Integrated Proteogenomic Characterization across Major Histological Types of Pediatric Brain Cancer.

We report a comprehensive proteogenomics analysis, including whole-genome sequencing, RNA sequencing, and proteomics and phosphoproteomics profiling, of 218 tumors across 7 histological types of childhood brain cancer: low-grade glioma (n = 93), ependymoma (32), high-grade glioma (25), medulloblastoma (22), ganglioglioma (18), craniopharyngioma (16), and atypical teratoid rhabdoid tumor (12). Proteomics data identify common biological themes that span histological boundaries, suggesting that treatments used for one histological type may be applied effectively to other tumors sharing similar proteomics features. Immune landscape characterization reveals diverse tumor microenvironments across and within diagnoses. Proteomics data further reveal functional effects of somatic mutations and copy number variations (CNVs) not evident in transcriptomics data. Kinase-substrate association and co-expression network analysis identify important biological mechanisms of tumorigenesis. This is the first large-scale proteogenomics analysis across traditional histological boundaries to uncover foundational pediatric brain tumor biology and inform rational treatment selection.

MitoScape: A big-data, machine-learning platform for obtaining mitochondrial DNA from next-generation sequencing data.

The growing number of next-generation sequencing (NGS) data presents a unique opportunity to study the combined impact of mitochondrial and nuclear-encoded genetic variation in complex disease. Mitochondrial DNA variants and in particular, heteroplasmic variants, are critical for determining human disease severity. While there are approaches for obtaining mitochondrial DNA variants from NGS data, these software do not account for the unique characteristics of mitochondrial genetics and can be inaccurate even for homoplasmic variants. We introduce MitoScape, a novel, big-data, software for extracting mitochondrial DNA sequences from NGS. MitoScape adopts a novel departure from other algorithms by using machine learning to model the unique characteristics of mitochondrial genetics. We also employ a novel approach of using rho-zero (mitochondrial DNA-depleted) data to model nuclear-encoded mitochondrial sequences. We showed that MitoScape produces accurate heteroplasmy estimates using gold-standard mitochondrial DNA data. We provide a comprehensive comparison of the most common tools for obtaining mtDNA variants from NGS and showed that MitoScape had superior performance to compared tools in every statistically category we compared, including false positives and false negatives. By applying MitoScape to common disease examples, we illustrate how MitoScape facilitates important heteroplasmy-disease association discoveries by expanding upon a reported association between hypertrophic cardiomyopathy and mitochondrial haplogroup T in men (adjusted p-value = 0.003). The improved accuracy of mitochondrial DNA variants produced by MitoScape will be instrumental in diagnosing disease in the context of personalized medicine and clinical diagnostics.

A proteogenomic surfaceome study identifies DLK1 as an immunotherapeutic target in neuroblastoma.

Cancer immunotherapies have produced remarkable results in B-cell malignancies; however, optimal cell surface targets for many solid cancers remain elusive. Here, we present an integrative proteomic, transcriptomic, and epigenomic analysis of tumor specimens along with normal tissues to identify biologically relevant cell surface proteins that can serve as immunotherapeutic targets for neuroblastoma, an often-fatal childhood cancer of the developing nervous system. We apply this approach to human-derived cell lines (N=9) and cell/patient-derived xenograft (N=12) models of neuroblastoma. Plasma membrane-enriched mass spectrometry identified 1,461 cell surface proteins in cell lines and 1,401 in xenograft models, respectively. Additional proteogenomic analyses revealed 60 high-confidence candidate immunotherapeutic targets and we prioritized Delta-like canonical notch ligand 1 (DLK1) for further study. High expression of DLK1 directly correlated with the presence of a super-enhancer spanning the DLK1 locus. Robust cell surface expression of DLK1 was validated by immunofluorescence, flow cytometry, and immunohistochemistry. Short hairpin RNA mediated silencing of DLK1 in neuroblastoma cells resulted in increased cellular differentiation. ADCT-701, a DLK1-targeting antibody-drug conjugate (ADC), showed potent and specific cytotoxicity in DLK1-expressing neuroblastoma xenograft models. Moreover, DLK1 is highly expressed in several adult cancer types, including adrenocortical carcinoma (ACC), pheochromocytoma/paraganglioma (PCPG), hepatoblastoma, and small cell lung cancer (SCLC), suggesting potential clinical benefit beyond neuroblastoma. Taken together, our study demonstrates the utility of comprehensive cancer surfaceome characterization and credentials DLK1 as an immunotherapeutic target. Highlights: Plasma membrane enriched proteomics defines surfaceome of neuroblastomaMulti-omic data integration prioritizes DLK1 as a candidate immunotherapeutic target in neuroblastoma and other cancersDLK1 expression is driven by a super-enhancer DLK1 silencing in neuroblastoma cells results in cellular differentiation ADCT-701, a DLK1-targeting antibody-drug conjugate, shows potent and specific cytotoxicity in DLK1-expressing neuroblastoma preclinical models.

OpenPBTA: The Open Pediatric Brain Tumor Atlas.

Pediatric brain and spinal cancers are collectively the leading disease-related cause of death in children; thus, we urgently need curative therapeutic strategies for these tumors. To accelerate such discoveries, the Children's Brain Tumor Network (CBTN) and Pacific Pediatric Neuro-Oncology Consortium (PNOC) created a systematic process for tumor biobanking, model generation, and sequencing with immediate access to harmonized data. We leverage these data to establish OpenPBTA, an open collaborative project with over 40 scalable analysis modules that genomically characterize 1,074 pediatric brain tumors. Transcriptomic classification reveals universal TP53 dysregulation in mismatch repair-deficient hypermutant high-grade gliomas and TP53 loss as a significant marker for poor overall survival in ependymomas and H3 K28-mutant diffuse midline gliomas. Already being actively applied to other pediatric cancers and PNOC molecular tumor board decision-making, OpenPBTA is an invaluable resource to the pediatric oncology community.

Inducible Sbds deletion impairs bone marrow niche capacity to engraft donor bone marrow after transplantation.

Bone marrow (BM) niche-derived signals are critical for facilitating engraftment after hematopoietic stem cell (HSC) transplantation (HSCT). HSCT is required for restoration of hematopoiesis in patients with inherited BM failure syndromes (iBMFSs). Shwachman-Diamond syndrome (SDS) is a rare iBMFS associated with mutations in SBDS. Previous studies have demonstrated that SBDS deficiency in osteolineage niche cells causes BM dysfunction that promotes leukemia development. However, it is unknown whether BM niche defects caused by SBDS deficiency also impair efficient engraftment of healthy donor HSC after HSCT, a hypothesis that could explain morbidity noted after clinical HSCT for patients with SDS. Here, we report a mouse model with inducible Sbds deletion in hematopoietic and osteolineage cells. Primary and secondary BM transplantation (BMT) studies demonstrated that SBDS deficiency within BM niches caused poor donor hematopoietic recovery and specifically poor HSC engraftment after myeloablative BMT. We have also identified multiple molecular and cellular defects within niche populations that are driven by SBDS deficiency and are accentuated by or develop specifically after myeloablative conditioning. These abnormalities include altered frequencies of multiple niche cell subsets, including mesenchymal lineage cells, macrophages, and endothelial cells; disruption of growth factor signaling, chemokine pathway activation, and adhesion molecule expression; and p53 pathway activation and signals involved in cell cycle arrest. Taken together, this study demonstrates that SBDS deficiency profoundly impacts recipient hematopoietic niche function in the setting of HSCT, suggesting that novel therapeutic strategies targeting host niches could improve clinical HSCT outcomes for patients with SDS.