Traditional gene expression deconvolution methods assess a limited number of cell types, therefore do not capture the full complexity of the tumor microenvironment (TME). Here, we integrate nine deconvolution tools to assess 79 TME cell types in 10,592 tumors across 33 different cancer types, creating the most comprehensive analysis of the TME. In total, we found 41 patterns of immune infiltration and stroma profiles, identifying heterogeneous yet unique TME portraits for each cancer and several new findings. Our findings indicate that leukocytes play a major role in distinguishing various tumor types, and that a shared immune-rich TME cluster predicts better survival in bladder cancer for luminal and basal squamous subtypes, as well as in melanoma for RAS-hotspot subtypes. Our detailed deconvolution and mutational correlation analyses uncover 35 therapeutic target and candidate response biomarkers hypotheses (including CASP8 and RAS pathway genes).

Table S4 includes immune signature and iCluster results.

Table S7 contains microbe screening results.

Table S1 contains cohort description, Master Patient Table and MutSigCV results.

Table S2 contains BAP1 analysis results, as well as detailed lists of YY1 and IRF8 target genes.

Table S6 contains results from the analysis of DNA methylation in SETD2 mutated and BAP1 inactivated samples.

Supplementary materials and methods, as well as supplementary figures S1-S7.

Table S3 contains the karyotypes of 16 genome-wide LOH MPM cases from the BWH cohort.

Table S5 contains detailed miRs and lncRNAs results.

Background Tumor molecular profiling from patients experiencing exceptional responses to systemic therapy may provide insights into cancer biology and improve treatment tailoring. This pilot study evaluates the feasibility of identifying exceptional responders retrospectively, obtaining pre-exceptional response treatment tumor tissues, and analyzing them with state-of-the-art molecular analysis tools to identify potential molecular explanations for responses. Methods Exceptional response was defined as partial (PR) or complete (CR) response to a systemic treatment with population PR or CR rate less than 10% or an unusually long response (eg, duration >3 times published median). Cases proposed by patients' clinicians were reviewed by clinical and translational experts. Tumor and normal tissue (if possible) were profiled with whole exome sequencing and, if possible, targeted deep sequencing, RNA sequencing, methylation arrays, and immunohistochemistry. Potential germline mutations were tracked for relevance to disease. Results Cases reflected a variety of tumors and standard and investigational treatments. Of 520 cases, 476 (91.5%) were accepted for further review, and 222 of 476 (46.6%) proposed cases met requirements as exceptional responders. Clinical data were obtained from 168 of 222 cases (75.7%). Tumor was provided from 130 of 168 cases (77.4%). Of 117 of the 130 (90.0%) cases with sufficient nucleic acids, 109 (93.2%) were successfully analyzed; 6 patients had potentially actionable germline mutations. Conclusion Exceptional responses occur with standard and investigational treatment. Retrospective identification of exceptional responders, accessioning, and sequencing of pretreatment archived tissue is feasible. Data from molecular analyses of tumors, particularly when combining results from patients who received similar treatments, may elucidate molecular bases for exceptional responses.

A small fraction of cancer patients with advanced disease survive significantly longer than patients with clinically comparable tumors. Molecular mechanisms for exceptional responses to therapy have been identified by genomic analysis of tumor biopsies from individual patients. Here, we analyzed tumor biopsies from an unbiased cohort of 111 exceptional responder patients using multiple platforms to profile genetic and epigenetic aberrations as well as the tumor microenvironment. Integrative analysis uncovered plausible mechanisms for the therapeutic response in nearly a quarter of the patients. The mechanisms were assigned to four broad categories-DNA damage response, intracellular signaling, immune engagement, and genetic alterations characteristic of favorable prognosis-with many tumors falling into multiple categories. These analyses revealed synthetic lethal relationships that may be exploited therapeutically and rare genetic lesions that favor therapeutic success, while also providing a wealth of testable hypotheses regarding oncogenic mechanisms that may influence the response to cancer therapy.

We present a systematic analysis of the effects of synchronizing a large-scale, deeply characterized, multi-omic dataset to the current human reference genome, using updated software, pipelines, and annotations. For each of 5 molecular data platforms in The Cancer Genome Atlas (TCGA)—mRNA and miRNA expression, single nucleotide variants, DNA methylation and copy number alterations—comprehensive sample, gene, and probe-level studies were performed, towards quantifying the degree of similarity between the ‘legacy’ GRCh37 (hg19) TCGA data and its GRCh38 (hg38) version as ‘harmonized’ by the Genomic Data Commons. We offer gene lists to elucidate differences that remained after controlling for confounders, and strategies to mitigate their impact on biological interpretation. Our results demonstrate that the hg19 and hg38 TCGA datasets are very highly concordant, promote informed use of either legacy or harmonized omics data, and provide a rubric that encourages similar comparisons as new data emerge and reference data evolve.

Sarcomas are a broad family of mesenchymal malignancies exhibiting remarkable histologic diversity. We describe the multi-platform molecular landscape of 206 adult soft tissue sarcomas representing 6 major types. Along with novel insights into the biology of individual sarcoma types, we report three overarching findings: (1) unlike most epithelial malignancies, these sarcomas (excepting synovial sarcoma) are characterized predominantly by copy-number changes, with low mutational loads and only a few genes (TP53, ATRX, RB1) highly recurrently mutated across sarcoma types; (2) within sarcoma types, genomic and regulomic diversity of driver pathways defines molecular subtypes associated with patient outcome; and (3) the immune microenvironment, inferred from DNA methylation and mRNA profiles, associates with outcome and may inform clinical trials of immune checkpoint inhibitors. Overall, this large-scale analysis reveals previously unappreciated sarcoma-type-specific changes in copy number, methylation, RNA, and protein, providing insights into refining sarcoma therapy and relationships to other cancer types.

Abstract Introduction: Preliminary data show that up to 10% of patients on early clinical trials have complete (CR) or partial (PR) responses. Exceptional responses (ER) are also observed in certain clinical cases using standard chemotherapy. The ERI Pilot study is evaluating the molecular alterations found in tumors from cancer patients who respond to a systemic treatment (standard or investigational) that is ineffective in at least 90% of patients. The study is available through the Cancer Trials Support Unit (www.ctsu.org). Experimental Procedures Cases of patients who achieve a CR or PR > 6 months are proposed by email at NCIExceptionalResponders@mail.nih.gov. Once provisionally approved, the proposer sends tissue to a central repository, where the tissue is evaluated for percent tumor &necrosis; nucleic acids are isolated & sent for whole exome, mRNA & deep targeted sequencing. Tissue may be frozen or formalin fixed paraffin embedded but must be at least an entire core biopsy. Data will be correlated with mechanism of action of the treatment given. The ERI pilot study will characterize > 100 ERs. If sufficient tissue is available, whole genome sequencing &other molecular analysis may be done. We anticipate that some specimens will not be of sufficient quality/quantity; thus many more tumors will need to be submitted to obtain these results. Six months after the genomic data are generated they will be released to a publicly available, controlled access database to facilitate research by qualified investigators. The ERI study accepts all types of malignant disease in adults or children if adequate tissue obtained prior to the ER exists. The ER could have been to an approved or investigational, targeted, or non-targeted systemic treatment. The feasibility of retrospectively collecting tumor tissue & clinical data for genomic analysis is a goal of the ERI study. Results: The ER study was activated on September 24, 2014. As of December 2, > 80 cases have been proposed, 46 are provisionally approved & 26 cases were not ERs or were withdrawn. Accepted Cases: solid & hematologic tumors (2 brain, 4 breast, 2 colon, 6 gastric, 1 hepatocellular, 7 head/neck, 2 renal, 2 lung cancer, 1 melanoma, 2 ovarian, 6 pancreatic, 2 prostate and 4 sarcomas; 2 acute leukemia, 2 lymphoma, 1 myeloma). Approximately 75% involved standard & 25% involved targeted drugs or combinations. Tissue analysis is ongoing & results will be presented. Conclusions: Collection of archived tissue from ERs for genomic evaluation is feasible. Efficient ER case identification occurs through expert review and consensus-based case selection. Citation Format: Barbara A. Conley, S Percy Ivy, James V. Tricoli, Jean-Claude Zenklusen, Roy Tarnuzzer, Irina Lubensky, Naoko Takebe, Paul M. Williams, JoAnne Zujewski, Richard Little, Jeffrey White, Elise Kohn, Shakun Malik, Ben Kim, Erin Souhan, Lou Staudt. The NCI exceptional responders initiatives: Initial feasibility result. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 612. doi:10.1158/1538-7445.AM2015-612

BACKGROUND Diffuse low-grade and intermediate-grade gliomas (which together make up the lower-grade gliomas, World Health Organization grades II and III) have highly variable clinical behavior that is not adequately predicted on the basis of histologic class. Some are indolent; others quickly progress to glioblastoma. The uncertainty is compounded by interobserver variability in histologic diagnosis. Mutations in IDH, TP53, and ATRX and codeletion of chromosome arms 1p and 19q (1p/19q codeletion) have been implicated as clinically relevant markers of lower-grade gliomas. METHODS We performed genomewide analyses of 293 lower-grade gliomas from adults, incorporating exome sequence, DNA copy number, DNA methylation, messenger RNA expression, microRNA expression, and targeted protein expression. These data were integrated and tested for correlation with clinical outcomes. RESULTS Unsupervised clustering of mutations and data from RNA, DNA-copy-number, and DNA-methylation platforms uncovered concordant classification of three robust, nonoverlapping, prognostically significant subtypes of lower-grade glioma that were captured more accurately by IDH, 1p/19q, and TP53 status than by histologic class. Patients who had lower-grade gliomas with an IDH mutation and 1p/19q codeletion had the most favorable clinical outcomes. Their gliomas harbored mutations in CIC, FUBP1, NOTCH1, and the TERT promoter. Nearly all lower-grade gliomas with IDH mutations and no 1p/19q codeletion had mutations in TP53 (94%) and ATRX inactivation (86%). The large majority of lower-grade gliomas without an IDH mutation had genomic aberrations and clinical behavior strikingly similar to those found in primary glioblastoma. CONCLUSIONS The integration of genomewide data from multiple platforms delineated three molecular classes of lower-grade gliomas that were more concordant with IDH, 1p/19q, and TP53 status than with histologic class. Lower-grade gliomas with an IDH mutation either had 1p/19q codeletion or carried a TP53 mutation. Most lower-grade gliomas without an IDH mutation were molecularly and clinically similar to glioblastoma. (Funded by the National Institutes of Health.).

BACKGROUNDPapillary renal-cell carcinoma, which accounts for 15 to 20% of renal-cell carcinomas, is a heterogeneous disease that consists of various types of renal cancer, including tumors with indolent, multifocal presentation and solitary tumors with an aggressive, highly lethal phenotype. Little is known about the genetic basis of sporadic papillary renal-cell carcinoma, and no effective forms of therapy for advanced disease exist.METHODSWe performed comprehensive molecular characterization of 161 primary papillary renal-cell carcinomas, using whole-exome sequencing, copy-number analysis, messenger RNA and microRNA sequencing, DNA-methylation analysis, and proteomic analysis.RESULTSType 1 and type 2 papillary renal-cell carcinomas were shown to be different types of renal cancer characterized by specific genetic alterations, with type 2 further classified into three individual subgroups on the basis of molecular differences associated with patient survival. Type 1 tumors were associated with MET alterations, whereas type 2 tumors were characterized by CDKN2A silencing, SETD2 mutations, TFE3 fusions, and increased expression of the NRF2-antioxidant response element (ARE) pathway. A CpG island methylator phenotype (CIMP) was observed in a distinct subgroup of type 2 papillary renal-cell carcinomas that was characterized by poor survival and mutation of the gene encoding fumarate hydratase (FH).CONCLUSIONSType 1 and type 2 papillary renal-cell carcinomas were shown to be clinically and biologically distinct. Alterations in the MET pathway were associated with type 1, and activation of the NRF2-ARE pathway was associated with type 2; CDKN2A loss and CIMP in type 2 conveyed a poor prognosis. Furthermore, type 2 papillary renal-cell carcinoma consisted of at least three subtypes based on molecular and phenotypic features. (Funded by the National Institutes of Health.).

Papillary thyroid carcinoma (PTC) is the most common type of thyroid cancer. Here, we describe the genomic landscape of 496 PTCs. We observed a low frequency of somatic alterations (relative to other carcinomas) and extended the set of known PTC driver alterations to include EIF1AX, PPM1D, and CHEK2 and diverse gene fusions. These discoveries reduced the fraction of PTC cases with unknown oncogenic driver from 25% to 3.5%. Combined analyses of genomic variants, gene expression, and methylation demonstrated that different driver groups lead to different pathologies with distinct signaling and differentiation characteristics. Similarly, we identified distinct molecular subgroups of BRAF-mutant tumors, and multidimensional analyses highlighted a potential involvement of onco-miRs in less-differentiated subgroups. Our results propose a reclassification of thyroid cancers into molecular subtypes that better reflect their underlying signaling and differentiation properties, which has the potential to improve their pathological classification and better inform the management of the disease.

PURPOSEWe hypothesize that remodeling of the scleral extracellular matrix, involving collagen and proteoglycan synthesis and turnover, is a key process involved in ocular growth. Decreased axial elongation is observed following neonatal removal of the crystalline lens in a rhesus monkey model of congenital cataract. We wanted to determine changes in gene expression in the operated and companion eye following lensectomy, especially for extracellular matrix in the sclera.METHODSBetween 4 and 7 days of age, infant monkeys underwent surgical removal of the lens from the right eye. Axial lengths of the operated and unmanipulated fellow eyes were measured and when interocular differences of >0.4 mm were achieved, monkeys were sacrificed and RNA was isolated from sclera. In order to determine changes in scleral gene expression in aphakic versus control eyes, we used Clontech's Atlas Gene Array (Human Cancer Array version 1.2) hybridized with total RNA from three monkeys.RESULTSAtlas Gene Array analysis demonstrated differential expression of several genes in the operated versus the unmanipulated eye. Most notably, there was a statistically significant increase in expression of several extracellular matrix (ECM) genes including: aggrecan, decorin, biglycan, several collagens, and tenascin in the RNA from the sclera of the aphakic eyes when compared to the unmanipulated eyes. Genes for several matrix metalloproteinases (MMPs) showed no significant change following lens removal although there was a trend towards decreased expression. There were also statistically significant changes in the pattern of gene expression in the operated eye relative to the unmanipulated eye for cell adhesion, cell cycle, apoptosis, and cytoskeleton transcripts.CONCLUSIONSOur results suggest that removal of the crystalline lens alters gene expression in the sclera with a prominent upregulation of ECM transcripts. These data support recent evidence that remodeling of the ECM composition of the sclera may be an important regulator of ocular growth.

The ability of engineered cerium oxide nanoparticles to confer radioprotection was examined. Human normal and tumor cells were treated with nanoceria and irradiated, and cell survival was measured. Treatment of normal cells conferred almost 99% protection from radiation-induced cell death, whereas the same concentration showed almost no protection of tumor cells. For the first time, nanoceria is shown to confer radioprotection to a normal human breast line but not to a human breast tumor line, MCF-7.

cDNA rat stress microarrays were used to test the general hypothesis that atypical gene expression patterns exist in the brains of Hydrocephalic-Texas (H-Tx) compared to normal Sprague-Dawley (SD) rats on embryonic day 18. Sixty-two percent of the 216 target transcripts were detected in at least 2 of 3 replicates, with maximum mean fold change (MFC) ratios (H-Tx:SD) in Bcl-2-related ovarian killer protein (BOK, 3.07) and peroxisome proliferator-activated receptor-alpha (PPAR-alpha, 0.04). Five (3.73%) of the 134 detected transcripts were elevated and 20 (17.2%) were suppressed more than twofold in H-Tx. MFC ratios for stress response, cytoskeleton-motility, and intracellular transducer-effector-modulator functional classifications were elevated, while MFC ratios for transcription and apoptosis groups were suppressed in H-Tx. K-means clustering revealed several patterns of gene expression with potential biological relevance in apoptosis, intracellular transducer-effector-modulator, metabolism, cell cycle, and stress response transcripts. Multiplex RT-PCR methodology, used to corroborate the cDNA data, captured four distinct temporal expression patterns on embryonic days 16-20 (E16-E20) for HSP27, DnaJ2, HSP47, HSP60, HSP70, HIP, HSP90A, and HSP90beta. The discovery of unique chaperone/heat shock expression profiles in the embryonic brains of H-Tx and SD rats is a powerful step towards the development of novel mechanistic hypotheses in the study of hydrocephalus disorders. This is the first study to associate early stress responses with the differential expression of chaperones/heat shock protein-related genes using the H-Tx model of congenital hydrocephalus.