We found that, similar to Yki, phosphorylation resistant human Yap is able to induce ovarian tumors when expressed in the Drosophila ovary, suggesting that the ability of Yap and Yki to induce ovarian tumorigenesis is conserved. Although we found no evidence of activating mutations in Yap in ovarian cancers, or for Yap overexpression in cell lines, we found that sub-cellular levels of Yap and pYap correlate and tend to associate, respectively, with survival of ovarian cancer patients. Consistent with a role for Yap in ovarian cancer progression, we find that overexpression of Yap2 and phosphorylation-defective Yap2-5SA in 80T cells resulted in several phenotypes associated with cancer including increased proliferation, resistance to cisplatin-induced apoptosis, faster cell migration, and anchorage independent growth, while knockdown of Yap in these cells resulted in increased sensitivity to cisplatin-induced apoptosis.
Paradoxically, Yap has been shown to act as both a tumor suppressor and as an oncogene in breast cancer (20
). However, neither of these studies attempted to correlate Yap expression with patient outcome, which would help resolve this controversy. Three lines of evidence refute a role for Yap as a tumor suppressor in ovarian cancer. First, low nYap expression in ovarian tumors correlated with better disease-specific survival. Second, Yap was not lost or dramatically down-regulated in 10 ovarian cancer cell lines. Third, Yap knockdown in 80T cells did not result in phenotypes associated with cancer. In fact, Yap knockdown cells were more sensitive to cisplatin-induced apoptosis than control cells ().
Although the sample size is modest and should be interpreted cautiously, Yap appears to be a powerful prognostic marker of poor survival in ovarian cancer. Cox regression analysis of the nY/cpY Category (3 vs 0, 1, 2,) revealed an exceptionally high hazard ratio of 7.7(comparable to the hazard ratio of 7.0 for stage), indicating that, in our data set, patients with both high nYap and low cpYap are at nearly eight times greater risk of death from the disease than other patients. This hazard ratio is substantially higher than the hazard ratio for nYap staining alone (HR=3.3). There could be two explanations for this finding. First, immunohistochemical studies of archival tumor samples can be confounded by both systematic and random errors such as inter- and intra-sample heterogeneity and differences in the age, fixation, processing, staining, and scoring of tumor samples. Therefore, two independent measures of the same marker would be expected to reduce error. Second, the nY/cpY Category might reflect the activity of Yap in the tumor. Yap is phosphorylated at S127 by Lats kinases and retained in the cytoplasm by 14-3-3(8
). When Yap is localized in the nucleus, it is likely transcriptionally active and thus intense nYap staining suggests high Yap activity. In contrast, when Yap is phosphorylated and in the cytoplasm, Yap is presumably transcriptionally inactive and thus intense cpYap staining suggests low Yap activity. Thus, tumors in nY/cpY Category 3, which have high levels of nYap and low levels of cpYap, might be expected to have the highest Yap activity. Analysis of the levels of Yap target genes in each nY/cpY Category might shed light on these two possibilities.
An important question that remains is the mechanism of Yap misregulation in ovarian cancer. We found no evidence for Yap protein amplification or for Yap activating mutations. Further, pYap staining was detectable in 63/70 ovarian tumors and all ovarian cancer cell lines (Supplementary Fig. 1A
), indicating that increased nYap does not likely result from complete loss of phosphorylation. A variety of mechanisms have been described Drosophila
that alter Yki levels and subcellular localization (8
), providing several possible mechanisms by which nYap levels may increase in human ovarian cancer. Elucidation of these mechanisms will likely provide insight for therapeutic intervention targeting Yap.
Our results support the notion that Yap might be an excellent therapeutic target for the treatment of ovarian cancer. Though our sample size is modest and our analysis needs to be confirmed in a larger patient population, Yap distribution appears to be a very strong predictor of survival from ovarian cancer. Even if further analysis reveals that the hazard ratio for nY/cpY Category 3 is at the low end of our 95% confidence interval, (2.1–28.9), this would be on par with EGF and Her-2/neu, which are currently targeted in clinical trials for ovarian cancer treatments (22
). Further, Yap activation results in several cancer-associated phenotypes while reduction of Yap resulted only in increased sensitivity to chemotherapy, indicating that therapies targeting Yap could affect multiple cancer processes with minimal deleterious effects.
ovarian follicular epithelium and human OSE are female specific tissues that share many important properties including mesodermal origin, monolayer cuboidal epithelial organization, hormone dependency, and the ability to undergo dynamic remodeling (7
). The results presented here support the notion that the Drosophila
follicular epithelium is a useful tool for the discovery of genes crucial to the pathogenesis of ovarian cancer.