Poster Presentation 37th Lorne Cancer Conference 2025

MLH1 promoter methylation detection in Endometrial Cancer through ctDNA analysis (#205)

Julia Matas 1 2 , Lesley Arends 1 2 , Maxwell Balden 1 2 , Sarah Ftouni 1 2 , Tarek Elsabeh 1 2 , Michelle M Cummins 3 , Kristy Robledo 3 , Martin Stockler 3 , Peey-Sei Kok 3 , Yeh Chen Lee 3 , Eric Joo 2 , Daniel Buchanan 2 , Linda Mileshkin 1 2 , Yoland Antill 4 , Sarah-Jane Dawson 1 2 , on behalf of the Australian New Zealand Gynaecology Oncology Group (ANZGOG) 5
  1. Peter MacCallum Cancer Centre, Melbourne, VIC, Australia
  2. University of Melbourne, Melbourne, VIC, Australia
  3. NHMRC Clinical Trials Centre, University of Sydney, Camperdown, NSW, Australia
  4. Dept Medical Oncology, Cabrini Health, Malvern, VIC, Australia
  5. ANZGOG, NSW, Australia

Advanced endometrial cancer (EC) management is challenging and primarily reliant on tissue biopsy. Circulating tumour DNA (ctDNA) analysis has emerged as a liquid biopsy approach offering minimally invasive and real-time assessment of tumour molecular alterations to guide cancer management. Hypermethylation at the promotor of the tumour suppressor gene MLH1 results in mismatch repair deficiency (dMMR) in EC. Characterising MLH1 methylation holds potential as a predictive biomarker for response to immunotherapy, however, historically in EC, this has only been possible by tumour tissue analysis which is not routinely performed in clinical practice.

We aimed to develop a non-invasive ctDNA assay to detect MLH1 promoter methylation and investigate genome-wide methylation profiles to predict immunotherapy response in EC. We developed a novel assay that combines methylation-sensitive restriction enzyme (MSRE) digestion coupled with ddPCR to evaluate ctDNA MLH1 promoter methylation levels. In addition, we used a new sequencing technology (BioModal duet +modC) to validate our results by capturing tumour whole genome methylation patterns.

Plasma for assay screening was obtained from 20 healthy donors and 65 participants in the PHAEDRA clinical trial, a phase 2 study assessing the activity of durvalumab, anti PD-L1 therapy, in advanced EC. To date, using our novel ctDNA assay, hypermethylated MLH1 has been detected in 8 patients with known tumour tissue hypermethylation and one additional patient who may have acquired methylation during prior chemotherapy. Nine patients with absent tissue methylation and four healthy donors tested negative for methylation of the MLH1 promoter, confirming high specificity of our approach. The remaining cohort is currently undergoing analysis, alongside genome-wide methylation assessments to allow identification of global methylation patterns predictive of immunotherapy response.

Our research demonstrates the feasibility of a non-invasive ctDNA approach for rapidly identifying MLH1 promoter methylation status in advanced EC. This assay holds significant translational potential to guide treatment decisions in this disease, where accessing tumour tissue can be challenging.