EACR25-0968

M2 macrophages promote survival and proliferation of patient-derived gastric cancer organoids in a tumor biology-dependent manner

M. Cabeza-Segura1, A. Aguilar-Cifre1, A. González-Vilanova2, A. Marín-Alonso1, B. Palomar3, R. Villagrasa4, C. Martínez-Ciarpaglini3,5, A. Cervantes1,5, T. Fleitas1,5, J. Castillo6,1,5
1INCLIVA Biomedical Research Institute, Oncology Department, Valencia, Spain
2INCLIVA Biomedical Research Institute, Bioinformatics Unit, Valencia, Spain
3INCLIVA Biomedical Research Institute, Pathology Department, Valencia, Spain
4Hospital Clínico Universitario de Valencia, Gastroenterology and Hepatology Department, Valencia, Spain
5Carlos III Health Institute, CIBERONC, Madrid, Spain
6University of Valencia, Biochemistry and Molecular Biology Department, Valencia, Spain
Introduction:

Gastric cancer (GC) is one of the leading causes of cancer-related mortality worldwide, with high incidence rates and poor prognosis due to its molecular heterogeneity and therapeutic resistance. In this regard, the use of functional models such as patient-derived tumor organoids can shed light on and allowing these issues to be addressed on a patient-specific basis. In addition, tumor microenvironment plays a critical role in GC progression by modulating the immune response and facilitating tumor evasion. In particular, tumor-associated macrophages can adopt pro-inflammatory (M1) or anti-inflammatory (M2) phenotypes, with the latter promoting tumor aggressiveness

Material and method:

A prospective biobank of gastric tumor organoids (GTOs) derived from advanced GC patients was generated according to an in-house protocol from January 2019. The human monocytic THP1 cell line was used and differentiated into macrophages, which were subsequently polarized into M1 macrophages using IFNγ + LPS, and M2 macrophages using IL4. Macrophage polarization was assessed measuring M1 (TNF; CXCL10) and M2 (CD206; MS4A4A) markers by real-time quantitative PCR (RT-qPCR). The conditioned media from M1 (M1CM) and M2 (M2CM) macrophages were used to culture the organoids. To assess the effect of each conditioned medium on organoid viability and proliferation, flow cytometry apoptosis/necrosis and cell cycle assays were performed, respectively. M2 macrophages were repolarized into M1 phenotype by treatment with IFNγ and LPS

Result and discussion:

A biobank of more than 40 GTO lines has been generated covering all gastric cancer subtypes. For this study, 7 GTOs were used: 3 intestinal, 2 diffuse, 1 mixed and 1 undifferentiated, according to Lauren's classification; with 2 of them being HER2-positive and 2 others presenting microsatellite instability. THP1 cells were differentiated into macrophages and properly polarized into M1 and M2 macrophages. A differential effect of M1CM and M2CM on GTOs viability and proliferation were observed. Concerning GTOs viability, M1CM promoted organoid cell death, significantly increasing the number of cells undergoing apoptosis and necrosis. With regard to proliferation, distinct effects were observed. While most GTOs exhibited an increase in the number of cells in the G2+S cell cycle phases in M2CM compared to M1CM, others experienced no difference, suggesting that the influence of M2 macrophages on tumor cell proliferation depends on tumor-specific biology. Treatment of M2 macrophages with IFNγ and LPS repolarized them toward M1 phenotype. Conditioned medium from these repolarized macrophages reversed the observed effects of M2CM on GTOs

Conclusion:

M2 macrophages promote survival and proliferation in most GTOs but showing an interesting heterogeneous effect possibly due to tumor-specific biology. Therapeutic strategies focused on repolarizing M2 macrophages into an M1 phenotype may improve GC management