Abstract 5553: Diagnostic aqueous humor proteome predicts metastatic potential in uveal melanoma
Authors: Peng Chen-Ching , Xu Liya , Berry Jesse 
Published: June 2023

Abstract
Introduction: Approximately half of patients with uveal melanoma (UM), a primary eye cancer, will develop metastatic disease. Gene expression profiling (GEP) is clinically validated to stratify risk of metastasis by assigning UM patients to two highly prognostic molecular classes: class 1 (low metastatic risk) and class 2 (high metastatic risk). However, GEP requires intraocular tumor biopsy which are limited by small tumor size and tumor heterogeneity; furthermore, there are small risks of retinal hemorrhage, bleeding or tumor dissemination. Thus, liquid biopsy has emerged as a significantly less invasive alternative. Blood biopsy has largely been unsuccessful for clinical use in UM due to low detection rates in the setting disease limited to the eye, however eye-specific aqueous humor (AH) liquid biopsy may be a better alternative. In this study, we seek to determine the AH proteome related to the advanced GEP class 2 using the diagnostic AH specimens.
Method: Twenty UM treatment naive AH were collected before plaque brachytherapy. Patients were sub-grouped by GEP classes into GEP 1 (n=12) and GEP 2 (n=5). Three patients were classified as GEP unknown (GEP NA, n=3) due to the unavailability of tumor biopsy. Ten microliters of AH samples were analyzed by proximity extension assay-derived multiplexed Olink platform. Protein expression levels of 1472 targets were analyzed, compared between GEP classes and correlated with clinical features. Significant differentially expressed proteins (DEPs, fold-change (FC) &gt; 2 or FC &lt; 0.5, P &lt; 0.01) were subjected to Qiagen ingenuity pathway analysis for cellular pathway and upstream regulator identification.
Results: GEP 2 class was correlated with AJCC stages (P = 0.012), advanced clinical tumor stages (P = 0.007) and mutated BAP1 (P = 0.018). 45 DEPs were identified when comparing GEP classes. Among them, 31 are up-regulated DEPs [fold-change (FC) &gt;2, P &lt; 0.01] and 14 are down-regulated DEPs (FC &lt; 0.5, P &lt; 0.01) in GEP 2 compared to GEP1. The unsupervised clustering analysis showed that the 45 DEPs well-differentiate AH samples by GEP classes, and the 3 GEP NA samples were clustered with GEP1 class. Pathway analysis showed that 45 DEPs contribute to metastatic-related pathways including cellular movement, cellular proliferation and epithelial to mesenchymal transition. Two upstream regulators, IL1 receptor and SPRY2, were predicted to regulate 8 out of the 45 DEPs. IL1R2 (FC = 3.4, P = 0.021) and SPRY2 (FC = 0.6, P = 0.052) protein expression levels were found matched as prediction in our Olink dataset.
Conclusions: 45 AH DEPs could differentiate GEP class 1 and 2 at the diagnostic stage and could be detected even when the tumor was too small to biopsy. IL1R and SPRY2 are potential upstream regulators for the 8/45 DEPs that contribute to metastasis-related pathways. AH liquid biopsy offers a new opportunity to determine metastatic potential for patients in the absence of tumor biopsy.
Citation Format: Chen-Ching Peng, Liya Xu, Jesse Berry. Diagnostic aqueous humor proteome predicts metastatic potential in uveal melanoma. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5553.