Abstract 2620 Culture in Physiological Oxygen and Plasma-Like Media Modulate Breast Cancer Cell Hallmarks

Standard cell culture conditions often fail to replicate the in vivo microenvironment, limiting the reproducibility and physiological relevance of experimental findings.For example, commonly used media such as DMEM contain supraphysiological levels of key metabolites like glucose while lacking others entirely.Additionally, standard cell culture incubators maintain near-atmospheric oxygen levels (18% O2), far exceeding the physiological oxygen levels (2-9% O2) typical of most tissues.Despite their significance, these factors are often studied independently.Here, we investigated the effects of both media composition and oxygen levels on breast cancer cell physiology.Using MCF7 cells cultured in 18% O2 or 5% O2 (physioxia) and in either DMEM or Plasmax-a physiological medium modeled after the human plasma metabolome-we performed transcriptomic, proteomic, and functional analyses.RNA sequencing revealed oxygen-and media-dependent changes in gene expression, particularly in pathways related to cell cycle regulation, metabolism, and redox homeostasis.While the overlap between differentially expressed genes and proteins was low, pathways related to proliferation and energy metabolism where also enriched in the proteomic analysis.Consequently, functional assays revealed differences in proliferation, migration, metabolic activity, glucose consumption, and reactive oxygen species production driven by different culture conditions.Generally, physiological culture conditions promoted proliferation, migration, and metabolic activity in MFC7 cells.Collectively, our results demonstrate that both oxygen levels and media composition profoundly influence cancer cell behavior, underscoring the necessity of implementing physiologically relevant culture conditions in in vitro cancer research.