The RNA binding protein LIN28 and its paralog LIN28B function at a critical junction of pleuripotency, metabolism and metastasis. Reactivation of LIN28 has been recently reported in epithelial ovarian cancer (EOC), where it is hypothesized to play a key role in maintaining tumor stem cells. Associations between single nucleotide polymorphisms in the LIN28B promoter and ovarian cancer susceptibility have also been identified. However, the clinical significance of these observations and the mechanisms by which either LIN28 gene contributes to ovarian cancer have not been explored.
Using Western blot, qPCR and immunohistochemistry, we found that a) LIN28 but not LIN28B is highly expressed in epithelia lining the distal fallopian tube and b) reactivation of LIN28 (1/8 specimens) and dysregulated expression of LIN28B (4/8 specimens) occur in EOC. To assess clinical significance of these events, we interrogated the TCGA ovarian cancer database, examining correlations between LIN28, LIN28B expression and outcomes by Kaplan-Meier analysis (n=581). Our results indicate that LIN28 expression is associated with shorter disease free interval in women with optimally debulked high grade serous ovarian cancers (p<0.05). Significantly higher levels of LIN28B were also noted in young (age <50) women diagnosed with ovarian cancer. Using serial logistic regression with L1 normalization (Lasso analysis), we discovered LIN28 levels correlated most robustly with multiple genes rather than miRNAs: APOC3 (β =0.74), FGG (β =0.69), HBG1(β=0.67) and HEMGN (β =0.67). Although associations between LIN28 and miRNAs predicted by current models were not observed, robust correlations between LIN28B and multiple miRNAs were detected: let-7b (β =-0.47), let-7d (β =-0.34), let-7i (β =-0.31) as well as others [miR-222 (β=-0.29), let-7e, miR-222, miR-324-5p] not previously known to be regulated by either LIN28 gene. Lastly, we found that expression of LIN28B but not LIN28 was induced by culturing A2780 and TOV112D ovarian cancer cells in media that promotes self-assembly into spheroids. Knockdown of LIN28B expression confirms that LIN28B directly regulates the let-7 and other miRNAs identified by our analyses, promotes spheroid assembly and in vivo tumorigenicity of ovarian cancer cell lines.
Collectively, these observations indicate that LIN28 likely plays an important role in regenerating epithelia that normally line the distal fallopian tube. Although our data also suggest that reactivation of LIN28 plays a key role in promoting ovarian cancer recurrences, LIN28 does not contribute to this process by targeting the differentiation of pluripotent cells via a let-7-mediated mechanism. Rather, this role appears to be most robust for its paralog LIN28B. Future work will focus on further dissecting the unique roles of these two gene products in ovarian cancer initiation and metastasis as well as their response to treatment.
Citation Format: Claire Mach, Ying-Wooi Wan, Zhandong Liu, Matthew L. Anderson. LIN28 paralogs impact ovarian cancer predisposition and tumorigenicity via distinct molecular pathways. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1955. doi:10.1158/1538-7445.AM2013-1955