Thy Leung

GRO-α and IL-8 enhance ovarian cancer metastatic potential via the CXCR2-mediated TAK1/NFκB signaling cascade

Intraperitoneal metastasis is a common occurrence and is usually involved in the poor prognosis of ovarian cancer. Its specific metastatic pattern implies that certain indispensable microenvironmental factors secreted in the peritoneal cavity can direct metastatic ovarian cancer cells to permissive niches for secondary lesion formation. However, the underlying molecular mechanisms are ill defined. Herein, we report that GRO-α and IL-8 are predominately upregulated in culture media derived from either normal or cancerous omenta and are associated with increased ovarian cancer aggressiveness. Methods: OCM was established from culture medium of fresh human omental tissues. Primary and metastatic ovarian cancer cell lines were generated from human tumor tissues and verified by specific antibodies. The functional roles of GRO-α, IL-8, and their specific receptor CXCR2 were examined by neutralizing antibodies, shRNA gene knockdown, CRISPR/Cas9 gene knockout and pharmaceutical CXCR2 inhibitor SB225002. The oncogenic properties of ovarian cancer cells were examined by in vitro and in vivo mouse models. Results: Both GRO-α and IL-8 can activate TAK1/NFκB signaling via the CXCR2 receptor. Intriguingly, TAK1/NFκB signaling activity was higher in metastatic ovarian cancer cells; this higher activity makes them more susceptible to OCM-induced tumor aggressiveness. Treatment of ovarian cancer cells with GRO-α and IL-8 neutralizing antibodies or ablation of CXCR2 by shRNA gene knockdown, CRISPR/Cas9 gene knockout, or CXCR2 inhibitor SB225002 treatment significantly attenuated TAK1/NFκB signaling and decreased in vitro and in vivo oncogenic and metastatic potential, suggesting CXCR2 plays a key role in the GRO-α and IL-8-governed metastatic spreading of ovarian cancer cells in the intraperitoneal cavity. Conclusion: This study highlights the significance of GRO-α and IL-8 as the key chemokines in the peritoneal tumor microenvironment and suggests the utility of targeting their receptor CXCR2 as a potential target-based therapy for peritoneal metastases of ovarian cancer.

PO-296 Elucidating the potential role of cd109 as a biomarker for cancer stem-like cells in cervical cancer

Introduction Cervical cancer is a common genital tract cancer. Radiotherapy is the mainstay of management for advanced cervical cancer. Response to radiation varies widely which may be explained by the existence of cancer stem-like cells (CSCs). Since CSCs is implicated in cervical cancer and demonstrated a high degree of resistance to radiation, the identification of novel CSC markers could be critical to specifically target the cervical CSC. In our pilot study, we established attached and spheroid cells from primary cervical tumour tissue. Multiple ‘stemness’ genes were detected in spheroid cells which indicated primary cervical cancer tissue harboured CSCs population. cDNA microarray analysis was performed to compare cDNA expression profile and CD109 was significantly up-regulated in spheroid cells. Therefore, we hypothesise that CD109 may serve as a potential cervical CSCs marker. Material and methods Flow cytometry was performed to analyse the CD109 expression and isolate the CD109 positive and negative sub-populations in cervical cancer cells. Following the sorting, cell proliferation and migration assay were performed. In order to investigate the effect on the blockade of CD109, SiHa, Caski and C4-1 cells were transfected with CD109 siRNAs. XTT assay, migration and invasion assay and spheroid formation assay were performed. Immunohistochemistry (IHC) was performed for the detection of CD109 expression in cervical cancer tissue microarray (TMA). Results and discussions The post-sorted CD109 (+) cells grew remarkably faster and have stronger migration capability than CD109(-) cells in Caski and C4-1. The CD109 knockdown cells with siRNA exhibited a slower cell growth, decreasing migration and self-renewal ability, as compared with the control group in SiHa, Caski and C4-1 cells. IHC of TMA indicated that CD109 was highly expressed in cancer cases than that in normal/benign cases. Conclusion CD109 increased cell proliferation rate and migration ability in post-sorted cervical cancer cell lines Caski and C4-1. On the contrary, CD109 knockdown reduced cell growth, migration and self-renewal capability in cervical cancer SiHa, Caski and C4-1 cells. Cervical carcinoma showed high expression of CD109 protein by IHC. Further in vivo and in vitro functional assays are essential to characterise the CD109-positive sub-population in cervical cancer which may provide more information of cervical CSCs-related properties and resistance of radiation therapy, with the underlying molecular mechanism involved.

HPV-E6 protein enriches the CD55(+) population in cervical cancer cells promoting radio-resistance and cancer aggressiveness

Accumulating evidence indicates that the human papillomavirus (HPV) E6 protein plays a crucial role in the development of cervical cancer. Subpopulations of cells that reside within tumours are responsible for tumour resistance to cancer therapy and recurrence. However, the identity of such cells residing in cervical cancer and their relationship with the HPV‐E6 protein have not been identified. Here, we isolated sphere‐forming cells, which showed self‐renewal ability, from primary cervical tumours. Gene expression profiling revealed that cluster of differentiation (CD) 55 was upregulated in primary cervical cancer sphere cells. Flow‐cytometric analysis detected abundant CD55(+) populations among a panel of HPV‐positive cervical cancer cell lines, whereas few CD55(+) cells were found in HPV‐negative cervical cancer and normal cervical epithelial cell lines. The CD55(+) subpopulation isolated from the C33A cell line showed significant sphere‐forming ability and enhanced tumourigenicity, cell migration, and radioresistance. In contrast, the suppression of CD55 in HPV‐positive CaSki cells inhibited tumourigenicity both in vitro and in vivo, and sensitized cells to radiation treatment. In addition, ectopic expression of the HPV‐E6 protein in HPV‐negative cervical cancer cells dramatically enriched the CD55(+) subpopulation. CRISPR/Cas9 knockout of CD55 in an HPV‐E6‐overexpressing stable clone abolished the tumourigenic effects of the HPV‐E6 protein. Taken together, our data suggest that HPV‐E6 protein expression enriches the CD55(+) population, which contributes to tumourigenicity and radioresistance in cervical cancer cells. Targeting CD55 via CRISPR/Cas9 may represent a novel avenue for developing new strategies and effective therapies for the treatment of cervical cancer. Copyright