Abstract Cancer stem cells (CSCs) are amongst the group of cells constituting tumors, being characterized by their strong self-renewal and survival properties. Cancer cells, CSCs included, are thought to rely mostly on glycolysis, even in the presence of oxygen, which confers them adaptative advantages. Adenine nucleotide translocator 2 (ANT2), responsible for the exchange of ADP and ATP in the mitochondrial inner membrane, has been correlated with a higher glycolytic metabolism and is known to be overexpressed in cancer cells. Using P19 embryonal carcinoma stem cells (P19SCs) as a CSCs model, we inhibit ANT2 translation by using siRNA. ANT2 protein levels were shown to be overexpressed in P19SC when compared to their differentiated counterparts. Furthermore, we showed here that the OXPHOS machinery and mitochondrial membrane potential are compromised after ANT2 depletion, exhibiting a metabolic adaptation towards a less oxidative phenotype. Interestingly, hexokinase II levels were downregulated, which was also accompanied by decreased cell growth, and decreased ability to form spheroids. Our findings underscore ANT2 as a key regulator of metabolic remodeling and cell survival of CSCs, suggesting its potential as a therapeutic target for controlling CSC-driven tumor progression. Highlights ANT2 silencing promotes cell growth arrest and metabolic remodeling in CSCs. ANT2 depletion modulates HKII protein levels. ANT2 induce anoikis resistance in P19SCs

Adenine nucleotide translocase 2 silencing promotes metabolic adaptations and anoikis in P19 embryonal carcinoma stem cells