Aqueous Humor as a Liquid Biopsy for Uveal Melanoma

Abstract

Purpose : Metastatic Uveal Melanoma (UM) is usually fatal within one year of symptom onset. There are a variety of biomarkers that can stratify patients’ metastatic risk, but identifying them requires intraocular tumor biopsy. Liquid biopsy offers a less invasive alterative without the risks of retinal detachment and intraocular hemorrhage. We investigated whether the aqueous humor (AH) has sufficient tumor-derived cell-free DNA (cfDNA) to function as a liquid biopsy for UM.

Methods : This case series includes 7 eyes of 7 UM patients from whom 12 AH samples were taken during routine brachytherapy plaque placement and/or removal. Double stranded DNA (dsDNA), single stranded DNA (ssDNA), RNA, and microRNA (miRNA) were quantified with respective Qubit High Sensitivity Assay Kits (Thermo Fischer) for each sample. CfDNA was then isolated and sequenced on an Illumina platform to assess genome-wide somatic copy number alterations (SCNAs).

Results : AH derived cfDNA was detected in all samples (dsDNA mean = 0.7 ng/µL, range 0.01-7.5 ng/µL; ssDNA mean = 2.0 ng/µL, range 0-21.3 ng/µL). RNA was detected in 2 of 12 samples (mean = 1.6 ng/µL, range = 0.7-2.5 ng/µL), and miRNA was detected in 11 of 12 samples (mean = 1.7 ng/µL, range = 0.094-16.2 ng/µL). There was a significantly higher concentration of all nucleic acids (dsDNA, ssDNA, RNA, and miRNA) in post-radiation samples than pre-radiation samples (median = 0.085 ng/µL in pre-radiation, 0.333 ng/µL in post-radiation, W = 1, z = -3.46, p = 0.00027, Wilcoxon Signed-rank test). Confirmed UM genomic alterations of 6p gain, 6q loss, and 8q gain were identified in one post-radiation AH sample from an eye with an iridociliary body tumor. These SCNAs are characteristic of UM. No SCNAs were found in the AH from the pre-radiation sample in the same eye, or any of the other AH samples sequenced to date.

Conclusions : The AH is a source of cfDNA in eyes with UM, with a higher yield of all nucleic acids detected after brachytherapy radiation treatment. UM associated SCNAs were identified in the AH of one eye with an iridociliary tumor after radiation only; we hypothesize this may be due to intratumoral necrosis from the radiation as well as the proximity of the primary tumor to the AH. These results represent the first time that UM SCNAs have been identified in cfDNA isolated from the AH, and this provides preliminary evidence that the AH can serve as a liquid biopsy for UM.

This is a 2021 ARVO Annual Meeting abstract.