Identifying Molecular Predictors of Response for Treatment of Early Stage Endometrial Cancer with the Mirena®

Endometrial Cancer (EC) is the most common gynaecologic malignancy in the developed world, and in Aotearoa New Zealand, incidence is increasing in premenopausal women. The Levonorgestrel Intrauterine System (LNG-IUS), also known as Mirena®, is gaining traction as an alternative treatment for hyperplasia and early stage EC for women wanting to retain their fertility, or who are unable to undergo surgery. However, 30 to 50% of women do not respond to treatment, and the mechanisms for resistance are not understood. The aim of this project was to investigate predictive biomarkers for Mirena® treatment of early stage endometrial cancer. Two levonorgestrel (LNG) resistant endometrial cancer cell lines (MFE296R and MFE319R) and cultures from three early stage endometrial cancer patients (GB#13, GB#16, GB#23) were developed. A literature search was conducted to identify possible candidate biomarkers of LNG resistance. RT-qPCR was used to analyse the relative mRNA expression of 15 candidate biomarkers from MFE296R and MFE319R. mRNA expression of the top six differentially expressed genes was then measured in primary cultures. The behavioural profile of MFE296R and MFE319R were analysed using proliferation, adhesion, migration (wound healing and transwell) and invasion (spheroid) assays. Relative mRNA expression of CRISPLD1, KLF4, SATB2, SOX17, ANO1 and HE4 were significantly amplified in MFE296R and MFE319R cells compared to their LNG sensitive clones. Relative mRNA expression of SOX17, ANO1, SATB2, KLF4 and ER in the GB#13R cell line was increased compared to the GB#13S cells. In the GB#16R cell line, results show increased relative mRNA expression of ANO1, SATB2, KLF4, CRISPLD1 and HE4 compared to the GB#16S cells. Relative mRNA expression of SOX17, KLF4, ER, CRISPLD1 and HE4 in the GB#23R cell line was increased compared to the GB#23S cells. LNG resistance had no effect on proliferation however significantly increased transwell migration in MFE319R cells (p=0.03), decreased cellular invasion in MFE296R cells, and decreased cellular adhesion to collagen in MFE29R cells (p=0.012) and MFE319R cells (p=0.04). Relative expression of CRISPLD1, KLF4, SATB2, SOX17, ANO1 and HE4 were significantly upregulated in resistant immortalised cell lines. This study suggests that these genes may serve as predictive biomarkers for response to the Mirena® and could be used to guide the treatment paradigm.