Kanako Hayashi, PhD
My research program is focused on understanding the mechanisms of female reproductive disorders. Specifically, my long-term goal is to discover novel mechanisms and targets for developing therapeutic strategies to suppress ovarian and endometrial cancer progression and metastasis, as well as improve female fertility (uterine function and endometriosis).
Ovarian cancer is the most lethal gynecological cancer due to the advanced stage of the disease at presentation and the inherent limitations of current therapeutic options. The canonical pathway of WNT signaling results in nuclear accumulation of β-catenin and transcriptional activation of target genes. Thus, activation of β-catenin promotes further downstream targets leading to tumor progression and metastasis. When I started working on WNT signaling in ovarian cancer, activation of β-catenin by gene mutation had been reported, but only in endometrioid subtype of ovarian cancer which only represents 3-5% of all ovarian cancers. The importance of WNT/β-catenin signaling in serous carcinomas, which are the most common subtype of ovarian carcinomas, was not described. We have first reported that the expression of WNT7A during malignant transformation of ovarian cancer plays a critical role in tumor progression mediated by the WNT/β-catenin signaling pathway. We also found that abundant WNT7A is present especially in serous carcinomas. WNT7A in ovarian cancer cells activates the WNT/β-catenin signaling pathway, and directly promotes cell functions associated with tumor growth in vivo. The pattern of diffuse tumor growth and progression is correlated to the relative level of WNT7A. These effects can be mediated by upregulation of Cyclin D1 and MMP7, downstream targets of WNT/β-catenin signaling, leading to stimulation of both cell cycle and migratory/invading programs in ovarian cancer cells. Thus, WNT7A-activated β-catenin signaling plays a major role in the primary tumorigenesis and the metastatic progression of serous ovarian carcinomas. These results were published in Mol Cancer Res 2012, and received “Best of American Association for Cancer Research 2012” (one of the most highly-cited Mol Cancer Res articles published in 2012).
Our next logical step was to identify novel WNT7A regulated genes. Our microarray approach, as a strategy for identifying novel WNT7A signaling target genes, highlighted FGF1, a member of the fibroblast growth factor family. We found that high expression of WNT7A and FGF1 are correlated in ovarian carcinomas and poor overall patient survival. A chromatin immunoprecipitation assay demonstrated that WNT7A/β-catenin signaling directly regulates FGF1 expression via TCF binding elements in the FGF1-1C promoter locus. In vitro gene manipulation studies revealed that FGF1 is sufficient to drive the tumor promoting effects of WNT7A. In vivo xenograft studies confirmed that the stable overexpression of WNT7A or FGF1 induced a significant increase in tumor incidence, while FGF1 knockdown in WNT7A overexpressing cells caused a significant reduction in tumor size. Furthermore, niclosamide most efficiently abrogated WNT7A/β-catenin signaling in our model, inhibited β-catenin transcriptional activity and cell viability, and increased cell death. Niclosamide decreased cell migration following an increase in E-cadherin subsequent to decreased levels of SLUG. The effects of niclosamide on cell functions were more potent in WNT7A overexpressing cells. Oral administration of niclosamide inhibited tumor growth and progression in an intraperitoneal xenograft mouse model representative of human ovarian cancer. Thus, these results indicate that FGF1 is a direct downstream target of WNT7A/β-catenin signaling and this pathway has potential as a therapeutic target in ovarian cancer. Moreover, niclosamide is a promising inhibitor of this pathway and may have clinical relevance. These results were published in Oncogene 2015.
Now, we are further trying to understand the mechanism of niclosamide’s direct action in chemoresistant ovarian cancer and identifying novel therapeutic targets. One of the reasons of the high mortality related to ovarian cancer is the inherent limitation of current therapeutic options (i.e. high incidence of recurrence with chemoresistant tumors). Niclosamide exhibits the most significant impact on inhibition of WNT/β-catenin signaling, and is able to target chemoresistant stem-like cancer cells. Therefore, a clear understanding of niclosamide actions, and how it may be manipulated to develop a new therapeutic drug for ovarian cancer, will improve the survival of ovarian cancer patients.
Endometrial cancer is the most common malignancy of the female genital tract. Two types of endometrial carcinoma have been distinguished, type I and type II. Type I (endometrioid) tumors, which account for 80% of all endometrial carcinomas, have a good prognosis, whereas Type II (serous) tumors are very aggressive, and more than 50% of patients present with recurrent disease shortly after primary treatment. A number of studies have reported that TP53 mutations are associated with poor prognosis, and approximately 80% of type II endometrial carcinomas harbor TP53 mutations. In addition to TP53 mutation, inactivation of CDH1 is also a common molecular feature in type II endometrial carcinomas. While TP53 mutation and CDH1 inactivation occur in 80-90% of endometrial type II carcinomas, the mechanisms underlying the aggressive phenotype in type II tumors have not been studied. In addition, the existing animal models had not sufficiently modeled or recapitulated type II human endometrial cancer. Therefore, we generated mice with conditional ablation of Trp53 and Cdh1 in the mouse uterus using the innovative Pgr-Cre mice to understand the mechanisms of tumorigenesis necessary for early stage diagnosis as well as rational design of therapies to increase long-term survival. While the uteri of mice lacking Trp53 or Cdh1 alone did not induce any tumor formation (initial characterization of Cdh1 cko mice was published in Biol Reprod 2012), conditional ablation of Cdh1 and Trp53 (Cdh1d/d Trp53d/d) clearly demonstrate architectural features characteristic of type II endometrial carcinomas. Cdh1d/d Trp53d/d tumors in 12-mo old mice were highly aggressive, and metastasized to nearby and distant organs within the peritoneal cavity, such as abdominal lymph nodes, mesentery and peri-intestinal adipose tissues, demonstrating that tumorigenesis in this model proceeds through the universally recognized morphologic intermediates associated with type II endometrial neoplasia. We also observed abundant cell proliferation and complex angiogenesis in the uteri of Cdh1d/d Trp53d/d mice. Our microarray analysis found that most of the genes differentially regulated in the uteri of Cdh1d/d Trp53d/d mice were involved in inflammatory responses. CD163 and Arg1, markers for tumor-associated macrophages, were also detected and increased in the uteri of Cdh1d/d Trp53d/d mice, suggesting that an inflammatory tumor microenvironment with immune cell recruitment is augmenting tumor development in Cdh1d/d Trp53d/d uteri. Furthermore, inflammatory mediators secreted from CDH1 negative, TP53 mutant endometrial cancer cells induced normal macrophages to express inflammatory related genes through activation of NFκB signaling. These results indicate that absence of CDH1 and TP53 in endometrial cells initiates chronic inflammation, promotes tumor microenvironment development following the recruitment of macrophages, and promotes aggressive endometrial carcinomas. The results were published in Oncogene 2014. We also characterized conditional ablation of Cdh1 and Pten in the mouse uterus. Although the uteri of Cdh1d/d Ptend/d mice were abnormally structured with curly horns, disorganized epithelial structure, and accelerated cellular invasiveness and angiogenesis, these mice died at postnatal day 15-19 with massive blood loss. Therefore, we could not further examine the mechanisms of invasiveness and angiogenesis in the uteri of Cdh1d/d Ptend/d mice. These results were published in Biol Reprod 2013.
"Development of New Therapeutic Strategies for Endometriosis" NICHHD $184,375
“Mechanisms of WNT7A-FGF1 signaling and their therapeutic potential in ovarian cancer” NIH/NCI $442,500