Abstract 2355: Marrow-derived macrophages mediate invasion of pancreatic adenocarcinoma by RET activation

Abstract

Abstract Introduction: Perineural invasion of cancer cells (CPNI) is found in most patients with pancreatic adenocarcinomas (PDAs). Although the presence of M2 macrophages in the perineural environment has been associated with shortened survival rates in PDA patients, the mechanism underlying this phenomenon has not been elucidated. It has been suggested that secretion of the RET ligand glial-derived neurotrophic factor (GDNF) by M2 macrophages causes cancerous RET-expressing cells to migrate along affected nerves. This concept, however, has not been demonstrated in vivo. This study was aimed at understanding the contribution of macrophages to perineural invasion for in vivo models of PDA, and to characterize the role of the GDNF-RET axis in this unique microenvironmental niche. Methods: Immunohistochemical analysis of PDA specimens of a novel Pdx-1-Cre/KrasG12D /p53R172H (KPC) mouse model, revealed that CD163+ (M2) endoneurial macrophages (EMΦs) predominate the endoneural space of invaded nerves. In a bone marrow transplant (BMT) from CX3CRGFP/+ mice, a 23- fold increase in the recruitment of GFP+ macrophages to the perineural space was witnessed as compared to GFP- cells, indicating recruitment of bone marrow- derived macrophages to the perineural niche, rather than resident tissue macrophages. A BMT from CCR2-/- mice lacking macrophage recruitment capacity resulted in a 6-fold decrease of the invasion severity score, and a 2.5-fold decrease in perineural invasion index. Analysis by fluorescence microscopy indicated that EMΦs expressed higher levels of GDNF compared to EMΦs in premalignant lesion and normal controls. In order to differentiate between the contributions of GDNF secreted by nerves to the invasion presses and its secretion from recruited M2 macrophages, we performed a BMT from GDNF+/- mice. The BMT led to a 1.5-fold improvement in both the invasion severity score and the perineural invasion index. Similarly, mice treated with PP1, a RET receptor inhibitor, caused a 1.5-fold decrease in the invasion severity score, and a 1.3-fold decrease in the perineural invasion index. Finally, xenografts of PDAC cells, in which RET was silenced using siRNA, resulted in a 3.5-fold decrease in the invasion severity score and a 3.2-fold decrease in the perineural invasion index. Discussion: Taken together, our results suggest a paracrine response between bone marrow-derived tumor-associated EMΦs and PDA cells. This loop probably orchestrates the formation of cancer nerve invasion. Further studies using novel transgenic animal models aimed at specifying the specific molecular signals that induce disease invasion and proliferation in the perineural microenvironment are needed. Citation Format: Moran Amit, Shorook Na'ara, Yoav Binenbaum, Ziv Gil. Marrow-derived macrophages mediate invasion of pancreatic adenocarcinoma by RET activation. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2355. doi:10.1158/1538-7445.AM2015-2355