Mechanism of urothelial cancers associated with endemic nephropathy.

Objectives and study: Approximately 50% of endemic nephropathy (EN) patients develop upper urothelial tract cancers (UUC) at some point during the course of their disease. Aristolochic acid (AA) was proven as the etiological factor both for EN and UUC. The goal of his study was to establish patterns of gene expression and posttranscriptional gene regulation mechanisms involved in AA induced carcinogenesis. Methods: Paired samples of tumors and adjacent normal urothelial tissues of 10 patients from Croatian endemic region were analyzed. miRNA profiling was performed by high-capacity qPCR using Applied Biosystems megaplex RT primer pools for 754 human miRNAs and the microfluidisc TagMan Low Density Arrays, while miRNA profiling of the same RNAs was performed using Affymetrix HG U133 Plus 2.0 arrays. Results: A signature of 50 miRNAs differentially modulated (19 elevated and 31 reduced) and 4801 significantly modulated mRNAs (2005 elevated and 2796 reduced) were identified in tumor vs. unaffected tissue. Using R-scripting, miRNA and mRNA dana were integrated, identifying 643 predicted, inversely correlated targets oft he 19 upregulated miRNAs and 483 predicted, inversely correlated targets oft he 31 downregulated miRNAs. Cancer-related miRNAs were upregulated in UUC while their tumor suppressor targets (PDCD4, PTEN, TPM1, PTCH1) were reduced. Additionally, the anti- metastatic miR-200 family was elevated in tumors, concurrently with low levels of its targets, the ZEB1/2 repressors of E cadherin. Inhibition of invasive/migratory process was documanted by increased miRNAs targeting cellular movement, migration and invasion (FGF1, COLIA2, CTGF, IGF1, MMP2, PTEN, RHOB, THBSI and TGFBR2). Comprehensive biological interpretation and mining oft he integrated miRNA:mRNA dana set (Ingenuity Pathway Analysis) identified high-scoring pathways of renal necrosis, renal damage and failure, cancer regulated cell growth and proliferation, G2/M DNA damage checkpoint regulation and DNA repair, inflammatory disease and suppression of cellular movement and migration. Conclusion: UUC tumorigenesis appears to involve primarily miRNA- mediated process of repression of tumor suppressor genes, tumor growth, induction of inflammatory processes and repression of anti- invasion/metastasis programs rendering the non- metastatic nature of these carcinomas.