Abstract 554: Aqueous humor as a liquid biopsy for uveal melanoma
Authors: Kim Mary E. , Xu Liya , Peng Chen-Ching , Kuhn Peter , Hicks James , Berry Jesse L. 
Published: July 2021

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 12 AH samples from 7 eyes of 7 UM patients from whom AH was sampled 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. The lower limit of detection was 0.01 ng/µL for dsDNA, 0.05 ng/µL for ssDNA, 0.25 ng/µL for RNA, and 0.05 ng/µL for miRNA. CfDNA was then isolated and sequenced on an Illumina platform to assess genome-wide somatic copy number alterations (SCNAs).

Results: Aqueous humor derived cfDNA was detected in all samples, ranging from 0.085-28.8 ng/µL (mean = 2.7 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 cfDNA concentration in post-radiation samples than pre-radiation samples, (median = 0.082 ng/µL in pre-radiation, 0.21 ng/µL in post-radiation, W = 1, z = -2.55, p &lt; 0.05, Wilcoxon Signed-rank test). This finding was similar when comparing all nucleic acid concentrations (dsDNA, ssDNA, RNA, and miRNA) between pre- and post-radiation (median = 0.085 ng/µL in pre-radiation, 0.333 ng/µL in post-radiation, W = 1, z = -3.46, p = 0.00027). 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 in the case where SCNAs were identified. 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.

Citation Format: Mary E. Kim, Liya Xu, Chen-Ching Peng, Peter Kuhn, James Hicks, Jesse L. Berry. Aqueous humor as a liquid biopsy for uveal melanoma [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 554.