CLONAL EVOLUTION IN HIGH RISK PEDIATRIC CANCERS

Orateur : Gudrun SCHLEIERMACHER (Institut Curie, Paris)

Although 80% of all pediatric patients with cancer can be cured based on modern multimodal therapeutic strategies, at least 20% of these patients will undergo tumor progression or relapse. Understanding the molecular mechanisms underlying tumour progression and resistance to treatment is crucial to the development of new treatment approaches. In high risk pediatric cancers, somatic DNA alterations such as genomic amplifications, copy number alterations, translocations or mutations play an important role as molecular diagnostic , prognostic and predictive biomarkers, which might further define targeted treatment approaches. However it is now known that in many high risk pediatric cancers, genetic alterations in a given tumor are not static, but undergo modifications, with clonal evolution most likely playing a major role in high risk tumor progression. The study of clonal evolution has been rendered feasible based on innovative technologies of liquid biopsies, as circulating tumour DNA, a fraction of cell free DNA can be readily isolated from plasma obtainted from sequential blood samples. In order to study clonal evolution in neuroblastoma (NB), we performed whole exome sequencing on primary tumours and on sequential ctDNA samples from 19 NB patients using a modified library contruction and capture approach, enabling a satisfactory depth of coverage in all cases. SNVs and structural variations were called using a standardized bioinformatics pipeline. At diagnosis, a majority of observed SNVs were common to the primary NB and corresponding diagnostic ctDNA of a given patient (mean number of SNVs : 19 ; range 9-69). At diagnosis, few SNVs specific to the NB (mean : 6 ; range 0-18) or specific to ctDNA (mean:22 ; range 9-69) were observed, suggesting spatial heterogeneity with different ctDNA amounts released by different clones. ctDNA samples obtained during disease progression harboured a higher numbers of SNVs, with additional relapse-specific SNVs (mean : 22 ; range 0-55) targeting, amongst others, the protein kinase A signaling pathway. Significant ctDNA quantities extracted from plasma and enabling whole exome sequening were also observed in the majority of other high risk pediatric cancers, such as alveolar rhabdomyosarcoma, Ewing tumour, other high grade sarcomas,as documented in our NGSkids protocol (ClinicalTrials.gov Identifier : NCT02546453). In non-metastatic brain tumours, ctDNA isolated from CSF resulted in a higher sensitivity and specificity of detection to tumor cell specific genetic alterations. A French national program MICCHADO (PI Gudrun Schleiermacher I Curie ; co PI Claudia PAsqualini, IGR) now aims at studying clonal evolution based on sequential ctDNA analysis from the time of diagnosis in all high risk pediatric cancers. Altogether the presence of tumor genetic abnormalities in ctDNA obtainted either from plasma or CSF can be documented in most patients with high risk pediatric cancer. Tumor- or ctDNA specific genetic alterations frequently suggest spatial and temporal heterogeneity. Sequential studies will further elucidate mechanisms of clonal evolution, tumor progression and therapy resistance. Thus, sequential studies of ctDNA should now be integrated into the development and optimization of targeted treatment strategies.

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