Dr. Vrije Universiteit Brussel (VUB) Jette, Brussels Hoofdstedelijk Gewest, Belgium
Abstract: Human pluripotent stem cells (hPSCs) are increasingly recognized for their potential in cell-based regenerative therapies for injuries and chronic conditions. However, hPSCs in culture frequently acquire chromosomal abnormalities (CA), raising safety concerns regarding their therapeutic use. Strikingly, these CAs resemble genetic aberrations found in cancers. It remains unclear whether CAs can predispose differentiated cells to oncogenic transformation, a gap stemming from a lack of suitable research models and systematic studies. In this work, we hypothesize that CAs represent a first hit in the oncogenic process, enhancing the potential of differentiated hPSCs to transform upon subsequent oncogenic hits. We used an in vitro organoid-based model of brain tumorigenesis, where we subject set of genetically balanced lines (VUB01, VUB02, VUB03, VUB04, VUB07, VUB14 and VUB19) as controls, and lines with well-characterized recurrent CAs (gains of 1q, 12p, 17q, 20q and losses of 18q), to directed mutagenesis. This approach allowed investigating genetic defects that could enhance the tumorigenic potential of hPSC-derived cells. Our findings indicate that hESCs with CAs, particularly those with gains on 20q11.21 (commonly found in glioblastomas), 1q24.2, and 17q, are more likely to form tumorous overgrowth in brain organoids (95-100% transformation rate) following mutagenic transformation via cMyc overexpression, compared to hESCcontrol organoids (20-30%). Organoids from hESCs with gains of 12p or losses of 18q exhibited lower transformation rates (20-40%). Notably, organoids with transformed overgrowth retained viability and expanded upon intracranial transplantation in immunodeficient mice, with transformed GFP+ cells proliferating and generating CD99+/SOX2+/GFP+ neoplastic-like regions. Moreover, hESCs with CAs, particularly those with gains on 12p, 20q, and losses on 18q, formed organoids with disorganized shapes compared to control organoids. This suggests that CAs not only enhance the transformation capacity of hPSCs but also impair their ability to develop into normal brain organoids. Our work provides insights into the functional impact of CAs in hPSCs and valuable knowledge for assessing the long-term risks associated with transplanting hPSC-derived cells with genetic defects.
