LAUSR

Pancreatic ductal adenocarcinoma (PDAC) is a relatively uncommon malignancy; however, its incidence is rising worldwide, and it is expected to become the second-leading cause of cancer-related death by 2030. From a histological point of view, PDAC is characterized by a prominent desmoplastic reaction that compresses blood vessels, limiting oxygen and nutrient availability in the tumor microenvironment. Despite that, PDAC cells are capable to adapt dynamically to these stress conditions, adopting different strategies that includes the strict regulation of the autophagic flux. Thus, studying these adaptive mechanisms is crucial to understand PDAC progression and to establish new therapeutic modalities to tackle it. Here, we explore the role of the tumor suppressor miR-15a in the regulation of its putative target Fra-2, a transcription factor, commonly activated by cell stress. By employing IPA on PDAC tumors from TCGA dataset, we found that both miR-15a and its target Fra-2 are predicted to regulate the IGF-1 signaling pathway. In an independent cohort of 44 PDAC samples, miR-15a levels inversely correlated with both Fra-2 and IGF1R expression; conversely, Fra-2 significantly correlated with IGF1R. By chromatin immunoprecipitation and luciferase assay, we assessed that miR-15a directly targeted Fra-2 and IGF1R that, in turn, is transcriptionally regulated by Fra-2 activity. Then, we investigated the role of miR-15a/Fra-2 regulation of IGF1R in the response to starvation-induced cell stress. In starved PDAC cell lines, Fra-2 transcriptional activity triggered IGF1R promoter, causing IGF1R overexpression in control cells but not in miR-15a-overexpressing cells. Consistently, the IGF1 release after starvation induced phosphorylation of IGF1R and activation of the downstream mTOR pathway in control cells but not in miR-15a-overexpressing cells. Therefore, TEM and western blot analysis demonstrated that activation of mTOR via IGF1 release reduced the autophagic flux of PDAC control cell lines compared to miR-15a overexpressing cells under starvation. To assess our results in vivo, we injected PDAC cells wild type or Fra-2 knockout into the flank of nude mice. At tumor onset, mice were randomly divided in two groups and fed with control or hypoproteic diet for three weeks. Hypoproteic diet did not interfere with the growth of wild-type tumors, by contrast, significantly impaired Fra-2KO tumors growth rate. Tumor analysis revealed that hypoproteic diet potently induced IGF1R overexpression and mTOR pathway activation in wild-type tumors but not in Fra-2KO tumors. Our findings demonstrate that IGF1R expression is regulated by miR-15a directly and indirectly via Fra-2 in PDAC. This novel miR-15a/Fra-2/IGF1R axis, triggered by starvation, regulates the autophagic flux and growth of PDAC cells in stress condition, and could be targeted by specific small inhibitors. Citation Format: Gian Luca Rampioni Vinciguerra, Marina Capece, Luca Reggiani Bonetti, Paolo Magistri, Federica Calore, Giovanni Nigita, Rosario Distefano, Roberto Ballarin, Fabrizio Di Benedetto, Andrea Vecchione, Barbara Belletti, Gustavo Baldassarre, Francesca Lovat, Carlo M. Croce. The novel miR-15a/Fra-2/IGF1R axis drives response to starvation-induced cell stress in pancreatic ductal adenocarcinoma. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4823.