PRODH promotes osimertinib drug-tolerant persistence in EGFR-mutant NSCLC by activating mitochondrial transcription.

The emergence of drug-tolerant persister (DTP) cells poses a major obstacle to durable responses to EGFR tyrosine kinase inhibitors (TKIs) in EGFR-mutant non-small cell lung cancer (NSCLC). However, how metabolic programming orchestrates this transient and reversible adaptive resistance remains largely unexplored. Here we report that proline dehydrogenase 1 (PRODH) promotes the persistence of osimertinib tolerance in NSCLC by increasing mitochondrial respiration in an enzyme activity-independent manner. Mechanistically, PRODH facilitates the recruitment of sirtuin-3 (SIRT3) to mitochondrial transcription factor B2 (TFB2M), resulting in the deacetylation of TFB2M at lysine 127 and lysine 153. This post-translational modification is indispensable for the mtDNA-binding capacity of TFB2M and subsequent initiation of mitochondrial transcription. Loss of PRODH, or increased TFB2M acetylation, impairs DTP formation and delays tumor relapse. Collectively, these data reveal a previously unrecognized role for PRODH in linking mitochondrial metabolism to drug-tolerant persistence, suggesting that targeting PRODH or TFB2M deacetylation may offer a promising strategy to overcome osimertinib resistance and suppress tumor relapse in EGFR-mutant NSCLC.