Modeling Epithelial Ovarian Cancer In The Mouse

For over five decades epithelial ovarian cancer (EOC) is identified as the leading cause of death from gynecological malignancies in the USA. Unfortunately, due to latent progression of EOC and lack of accurate animal models studies of this disease's pathogenesis have been challenging. Not surprisingly, establishment of adequate screening tests for early disease stages, at which point the cancer would be curable, and advances in treatment approaches have been slow. Deficiency of tumor suppressor genes p53 and RB and overexpression of c-MYC oncogene occur frequently in the most common form of sporadic EOC, high grade serous adenocarcinoma. Thus, we have initiated development of several mouse models of EOC based on alterations of these genes in the presumed EOC tissue of origin, the ovarian surface e pithelium (OSE). By using a new approach of in situ targeting of OSE we have demonstrated that disruption of p53 and Rb leads to high grade serous adenocarcinoma. We have also established new models of EOC intraperitoneal spreading and have shown that c-MYC promotes tumorigenicity of p53-and p53/Rb-deficient OSE. Next, we have shown that multiphoton microscopy and second harmonic generation (MPM/SHG) imaging are well suited for studies of cell motility in vivo and can complement pathological characterization EOC by allowing optical biopsies of native human and mouse tissues at resolution comparable to that of routinely processed histological sections. We have also shown the feasibility of laparoscopic MPM/SHG. Moreover, we identified layered nanohybrids as suitable particles for combination of intravital biomedical imaging with controlled drug release. As the first step to determine whether EOC arises from the stem cell compartment we have identified an OSE subpopulation which is positive for stem cell marker aldehyde dehydrogenase 1, efficiently forms clonogenic spheres and displays extended self-renewal properties in serial sphere generation assays. These putative OSE stem cells are slow cycling according to BrdU label retention assays and are mainly located in the hilum area of the mouse ovary. These findings will direct our future search for better approaches to target EOC cells with stem cell properties

Strategies for High-Resolution Imaging of Epithelial Ovarian Cancer by Laparoscopic Nonlinear Microscopy1

Ovarian cancer remains the most frequently lethal of the gynecologic cancers owing to the late detection of this disease. Here, by using human specimens and three mouse models of ovarian cancer, we tested the feasibility of nonlinear imaging approaches, the multiphoton microscopy (MPM) and second harmonic generation (SHG) to serve as complementary tools for ovarian cancer diagnosis. We demonstrate that MPM/SHG of intrinsic tissue emissions allows visualization of unfixed, unsectioned, and unstained tissues at a resolution comparable to that of routinely processed histologic sections. In addition to permitting discrimination between normal and neoplastic tissues according to pathological criteria, the method facilitates morphometric assessment of specimens and detection of very early cellular changes in the ovarian surface epithelium. A red shift in cellular intrinsic fluorescence and collagen structural alterations have been identified as additional cancer-associated changes that are indiscernible by conventional pathologic techniques. Importantly, the feasibility of in vivo laparoscopic MPM/SHG is demonstrated by using a “stick” objective lens. Intravital detection of neoplastic lesions has been further facilitated by low-magnification identification of an indicator for cathepsin activity followed by MPM laparoscopic imaging. Taken together, these results demonstrate that MPM may be translatable to clinical settings as an endoscopic approach suitable for high-resolution optical biopsies as well as a pathology tool for rapid initial assessment of ovarian cancer samples

miR-34 Cooperates with p53 in Suppression of Prostate Cancer by Joint Regulation of Stem Cell Compartment

The miR-34 family was originally found to be a direct target of p53 and is a group of putative tumor suppressors. Surprisingly, mice lacking all mir-34 genes show no increase in cancer formation by 18 months of age, hence placing the physiological relevance of previous studies in doubt. Here, we report that mice with prostate epithelium-specific inactivation of mir-34 and p53 show expansion of the prostate stem cell compartment and develop early invasive adenocarcinomas and high-grade prostatic intraepithelial neoplasia, whereas no such lesions are observed after inactivation of either the mir-34 or p53 genes alone by 15 months of age. Consistently, combined deficiency of p53 and miR-34 leads to acceleration of MET-dependent growth, self-renewal, and motility of prostate stem/progenitor cells. Our study provides direct genetic evidence that mir-34 genes are bona fide tumor suppressors and identifies joint control of MET expression by p53 and miR-34 as a key component of prostate stem cell compartment regulation, aberrations in which may lead to cancer

Ovarian surface epithelium at the junction area contains a cancer-prone stem cell niche

Epithelial ovarian cancer (EOC) is the fifth leading cause of cancer deaths among women in the United States, but its pathogenesis is poorly understood. Some epithelial cancers are known to occur in transitional zones between two types of epithelium, whereas others have been shown to originate in epithelial tissue stem cells. The stem cell niche of the ovarian surface epithelium (OSE), which is ruptured and regenerates during ovulation, has not yet been defined unequivocally. Here we identify the hilum region of the mouse ovary, the transitional (or junction) area between the OSE, mesothelium and tubal (oviductal) epithelium, as a previously unrecognized stem cell niche of the OSE. We find that cells of the hilum OSE are cycling slowly and express stem and/or progenitor cell markers ALDH1, LGR5, LEF1, CD133 and CK6B. These cells display long-term stem cell properties ex vivo and in vivo, as shown by our serial sphere generation and long-term lineage-tracing assays. Importantly, the hilum cells show increased transformation potential after inactivation of tumour suppressor genes Trp53 and Rb1, whose pathways are altered frequently in the most aggressive and common type of human EOC, high-grade serous adenocarcinoma. Our study supports experimentally the idea that susceptibility of transitional zones to malignant transformation may be explained by the presence of stem cell niches in those areas. Identification of a stem cell niche for the OSE may have important implications for understanding EOC pathogenesis

Local Mesenchymal Stem/Progenitor Cells Are a Preferential Target for Initiation of Adult Soft Tissue Sarcomas Associated with p53 and Rb Deficiency

The cell of origin and pathogenesis of the majority of adult soft tissue sarcomas (STS) remains poorly understood. Because mutations in both the P53 and RB tumor suppressor genes are frequent in STS in humans, we inactivated these genes by Cre-loxP–mediated recombination in mice with floxed p53 and Rb. Ninety-three percent of mice developed spindle cell/pleomorphic sarcomas after a single subcutaneous injection of adenovirus carrying Cre-recombinase. Similar to human STS, these sarcomas overexpress Cxcr4, which contributes to their invasive properties. Using irradiation chimeras generated by transplanting bone marrow cells from mice carrying either the Rosa26StoploxPLacZ or the Z/EG reporter, as well as the floxed p53 and Rb genes, into irradiated p53loxP/loxPRbloxP/loxP mice, it was determined that sarcomas do not originate from bone marrow–derived cells, such as macrophages, but arise from the local resident cells. At the same time, dermal mesenchymal stem cells isolated by strict plastic adherence and low levels of Sca-1 expression (Sca-1low, CD31negCD45neg) have shown enhanced potential for malignant transformation according to soft agar, invasion, and tumorigenicity assays, after the conditional inactivation of both p53 and Rb. Sarcomas formed after transplantation of these cells have features typical for undifferentiated high-grade pleomorphic sarcomas. Taken together, our studies indicate that local Sca-1low dermal mesenchymal stem/progenitor cells are preferential targets for malignant transformation associated with deficiencies in both p53 and Rb