Genomic characterization and development of therapies for pediatric sarcoma

儿科肉瘤的基因组表征和疗法开发

• 批准号: 10262419
• 负责人: John Shern
• 金额: $ 74.38万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10262419
• 关键词: ATAC-seq; Adolescent; Affect; Alveolar; American Society of Clinical Oncology; Benign; Biochemical; Biological; Biological Assay; Biological Models; CLIA certified; CRISPR/Cas technology; CTNNB1 gene; Categories; Cell Cycle; Cells; Chemotherapy and/or radiation; Chemotherapy-Oncologic Procedure; Child; Childhood; Childhood Soft Tissue Sarcoma; Clinical; Clinical Data; Clinical Trials; Collaborations; Complex; Consensus; Data; Data Set; Development; Diagnosis; Disease; Embryonal Rhabdomyosarcoma; Enrollment; Epigenetic Process; Event; Evolution; Ewings sarcoma; FBXW7 gene; FGFR4 gene; FOXO1A gene; Foundations; Frequencies; Gene Dosage; Genes; Genetic; Genetic Markers; Genetic Transcription; Genomics; Germ-Line Mutation; Goals; Histologic; Histology; Informatics; KRAS2 gene; Knowledge; Lesion; Localized Disease; Malignant Neoplasms; Molecular Biology; Molecular Genetics; Morbidity - disease rate; Mutation; NF1 gene; Neurofibromatosis 1; Neurofibrosarcoma; Operative Surgical Procedures; Outcome; PIK3CA gene; Pathology; Pathway Analysis; Pathway interactions; Patients; Pediatric Oncology; Pediatric Oncology Group; Pilot Projects; Plexiform Neurofibroma; Polycomb; Pre-Clinical Model; Prognostic Marker; Publications; RAS genes; RNA; Recurrence; Relapse; Reproducibility; Resolution; Rhabdomyosarcoma; Risk; Risk stratification; SNP array; Sampling; Signal Transduction; Soft tissue sarcoma; Survival Rate; TP53 gene; Techniques; Toxic effect; Transcription Repressor; Treatment Protocols; United States; Visualization; WNT Signaling Pathway; Work; aggressive therapy; base; bioinformatics pipeline; childhood sarcoma; clinical decision-making; clinical risk; clinically significant; cohort; disorder subtype; exome; falls; functional genomics; gene discovery; high risk; high-throughput drug screening; improved; in vitro Model; interest; meetings; member; mortality; multimodality; neoplastic cell; novel therapeutics; patient population; patient stratification; patient subsets; prognostic; prognostic significance; prospective; response; sarcoma; therapeutic target; therapy development; transcription factor; transcriptome sequencing; transcriptomics; tumor; user-friendly; whole genome

1. Efforts in Rhabdomyosarcoma Rhabdomyosarcoma (RMS) is a myogenic cancer that is the most common soft tissue sarcoma of childhood. With the development of multimodal chemotherapy regimens, relapse-free survival rates have improved to 70-80% in patients with localized disease, albeit with significant toxicity. Unfortunately, despite aggressive therapy, the 5-year survival rate for patients with metastatic disease remains only 30%. Therapy assignment is currently based on clinicopathologic features and using these criteria, three distinct subgroups of patients can be identified (low, intermediate and high risk). However, many patients fall into the intermediate risk category (which accounts for about 50% of all patients) and have a heterogeneous clinical outcome. This suggests that some of these children could be treated with less aggressive therapy or alternatively should be considered to have more aggressive disease. In an effort to further characterize the genetic events underlying this tumor type, our group in collaboration with the Children's Oncology Group performed a large sequencing effort using a combination of whole-genome, whole-exome and whole-transcriptome sequencing along with high resolution SNP arrays to characterize the landscape of somatic alterations in 147 tumor/normal pairs. Our findings describe a heterogenous group of genetic events appears to drive RMS most notably the PAX 3/7-FOXO1 fusion in the alveolar subtype and mutation of multiple RAS pathway genes in fusion negative tumors including recurrent genetic lesions in 10 cancer consensus genes (NRAS, KRAS, HRAS, PIK3CA, BCOR, TP53, NF1, FGFR4, FBXW7, CTNNB1). While the majority of these mutations appear to be mutually exclusive, a subset of tumors appears to have coexisting lesions within the same tumor; perhaps indicating a biologically relevant progression in these tumors. Unfortunately, much of the clinical annotation for these cases was incomplete, severely limiting our ability to derive prognostic information from this data set. To overcome this, a more focused retrospective analysis is needed to determine the prognostic significance of the discovered mutations. In collaboration with the Children's Oncology Group, we have sequenced 675 clinically annotated rhabdomyosarcoma cases. In this project, we are using high-throughput capture based sequencing to determine the frequency with which the observed mutations occur and determine whether these mutations could be used as markers of prognosis or response to therapy. The specific objectives of the study are the following: 1. To validate our pilot study and accurately estimate the frequency of NRAS, KRAS, HRAS, PIK3CA, BCOR, TP53, NF1, FGFR4, FBXW7, CTNNB1 mutations in low, intermediate and high-risk embryonal rhabdomyosarcomas. 2. To estimate the frequency of NRAS, KRAS, HRAS, PIK3CA, BCOR, TP53, NF1, FGFR4, FBXW7, CTNNB1 mutations in a small cohort of alveolar histology tumors. 3. To determine the clinical significance of NRAS, KRAS, HRAS, PIK3CA, BCOR, TP53, NF1, FGFR4, FBXW7, CTNNB1 mutations in patients with central pathology confirmed ERMS cases. 4. To develop an accurate and reproducible assay for assessing mutations in NRAS, KRAS, HRAS, PIK3CA, BCOR, TP53, NF1, FGFR4, FBXW7, CTNNB1 that can be used in prospective clinical trials. We have validated that our assay can reliably detect mutations even at low frequency and that we can accurately detect gene copy number changes. In addition, we have developed a bioinformatics pipeline and visualization portal that enables a user-friendly interface with the generated data. Clinical outcome correlation analysis has been performed in concert with the Childrens Oncology Group and we have demonstrated the clinical impact of the presence or absence of each genetic lesion. In the current year, we have defined that mutations of MYOD1 and TP53 were found to be associated with clinical outcome in fusion negative Rhabdomyosarcoma and these results were presented at the American Society of Clinical Oncology Annual Meeting as well as the Childrens Oncology Group annual meeting and this data is submitted for publication. Current efforts are focused on using these results to enlighten prospective clinical trials. This includes implementation of a CLIA certified assay and informatics pipeline to centrally profile all Rhabdomyosarcoma patients enrolled on Childrens Oncology Group trials. In additional we performed additional sequencing of retrospective samples collected on intermediate and high risk clinical trials to provide foundational data for clinical trials currently under development that will use genetic markers for improved clinical risk stratification and clinical decision making. 2. Efforts in Malignant Peripheral Nerve Sheath Tumor (MPNST) The goal of this work is to provide further understanding of the genetic and transcriptomic mechanisms of the tumor evolution underlying the transformation of a Plexiform Neurofibroma to a Malignant Peripheral Nerve Sheath Tumor (MPNST). MPNSTs are a devastating risk that predominantly affects patients with a germline mutation in the gene NF1 (Neurofibromatosis Type 1). Leveraging the expertise and patient population within the Pediatric Oncology Branch, in this project we are using preclinical model systems to dissect the genetic and epigenetic changes that occur as NF1 tumors transition from benign precursors to the aggressive MPNST. Specifically, we are studying the effects of mutation or deletion in one of the polycomb complex (PRC2) members SUZ12 or EED as the final step in the transformation to MPNST. The PRC2 complex is a major transcriptional repressor within the cell and recurrent alterations of members of this complex have been discovered in genomic sequencing studies from patient samples. Within the current fiscal year, we used inducible in vitro model systems and performed ChIPseq, RNAseq and ATACseq to nominate genes whose expression is altered upon loss of these genes with the primary objective to identify genes downstream of the PRC2 loss that are potential therapeutic targets. Not surprisingly, there are hundreds of genes whose expression is altered upon reassembly of the PRC2 complex and includes groups of genes whose expression decreases and those whose expression increases. Pathway analysis of this group of genes demonstrated a marked enrichment for genes involved in PI3K signaling, WNT signaling and cell cycle. Interestingly, the transcriptional changes driven by induction of competent PRC2 imperfectly overlapped with gain of the H3K27me3 mark, whereby only a distinct subset of genes appeared to be directly regulated at the RNA level, by the gain of the repressive mark. Analysis of this interesting group of 145 genes discovered a remarkable enrichment of bivalent genes as the primary downstream targets of PRC2 whose expression changes in response to reintroduction of the competent SUZ12. Of interest are several lineage specific transcription factors that appear to be unique vulnerabilities in MPNST cells. We are currently further interrogating these genes for functional relevance to MPNST using biochemical, molecular biology and CRISPR technologies.