Phosphodiesterase Type 5 Promotes The Invasive Potential Of Breast Cancer Cells Through Rho Gtpase Activation

The impairment of cyclic guanosine monophosphate (cGMP) signaling by overexpression of PDE5 isoform has been recently described in multiple human carcinomas. In addition, accumulating evidences indicate that PDE5 inhibitors could have direct anti-cancer activities as well as they may enhance the sensitivity of certain types of cancer to standard chemotherapeutic drugs. However, despite these studies, neither the expression of PDE5 in breast cancer subtypes nor the underlying regulatory molecular mechanisms by which PDE5 expression may contribute to breast cancer progression have been deeply studied. We demonstrated that PDE5 was expressed in different subtypes of breast cancer cell lines at higher levels than in non tumorogenic human epithelial breast cell lines. Increased levels were detected in more aggressive endocrine non responsive basal-like breast cancer cells. Interestingly, PDE5 was expressed at very low levels in luminal A-type breast cancer cell lines, which display low ki67 expression, weak invasive behavior and endocrine responsiveness (MCF-7 and T47D cells) compared to luminal B-like cells (such as ZR-75 cells). These results well correlated with data obtained in immunohistochemistry analyses of human breast cancer tissues, showing PDE5 expression in 30 of 35 tumor entities analyzed, with the highest intensity staining in high-grade tumors. Concomitantly, no cytoplasmic PDE5 staining was observed in non neoplastic tissues examined (n=5). In addition, retrospective analyses (n=1959, median follow-up time: 25 years) showed that high PDE5 expression in breast cancer patients was correlated with a statistically significant poorer survival compared to low PDE5-expressing patients. A more relevant discrimination is achieved in lymphnode-negative patients, suggesting a role of PDE5 for identifying early patients at high risk of rapid progression. In order to better ascertain the role of PDE5 in breast tumorogenesis, we selected a breast tumor cell line that express low levels of this enzyme, MCF-7 and engineered stable clones for overexpression studies. Both vector- and PDE5-stable MCF-7 clones demonstrated comparable proliferation rates; whereas, cell motility and invasion were dramatically increased in PDE5-overexpressing cells. RNA sequencing to compare the transcriptomes of vector- and PDE5-overexpressing MCF-7 cells identified differential expression of genes involved in cell migration and invasion. Particularly, based on pathway analysis we found marked changes in the expression of Rho GTPase family members, proteins involved in cell cytoskeleton organization, migration, and metastasis dissemination (Rho A, cdc42 and Rac signaling, activation score= 1.9, 1.342, and 0.302, respectively). Indeed, Rho and cdc42 pull-down assays revealed increased Rho GTPase activity in cells overexpressing PDE5. Moreover, the selective ROCK inhibitor Y-27632 as well as the PDE5 inhibitor sildenafil were able to significantly reduce both migration and invasion of PDE5 clones. Our data reveal that PDE5 expression enhances motility and invasiveness of breast cancer cells through the activation of the Rho family of GTPases, and highlight, for the first time, a novel role for PDE5 as a marker of poor outcome in breast cancer patients. Citation Format: Barone I, Campana A, Giordano C, Tarallo R, Rinaldi A, Bruno G, Gyorffy B, Lanzino M, Bonofiglio D, Catalano S, Ando9 S. Phosphodiesterase type 5 promotes the invasive potential of breast cancer cells through Rho GTPase activation. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P5-04-10.