Abstract 480: Aberrant promoter hypermethylation of HIC1 and SOX1 in circulating cell free DNA as a potential biomarker for noninvasive early detection of ovarian cancer
Authors: Singh Alka , Sachan Manisha 
Published: July 2021

Background: Aberrant CpG island hypermethylation is an early event during carcinogenesis and a significant mechanism of tumor suppressor gene silencing. Assessment of tumor specific methylation changes in tissue or body fluids might provide insights into disease mechanisms and discovery of biomarkers. Ovarian cancer remains the most lethal gynaecological malignancy. Non-specific symptoms at early stage, presentation of disease at advanced stage and lack of sufficiently sensitive screening modalities have rendered significant challenge to its prompt diagnosis. To date, no single biomarker is able to accurately detect early OC in either tissue or body fluid. Aberrant promoter hypermethylation in cell free tumor-specific DNA can be indicative of highly specific cancer signals.

Methods: Herein, we concurrently evaluated the promoter methylation of HIC1 and SOX1 gene in 120 ovarian tissue samples and validated in 70 matched serum cell free DNA of cancerous and non cancerous samples by multiplex MethyLight assay (a high throughput quantitative, TaqMan based real time PCR technique). Additionally, promoter methylation pattern of these genes were validated by clonal bisulfite sequencing. The ROC (Receiver-operator characteristic) curves were constructed to evaluate the diagnostic performances of both individual and combined gene panel.

Results: In singleplex assay, high methylation sensitivity and specificity was evident for SOX1 (78.8% and 80.0%, respectively; AUC= 0.88) and HIC1 (80.0% and 77.14%, respectively; AUC=0.85) in tissue samples. In the multiplex assay, when either or both gene promoter showed methylation, the highly sensitivity and specific discrimination of EOC patients from normal controls was observed for SOX1 + HIC1 (83.53% and 88.57%, respectively; AUC=0.89). We identified DNA methylation of HIC1 and SOX1 as best discriminator between cancer and non-neoplastic tissue. Methylated SOX1 and HIC1 occurred in 53.3% and 71.1% of OC serum samples when their promoter methylation was analyzed in singleplex assay (AUC=0.81 and 0.88, respectively). In cell free DNA, this novel panel achieved a sensitivity of 80.0% and a specificity of 96.0% with an area under the ROC curve of 0.93 for discriminating OC samples from normal control. The results of MethyLight were highly concordant with those of clonal bisulfite sequencing. Methylated Individual gene as well as in combined panel exhibited better discriminatory efficiencies to identify ovarian cancer at various stages of disease when analyzed in tissue and serum cell free DNA.

Conclusion: We report a QPCR based non-invasive epigenetic biomarker assay with high sensitivity and specificity for OC screening. Our findings also reveal the potential utility of methylation based detection of circulating cell free tumor DNA in clinical management of ovarian cancer.

Citation Format: Alka Singh, Manisha Sachan. Aberrant promoter hypermethylation of HIC1 and SOX1 in circulating cell free DNA as a potential biomarker for noninvasive early detection of ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 480.