Development and characterization of human fetal female reproductive tract organoids to understand Müllerian duct anomalies.

Müllerian ducts are paired tubular structures that give rise to most of the female reproductive organs. Any abnormalities in the development and differentiation of these ducts lead to anatomical defects in the female reproductive tract organs categorized as Müllerian duct anomalies. Due to the limited access to fetal tissues, little is understood of human reproductive tract development and the associated anomalies. Although organoids represent a powerful model to decipher human development and disease, such organoids from fetal reproductive organs are not available. Here, we developed organoids from human fetal fallopian tubes and uteri and compared them with their adult counterparts. Our results demonstrate that human fetal reproductive tract epithelia do not express some of the typical markers of adult reproductive tract epithelia. Furthermore, fetal organoids are grossly, histologically, and proteomically different from adult organoids. While external supplementation of WNT ligands or activators in culture medium is an absolute requirement for the adult reproductive tract organoids, fetal organoids are able to grow in WNT-deficient conditions. We also developed decellularized tissue scaffolds from adult human fallopian tubes and uteri. Transplantation of fetal organoids onto these scaffolds led to the regeneration of the adult fallopian tube and uterine epithelia. Importantly, suppression of Wnt signaling, which is altered in patients with Müllerian duct anomalies, inhibits the regenerative ability of human fetal organoids and causes severe anatomical defects in the mouse reproductive tract. Thus, our fetal organoids represent an important platform to study the underlying basis of human female reproductive tract development and diseases.

Bovine and human endometrium-derived hydrogels support organoid culture from healthy and cancerous tissues.

Organoid technology has provided unique insights into human organ development, function, and diseases. Patient-derived organoids are increasingly used for drug screening, modeling rare disorders, designing regenerative therapies, and understanding disease pathogenesis. However, the use of Matrigel to grow organoids represents a major challenge in the clinical translation of organoid technology. Matrigel is a poorly defined mixture of extracellular matrix proteins and growth factors extracted from the Engelbreth-Holm-Swarm mouse tumor. The extracellular matrix is a major driver of multiple cellular processes and differs significantly between tissues as well as in healthy and disease states of the same tissue. Therefore, we envisioned that the extracellular matrix derived from a native healthy tissue would be able to support organoid growth akin to organogenesis in vivo. Here, we have developed hydrogels from decellularized human and bovine endometrium. These hydrogels supported the growth of mouse and human endometrial organoids, which was comparable to Matrigel. Organoids grown in endometrial hydrogels were proteomically more similar to the native tissue than those cultured in Matrigel. Proteomic and Raman microspectroscopy analyses showed that the method of decellularization affects the biochemical composition of hydrogels and, subsequently, their ability to support organoid growth. The amount of laminin in hydrogels correlated with the number and shape of organoids. We also demonstrated the utility of endometrial hydrogels in developing solid scaffolds for supporting high-throughput, cell culture-based applications. In summary, endometrial hydrogels overcome a major limitation of organoid technology and greatly expand the applicability of organoids to understand endometrial biology and associated pathologies.

Anogenital High-Grade Squamous Intraepithelial Lesion Comorbid With Vulvar Lichen Sclerosus and Lichen Planus.

OBJECTIVE: The aim of the study was to describe the clinicopathologic features of vulvovaginal or anal high-grade squamous intraepithelial lesion (HSIL) comorbid with lichen sclerosus and/or lichen planus (LS/LP). METHODS: The local pathology database identified 37 consecutive cases from 2007 to 2019 of vulvar, vaginal, or anal HSIL among women who had a histopathologic diagnosis of vulvar LS/LP. Cases had p16 and p53 immunoperoxidase stains. Clinical data included age, relative location of HSIL and LS/LP, immune-modifying conditions, tobacco use, treatment type, and follow-up. Histopathologic data included HSIL morphology categorized as warty-basaloid or keratinizing, p16 and p53 patterns within HSIL, and features of LS/LP. RESULTS: The mean age was 69 years with a median follow-up up 42 months. Lichen sclerosus, alone or in combination with LP, was the comorbid dermatosis in 89%. Lichen sclerosus/lichen planus was overlapping or adjacent to HSIL in two-thirds of cases and located separately in the remainder. Rates of tobacco use and immunologic dysfunction were each 40%. In cases of co-located LS and HSIL, sclerosis was absent under the neoplasia in 57%. Twenty-four percent of HSIL cases showed keratinizing morphology; block-positive p16 and suprabasilar-dominant p53 helped distinguish HSIL from human papillomavirus-independent neoplasia. CONCLUSIONS: Histopathologic identification of comorbid HSIL and LS/LP may be challenging because of keratinizing morphology and loss of diagnostic features of LS. Clinicopathologic correlation and use of p16 and p53 are essential to achieve an accurate diagnosis and enact disease-specific management plans.

Stage IB2 adenosquamous cervical cancer diagnosed at 19-weeks' gestation.

Neoadjuvant chemotherapy (NACT) for advanced cervical cancer in pregnancy has been shown to increase operability and be effective against spread of disease. In all reported cases of advanced disease, residual tumour has been found at surgery following NACT. We present a case of a 27-year old diagnosed with stage IB2 adenosquamous cervical carcinoma at 19-weeks' gestation who was treated with NACT. Following caesarean section and radical hysterectomy, histopathology showed no evidence of residual tumour in the cervix and negative pelvic lymph nodes.

Proteomic and functional characterization of intra-tumor heterogeneity in human endometrial cancer.

Endometrial cancer is one of the most frequently diagnosed gynecological cancers worldwide, and its prevalence has increased by more than 50% over the last two decades. Despite the understanding of the major signaling pathways driving the growth and metastasis of endometrial cancer, clinical trials targeting these signals have reported poor outcomes. The heterogeneous nature of endometrial cancer is suspected to be one of the key reasons for the failure of targeted therapies. In this study, we perform a sequential window acquisition of all theoretical fragment ion spectra (SWATH)-based comparative proteomic analysis of 63 tumor biopsies collected from 20 patients and define differences in protein signature in multiple regions of the same tumor. We develop organoids from multiple biopsies collected from the same tumor and show that organoids capture heterogeneity in endometrial cancer growth. Overall, using quantitative proteomics and patient-derived organoids, we define the heterogeneous nature of endometrial cancer within a patient's tumor.

Adipose-Derived VEGF-mTOR Signaling Promotes Endometrial Hyperplasia and Cancer: Implications for Obese Women.

Obesity is responsible for increased morbidity and mortality in endometrial cancer. Despite the positive correlation of body mass index (BMI) or obesity in endometrial carcinogenesis, the contribution of adipose tissue to the pathogenesis of endometrial hyperplasia and cancer is unclear. This study clarifies the role of adipocytes in the pathogenesis of endometrial cancer by demonstrating that adipocyte-conditioned medium (ACM) increases proliferation, migration, and survival of endometrial cancer cells compared with preadipocyte-conditioned medium (PACM). Comparative cytokine array analysis of ACM and PACM reveal upregulation of a group of cytokines belonging to the VEGF signaling pathway in ACM. VEGF protein expression is upregulated in visceral adipose tissue (VAT) in obese patients, which is correlated with increased tumor growth in an in vivo xenograft model. The increased tumor size is mechanistically associated with the activation of the PI3K/AKT/mTOR pathway, a downstream target of VEGF signaling, and its suppression decreased the growth-promoting effects of VAT on endometrial cancer cells. Similar to the human model systems, pathologic changes in endometrial cells in a hyperphagic obese mouse model are associated with increased body weight and hyperactive mTOR signaling. Analysis of human tissue specimens depicts increased in tumor vasculature and VEGF-mTOR activity in obese endometrial cancer patients compared with nonobese patients. Collectively, these results provide evidence that VEGF-mTOR signaling drives endometrial cell growth leading to hyperplasia and cancer.Implications: Adipocyte-derived VEGF-mTOR signaling may be an attractive therapeutic target against endometrial cancer in obese women. Mol Cancer Res; 16(2); 309-21. ©2017 AACR.