Oxidative stress contributes to the functional decline of the myocardium and is correlated with unfavourable metabolic perturbations, including the switch from oxidative metabolism to glycolysis for energy production. Oxidative post-translational modifications (oxPTMs) might play a significant role in this process as they can influence both structure as well as the function of proteins. We already demonstrated that failing human hearts endure higher levels of oxidative stress and exhibit higher oxidation of several different enzymes, namely related to glucose metabolism. To unravel which of these oxidative changes might be connected with the early onset of oxidative damage, we simulated oxidative injury using cultured cardiomyocytes as well as stem-cell derived and metabolically matured cardioids, followed by redox analysis. As a result, we report pyruvate kinase M (PKM) as one of the most redox-sensitive enzymes. PKM might be a critical target as a) its fetal isoform, PKM2 was increased in failing hearts, and b) PKM2 was proven susceptible to oxPTM which can lead to inhibition of the enzyme. Treatment of metabolically matured cardioids with TEPP46, a potent PKM2 activator, was unable to hinder the activation of apoptosis upon oxidative injury; however, TEPP46 overall seems to improve cellular antioxidative defence as well as lead to a higher reliance on lipid metabolism (even under hypoxic conditions). Collectively, our data suggest a complex role of PKM in the heart and hints that its enzymatic activity might also be regulated on the post-translational level in cardiac cells.