Orthoxenografts of Testicular Germ Cell Tumors Demonstrate Genomic Changes Associated with Cisplatin Resistance and Identify PDMP as a Resensitizing Agent

Abstract

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. .

Testicular cancer: Determinants of cisplatin sensitivity and novel therapeutic opportunities

“…So far, only 14 orthotopically established TC PDX models and 14 subcutaneous TC PDX models have been reported [92,111,112], with limited data available of their establishment [113]. One study described the establishment of 14 non-seminoma PDX models, specifically from the CC, EC and YSC subtypes, as well as mixed tumors with YSC, teratoma and EC components [113]. In this study, orthotopic implantation of the tumor pieces was more successful than subcutaneous implantation.…”

“…Patient-derived xenograft models are increasingly used in current cancer research due to several advantages over cell line-based xenografts, including maintenance of tumor heterogeneity, high similarity to human tumors [107,108] and increased predictive power of drug response [109,110]. So far, only 14 orthotopically established TC PDX models and 14 subcutaneous TC PDX models have been reported [92,111,112], with limited data available of their establishment [113]. One study described the establishment of 14 non-seminoma PDX models, specifically from the CC, EC and YSC subtypes, as well as mixed tumors with YSC, teratoma and EC components [113].…”

“…In this study, orthotopic implantation of the tumor pieces was more successful than subcutaneous implantation. These PDX models were used to study cisplatin resistance and to test novel combinatorial strategies using a glucosylceramide synthase (GCS) inhibitor, DL-threo-PDMP, and a multi-targeted receptor tyrosine kinase inhibitor, sunitinib, in combination with cisplatin [113,114]. Recently, we established and characterized three subcutaneous TC PDX models.…”

Molecular Biology of Pediatric and Adult Male Germ Cell Tumors

“…Other strategies to investigate the mechanisms associated with the resistance acquired by tumor cells to cisplatin include the use of preclinical models, in vitro and in vivo, obtained from the cultivation and exposure of TGCT cell lines to incremental doses of the drug, for long periods of time [ 101 ], in addition to the use of animal models that reproduce the phenotypic properties of the human tumor [ 102 ]. Although in vitro cell culture systems have been used extensively for decades, they represent oversimplified models, which are characterized by the absence of heterogeneity and lack of microenvironment components [ 103 ].…”

Cisplatin Resistance in Testicular Germ Cell Tumors: Current Challenges from Various Perspectives

“…Importantly, dimethyl sulfoxide (DMSO), commonly used as a drug vehicle, was demonstrated to induce cisplatin resistance and differentiation in embryonal carcinoma cells, which should be taken into consideration when designing biological studies [ 39 ]. Moreover, in vivo mouse models of resistant disease can also be used by implanting cisplatin-resistant human tumor specimens or, more commonly, by injecting the resistant clones of (T)GCT cell lines available [ 40 , 41 , 42 ].…”