Endometrial Cancer (EC) is the most common gynecologic malignancy globally. The Levonorgestrel Intrauterine System (LNG-IUS) is an alternative treatment for atypical hyperplasia (AEH) and early-stage EC for women who are unable to undergo surgery. Thirty to 50% of women do not respond to this treatment, making prediction of Levonorgestrel (LNG) resistance a critical obstacle to overcome. This study aims to identify predictive biomarkers of resistance in Extracellular Vesicles (EVs) to guide LNG-IUS treatment of early-stage EC and AEH.
Differentially expressed genes (DEGs) and HUB proteins were previously identified in two LNG resistant and sensitive EC cell lines using whole transcriptome sequencing. EVs were isolated from conditioned culture media (CCM) using size exclusion chromatography with qEV10/70nm columns. RNA was extracted from CCM EVs, libraries were prepared, small RNA sequencing was carried out and differentially abundant miRNAs were identified. Patients with early-stage EC or AEH treated with the LNG-IUS were recruited and plasma samples were obtained with informed consent. EVs were extracted using qEVoriginal/70nm columns and characterised by tunable resistive pulse sensing, TEM and Western Blot. Candidate miRNA and protein markers were analysed in plasma EVs via Western Blot and RT-qPCR.
The mean concentration and size of plasma EVs was 3.299 particles/mL and 181 ± 17nm, respectively. Previously identified proteins, ALDHA1A, MAOA, KLF4 were not present in plasma EVs. CD133 was present in plasma EVs. miR-504-3p was identified as a potential marker of LNG resistance.
This research investigated potential novel markers of resistance to the LNG-IUS in patient plasma EV samples.