demonstrated that trends of karyotypic abnormalities are culture condition-dependent 15 . Specifically, we noted an increase in prevalence of chromosome 1q gains in recent years, associated with increased use of feeder-free cultures 15 . Our work 15 and that of others 16 , showed that MDM4 is a likely driver gene for the selective advantage of variant 1q cells, allowing variants to rapidly take over cultures through faster proliferation and reduced apoptosis.…”

Gain of chromosome region 1q31.3 in human iPSCs confers growth advantage and alters contraction in derivative cardiomyocytes

“…Over extended passages, the cells harbouring an interstitial duplication in amplicon 1q31.3 gained a selective growth advantage and eventually dominated the hiPSC culture, although it was possible to isolate and maintain the wild type hiPSCs prior to this occurring. Aberrations in chromosome 1q are recurrent in hPSCs, with the prevalence of this abnormality increasing in recent years and correlating with a shift from the culturing of hPSCs on mouse embryonic fibroblasts (Stavish et al, 2023).…”

“…Recent studies have reported that the anti-apoptotic gene MDM4, located in the duplicated chromosomal region, as the likely driver for the competitive advantage of hPSCs containing a gain of chromosome 1q (Krivec et al, 2023;Stavish et al, 2023).…”