Hypoxia induces alterations in the volatile signature of pleural mesothelioma cells

Volatile organic compounds in exhaled breath are promising, non-invasive biomarkers for pleural mesothelioma diagnosis. Studying headspace volatile patterns at the cellular level is valuable to elucidate the biological origin and underlying metabolic pathways of these potential biomarkers. However, it is important to mimic the in vivo physiological conditions as closely as possible. This study investigated the impact of an in vivo-like hypoxic microenvironment on the volatile profile of pleural mesothelioma cells. NCI-H2818 (epithelioid), NCI-H2731 (sarcomatoid) and NKI04 (biphasic) cell lines were cultured under normoxic (21 % O2) and hypoxic (1 % O2) conditions before characterizing their headspace volatile organic compound profiles using gas chromatography-mass spectrometry. A combination of multivariate and univariate statistics was applied to identify significant alterations in volatile patterns between both oxygen conditions and between the prognostically more favorable epithelioid subtype and the less favorable non-epithelioid subtype. Eighty-six volatiles significantly changed between hypoxic and normoxic mesothelioma cells. Increased degradation of aldehydes was observed upon hypoxic exposure alongside increased production of several hydrocarbons, alcohols and ketones. Additionally, different volatiles were significantly altered between the epithelioid and non-epithelioid subtype, with most differential metabolites being specific to hypoxia. This study demonstrated a significant impact of hypoxia on the pleural mesothelioma volatile profile, providing mechanistic insights into how hypoxia alters cellular metabolism in mesothelioma, thereby more closely reflecting patterns observed in patient breath. This stresses the importance of using in vivo-like hypoxic cell culture conditions to identify cancer-related volatile organic compounds for future clinical biomarker development.