Abstract 6538: Modulation of human macrophage differentiation, phenotype and function in vitro as a strategy to characterize novel tumor microenvironment modulators

Abstract Macrophage infiltration of the tumor microenvironment is a key process that can determine the success or failure of anti-tumor therapies. Thus, many immunooncology therapies are being developed to modulate the macrophage differentiation program towards the anti-tumor M1 subset, while hindering the M2 subset associated with tumor progression. Monocyte-derived macrophages comprise a crucial in vitro cellular tool to evaluate the plasticity potential between the different polarization states. Several hurdles like their in vitro adherence, low cell number yield and the donor-to-donor variability pose technical challenges when using in vitro differentiated macrophages for compound screening and titration or functional assays. At Reaction Biology we have established human macrophage assay pipelines to evaluate the novel macrophage polarization compounds in vitro. By isolating monocytes from frozen PBMC stocks it is possible to have access to the same donor or group of donors PBMC for repetitive analysis. We have developed a differentiation protocol that allows us to use different plate format from 6-well to 48 well plates, and thus increase and modulate the throughput according to the required readout. The effect of therapeutic compounds can be analyzed by flow cytometry. Our data shows that human primary monocytes grown under M1 polarizing conditions express high levels of CD86 and CD80 in contrast to those cultivated under M2 polarizing conditions, with high expression of CD163, CD206, and CD209. Cell culture supernatant can be used to determine cytokine secretion. The functionality of the different macrophage subtypes can be further investigated using a pH-sensitive phagocytosis tracker, like pHRodo-Zymosan or pHRodo-Ecoli, which shows differential uptake according to differentiation profile. Moreover, treated macrophages can be used in coculture with T cells to investigate their capability to foster or inhibit an immune response. Our data shows the value of monocyte-derived macrophage assays in the evaluation and efficacy assessment of TME modulators. Citation Format: Carla N. Castro, Tamara Sahner, Cynthia Obodozie, Philipp Metzger, Holger Weber. Modulation of human macrophage differentiation, phenotype and function in vitro as a strategy to characterize novel tumor microenvironment modulators [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6538.