Molecular classification of endometrial carcinoma applied to endometrial atypical hyperplasia biopsy specimens.
Authors: Corey Logan , Wu Sharon , Brodskiy Pavel , Hodges Kurt , Oberley Matthew James , Musallam Rami , Kheil Mira , Bandyopadhyay Sudeshna , Wallbillich John , Winer Ira Seth , Morris Robert , Ali-Fehmi Rouba 
Published: June 2022

e17622

Background: 50% of endometrial atypical hyperplasia (AH) cases on endometrial sampling have concurrent endometrial cancer (EC) on hysterectomy. Little data exists regarding molecular signatures that can predict the presence of concurrent endometrial cancer in patients with AH on biopsy. Therefore, we set to characterize molecular landscapes of endometrial samples diagnosed with AH with EC on the final hysterectomy specimen, using matched controls. Methods: Karmanos Cancer Institute pathology database was searched for patients with AH on endometrial curettage or biopsy who had a hysterectomy within 6 months. Only samples with adequate tissue for next generation sequencing were included. In total, 59 tissue samples from 34 patients were included: there were 15 AH endometrial biopsy samples, 18 AH final hysterectomy samples (13 matched pairs), 13 EC endometrial biopsies and 13 samples that were EC on final hysterectomy samples (12 matched pairs) that were analyzed using next-generation sequencing (WES) and/or whole transcriptome sequencing (WTS) (NovaSeq). TMB was measured by counting all somatic mutations found per tumor (TMB-high: >10 mutations per MB). Immune cell infiltrates were calculated by Quantiseq. Significance was determined using Fisher exact, Chi-square and Mann-Whitney U test and adjusted for multiple comparisons: p < 0.05 but q > 0.05 was considered a trend. Results: Clinical analysis demonstrated 15/34 patients with endometrial AH on initial sampling and EC on the final hysterectomy. 2 out of 15 patients were > stage II at time of surgery. Molecularly, fewer PTEN mutations were found between AH (2/8) and EC (14/17) on final hysterectomy (25% vs 82.4%, p = 0.01). Samples of EC origin were MSI-H by NGS-MSI (3/18) and had no mutations in PPP2R1A (0/16). Comparing hysterectomy samples, EC trended toward increased CTLA4 (FC: 6.97-fold) expression and immune cell infiltration of Macrophage M1 (+1.19%), NK cells (+2.13%), CD8+ T Cells (+1.27%), regulatory T cells (+2.17%) and Dendritic cells (+2.87%) compared to AH samples (all p < 0.05). Similarly, when comparing endometrial biopsies, EC samples trended toward increased expression of CTLA4 (12.6-fold), HAVCR2/TIM3 (FC: 2.59-fold) and IFNG (FC: 17.8-fold) immune checkpoint genes, as well as increased immune cell infiltration of Neutrophils (+11.7%), CD8+ T cells (+2.39%) and regulatory T cells (+2.43%) compared to AH samples (all p < 0.05). Conclusions: There are molecular and tumor microenvironment differences seen between AH endometrial biopsies that have concurrent EC compared to those that do not on the final hysterectomy specimens. These differences may lead to advances in identifying appropriate patients for fertility sparing treatments, versus those that can be managed surgically by a benign gynecologist or a gynecologic oncologist. Analyses with larger sample sizes are needed and are ongoing.