3 minute lightning oral presentation (and poster) ESA-SRB-APEG-NZSE 2022

Using a quantitative high-throughput screening platform to identify novel molecular targets and repurposable compounds for endometriosis treatment (#106)

Jacqueline F Donoghue 1 , Molly L Churchill 1 , Sarah J Holdsworth-Carson 1 2 , Karla J Cowley 3 , Jennii Luu 3 , Kaylene Simpson 3 4 , Martin Healey 1 5 , Peter AW Rogers 1
  1. Obstetrics and Gynaecology Department and Gynaecology Research Centre, University of Melbourne and The Royal Women's Hospital, Parkville, Victoria, Australia
  2. Julia Argyrou Endometriosis Centre Epworth HealthCare, East Melbourne, Victoria, Australia
  3. Victorian Centre for Functional Genomics , Peter MacCallum Cancer Centre , Parkville, Victoria, Australia
  4. Sir Peter MacCallum Department of Oncology, University of Melbourne, Parkville, Victoria, Australia
  5. Gynaecology Endometriosis and Pelvic Pain Unit, Royal Women’s Hospital, Parkville, Victoria, Australia

Endometriosis is defined as a chronic pro-inflammatory estrogen dependent disease where endometrial-like tissue grows outside of the uterine cavity of reproductive aged women. Endometriosis impacts 11% of Australian women causing chronic pelvic pain, dysmenorrhea and subfertility. Current pharmaceutical and surgical interventions are not always curative highlighting a significant need for the discovery of new compounds that disrupt estrogen-driven endometriosis pathways.  

Using quantitative high-throughput screening (qHTS) technology, 3,700 clinically approved compounds were screened in 384 well formats using 3 immortalised endometrial stromal cell lines.  Cells were screened under estrogenic conditions to mimic endometriosis environment and after 72 hours, cells were washed, fixed, stained, and imaged using the high Cell-Insight CX7 High Content Imaging system. Compounds with >70% growth inhibition were selected for dose response curve validation where an IC50 ≤1 µM triaged a compound for cell-based functional assay validation.

The initial qHTS identified 40 compounds that significantly inhibited cell growth during estrogen treatment compared to vehicle.  This demonstrated a specific inhibitory function related to estrogen stimulated signalling.  From these compounds, 23 exhibited IC50 ≤1µM demonstrating a physiological relevant (non-toxic) concentration.  Six compounds with analgesic or anti-inflammatory functions (Cytidine triphosphate disodium, 7-8 Dimethoxyflavone, Hydroxyzine pamoate, Pregabalin, Repaglinide, and Sildenafil HCl) were then characterised in cell based functional assays with each compound inhibiting cell migration, invasion and adhesion along with down regulation of ERK signalling and compound target expression.

New therapeutic discoveries for non-hormonal treatments for endometriosis are essential to improve patient lives and repurposing existing compounds is a faster pathway to clinical translation. This study demonstrates the feasibility of qHTS for therapeutic discovery in endometrial disease and identified several compounds that are prioritised for preclinical endometriosis model investigations and future clinical trials.  Future studies using this technology are also being designed to accommodate organoid cell lines, molecular screens and novel compounds.