AGAP2-AS1 as a potential marker for development of distant metastases in surgically treated low-risk clear cell renal cell carcinoma.

Abstract

732 Background: Historically, 30 % of patients with localized kidney cancer develop distant metastases during follow-up. There is an urgent need to improve the individual risk assessment for clear cell renal cell carcinoma (ccRCC) patients. We therefore aim to characterize the gene expression profile of low-risk patients both with and without progressive disease to define predictive outcome candidate markers. Methods: Formalin-fixed tissue blocks from ccRCC patients (n=24, eight progressors and 16 non-progressors) with a low Leibovich score were collected. Patients had a mean age of 65 years (5 females and 19 males). The non-progressors were matched 2:1 to the progressors for gender, age, pT tumor stage, size, Fuhrman grade, and eGFR. Total RNA was extracted(miRNeasy FFPE Kit, Qiagen) and sequenced (TruSeq RNA Access Library Kit, Illumina). RNA-seq results were analyzed by ingenuity pathway analysis, K Nearest Neighbors algorithm, and survival analysis. Results: 1167 differentially expressed genes (abs.FC≥2, p≤0.05) were detected. Progressors overexpressed genes related to cancer, B-cell infiltration and other immune-system related pathways. Principal component analyses and hierarchical clustering depicted a systematic transcriptomic difference between progressors and non-progressors. Combinations of up to 10 genes were evaluated as classifiers. The AGAP2-AS1 mRNA classified 23 out of 24 samples correctly, without the need for a larger gene panel. The trend of expression was confirmed with RT-PCR.The correlation between sample status as either progressor or non-progressor and AGAP2-AS1 level was R2 =0.69, p <0.01. Patients were split into groups based on AGAP2-AS1 expression (cut-off log2cpm>1), where higher expression correlated with shorter survival; Wilcoxon (p<0.0001),Log-rank test (p<0.0001), Hazard ratio; 9.24E-11. Immunohistochemistry of AGAP2, USP10 and KI-67 confirmed results from the mRNA level. Conclusions: RNA-seq results show a transcriptomic difference between low-risk ccRCC progressors and low-risk non-progressors. AGAP2-AS1 may serve as a potential classifier for the identification of low-risk progressors.