Microtentacle Formation in Ovarian Carcinoma

Simple Summary Ovarian cancer commonly spreads throughout the peritoneal cavity by exfoliation of malignant cells into ascites. Chemoresistance remains an important therapeutic obstacle. Microtentacles (McTNs) are microtubule-based protrusions that may influence the metastatic potential and chemoresistance profile of free-floating cells. In this study, we define the various microtentacle morphologies that can be observed in detached ovarian cancer cells, and their clustering behavior in relationship to histology, alpha-tubulin post-translational modifications, beta-tubulin isotype, modulators of cortical stiffness, and sensitivity to clinically relevant microtubule-targeting agents. McTNs represent a new therapeutic target for this disease, and an understanding of their biology could have implications for the refinement of therapies, including intraperitoneal drug delivery. Background: The development of chemoresistance to paclitaxel and carboplatin represents a major therapeutic challenge in ovarian cancer, a disease frequently characterized by malignant ascites and extrapelvic metastasis. Microtentacles (McTNs) are tubulin-based projections observed in detached breast cancer cells. In this study, we investigated whether ovarian cancers exhibit McTNs and characterized McTN biology. Methods: We used an established lipid-tethering mechanism to suspend and image individual cancer cells. We queried a panel of immortalized serous (OSC) and clear cell (OCCC) cell lines as well as freshly procured ascites and human ovarian surface epithelium (HOSE). We assessed by Western blot beta-tubulin isotype, alpha-tubulin post-translational modifications and actin regulatory proteins in attached/detached states. We studied clustering in suspended conditions. Effects of treatment with microtubule depolymerizing and stabilizing drugs were described. Results: Among cell lines, up to 30% of cells expressed McTNs. Four McTN morphologies (absent, symmetric-short, symmetric-long, tufted) were observed in immortalized cultures as well as ascites. McTN number/length varied with histology according to metastatic potential. Most OCCC overexpressed class III ss-tubulin. OCCC/OSC cell lines exhibited a trend towards more microtubule-stabilizing post-translational modifications of alpha-tubulin relative to HOSE. Microtubule depolymerizing drugs decreased the number/length of McTNs, confirming that McTNs are composed of tubulin. Cells that failed to form McTNs demonstrated differential expression of alpha-tubulin- and actin-regulating proteins relative to cells that form McTNs. Cluster formation is more susceptible to microtubule targeting agents in cells that form McTNs, suggesting a role for McTNs in aggregation. Conclusions: McTNs likely participate in key aspects of ovarian cancer metastasis. McTNs represent a new therapeutic target for this disease that could refine therapies, including intraperitoneal drug delivery.