Presentation Authors: Claudio Murta*, Sao Paulo, Brazil, José Pontes Jr, Tatiane Furuya, Miyuki Uno, Alexis Carrasco, Rafael Coelho, Giuliano Guglielmetti, Maurício Cordeiro, Sheila Faraj, Katia Leite, Laura Sichero, Luisa Villa, Miguel Srougi, Roger Chammas, William Nahas, São Paulo, Brazil
Introduction: Penile cancer (PeC) is a rare disease with high morbidity and mortality. Few studies have investigated miRNAs and their role in regulating gene networks and signaling pathways in PeC. Therefore, we aimed to identify a molecular signature based on miRNA and gene expression profiling related to carcinogenesis and prognosis in order to describe new biomarkers in PeC.
Methods: We prospectively collected fresh samples of primary tumors (TT) and adjacent non-neoplastic tissue (NT) from 11 patients who underwent surgical treatment. 5 patients had localized disease (non-metastatic group) and 6 had inguinal lymph node metastases (metastatic group). RNA was purified and microarray analysis was performed using miRNA 4.0 Genechip (Affymetrix). We identified the differentially expressed miRNAs (DEmiRs) comparing TT of metastatic to non-metastatic patients and the DEmiRs comparing TT to NT of the 11 patients using TAC Software. The DEmiRs expression were validated in an expanded sample of 24 patients by qRT-PCR using TaqMan assays. The DEmiRs predicted targets of both comparisons were investigated using miRTarBase, mirWalk and MetaCore softwares. We evaluated these target transcripts expression using a high-throughput nanofluidic qRT-PCR platform. We correlated the DEmiR and mRNA expression to the classic prognostic factors and cancer specific survival.
Results: The mean follow-up time was 16.3 months. Half of patients developed metastasis to inguinal nodes and 7 have died of the disease. We identified and validated 8 DEmiRs (miR31-5p, miR31-3p, miR224-5p, miR200a-5p, miR432-5p, miR487b-3p, miR30a-5p, miR149-5p) and 35 differentially expressed genes (DEGs) when comparing TT vs NT. We validated the DEmiRs miR421 and miR744-5p and 7 DEGs (CCND1, EGFR, ENTPD5, HOXA10, IGF1R, MYC, SNAI2) at the metastatic vs non-metastatic comparison. The miR31-3p down expression was associated with metastasis (p=0.043). No miRNA was able to predict mortality, although miR421 showed a marginal association (p=0.07). Overexpression of MMP1 was associated with higher size (p=0.042), grade (p=0.045), T stage (p=0.018), perineural invasion (p=0.007), and metastasis (p=0.018). MMP1 is regulated by miR145-5p which is down-regulated in TT (FC=28). The overexpression of CCND1 (p=0.006) and EGFR (p=0.001) was related to higher grade; and MMP9 overexpression with microvascular invasion (p=0.044). The CCND1, EGFR, MYC and SNAI2 expressions were significantly associated with worst results at specific survival curves. We also observed that SNAI2 expression was associated with cancer-specific survival HR 9.93; 95% CI, 1.153-85.542; p=0.037).
Conclusions: We described a miRNA and mRNA expression profile associated to carcinogenesis and worst prognosis of penile cancer. We postulated that miR31-5p, MMP1, CCND1, EGFR, MYC, MMP9 and SNAI2 expressions are potential biomarkers for prognosis in PeC.
Source of Funding: FAPESP 2015/17097-2