Proline Dehydrogenase as new marker and target in EGFR mutant Non-Small Cell Lung carcinoma

Non-Small Cell Lung Cancer (NSCLC) is one of the most frequent types of tumor for incidence and the leading cause of cancer mortality. Identification of markers to better define diagnosis, prognosis and therapeutic options of NSCLC is needed. Proline dehydrogenase (PRODH), a mitochondrial flavoenzyme catalyzing the key step in proline degradation, is expressed in lung adenocarcinomas and expression appears to correlate with EGFR mutations. PRODH expression is also regulated by AMPK. Observing the synergism between metformin and EGFR-inhibitors, we propose that PRODH may be a mediator of metformin growth-inhibitory and proapoptotic effects, alone or in combination with tyrosine kinase inhibitors. The relation between EGFR mutations and PRODH expression was investigated by modulating EGFR or specific downstream signaling pathways, in particular JAK-STAT3, as we identified putative STAT3 binding sites in the PRODH promoter. We developed two cellular models with in vitro acquired resistance to Osimertinib (EGFR inhibitor): we selected PC9 and H1975 cell lines, harbouring an in-frame deletion in exon 19 and T790M and L858R mutations of the EGFR gene, respectively. Preliminary results show that PRODH expression is modulated upon STAT3 activation or transfection of constitutively active STAT3 construct. We will investigate if modulation occurs by direct promoter regulation. We hypothesize that STAT3 may be implicated in PRODH activation and that PRODH may be a new marker and therapeutic target. In parallel, we investigated expression of PRODH in Osimertinib resistant cell lines (PC9-OR and H1975-OR) compared to corresponding parental cells: we detected increased level of PRODH in resistant cells, suggesting that it may represent an escape mechanism to EGFR inhibition. Based on the molecular connection of PRODH with targets modulated by metformin, we tested the effect of metformin on PRODH levels and we show that metformin reduces PRODH protein levels in vitro in NSCLC cells. Also, considering the synergism of metformin with EGFR-inhibitors and the inter-dependence between EGFR and PRODH, we speculate that PRODH activation may be an EGFR resistance mechanism reversible by metformin also in OR-cells, thus suggesting a new combinational strategy. We propose the involvement of PRODH as escape mechanism to EGFR and its role in the mechanism of antineoplastic activity of metformin.