Whole-proteome analysis of mesonephric-derived cancers describes new potential biomarkers.

Mesonephric carcinomas (MEs) and female adnexal tumors of probable Wolffian origin (FATWO) are derived from embryologic remnants of Wolffian/mesonephric ducts. Mesonephric-like carcinomas (MLCs) show identical morphology to ME of the cervix but occur in the uterus and ovary without convincing mesonephric remnants. ME, MLC, and FATWO are challenging to diagnose due to their morphologic similarities to Müllerian/paramesonephric tumors, contributing to a lack of evidence-based and tumor-specific treatments. We performed whole-proteomic analysis on 9 ME/MLC and 56 endometrial carcinomas (ECs) to identify potential diagnostic biomarkers. Although there were no convincing differences between ME and MLC, 543 proteins showed increased expression in ME/MLC relative to EC. From these proteins, euchromatic histone lysine methyltransferase 2 (EHMT2), glutathione S-transferase Mu 3 (GSTM3), eukaryotic translation elongation factor 1 alpha 2 (EEF1A2), and glycogen synthase kinase 3 beta were identified as putative biomarkers. Immunohistochemistry was performed on these candidates and GATA3 in 14 ME/MLC, 8 FATWO, 155 EC, and normal tissues. Of the candidates, only GATA3 and EHMT2 were highly expressed in mesonephric remnants and mesonephric-derived male tissues. GATA3 had the highest sensitivity and specificity for ME/MLC versus EC (93% and 99%) but was absent in FATWO. EHMT2 was 100% sensitive for ME/MLC & FATWO but was not specific (65%). Similarly, EEF1A2 was reasonably sensitive to ME/MLC (92%) and FATWO (88%) but was the least specific (38%). GSTM3 performed intermediately (sensitivity for ME/MLC and FATWO: 83% and 38%, respectively; specificity 67%). Although GATA3 remained the best diagnostic biomarker for ME/MLC, we have identified EHMT2, EEF1A2, and GSTM3 as proteins of interest in these cancers. FATWO's cell of origin is uncertain and remains an area for future research.

Single cell transcriptomes of normal endometrial derived organoids uncover novel cell type markers and cryptic differentiation of primary tumours.

Endometrial carcinoma, the most common gynaecological cancer, develops from endometrial epithelium which is composed of secretory and ciliated cells. Pathologic classification is unreliable and there is a need for prognostic tools. We used single cell sequencing to study organoid model systems derived from normal endometrial endometrium to discover novel markers specific for endometrial ciliated or secretory cells. A marker of secretory cells (MPST) and several markers of ciliated cells (FAM92B, WDR16, and DYDC2) were validated by immunohistochemistry on organoids and tissue sections. We performed single cell sequencing on endometrial and ovarian tumours and found both secretory-like and ciliated-like tumour cells. We found that ciliated cell markers (DYDC2, CTH, FOXJ1, and p73) and the secretory cell marker MPST were expressed in endometrial tumours and positively correlated with disease-specific and overall survival of endometrial cancer patients. These findings suggest that expression of differentiation markers in tumours correlates with less aggressive disease, as would be expected for tumours that retain differentiation capacity, albeit cryptic in the case of ciliated cells. These markers could be used to improve the risk stratification of endometrial cancer patients, thereby improving their management. We further assessed whether consideration of MPST expression could refine the ProMiSE molecular classification system for endometrial tumours. We found that higher expression levels of MPST could be used to refine stratification of three of the four ProMiSE molecular subgroups, and that any level of MPST expression was able to significantly refine risk stratification of the copy number high subgroup which has the worst prognosis. Taken together, this shows that single cell sequencing of putative cells of origin has the potential to uncover novel biomarkers that could be used to guide management of cancers. © 2020 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

c-KIT Analysis and Targeted Molecular Sequencing of Mesonephric Carcinomas of the Female Genital Tract.

Mesonephric carcinoma is a rare malignancy, thought to derive from Wolffian remnants. To date, no targeted molecular therapeutic options have been identified. On the basis of limited case reports, c-KIT immunohistochemical expression has been reported in female adnexal tumors of Wolffian origin, and targeted therapy with Imatinib has been attempted with mixed success. Currently, it is unclear whether c-KIT immunohistochemical expression is seen in mesonephric carcinoma, a tumor that is thought to be related to female adnexal tumors of Wolffian origin, and how this correlates with KIT mutational status. In this study, we assessed the immunohistochemical expression of c-KIT and KIT mutational status, in a series of 13 mesonephric neoplasms (5 cervical [including 2 cervical carcinosarcomas], 3 uterine corpora, 4 ovarian, and 1 vaginal/pelvic). The intensity of staining and proportion of cells showing cytoplasmic/membranous staining for c-KIT were recorded. KIT was sequenced using a next-generation sequencing panel that targeted 120 hotspots and 17 exons in 33 known actionable cancer genes. This panel included KIT exons 9, 11, and 13, and 6 hotspots (T670, D816, D820, N822, Y823, A829). Although c-KIT immunohistochemical expression was observed in the majority of mesonephric carcinomas (10/12 cases; 83%), no KIT mutations were detected. This cautions pathologists against the use of c-KIT immunohistochemistry as a surrogate marker for KIT-activating mutations in this setting. Consistent with previous studies, the majority of mesonephric neoplasms (10/13; 77%) harbored KRAS mutations. Additional mutations were found in CTNNB1 (2/13, 15%), TP53 (2/13, 15%), and PIK3CA (1/13, 8%).