Evaluating interaction between tumor educated platelets and cancer stem cells on breast cancer subtypes.

5 Background: Thrombocytosis has been reported to be responsible for poor prognosis of numerous malignancies. This swarm of platelets acts as foe for the body but as mate for the cancer cells by providing a protective shield around them, helping them to escape immunosurveillance process. A bidirectional interaction between platelets and tumor cells promote their conversion to an activated tumor educated platelet (TEP) state. The present report reveals the critical role played by TEPs to promote metastasis in BC subtypes by interacting with cancer stem cells (CSCs). Methods: Status of intra-tumoral and humoral TEPs (CD41 + /CD62P + ) was screened in luminal A (LumA) and triple negative (TNBC) BC subgroups. The impact of TEPs on lin - /CD44 + /CD24 - CSCs of both the categories in-vitro, was deciphered by performing mammosphere assay, clonogeneic and migration assays, with final validation in murine system. Effect on various genes and proteins related to stemness, metastasis and angiogenesis as a result of this interaction was elucidated by RT-PCR, flow cytometry and western blotting. Results: High percentage of TEPs in the peripheral blood is a potential biomarker and responsible for poor prognosis in BC subtypes. Both Lum A and TNBC patients had elevated TEP frequency, compared to healthy blood. Further, patients in the pre-carcinogenic stage had platelet count of >3.5 lakhs. Additionally, screening of breast tumor sections revealed intra-tumoral enrichment of TEPs, comparatively to a greater extent in TNBC than LumA, supporting their role in disease aggression. Also, a positive correlation was noted between TEP and CSC frequencies proving their synergistic interactions. Moreover, in-vitro, TEP influenced CSCs exhibited enormous clonogenic and tumorigenic potentialities. Their metastatic nature was confirmed by their enhanced migratory, invasive, angiogenic capacities with higher VIMENTIN and TWIST along with HIF1α, CD31 and MMP9 expression compared to E-CADHERIN. In line with in-vitro observations, in-vivo too, TEP~CSC produced metastatic colonies in murine lungs. Upon investigation, it was disclosed that it is through WNT-βcatenin-VEGFR2 axis that this mutual interdependency was functioning. Conclusions: This study advocates for the importance of TEPs as potential biomarker in BC diagnosis. Their alliance with vicious CSCs to promote disease advancement acquiesce them as a novel restorative agent.