Abstract 2735: Role of SEMA3A and SEMA3C in tumor progression and metastatic processes in PDAC

Abstract

Abstract Pancreatic ductal adenocarcinoma (PDAC) has a 5-years survival rate less than 8% and is predicted to become the second leading cause of cancer-related death by 2030. Recent genome-wide analyses have highlighted the involvement of genes belonging to axon guidance (AG) pathway in the progression of PDAC. Among AG genes, semaphorins were originally identified as axon guidance factors for the developing nervous system but a role for them is described also outside of the nervous system; several semaphorins have been found amplified in a subset of PDAC (around 18%) and elevated expression of SEMA3A, in particular, has been previously correlated to poor prognosis. The aim of this study is to assess the role of the axon guidance genes SEMA3A and SEMA3C in PDAC progression and metastasization using organoid models of the disease, in order to identify either biomarkers predictive of disease recurrence or metastases prevention strategies. Data mining has been performed using the ICGC (PACA-AU) and the TCGA (PAAD) databases; RNA in situ hybridization (ISH) has been performed on FFPE of non-neoplastic and tumor tissue sections using the RNAscope 2.5 HD Duplex Detection Kit. Perturbation analysis has been performed by overexpressing and silencing Semaphorins (Sema3a and Sema3c) by transduction of tumor-derived organoid cultures with ORF and inducible CRISPR/Cas9 vectors, respectively. For orthotopic transplantation, a small incision in the skin of the upper left quadrant of the abdomen of C57BL/6J and NSG mice has been made; 5 × 105 cells/mouse have been injected into the parenchyma of the pancreas. Tumor growth has been monitored once a week with the VEVO 2100 system.Expression of SEMA3A and SEMA3C was enriched in the basal like/squamous subtype and was correlated to poor survival in both the ICGC and TCGA cohorts. However, in silico microdissection of bulk sequencing data (ICGC and TGCA) revealed that Semaphorins were particularly expressed in the stromal compartment. Accordingly, mRNA in situ hybridization analyses of human PDAC tissues showed that SEMA3A is almost exclusively expressed in cancer-associated fibroblasts (CAFs), while SEMA3C is expressed both in the neoplastic and in CAFs. We further analyzed expression of semaphorins in human tumor-derived organoids and CAFs to confirm that those genes were abundant in CAFs and less expressed in squamous/basal-like organoids. Consistently with in silico and in vitro data, transplantation of mouse tumor organoids overexpressing semaphorins showed reduced tumor growth and lack of metastatic competence as compared to mock transfected cultures, while organoids lacking either Sema3a or Sema3c showed accelerated tumor development and metastasization.These data suggest that SEMA3A and SEMA3C are preferentially expressed by CAFs in PDAC and that their high stromal expression contributes to aggressive molecular subtype of PDAC. Citation Format: Francesca Lupo, Pietro Delfino, Dea Filippini, Caterina Vicentini, Vincenzo Corbo. Role of SEMA3A and SEMA3C in tumor progression and metastatic processes in PDAC [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2735.