Abstract 204: Reconstructing testicular germ cell cancer progression: Primary tumor heterogeneity and early metastatic clone selection
Authors: Dorssers Lambert C. , Gillis Ad J. , Stoop Hans , Marion Ronald Van , Nieboer Marleen M. , Riet Job Van , Werken Harmen J. Van de , Oosterhuis J Wolter , Ridder Jeroen de , Looijenga Leendert H. 
Published: August 2018

Abstract
Background. Testicular germ cell cancer (TGCC) is initiated from a primordial germ cell during early life and the most frequent malignancy in young Caucasian males. Although treatment of TGCC is generally curative, rare cases of therapy resistance occur. Little information is available on the presence of heterogeneity of the complex primary tumor and derived metastases after systemic treatment. The goals of this study are to determine the intratumor heterogeneity, and to unravel tumor progression from initiation till therapy-resistant metastases.
Methods. In this study, we have investigated 42 purified samples of four cases of nonseminoma with intrinsic resistance to chemotherapy including different histological elements (embryonal carcinoma, yolk sac tumor and teratoma), metastatic specimens and precursor lesions (germ cell neoplasia in situ, GCNIS) using whole genome-, targeted-, and RNA sequencing, as well as methylation profiling. Enriched tumor samples were prepared using laser-assisted micro-dissection (PALM) or macro dissection. Sequencing data were used to reconstruct the evolutionary history of these rare cases.
Results. A low frequency of somatic mutations (~0.1 per Mb) was found in these TGCC cases with a BRCA-like mutational signature without evidence for direct involvement of BRCA1 and BRCA2 genes. Intratumor molecular heterogeneity was observed and did not correspond to the supposed histological evolution of the primary tumor. Metastases after systemic treatment were derived from precursors frequently not identified in the primary cancer. GCNIS mostly lacked the molecular marks of the primary TGCC (including gain of chromosome 12p) and comprised dominant subclones that had failed to progress into a manifest malignancy. The low read frequencies of somatic variants in the enriched samples indicated early genome duplication.
Conclusions. Our data strongly support the hypothesis that TGCC is initiated by whole genome duplication, followed by copy number alterations, dynamic acquisition of chromosome 12p overrepresentation, and accumulation of low numbers of somatic mutations resembling a BRCA-like mutational signature. Moreover, metastatic clones may originate early in tumor development and follow an independent path of genetic evolution. The observations of heterogeneity at all stages of tumorigenesis should be considered when treating patients with clinically overt TGCC, or with GCNIS-only disease.
Citation Format: Lambert C. Dorssers, Ad J. Gillis, Hans Stoop, Ronald Van Marion, Marleen M. Nieboer, Job Van Riet, Harmen J. Van de Werken, J Wolter Oosterhuis, Jeroen de Ridder, Leendert H. Looijenga. Reconstructing testicular germ cell cancer progression: Primary tumor heterogeneity and early metastatic clone selection [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 204.