Tumor-stroma interactions influence the response to PI3K targeted agents in preclinical models of colorectal cancer (CRC)

Introduction: One of the main obstacle to the successful development of therapeutic strategies remains the identification of biomarker underlying drug resistance. Recently, investigators have become more aware the role of the tumor microenvironment (TME) in cancer and the potential therapeutic opportunities that derive from suppressing potential resistance mechanisms arising microenvironmental interactions. The aim of this study was to set-up multicellular culture models to uncover the molecular mechanisms by which stromal/endothelial cells modulate response to signaling inhibitors and to identify potential therapeutic targets in PTEN-loss contexts. Methods and Materials: Isogenic CRC cell lines (X-MAN™ HCT116 and HCT116 PTEN-/-) were treated with MAPKi and PI3K/mTORi alone or in combination, in the presence or absence of stromal fibroblasts or fibroblast/endothelial cell conditioned medium (CM). Cytofluorimetric analysis and Crystal Violet assay were used to analyse functional response to targeted agents; pathways activation and cytokine/chemokine profile were analysed using Western blot and ELISA assay respectively. Results and Discussion: In co-culture CRC models, the response to MAPK and PI3K inhibitors is the result of interaction between tumor cells and their surrounding stroma. The response to PI3K/mTORi is mainly influenced by microenvironmental interactions: direct cell-to-cell tumor/stroma contact renders stromal cells resistant to PI3K/mTORi, while the presence of stromal cell-derived soluble factors sensitizes PTEN-competent CRC cells to PI3K/mTORi-mediated growth inhibition. This effect was confirmed using CM from different types of stromal cells (fibroblast/endothelial) that similarly affected the response of CRC cell lines to signalling inhibitors; this is probably due to similar profile of cytokine/chemokine production in stromal cell and is subjected to a “saturation” effect. The presence of stromal CM upregulates MAPK activation regardless of PTEN status, whereas mTOR pathway upregulation is observed mainly in PTEN-competent CRC cellsin PTEN-competent cells soluble factors released by stromal elements paradoxically impair PTEN function, leading to downstream mTORC1 complex formation and pathway activation. This paradoxical mTORC1 activation upon exposure to stroma-derived soluble factors results in functional hypersensitivity of PTEN-competent CRC cells to the growth inhibitory effects of double PI3K/mTOR inhibitors. . Conclusions: The presence of stromal cells (fibroblasts/endothelium) profoundly influences CRC response to PI3K/mTOR-targeting agents. Understanding the mechanisms underlying microenvironmental interactions (tumor, stroma, soluble factors) may be of fundamental importance to overcome therapeutic resistance and develop more effective therapies for patients affected by cancer.