Investigating extracellular vesicles as predictive biomarkers of LNG-IUD treatment response in early endometrial cancer
Endometrial cancer (EC) is the most common gynaecological cancer in Aotearoa, with rates rising rapidly in premenopausal women. There is an increasing number of women who are not able to undergo the standard of care which is hysterectomy (removal of the uterus). The levonorgestrel intrauterine device (LNG-IUD) is an alternative treatment that allows women to retain their uterus. However, around half of women will not respond to this treatment and there is no way of predicting response. Predictive biomarkers will enable safer and more equitable access to this treatment. Extracellular vesicles (EVs) are particles found in bodily fluids enriched with miRNAs and are a novel opportunity to identify blood-based biomarkers of LNG-IUD treatment response. Two cell line culture models of LNG resistance were used to investigate the expression of miRNA biomarkers of EC within EVs. EVs were isolated from cell culture media, and their size, concentration and morphology characterised using tunable resistive pulse sensing (TRPS), transmission electron microscopy (TEM) and Western blotting. RT-qPCR was carried out on RNA extracted from EVs, followed by pathway enrichment analysis. miRNA expression was validated in EVs from EC patient plasma samples from the UOW gynaecological cancer biobank. EV size and concentration was similar between resistant and sensitive cells as found by TRPS. TEM showed isolated particles had the characteristic EV double membrane. Protein analysis through Western blotting (TSG101, CD9 and ApoB) demonstrated EV purity. Many candidate miRNA biomarkers were expressed at very low levels in culture media EVs, and high biological variation was observed. There were no statistically significant differences in expression of any of the miRNAs investigated in media EVs, although miR-93, miR-133a and miR-205 were close to significance (p=0.071, p=0.088, p=0.070 respectively). Pathway analysis indicated these miRNAs may be providing resistant cells with greater oncogenic potential. The miRNAs were expressed at higher levels in EC patient plasma EVs than the media EVs. This was the first study to investigate EV biomarkers of LNG-IUD treatment resistance. miRNAs currently thought to be involved in EC were expressed in low levels in culture media EVs. miR-93, miR-133a and miR-205 warrant further research. This study also acted as a proof of concept that miRNAs previously investigated in EC can be detected within patient plasma EVs. This study has laid the foundations for further EC EV research and has provided valuable preliminary investigations into EC and LNG resistance EV miRNA biomarkers.
Advisor: Henry, Claire; Danielson, Kirsty
Degree Name: Bachelor of Biomedical Sciences with Honours
Degree Discipline: Department of Obstetrics, Gynaecology and Women's Health
Publisher: University of Otago
Keywords: Endometrial cancer; extracellular vesicles; LNG-IUD; biomarkers; EV; miRNA; LNG resistance
Research Type: Thesis