Abstract P6-06-11: The inhibition of leptin receptor impairs macrophage recruitment in the tumor microenvironment blocking breast cancer growth and progression
Authors: Gelsomino Luca , Panza Salvatore , Augimeri Giuseppina , Naimo Giuseppina Daniela , Malivindi Rocco , La Camera Giusi , Paolì Alessandro , Giordano Cinzia , Barone Ines , Mauro Loredana , Bonofiglio Daniela , Catalano Stefania , Andò Sebastiano 
Published: March 2020

Background: Breast tumorigenesis results from the sequential acquisition of multiple genetic alterations in the epithelial cells and is also influenced by reciprocal interactions between the tumor cells and the surrounding stromal microenvironment. In this regard, an important mediator in driving mammary tumorigenesis is leptin, mainly secreted by adipocytes, but also by other cells in the tumor microenvironment. Several studies support the crucial involvement of leptin/leptin receptor (ObR) in the biology of breast cancer; however it still remains to be elucidated how ObR signaling may modulate tumor cell cytokine secretion favoring a permissive microenvironment required for tumor progression. Thus, we evaluated the impact of ObR lack in affecting breast cancer phenotype and in mediating epithelial/stromal tumor reciprocal interaction by using both in vitro and in vivo experimental approaches.

Methods: As experimental models we established stable ERα-positive MCF-7 with an empty lentiviral vector (MCF-7 EV) and a lentiviral vector to silence human ObR (MCF-7 shObR), triple-negative MDA-MB-231 and human monocyte cell line derived from an acute monocyte leukemia patient, THP-1. We ran cytokine array to study cytokine secretion pattern in both MCF-7 EV and MCF-7 shObR cells. qRT-PCR, immunofluorescence and ELISA were performed to detect MCP-1 expression and secretion. Co-culture experiments and Boyden Chamber Transmigration assays were assessed to study the recruitment of THP1. Tumor associated macrophage (TAM) markers were detected after co-colture experiments between MCF-7 stable clones, MDA-MB-231 cells and THP1. Orthotopic mammary fat pad implantation of MCF-7 stable clones was performed to test tumor growth and macrophage infiltration.

Results: Firstly we demonstrated a reduced activation of leptin signaling, and a decreased growth and motility in MCF-7 shObR cells compared to MCF-7 EV. Cytokine array revealed that the lack of ObR abrogated the release of monocyte chemoattractant protein (MCP1), and dramatically down-regulated interleukin 8 (IL-8) and intercellular adhesion molecule 1 (ICAM-1) secretion. Moreover, in MCF-7 shObR cells we found a diminution of MCP-1 in terms of mRNA and protein expression. As expected the suppression of MCP-1 secretion in these cells blocked the recruitment of monocytes and inhibited TAM development. The specific role of ObR/Ob signaling in regulating MCP-1 expression and secretion was further demonstrated in the presence of the ObR selective antagonist, the LDFI. LDFI treatment markedly down-regulated MCP-1 expression thus blocking macrophage recruitment and polarization in MDA-MB-231 cells. In vivo experiments revealed a decrease of tumor growth in MCF-7 shObR cells compared to MCF-7 EV wherein a dramatic reduction of MCP-1 expression was well reproduced in MCF-7 shObR clones.

Conclusions: Our results, for the first time, showed that Ob/ObR signaling pathway may directly control MCP-1 function and tumor cell-polarized macrophages. Thus these data suggest the use of leptin receptor specific antagonist able to block macrophage-mediated effects of tumor microenvironment in sustaining breast cancer growth and progression.

Citation Format: Luca Gelsomino, Salvatore Panza, Giuseppina Augimeri, Giuseppina Daniela Naimo, Rocco Malivindi, Giusi La Camera, Alessandro Paolì, Cinzia Giordano, Ines Barone, Loredana Mauro, Daniela Bonofiglio, Stefania Catalano, Sebastiano Andò. The inhibition of leptin receptor impairs macrophage recruitment in the tumor microenvironment blocking breast cancer growth and progression [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P6-06-11.