Molekularno profiliranje karcinoma prijelaznog epitela gornjega dijela mokraćnoga sustava u nefropatiji aristolohične kiseline [Molecular profiling of the upper urinary tract transitional cell cancers associated with aristolochic acid nephropathy]

Karanović, Sandra (2015) Molekularno profiliranje karcinoma prijelaznog epitela gornjega dijela mokraćnoga sustava u nefropatiji aristolohične kiseline [Molecular profiling of the upper urinary tract transitional cell cancers associated with aristolochic acid nephropathy]. PhD thesis, Sveučilište u Zagrebu.

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Abstract

INTRODUCTION: Upper urinary tract urothelial cancers (UUC) are rare in general population but occur in almost 50% of patients with endemic nephropathy (EN)/aristolochic acid nephropathy (AAN). Aristolochic acid (AA) was proven as the etiological factor both for EN/AAN and UUC. The goal of this study was to establish patterns of gene expression and posttranscriptional gene regulation mechanisms involved in AA induced carcinogenesis. MATERIALS AND METHODS: Paired samples of tumors and adjacent normal urothelial tissues of 13 patients from Croatian and Bosnian endemic regions were analyzed. MiRNA profiling was performed by RT-qPCR, while mRNA expression profiling was done using microarray technology. Immunohistochemistry was performed on tissue microarrays. Integrative analysis of data was done using number of bioinformatical programs. RESULTS: A signature of 138 miRNAs (74 up, 64 down) and 5438 (2021 up, 3417 down) significantly modulated mRNAs were identified. Integrating miRNA and mRNA data, 1159 predicted, inversely correlated targets of the 45 upregulated miRNAs and 703 predicted, inversely correlated targets of the 44 downregulated miRNAs were identified. Top ranked differentially expressed miRNA included miR-200 family, miR- 17-92 cluster, let-7, miR-143/145 cluster, and miR-23b. Comprehensive biological interpretation and mining of the integrated miRNA:mRNA data set identified pathways of cell cycle, DNA damage response and DNA repair, bladder cancer, deregulated oncogenic and developmental signaling pathways and remodeling of the chromatin structure. Molecular profiling data were validated by means of immunohistochemistry. CONCLUSIONS: UUC tumorigenesis appears to involve primarily miRNA-mediated process of oncogenic activation of tumor growth and repression of tumor suppressor genes, and chromatin silencing and repression of pro-metastasis programs rendering the non-metastatic nature of these carcinomas, what is in line with phenotype of this specific type of tumor.

Abstract in Croatian

UVOD: Karcinomi prijelaznog epitela gornjeg dijela mokraćnog sustava (UUC) rijetki su u općoj populaciji, ali se javljaju u gotovo 50% bolesnika s endemskom nefropatijom (EN)/nefropatijom aristolohične kiseline (AAN). Aristolohična kiselina (AA) je potvrđena kao etiološki čmbenik za razvoj EN/AAN i UUC. Cilj ovog istraživanja jest odrediti obrasce genske ekspresije i mehanizme posttranskripcijske regulacije genske ekspresije uključene u karcinogenezu uzrokovanu s AA. MATERIJAL I METODE: Analizirani su parni uzorci tumora i priležećeg normalnog tkiva od 13 bolesnika porijeklom iz hrvatskih i bosanskih endemskih područja. Profiliranje miRNA učinjeno je pomoću RT-qPCR uređaja, dok je analiza genske ekspresije učinjena na genskim čipovima. Imunohistokemijsko bojenje je učinjeno na tkivnim microarrayima. Integrirana analiza podataka učinjena je koristeći više bioinformatičkih programa. REZULTATI: Identificirani su potpisi od 138 miRNA (74 povišene, 64 snižene) i 5438 (2021 povišene, 3417 snižene) značajno modulirane mRNA. Integrirajući mRNA i miRNA podatke, detektirano je 1159 inverzno koreliranih mRNA, ciljnih molekula od 45 povišenih miRNA te 703 inverzno korelirane mRNA, ciljne molekule od 44 snižene miRNA. Među najviše rangiranim dismoduliranim miRNA istaknule su se miR-200 obitelj, miR-17-92 skupina, let-7, miR-143/45 skupina i miR-23b. Opsežna biološka analiza integriranih mRNA i miRNA podataka identificirala je sljedeće značajne kategorije: stanični ciklus, odgovor na DNA oštećenje, popravak DNA, karcinom mokraćnog mjehura, deregulacija onkogena, razvojni signalni putevi, remodeliranje strukture kromatina. Podaci dobiveni molekularnim profiliranjem potvrđeni su imunohistokemijski. 132 ZAKLJUČCI: Tumorigeneza UUC uključuje procese posredovane djelovanjem miRNA: rast tumora posljedično aktivaciji onkogena i represiji tumor supresor gena, odnosno utišavanje kromatina i represiju prometastatskih programa uz posljedičnu ograničenu metastatsku sposobnost ovog tipa tumora, što je u skladu s fenotipom ovih tumora.

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