Genome-scale functional genomics identify genes preferentially essential for multiple myeloma cells compared to other neoplasias.

Clinical progress in multiple myeloma (MM), an incurable plasma cell (PC) neoplasia, has been driven by therapies that have limited applications beyond MM/PC neoplasias and do not target specific oncogenic mutations in MM. Instead, these agents target pathways critical for PC biology yet largely dispensable for malignant or normal cells of most other lineages. Here we systematically characterized the lineage-preferential molecular dependencies of MM through genome-scale clustered regularly interspaced short palindromic repeats (CRISPR) studies in 19 MM versus hundreds of non-MM lines and identified 116 genes whose disruption more significantly affects MM cell fitness compared with other malignancies. These genes, some known, others not previously linked to MM, encode transcription factors, chromatin modifiers, endoplasmic reticulum components, metabolic regulators or signaling molecules. Most of these genes are not among the top amplified, overexpressed or mutated in MM. Functional genomics approaches thus define new therapeutic targets in MM not readily identifiable by standard genomic, transcriptional or epigenetic profiling analyses.

Smaller p-values in genomics studies using distilled auxiliary information.

Medical research institutions have generated massive amounts of biological data by genetically profiling hundreds of cancer cell lines. In parallel, academic biology labs have conducted genetic screens on small numbers of cancer cell lines under custom experimental conditions. In order to share information between these two approaches to scientific discovery, this article proposes a "frequentist assisted by Bayes" (FAB) procedure for hypothesis testing that allows auxiliary information from massive genomics datasets to increase the power of hypothesis tests in specialized studies. The exchange of information takes place through a novel probability model for multimodal genomics data, which distills auxiliary information pertaining to cancer cell lines and genes across a wide variety of experimental contexts. If the relevance of the auxiliary information to a given study is high, then the resulting FAB tests can be more powerful than the corresponding classical tests. If the relevance is low, then the FAB tests yield as many discoveries as the classical tests. Simulations and practical investigations demonstrate that the FAB testing procedure can increase the number of effects discovered in genomics studies while still maintaining strict control of type I error and false discovery rate.

Reconstruction of Metagenome-Assembled Genomes from Aquaria.

Here, we report 11 metagenome-assembled genomes (MAGs) reconstructed from freshwater and saltwater aquaria, including representatives of Polynucleobacter, Anaerolinea, Roseobacter, Flavobacteriia, Octadecabacter, Mycobacterium, and Candidate Phyla Radiation (CPR) members. These MAGs can serve as a resource for aquatic research and elucidating the role of CPR taxa in the built environment.

Genome-scale screens identify factors regulating tumor cell responses to natural killer cells.

To systematically define molecular features in human tumor cells that determine their degree of sensitivity to human allogeneic natural killer (NK) cells, we quantified the NK cell responsiveness of hundreds of molecularly annotated 'DNA-barcoded' solid tumor cell lines in multiplexed format and applied genome-scale CRISPR-based gene-editing screens in several solid tumor cell lines, to functionally interrogate which genes in tumor cells regulate the response to NK cells. In these orthogonal studies, NK cell-sensitive tumor cells tend to exhibit 'mesenchymal-like' transcriptional programs; high transcriptional signature for chromatin remodeling complexes; high levels of B7-H6 (NCR3LG1); and low levels of HLA-E/antigen presentation genes. Importantly, transcriptional signatures of NK cell-sensitive tumor cells correlate with immune checkpoint inhibitor (ICI) resistance in clinical samples. This study provides a comprehensive map of mechanisms regulating tumor cell responses to NK cells, with implications for future biomarker-driven applications of NK cell immunotherapies.

An Embryonic Diapause-like Adaptation with Suppressed Myc Activity Enables Tumor Treatment Persistence.

Treatment-persistent residual tumors impede curative cancer therapy. To understand this cancer cell state we generated models of treatment persistence that simulate the residual tumors. We observe that treatment-persistent tumor cells in organoids, xenografts, and cancer patients adopt a distinct and reversible transcriptional program resembling that of embryonic diapause, a dormant stage of suspended development triggered by stress and associated with suppressed Myc activity and overall biosynthesis. In cancer cells, depleting Myc or inhibiting Brd4, a Myc transcriptional co-activator, attenuates drug cytotoxicity through a dormant diapause-like adaptation with reduced apoptotic priming. Conversely, inducible Myc upregulation enhances acute chemotherapeutic activity. Maintaining residual cells in dormancy after chemotherapy by inhibiting Myc activity or interfering with the diapause-like adaptation by inhibiting cyclin-dependent kinase 9 represent potential therapeutic strategies against chemotherapy-persistent tumor cells. Our study demonstrates that cancer co-opts a mechanism similar to diapause with adaptive inactivation of Myc to persist during treatment.

Likelihood ratio statistics for gene set enrichment in Alzheimer's disease pathways.

INTRODUCTION: The study of Alzheimer's disease (AD) has revealed biological pathways with implications for disease neuropathology and pathophysiology. These pathway-level effects may also be mediated by individual characteristics or covariates such as age or sex. Evaluation of AD biological pathways in the context of interactions with these covariates is critical to the understanding of AD as well as the development of model systems used to study the disease. METHODS: Gene set enrichment methods are powerful tools used to interpret gene-level statistics at the level of biological pathways. We introduce a method for quantifying gene set enrichment using likelihood ratio-derived test statistics (gsLRT), which accounts for sample covariates like age and sex. We then use our method to test for age and sex interactions with protein expression levels in AD and to compare the pathway results between human and mouse species. RESULTS: Our method, based on nested logistic regressions is competitive with the existing standard for gene set testing in the context of linear models and complex experimental design. The gene sets we identify as having a significant association with AD-both with and without additional covariate interactions-are validated by previous studies. Differences between gsLRT results on mouse and human datasets are observed. DISCUSSION: Characterizing biological pathways involved in AD builds on the important work involving single gene drivers. Our gene set enrichment method finds pathways that are significantly related to AD while accounting for covariates that may be relevant to disease development. The method highlights commonalities and differences between human AD and mouse models, which may inform the development of higher fidelity models for the study of AD.

Opportunities and Challenges for Analyzing Cancer Data at the Inter- and Intra-Institutional Levels.

PURPOSE: Our goal was to identify the opportunities and challenges in analyzing data from the American Association of Cancer Research Project Genomics Evidence Neoplasia Information Exchange (GENIE), a multi-institutional database derived from clinically driven genomic testing, at both the inter- and the intra-institutional level. Inter-institutionally, we identified genotypic differences between primary and metastatic tumors across the 3 most represented cancers in GENIE. Intra-institutionally, we analyzed the clinical characteristics of the Vanderbilt-Ingram Cancer Center (VICC) subset of GENIE to inform the interpretation of GENIE as a whole. METHODS: We performed overall cohort matching on the basis of age, ethnicity, and sex of 13,208 patients stratified by cancer type (breast, colon, or lung) and sample site (primary or metastatic). We then determined whether detected variants, at the gene level, were associated with primary or metastatic tumors. We extracted clinical data for the VICC subset from VICC's clinical data warehouse. Treatment exposures were mapped to a 13-class schema derived from the HemOnc ontology. RESULTS: Across 756 genes, there were significant differences in all cancer types. In breast cancer, ESR1 variants were over-represented in metastatic samples (odds ratio, 5.91; q < 10-6). TP53 mutations were over-represented in metastatic samples across all cancers. VICC had a significantly different cancer type distribution than that of GENIE but patients were well matched with respect to age, sex, and sample type. Treatment data from VICC was used for a bipartite network analysis, demonstrating clusters with a mix of histologies and others being more histology specific. CONCLUSION: This article demonstrates the feasibility of deriving meaningful insights from GENIE at the inter- and intra-institutional level and illuminates the opportunities and challenges of the data GENIE contains. The results should help guide future development of GENIE, with the goal of fully realizing its potential for accelerating precision medicine.

Discovering the anti-cancer potential of non-oncology drugs by systematic viability profiling.

Anti-cancer uses of non-oncology drugs have occasionally been found, but such discoveries have been serendipitous. We sought to create a public resource containing the growth inhibitory activity of 4,518 drugs tested across 578 human cancer cell lines. We used PRISM, a molecular barcoding method, to screen drugs against cell lines in pools. An unexpectedly large number of non-oncology drugs selectively inhibited subsets of cancer cell lines in a manner predictable from the cell lines' molecular features. Our findings include compounds that killed by inducing PDE3A-SLFN12 complex formation; vanadium-containing compounds whose killing depended on the sulfate transporter SLC26A2; the alcohol dependence drug disulfiram, which killed cells with low expression of metallothioneins; and the anti-inflammatory drug tepoxalin, which killed via the multi-drug resistance protein ABCB1. The PRISM drug repurposing resource (https://depmap.org/repurposing) is a starting point to develop new oncology therapeutics, and more rarely, for potential direct clinical translation.

A case report of necrotizing fasciitis with growth of Actinomyces europaeus and Actinotignum schaalii.

Actinomyces europeaeus and Actinotignum schaalii are two facultative anaerobes that are common contaminants of human flora; namely the urinary tract, the female genital tract and the gastrointestinal tract. A. europeaeus has been linked with abscesses, decubitus ulcers and purulent urethritis, while A. schaalii has been associated with urinary tract infections, bacteremia and Fournier's gangrene. Here we present a case report of an 84-year-old female patient found to have a necrotizing soft tissue infection caused by A. europeaeus and A. schaalii. To our knowledge, this is the first case report that documents A. europeaeus as a causal agent of a necrotizing infection.

Improved estimation of cancer dependencies from large-scale RNAi screens using model-based normalization and data integration.

The availability of multiple datasets comprising genome-scale RNAi viability screens in hundreds of diverse cancer cell lines presents new opportunities for understanding cancer vulnerabilities. Integrated analyses of these data to assess differential dependency across genes and cell lines are challenging due to confounding factors such as batch effects and variable screen quality, as well as difficulty assessing gene dependency on an absolute scale. To address these issues, we incorporated cell line screen-quality parameters and hierarchical Bayesian inference into DEMETER2, an analytical framework for analyzing RNAi screens ( https://depmap.org/R2-D2 ). This model substantially improves estimates of gene dependency across a range of performance measures, including identification of gold-standard essential genes and agreement with CRISPR/Cas9-based viability screens. It also allows us to integrate information across three large RNAi screening datasets, providing a unified resource representing the most extensive compilation of cancer cell line genetic dependencies to date.

Interactive or static reports to guide clinical interpretation of cancer genomics.

Objective: Misinterpretation of complex genomic data presents a major challenge in the implementation of precision oncology. We sought to determine whether interactive genomic reports with embedded clinician education and optimized data visualization improved genomic data interpretation. Materials and Methods: We conducted a randomized, vignette-based survey study to determine whether exposure to interactive reports for a somatic gene panel, as compared to static reports, improves physicians' genomic comprehension and report-related satisfaction (overall scores calculated across 3 vignettes, range 0-18 and 1-4, respectively, higher score corresponding with improved endpoints). Results: One hundred and five physicians at a tertiary cancer center participated (29% participation rate): 67% medical, 20% pediatric, 7% radiation, and 7% surgical oncology; 37% female. Prior to viewing the case-based vignettes, 34% of the physicians reported difficulty making treatment recommendations based on the standard static report. After vignette/report exposure, physicians' overall comprehension scores did not differ by report type (mean score: interactive 11.6 vs static 10.5, difference = 1.1, 95% CI, -0.3, 2.5, P = .13). However, physicians exposed to the interactive report were more likely to correctly assess sequencing quality (P < .001) and understand when reports needed to be interpreted with caution (eg, low tumor purity; P = .02). Overall satisfaction scores were higher in the interactive group (mean score 2.5 vs 2.1, difference = 0.4, 95% CI, 0.2-0.7, P = .001). Discussion and Conclusion: Interactive genomic reports may improve physicians' ability to accurately assess genomic data and increase report-related satisfaction. Additional research in users' genomic needs and efforts to integrate interactive reports into electronic health records may facilitate the implementation of precision oncology.

Computational correction of copy number effect improves specificity of CRISPR-Cas9 essentiality screens in cancer cells.

The CRISPR-Cas9 system has revolutionized gene editing both at single genes and in multiplexed loss-of-function screens, thus enabling precise genome-scale identification of genes essential for proliferation and survival of cancer cells. However, previous studies have reported that a gene-independent antiproliferative effect of Cas9-mediated DNA cleavage confounds such measurement of genetic dependency, thereby leading to false-positive results in copy number-amplified regions. We developed CERES, a computational method to estimate gene-dependency levels from CRISPR-Cas9 essentiality screens while accounting for the copy number-specific effect. In our efforts to define a cancer dependency map, we performed genome-scale CRISPR-Cas9 essentiality screens across 342 cancer cell lines and applied CERES to this data set. We found that CERES decreased false-positive results and estimated sgRNA activity for both this data set and previously published screens performed with different sgRNA libraries. We further demonstrate the utility of this collection of screens, after CERES correction, for identifying cancer-type-specific vulnerabilities.

Low-affinity uptake of the fluorescent organic cation 4-(4-(dimethylamino)styryl)-N-methylpyridinium iodide (4-Di-1-ASP) in BeWo cells.

Understanding the mechanisms of transport processes in the placenta can improve the safety and efficacy of drug delivery during pregnancy. Functional studies of organic cation transporters (OCTs) are usually carried out using radioactivity, and a fluorescent marker would add flexibility to experimental methods. As a published substrate for OCT1 and OCT2, the fluorescent compound 4-(4-(dimethylamino)styryl)-N-methylpyridinium iodide (4-Di-1-ASP) was chosen as a candidate for studying placental OCT function in BeWo cells. The expression of OCT1 and OCT2 was also investigated in BeWo cells, an established human choriocarcinoma trophoblastic cell line frequently used as an in vitro model of the rate-limiting barrier for maternal-fetal exchange of drugs and nutrients within the placenta. 4-Di-1-ASP was taken up into BeWo cells by a low-affinity, carrier-mediated process exhibiting a Km of 580+/-110 microM and Vmax of 97+/-9 nmol/mg protein/30 min, and asymmetric transport was observed, with greater permeability in the apical to basolateral (maternal-to-fetal) direction. However, RT-PCR revealed no expression of OCT1 or OCT2 in either BeWo cells or primary cultured human cytotrophoblast cells, and OCT substrates such as TEA and choline did not inhibit the uptake of 4-Di-1-ASP. Although the uptake of this fluorescent compound in BeWo cells is not mediated by an OCT, the colocalization experiments with fluorescence microscopy and inhibition studies confirmed significant mitochondrial uptake of 4-Di-1-ASP. Transport of 4-Di-1-ASP into the nuclear region of BeWo cells was also observed, which is likely mediated by a nucleoside transporter.