Reprogramming the Pancreatic Adenocarcinoma Microenvironment integrating mutSEMA3A mRNA-Lipid Nanoparticles with a Vascularized Organ-on-Chip Model

Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest solid tumors due to late diagnosis, limited therapeutic options, and an immunosuppressive tumor microenvironment (TME). The PDAC TME is characterized by aberrant vasculature that hampers immune cell infiltration and drug delivery [1]. Semaphorin3A (Sema3A) has shown promising potential in normalizing tumor vasculature and activating cytotoxic CD8+ T cells, thereby enhancing anti-tumor immunity. The project aims to develop lipid nanoparticles (LNPs) encapsulating the mRNA of a mutated, constitutively active version of Sema3A (mutSEMA3A) to evaluate its impact on PDAC progression [2]. To reduce reliance on in vivo models and develop physiologically relevant platforms that mimic TME complexity [3], we established a vascularized PDAC-on-a-chip model using the Organoplate Graft system. This in vitro platform recapitulates key features of the PDAC TME by integrating endothelial cells (ECs), tumor spheroids, and eventually immune components within a 3D collagen matrix. It provides a controlled environment to investigate the inhibitory effects exerted by PDAC on the endothelium, as well as LNP transcytosis and the endothelium-normalizing effects of mutSEMA3A mRNA. mRNA transfection efficiency and mutSEMA3A protein secretion have been validated in PDAC and endothelial cell lines. The secreted mutSEMA3A reduces EC migration in haptotactic EC assays. We are currently investigating its effect on the vasculature. Preliminary tests with mutSEMA3A mRNA-LNPs, utilizing an in-house developed formulation [4], proved their ability to transfect both cell lines. This chip platform will be used to screen and refine LNP formulations, ultimately guiding the design of future in vivo studies in syngeneic orthotopic mouse models. By integrating advanced nanotherapy with a pathologically relevant in vitro model, this project aims to overcome PDAC resistance mechanisms and improve therapeutic outcomes. Keywords: PDAC-on-a-chip, LNPs, mRNA delivery, tumour microenvironment, semaphorin3A [1] Manji GA et al, Clin Cancer Res, 23(7):1670-8 (2017) [2] Gioelli N et al, Sci Transl Med, 10:eaah4807 (2018) [3] Bonanini F et al, Angiogenesis, 25(4):455-70 (2022) [4] Garbujo S et al, J Colloid Interface Sci, 680:104276 (2026)