Synthetic Lethal and Resistance Interactions with BET Bromodomain Inhibitors in Triple-Negative Breast Cancer.

BET bromodomain inhibitors (BBDIs) are candidate therapeutic agents for triple-negative breast cancer (TNBC) and other cancer types, but inherent and acquired resistance to BBDIs limits their potential clinical use. Using CRISPR and small-molecule inhibitor screens combined with comprehensive molecular profiling of BBDI response and resistance, we identified synthetic lethal interactions with BBDIs and genes that, when deleted, confer resistance. We observed synergy with regulators of cell cycle progression, YAP, AXL, and SRC signaling, and chemotherapeutic agents. We also uncovered functional similarities and differences among BRD2, BRD4, and BRD7. Although deletion of BRD2 enhances sensitivity to BBDIs, BRD7 loss leads to gain of TEAD-YAP chromatin binding and luminal features associated with BBDI resistance. Single-cell RNA-seq, ATAC-seq, and cellular barcoding analysis of BBDI responses in sensitive and resistant cell lines highlight significant heterogeneity among samples and demonstrate that BBDI resistance can be pre-existing or acquired.

Mitonuclear epistasis, genotype-by-environment interactions, and personalized genomics of complex traits in Drosophila.

Mitochondrial function requires the coordinated expression of dozens of gene products from the mitochondrial genome and hundreds from the nuclear genomes. The systems that emerge from these interactions convert the food we eat and the oxygen we breathe into energy for life, while regulating a wide range of other cellular processes. These facts beg the question of whether the gene-by-gene interactions (G x G) that enable mitochondrial function are distinct from the gene-by-environment interactions (G x E) that fuel mitochondrial activity. We examine this question using a Drosophila model of mitonuclear interactions in which experimental combinations of mtDNA and nuclear chromosomes generate pairs of mitonuclear genotypes to test for epistatic interactions (G x G). These mitonuclear genotypes are then exposed to altered dietary or oxygen environments to test for G x E interactions. We use development time to assess dietary effects, and genome wide RNAseq analyses to assess hypoxic effects on transcription, which can be partitioned in to mito, nuclear, and environmental (G x G x E) contributions to these complex traits. We find that mitonuclear epistasis is universal, and that dietary and hypoxic treatments alter the epistatic interactions. We further show that the transcriptional response to alternative mitonuclear interactions has significant overlap with the transcriptional response to alternative oxygen environments. Gene coexpression analyses suggest that these shared genes are more central in networks of gene interactions, implying some functional overlap between epistasis and genotype by environment interactions. These results are discussed in the context of evolutionary fitness, the genetic basis of complex traits, and the challenge of achieving precision in personalized medicine. © 2018 The Authors. IUBMB Life published by Wiley Periodicals, Inc. on behalf of International Union of Biochemistry and Molecular Biology, 70(12):1275-1288, 2018.

Heterogeneity and transcriptional drivers of triple-negative breast cancer.

Triple-negative breast cancer (TNBC) is a heterogeneous disease with limited treatment options. To characterize TNBC heterogeneity, we defined transcriptional, epigenetic, and metabolic subtypes and subtype-driving super-enhancers and transcription factors by combining functional and molecular profiling with computational analyses. Single-cell RNA sequencing revealed relative homogeneity of the major transcriptional subtypes (luminal, basal, and mesenchymal) within samples. We found that mesenchymal TNBCs share features with mesenchymal neuroblastoma and rhabdoid tumors and that the PRRX1 transcription factor is a key driver of these tumors. PRRX1 is sufficient for inducing mesenchymal features in basal but not in luminal TNBC cells via reprogramming super-enhancer landscapes, but it is not required for mesenchymal state maintenance or for cellular viability. Our comprehensive, large-scale, multiplatform, multiomics study of both experimental and clinical TNBC is an important resource for the scientific and clinical research communities and opens venues for future investigation.

Prolonged Survival in Patients with Metastatic HER2-Positive Inflammatory Breast Cancer: A Case Report and Review of the Literature.

Inflammatory breast cancer (IBC) is a rare type of breast cancer that is associated with poor outcomes compared with non-IBC. Overexpression of HER2 is enriched in IBC, and those with HER2-positive disease have a relatively favorable prognosis, with improved survival over the last two decades driven by the advent of novel targeted therapies. Here, we present two patients who have survived for over 10 years after being diagnosed with de novo metastatic HER2-positive IBC. We review the data for the treatments available for metastatic HER2-positive IBC and the evolving treatment recommendations for this disease.

Developing a pulse oximetry home monitoring protocol for patients suspected with COVID-19 after emergency department discharge.

OBJECTIVES: Patients with COVID-19 can present to the emergency department (ED) without immediate indication for admission, but with concern for decompensation. Clinical experience has demonstrated that critical illness may present later in the disease course and hypoxia is often the first indication of disease progression. The objectives of this study are to (a) assess feasibility and describe a protocol for ED-based outpatient pulse-oximetry monitoring with structured follow-up and (b) determine rates of ED return, hospitalisation and hypoxia among participants. METHODS: Prospective observational study of patients presenting to a single academic ED in Boston with suspected COVID-19. Eligible patients were adults being discharged from the ED with presumed COVID-19. Exclusion criteria included resting oxygen saturation <92%, ambulatory oxygen saturation <90%, heart rate >110 beats per minute or inability to use the device. Study personnel made scripted phone calls on postdischarge days 1, 3 and 7 to review the pulse-oximetry readings and to evaluate for decompensation. Return visit and admission information were collected via medical record and 28-day follow-up calls. RESULTS: 81 patients were enrolled of which 10 (12%) developed hypoxia after their initial discharge from the ED. Overall, 23 (28%) of the 81 patients returned to the ED at least once and 10 of those who returned (43%) were admitted. We successfully contacted 76/81 (94%) of subjects via phone at least once for follow-up assessment. DISCUSSION: Patients are eager and willing to participate in home monitoring systems and are comfortable with using technology, which will allow providers and health systems to extend our hospitals capabilities for tracking patient populations in times of crisis. CONCLUSIONS: It is feasible to implement an outpatient pulse-oximetry monitoring protocol to monitor patients discharged from the ED with confirmed or suspected COVID-19.

Acquired resistance to combined BET and CDK4/6 inhibition in triple-negative breast cancer.

BET inhibitors are promising therapeutic agents for the treatment of triple-negative breast cancer (TNBC), but the rapid emergence of resistance necessitates investigation of combination therapies and their effects on tumor evolution. Here, we show that palbociclib, a CDK4/6 inhibitor, and paclitaxel, a microtubule inhibitor, synergize with the BET inhibitor JQ1 in TNBC lines. High-complexity DNA barcoding and mathematical modeling indicate a high rate of de novo acquired resistance to these drugs relative to pre-existing resistance. We demonstrate that the combination of JQ1 and palbociclib induces cell division errors, which can increase the chance of developing aneuploidy. Characterizing acquired resistance to combination treatment at a single cell level shows heterogeneous mechanisms including activation of G1-S and senescence pathways. Our results establish a rationale for further investigation of combined BET and CDK4/6 inhibition in TNBC and suggest novel mechanisms of action for these drugs and new vulnerabilities in cells after emergence of resistance.

Mitochondrial DNA Fitness Depends on Nuclear Genetic Background in Drosophila.

Mitochondrial DNA (mtDNA) has been one of the most extensively studied molecules in ecological, evolutionary and clinical genetics. In its early application in evolutionary genetics, mtDNA was assumed to be a selectively neutral marker conferring negligible fitness consequences for its host. However, this dogma has been overturned in recent years due to now extensive evidence for non-neutral evolutionary dynamics. Since mtDNA proteins physically interact with nuclear proteins to provide the mitochondrial machinery for aerobic ATP production, among other cell functions, co-variation of the respective genes is predicted to affect organismal fitness. To test this hypothesis we used an mtDNA-nuclear DNA introgression model in Drosophila melanogaster to test the fitness of genotypes in perturbation-reperturbation population cages and in a non-competitive assay for female fecundity. Genotypes consisted of both conspecific and heterospecific mtDNA-nDNA constructs, with either D. melanogaster or D. simulans mtDNAs on two alternative D. melanogaster nuclear backgrounds, to investigate mitonuclear genetic interactions (G x G effects). We found considerable variation between nuclear genetic backgrounds on the selection of mtDNA haplotypes. In addition, there was variation in the selection on mtDNAs pre- and post- reperturbation, demonstrating overall poor repeatability of selection. There was a strong influence of nuclear background on non-competitive fecundity across all the mtDNA species types. In only one of the four cage types did we see a significant fecundity effect between genotypes that could help explain the respective change in genotype frequency over generational time. We discuss these results in the context of G x G interactions and the possible influence of stochastic environments on mtDNA-nDNA selection.

An alternative splicing switch in FLNB promotes the mesenchymal cell state in human breast cancer.

Alternative splicing of mRNA precursors represents a key gene expression regulatory step and permits the generation of distinct protein products with diverse functions. In a genome-scale expression screen for inducers of the epithelial-to-mesenchymal transition (EMT), we found a striking enrichment of RNA-binding proteins. We validated that QKI and RBFOX1 were necessary and sufficient to induce an intermediate mesenchymal cell state and increased tumorigenicity. Using RNA-seq and eCLIP analysis, we found that QKI and RBFOX1 coordinately regulated the splicing and function of the actin-binding protein FLNB, which plays a causal role in the regulation of EMT. Specifically, the skipping of FLNB exon 30 induced EMT by releasing the FOXC1 transcription factor. Moreover, skipping of FLNB exon 30 is strongly associated with EMT gene signatures in basal-like breast cancer patient samples. These observations identify a specific dysregulation of splicing, which regulates tumor cell plasticity and is frequently observed in human cancer.