Introduction

Clear cell renal cell carcinoma (CCC), the most lethal urologic tumor (250 000 cases, 110 000 deaths worldwide/year), is the main subtype of kidney cancer. It is characterized in 75% of cases by the loss of the von Hippel-Lindau (VHL) tumor suppressor gene leading to HIF? stabilization. Prostate cancer (PCa) is the most common cancer in men > 50 yo (1 100 000 cases, 300 000 deaths worldwide/year). CCC and PCa are resistant to therapies. DNPH1 (2'-deoxynucleoside 5'-phosphate N-hydrolase 1), a c-Myc target overexpressed in various cancers, regulates cell growth and angiogenesis and may thus behave as a new oncogene. Through alternative splicing, 4 isoforms can be generated with still unknown precise function. Our objective was to define the role of DNPH1 in these cancers.

Methods

We used a panel of cell lines i.e 786-0, A498 (VHL -) and Caki-1, Caki-2, ACHN (VHL +) for CCC and LNCaP, VCap, PC3, DU145, 22RV1 for PCa. Tumor/normal corresponding tissues pairs from 42 and 41 CCC and PCa patients, respectively, were also used. DNPH1 isoforms expression was measured by RTqPCR and Western blot. Since no specific chemical inhibitors are available, the role of DNPH1 isoforms in vitro and in vivo was evaluated using siRNAs and expressing vectors (wild-type or inactive, dead active site). In vivo, we used the tumor xenografted nude mice model to assess the role and the underlying mechanisms of DNPH1 in tumor growth.

Results

We show that CCC and PCa express only isoforms 1 and 2. These are 174 and 148 aminoacids long, respectively, and are identical till aminoacid 126. DNPH1 expression was deregulated in 71% of CCC cases but upregulated in 75% of PCa cases, regardless of the stage. Both isoforms behave similarly but isoform 1 represented 80% of total DNPH1 expression. In CCC cell lines, the transfection with wild-type DNPH1 expressing vectors decreased cell growth up to 60% by stimulating cell proliferation and inhibiting apoptosis, while the transfection with the inactive vector had no effect. In PCa cells, siRNAs targeting DNPH1 isoforms 1 and 2 decreased cell growth dose-dependently by up to 50% through inhibition of cell proliferation and induction of apoptosis. We are currently evaluating the effect of both isoforms in nude mice xenografted with CCC cell lines transfected with the DNPH1 expressing vector and in PCa-bearing nude mice treated with in vivo siRNAs. First results corroborate in vitro observations.

Conclusions

Our results show the opposite tumor suppressor/oncogene properties of DNPH1 in CCC and PCa and should allow to design new therapeutic options for these refractory diseases.

Funding

INSERM, University of Strasbourg, Fondation pour la Recherche Médicale