The effect of acute and prolonged acidosis on a panel of carcinoma cell lines

Due to cancer cell metabolism and disorganized tissue structure, the tumour microenvironment is associated with several pathophysiological conditions, including hypoxia, nutrient deprivation, accumulation of waste products, and acidification of the tumour microenvironment (tumour acidosis). Despite the belief that tumour acidosis drives tumorigenesis, it is still unclear how cancer cells respond to acidosis in the absence of other pathophysiological conditions. Here, we investigate how both acute and prolonged acidosis (pH 6.8) affects different epithelial features of a panel of carcinoma cell lines. We find that acute acidosis in all cell lines investigated represses cell growth and causes a disturbance of adherens junctions and apical-basal polarity, reminiscent of epithelial-to-mesenchymal transition (EMT). However, these changes did not coincide with altered expression of E− and N-cadherin. Neither did acute acidosis have a general effect on adhesion and migration in our panel of cell lines. Exposing our panel of carcinoma cell lines to acidosis for more than six weeks did not lead to adaptations restoring cell growth. On the contrary, prolonged acidosis caused one cell line (A431) to halt proliferation. Another cell line (A549) reacted to prolonged acidosis by gradually inducing the expression of ZEB2, which in turn orchestrated cadherin switching, possibly indicating a gradual induction of EMT. In the rest of the cell lines, we did not find any noticeable effect of prolonged acidosis. Lastly, all cell lines quickly restored their original phenotype and growth rate when returned to media with normal pH (pH 7.5). Collectively, our findings reveal that carcinoma cells exposed to moderate acidosis (pH 6.8) generally exhibit slower proliferation rates and a reduction in apical-basal polarity. Furthermore, we conclude that prolonged exposure to acidic conditions, depending on the specific cell line, may elicit responses that could influence tumour progression and the efficacy of cancer treatments. Key Scientific Contributions . • The study systematically investigates both acute and prolonged acidosis (pH 6.8) across a panel of carcinoma cell lines (A431, A549, HCT116, HT29, MDA-MB-231), providing a comparative and robust analysis of how different cancer cell types respond to acidosis. • It demonstrates that acute acidosis consistently reduces cell growth across all lines and disrupts epithelial features such as adherens junctions and apical-basal polarity, resembling early stages of epithelial-to-mesenchymal transition (EMT), but without classic cadherin switching. • The manuscript reveals that prolonged acidosis does not generally lead to adaptation but rather to cell line-specific outcomes: A431 cells enter a dormant, low-proliferation state, while A549 cells undergo a partial and reversible EMT mediated by upregulation of ZEB2 and SNAIL2, with corresponding changes in E− and N-cadherin expression. • These findings highlight that cancer cells do not universally adapt to acidosis and that responses are highly context-dependent. • The study provides evidence that the morphological and transcriptional changes induced by acidosis are reversible upon return to normal pH, suggesting plasticity in cancer cell responses to microenvironmental stress. • It underscores that acidosis-induced changes are not necessarily mediated by conventional transcriptional pathways but may arise from subtle, cumulative effects on protein function and cellular metabolism. Broader Scientific Impact . • The manuscript challenges the notion that acidosis universally drives tumorigenesis or EMT in all cancer cells. Instead, it shows that while acidosis can induce EMT-like changes in some contexts, this is not a general phenomenon and is reversible. • It also questions the idea that cancer cells readily adapt to acidosis, as adaptation was only observed in a minority of cell lines and was not sustained. • The findings have important implications for the development of therapies targeting the tumour microenvironment. They suggest that strategies exploiting acidosis may need to be tailored to specific cancer types or combined with other microenvironmental factors. • The study highlights the complexity of tumour microenvironmental interactions and the need for more nuanced, integrated approaches to cancer therapy.