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Endometrial carcinoma (ECa) is the fourth most common cancer among women. The oncogene PELP1 is frequently overexpressed in a variety of cancers, including ECa. We recently generated SMIP34, a small‐molecule inhibitor of PELP1 that suppresses PELP1 oncogenic signaling. In this study, we assessed the effectiveness of SMIP34 in treating ECa. Treatment of established and primary patient‐derived ECa cells with SMIP34 resulted in a significant reduction of cell viability, colony formation ability, and induction of apoptosis. RNA‐seq analyses showed that SMIP34‐regulated genes were negatively correlated with ribosome biogenesis and eukaryotic translation pathways. Mechanistic studies showed that the Rix complex, which is essential for ribosomal biogenesis, is disrupted upon SMIP34 binding to PELP1. Biochemical assays confirmed that SMIP34 reduced ribosomal biogenesis and new protein synthesis. Further, SMIP34 enhanced the efficacy of mTOR inhibitors in reducing viability of ECa cells. SMIP34 is also effective in reducing cell viability in ECa organoids in vitro and explants ex vivo. Importantly, SMIP34 treatment resulted in a significant reduction of the growth of ECa xenografts. Collectively, these findings underscore the potential of SMIP34 in treating ECa.

Abstract Of all gynecologic cancers, epithelial-ovarian cancer (OCa) stands out with the highest mortality rates. Despite all efforts, 90% of individuals who receive standard surgical and cytotoxic therapy experience disease recurrence. The precise mechanism by which leukemia inhibitory factor (LIF) and its receptor (LIFR) contribute to the progression of OCa remains unknown. Analysis of cancer databases revealed that elevated expression of LIF or LIFR was associated with poor progression-free survival of OCa patients and a predictor of poor response to chemotherapy. Using multiple primary and established OCa cell lines or tissues that represent five subtypes of epithelial-OCa, we demonstrated that LIF/LIFR autocrine signaling is active in OCa. Moreover, treatment with LIFR inhibitor, EC359 significantly reduced OCa cell viability and cell survival with an IC50 ranging from 5-50 nM. Furthermore, EC359 diminished the stemness of OCa cells. Mechanistic studies using RNA-seq and rescue experiments unveiled that EC359 primarily induced ferroptosis by suppressing the glutathione antioxidant defense system. Using multiple in vitro, ex vivo and in vivo models including cell-based xenografts, patient-derived explants, organoids, and xenograft tumors, we demonstrated that EC359 dramatically reduced the growth and progression of OCa. Additionally, EC359 therapy considerably improved tumor immunogenicity by robust CD45+ leukocyte tumor infiltration and polarizing tumor-associated macrophages (TAMs) toward M1 phenotype while showing no impact on normal T-, B-, and other immune cells. Collectively, our findings indicate that the LIF/LIFR autocrine loop plays an essential role in OCa progression and that EC359 could be a promising therapeutic agent for OCa.

Objectives: Endometrial cancer is the most common gynecologic malignancy in women. Of the two histological subtypes of endometrial cancer described, Type II tumors are biologically more aggressive and convey a less favorable prognosis when compared to Type I tumors. Novel therapeutics for treatment of Type II endometrial cancer are needed. Leukemia Inhibitory Factor Receptor (LIFR) and its ligand, LIF, play a critical role in cancer progression, metastasis, stem cell maintenance, and therapy resistance. Our recent studies revealed high levels of LIF and its receptor LIFR are associated with poor survival in endometrial cancer patients. We evaluated the efficacy of EC359, a novel small molecule inhibitor of the LIF receptor, against Type II Endometrial cancer cells. Methods: Primary and established cell lines were utilized in this study. Established cell lines were purchased from the American Type Culture Collection (ATCC CRL-1622). All primary samples were collected through the UT Health San Antonio tumor registry under IRB approval. The recently developed LIFR inhibitor, EC359, was used to elucidate the therapeutic utility of targeting the LIF/LIFR pathway in Type II endometrial cancer. In vitro activity was tested using MTT, colony formation, invasion, and apoptosis assays. Mechanistic studies were conducted using Western blot, reporter gene assays, and RT-qPCR analysis. Patient-derived xenograft (PDX) models were used for preclinical evaluation and toxicity. Results: Treatment of established and primary endometrial cancer cells with the LIFR inhibitor EC359 significantly reduced cell viability, long-term cell survival, invasion, and promoted apoptosis. Treatment with EC359 attenuated the activation of LIF/LIFR driven pathways including STAT3, mTOR, and AKT. EC359 significantly reduced tumor progression in patient derived xenograft model. Conclusions: These data support EC359 as a potential novel small molecule therapeutic for treatment of Type II Endometrial cancer.

Objectives: The purpose of this study is to describe the impact of paracrine signaling of plasminogen activator inhibitor-1 (PAI-1) and interleukin-6 (IL-6) on endometrial epithelial junctional proteins in an obese microenvironment in order to gain insight into non-hormonal mechanisms of endometrial oncogenesis in the obese population. Methods: Immortalized endometrial epithelial cells (EM-E6/E7/TERT) were cultured and treated with PAI-1 (0.5 nM) and IL-6 (10 ng/mL). Cells were assessed for changes in junctional complex gene transcription, protein expression, and cellular proliferation and migration, via quantitative real time polymerase chain reaction (qRT-PCR), Western blotting, and Incucyte-based functional assays of proliferation. Immunofluorescence (IF) staining and confocal microscopy were used to highlight changes in connexin 32 (Cx32) encoded by the gap junction gene GJB1. These experiments were repeated in another established endometrial epithelial cell line (hEM3). Results: Transcription of GJB1 and corresponding protein connexin 32 (Cx32) levels were reduced when EME-E6/E7/TERT cells were treated with both PAI-1 and IL-6. This reduction in Cx32 levels with PAI-1 and IL-6 treatments was also seen in the hEM3 cells. Conclusions: The decline of Cx32 levels in the setting of PAI-1 and IL-6 exposure indicates that PAI-1 and IL-6 contribute to important dysregulatory changes within the endometrial epithelial junctional complex proteins. This assertion is strengthened by concordant findings in a separate endometrial epithelial cell line. Reduction in Cx32 production has previously been associated with endometrial hyperplasia and carcinoma. The findings of this study link obesity-related paracrine signaling to this phenomenon, which provides further information about causality between obesity and endometrial cancer. More research is required to investigate if IL-6 or PAI-1 would serve as a suitable therapeutic target in the management of endometrial hyperplasia or malignancy in obesity.