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Rhabdoid tumors are rare and aggressive tumors that usually arise in very young children. They are characterized by a bi-allelic inactivation of the SMARCB1 gene. Although the majority of alterations of SMARCB1 are acquired in tumors, a heterozygous germline alteration is seen in one third of patients and defines the rhabdoid tumors predisposition syndrome. Penetrance is almost complete and the vast majority of germline alterations of SMARCB1 are acquired de novo but rare familial cases with a healthy carrier have also been described. Since the advent of more sensitive molecular analysis technologies such as next-generation sequencing (NGS), the number of mosaicisms of genes involved in genetic diseases discovered from blood samples has increased considerably. The aim of our study was to explore the mosaicisms of SMARCB1 in the blood 1/ of children with rhabdoid tumors with at least one alteration of SMARCB1 previously identified in the tumor but not found in the blood with old-fashioned low-sensitivity technologies and 2/ in parents of children with heterozygous germline alteration of SMARCB1. We analyzed a custom NGS panel which covers the SMARCB1 gene with an average depth of 1.500X on blood samples of 111 children with rhabdoid tumors and 32 parents collected at the Institut Curie since 1999. The mosaicism rate found in index cases was 11.7% (13/111) and 3.1% (1/32) in parents. The variant allele frequency vary from 0.8% to 12.9%. Our results also indicate to be cautious about the possible confounding effect of circulating tumor DNA. This hitherto underestimated SMARCB1 mosaicism rate should motivate an optimization of the genetic counseling as well as the oncological monitoring of these children and thus have a significant medical impact given the catastrophic prognosis of rhabdoid tumors.

Pediatric chordomas are rare malignant neoplasms, and few data are available for optimizing therapeutic strategies and outcome. This study aimed at evaluating how best to manage them and to identify prognostic factors. This multicentric retrospective study included 40 children diagnosed with chordomas between 1966 and 2012. Clinical, radiological, and histopathological data, treatment modalities, and outcomes were reviewed. The median age was 12 years old. Most chordomas were histologically classical forms (45.5%) and were mostly located at the skull base (72.5%). The overall survival (OS) was 66.6% and 58.6%, and progression-free survival (PFS) was 55.7% and 52% at 5 and 10 years, respectively. Total resection was correlated with a better outcome (p = 0.04 for OS and PFS, log-rank). A histopathological/immunohistochemical grading system recently crafted for adults was applied. In a multivariate analysis, it significantly correlated with outcome (PFS and OS, p = 0.004), and the loss of BAF47 immunoexpression appeared to be a significant independent prognostic factor (PFS, p = 0.033). We also identified clinical and histopathological parameters that correlated with prognosis. A new grading system combined with the quality of surgical resection could help classify patients to postpone radiotherapy in case of low risk. Targeted therapy and reirradiation at recurrence may be considered as potential therapeutic strategies.

Extra-cranial rhabdoid tumors (RT) are highly aggressive malignancies of infancy, characterized by undifferentiated histological features and loss of SMARCB1 expression. The diagnosis is all the more challenging that other poorly differentiated cancers lose SMARCB1 expression, such as epithelioid sarcomas (ES), renal medullary carcinomas (RMC) or undifferentiated chordomas (UC). Moreover, late cases occurring in adults are now increasingly reported, raising the question of differential diagnoses and emphasizing nosological issues. To address this issue, we have analyzed the expression profiles of a training set of 32 SMARCB1-deficient tumors (SDT), with ascertained diagnosis of RT (n = 16, all < 5 years of age), ES (n = 8, all > 10 years of age), UC (n = 3) and RMC (n = 5). As compared with other SDT, RT are characterized by an embryonic signature, and up-regulation of key-actors of de novo DNA methylation processes. Using this signature, we then analysed the expression profiling of 37 SDT to infer the appropriate diagnosis. Thirteen adult onset tumors showed strong similarity with pediatric RT, in spite of older age; by exome sequencing, these tumors also showed genomic features indistinguishable from pediatric RT. In contrary, 8 tumors were reclassified within carcinoma, ES or UC categories, while the remaining could not be related to any of those entities. Our results demonstrate that embryonic signature is shared by all RT, whatever the age at diagnosis; they also illustrate that many adult-onset SDT of ambiguous histological diagnosis are clearly different from RT. Finally, our study paves the way for the routine use of expression-based signatures to give accurate diagnosis of SDT.

The poor prognosis of rhabdoid tumors (RT) justifies the search for new therapies. By a high‐throughput drug screening and subsequent in vitro confirmation of selected drugs, we identified as good SMARCB1‐dependent therapeutic candidates, broad inhibitors of tyrosine kinase receptors (RTK) such as pazopanib, and the potassium channel inhibitor clofilium tosylate (CfT). Their combination significantly induced RT apoptotic cell death in vitro. A human phospho‐kinase antibody array revealed PDGFRα/β and FGFR2 as pazopanib targets, while these genes were the most highly expressed RTK in our primary tumor data set. Of note, combined genetic inhibition of at least two RTK was only partially able to recapitulate the effect of pazopanib, emphasizing the requirement of broad inhibition. We then showed that pazopanib treatment reduced tumor growth in vivo while its combination with CfT could at least recapitulate the efficiency of conventional chemotherapy. Altogether, these results may influence future clinical trials for RT.

Summary: Rhabdoid tumors (RTs) are aggressive tumors of early childhood characterized by SMARCB1 inactivation. Their poor prognosis highlights an urgent need to develop new therapies. Here, we performed a high-throughput screening of approved drugs and identified broad inhibitors of tyrosine kinase receptors (RTKs), including pazopanib, and the potassium channel inhibitor clofilium tosylate (CfT), as SMARCB1-dependent candidates. Pazopanib targets were identified as PDGFRα/β and FGFR2, which were the most highly expressed RTKs in a set of primary tumors. Combined genetic inhibition of both these RTKs only partially recapitulated the effect of pazopanib, emphasizing the requirement for broad inhibition. CfT perturbed protein metabolism and endoplasmic reticulum stress and, in combination with pazopanib, induced apoptosis of RT cells in vitro. In vivo, reduction of tumor growth by pazopanib was enhanced in combination with CfT, matching the efficiency of conventional chemotherapy. These results strongly support testing pazopanib/CfT combination therapy in future clinical trials for RTs. : Rhabdoid tumors (RTs) are aggressive pediatric tumors characterized by SMARCB1 inactivation. Chauvin et al. identify two SMARCB1-dependent targeted therapies for RT: pazopanib, which inhibits PDGFR and FGFR2, and the potassium channel inhibitor clofilium tosylate, which induces endoplasmic reticulum stress. Combining both drugs induces cell apoptosis and reduces PDX tumor growth. Keywords: rhabdoid tumors, SMARCB1, pazopanib, clofilium tosylate, high-throughput drug screening, tyrosine kinase inhibitors