Purpose: We examined whether PI3K-AKT or extracellular signal-regulated kinase (ERK) signaling pathways could play a role in the development of cisplatin (CDDP) resistance in testicular germ cell tumor (TGT) cells.Experimental Design: We compared AKT and ERK activation levels in CDDP-sensitive testicular tumor cells and in their corresponding CDDP-resistant-derived cells. We also analyzed these pathways in orthotopic testicular tumors and human patient samples.Results: Our results indicated that there was overactivation of AKT in CDDP-resistant cells compared with sensitive cells, but no effect on activated ERK levels. We observed an increase in mRNA and protein levels for platelet-derived growth factor (PDGF) receptor b and PDGF-B ligand. These were responsible for AKT overactivation in CDDP-resistant cells. When PDGFRb levels were decreased by short hairpin RNA (shRNA) treatment or its activation was blocked by pazopanib, CDDP-resistant cells behaved like sensitive cells. Moreover, CDDP-resistant cells were more sensitive to incubation with PDGFRb inhibitors such as pazopanib or sunitinib than sensitive cells, a finding consistent with these cells being dependent on this signaling pathway. We also found overexpression of PDGFRb and pAKT in CDDP-resistant choriocarcinoma orthotopic tumor versus their CDDP-sensitive counterparts. Finally, we found high PDGFRb levels in human testicular tumors, and overexpression in CDDP-resistant testicular choriocarcinomas compared with the CDDP-sensitive and nontreated tumors.Conclusions: The PDGFRb-AKT pathway plays a critical role in the development of CDDP resistance in testicular tumoral cells. Clin Cancer Res; 20(3); 658-67. Ó2013 AACR.

To investigate the genetic basis of cisplatin resistance as efficacy of cisplatin-based chemotherapy in the treatment of distinct malignancies is often hampered by intrinsic or acquired drug resistance of tumor cells. We produced 14 orthoxenograft transplanting human nonseminomatous testicular germ cell tumors (TGCT) in mice, keeping the primary tumor features in terms of genotype, phenotype, and sensitivity to cisplatin. Chromosomal and genetic alterations were evaluated in matched cisplatin-sensitive and their counterpart orthoxenografts that developed resistance to cisplatin in nude mice. Comparative genomic hybridization analyses of four matched orthoxenografts identified recurrent chromosomal rearrangements across cisplatin-resistant tumors in three of them, showing gains at 9q32-q33.1 region. We found a clinical correlation between the presence of 9q32-q33.1 gains in cisplatin-refractory patients and poorer overall survival (OS) in metastatic germ cell tumors. We studied the expression profile of the 60 genes located at that genomic region. and were the only two genes deregulated in resistant tumors harboring the 9q32-q33.1 gain. Moreover, other four genes ( and ) were deregulated in all five resistant tumors independently of the 9q32-q33.1 amplification. RT-PCRs in tumors and functional analyses in () indicate that the influence of 9q32-q33.1 genes in cisplatin resistance can be driven by either up- or downregulation. We focused on glucosylceramide synthase (GCS) to demonstrate that the GCS inhibitor DL-threo-PDMP resensitizes cisplatin-resistant germline-derived orthoxenografts to cisplatin. Orthoxenografts can be used preclinically not only to test the efficiency of drugs but also to identify prognosis markers and gene alterations acting as drivers of the acquired cisplatin resistance. .

BackgroundCisplatin (CDDP) resistance in testicular germ cell tumors (GCTs) is still a clinical challenge, and one associated with poor prognosis. The purpose of this work was to test pazopanib, an anti-tumoral and anti-angiogenic multikinase inhibitor, and its combination with lapatinib (an anti-ErbB inhibitor) in mouse orthotopic models of human testicular GCTs.MethodsWe used two different models of human testicular GCTs orthotopically grown in nude mice; a CDDP-sensitive choriocarcinoma (TGT38) and a new orthotopic model generated from a metastatic GCT refractory to first-line CDDP chemotherapy (TGT44). Nude mice implanted with these orthotopic tumors were treated with the inhibitors and the effect on tumoral growth and angiogenesis was evaluated.ResultsTGT44 refractory tumor had an immunohistochemical profile similar to the original metastasis, with characteristics of yolk sac tumor. TGT44 did not respond when treated with cisplatin. In contrast, pazopanib had an anti-angiogenic effect and anti-tumor efficacy in this model. Pazopanib in combination with lapatinib in TGT38, an orthotopic model of choriocarcinoma had an additive effect blocking tumor growth.ConclusionsWe present pazopanib as a possible agent for the alternative treatment of CDDP-sensitive and CDDP-refractory GCT patients, alone or in combination with anti-ErbB therapies.