Autocrine lymphotoxin signalling promotes ovarian tumor growth through CD74

Background: Ovarian cancer is the eighth most common cause of cancer and cancer death in women worldwide. The 5-year overall survival rate for advanced-stage ovarian cancer is 10-40%. A new therapy for advanced ovarian cancer is urgently needed. Identifying signalling pathways which promote tumor growth is crucial to developing targeted therapy. Lymphotoxin is overexpressed in ovarian cancer cells and mediates tumor-stromal interaction in ovarian tumor. However, the transcription regulation of lymphotoxin and the role of autocrine lymphotoxin signalling in ovarian cancer cells are still unclear. Methods: To investigate transcriptional regulation of lymphotoxin, correlation analysis and pharmacologic inhibitions was performed in human ovarian cancer cell lines. To investigate autocrine lymphotoxin signalling, lymphotoxin-beta (LTB) was edited in human ovarian cancer cells by CRISPR-cas9 knockout. On the contrary, lymphotoxin genes (Lta and Ltb) were overexpressed in mouse ovarian cancer cells by stable transfection. Tumor growth was examined by cancer spheroid in vitro model and preclinical metastatic tumor in vivo model. Involvement of lymphotoxin-beta receptor (LTBR) and noncanonical NF-κB pathway (RelB and p52) were investigated by pharmacological inhibitions. To identify NF-κB target genes, differentially expressed genes were analysed by NanoString nCounter pan-cancer pathways panel. Relationships between lymphotoxin, NF-κB and CD74 in ovarian tumor samples (TCGA and CUHK) were performed by correlation analysis. Function of CD74 was investigated by CRISPR-cas9 knockout in human ovarian cancer cells. Results: Positive correlations between IL33 and LTB expression, and between ST2 and LTA expression, are found in human ovarian cancer cell lines. Inhibition of IL33 and ST2 downregulates LTA transcription in human ovarian cancer cells. LTB knockout in human ovarian cancer cells induces cell cycle G2/M arrest and reduces tumor growth in vitro, whereas lymphotoxin overexpression in mouse ovarian cancer cells increases tumor growth in vitro and in vivo. Inhibition of LTBR and p52 reduces lymphotoxin-mediated ovarian tumor growth in vitro and in vivo. Cd74 is upregulated in mouse ovarian cancer cells after lymphotoxin overexpression and LTBR activation. In human ovarian cancer cells, CD74 transcription is regulated by lymphotoxin-LTBR-noncanonical-NF-κB pathway. Positive correlations between lymphotoxin, NFKB2, RELB and CD74 mRNA expressions are found in ovarian tumor samples. CD74 knockout in human ovarian cancer cells induces G2/M cell cycle arrest and reduces tumor growth in vitro. Conclusion: Our results demonstrated that IL33-ST2 pathway positively regulates lymphotoxin expression in ovarian cancer, and autocrine lymphotoxin signalling promotes ovarian tumor growth through CD74. These results are a foundation for developing targeted therapy for advanced ovarian cancer.