Abstract
Purpose :
Tissue biopsy of Rb can cause tumor spread, so it is contraindicated. We demonstrated that aqueous humor (AH), an ocular fluid, is a high-yield liquid biopsy source enabling in vivo detection of tumor-derived cell-free DNA (cfDNA), thus overcoming the contraindication to biopsy. In ~13% of Rb patients, tumor progression is driven by epigenetic deregulation of tumor-promoting pathways without detectable genomic alterations. However, epigenetic studies have been done only on primary tumors from surgically removed eyes. The frequency and effect of epigenomic regulation in eyes that have been saved with therapy are unclear due to a complete lack of access to in vivo tumor tissue. Therefore, epigenetic analysis of AH cfDNA is highly desired to understand the broader spectrum of Rb tumorigenesis and prognosis.
Methods :
We included 16 AH samples and 4 Rb tumors from 12 patients with 14 Rb eyes in the study. We conducted global DNA methylation profiling of tumor tissues from surgically removed Rb eyes and AH samples collected from different clinical stages and treatment outcomes using the Illumina Infinium EPIC DNA methylation BeadArray platform. Publicly-available DNA methylation data of normal retina, Rb eyes, and Rb patients were obtained from Gene Expression Omnibus (GEO, GSE57362) for cell-type DNA methylation comparisons.
Results :
Our preliminary studies revealed a high degree of concordance in genome-wide differential DNA methylation patterns between paired AH and tumor samples. Integrating our data with large public datasets, we identified reliable Rb DNA methylation signatures in AH cfDNA that have potential diagnostic and prognostic applications. We also identified DNA hypermethylation of the RB1 promoter, which may serve as an efficacious target of DNA methylation inhibitors. By integrating DNA methylation data with gene expression data, we identified over 300 differentially expressed genes potentially directly regulated by DNA methylation in Rb tumorigenesis.
Conclusions :
Our findings set the stage for exploring epigenetic markers using AH cfDNA specimens, including identifying potential prognostic markers and therapeutic targets to ultimately improve the clinical management of Rb patients.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.