Gene expression differences between species are driven by both cis and trans effects. Whereas cis effects are caused by genetic variants located on the same DNA molecule as the target gene, trans ...effects are due to genetic variants that affect diffusible elements. Previous studies have mostly assessed the impact of cis and trans effects at the gene level. However, how cis and trans effects differentially impact regulatory elements such as enhancers and promoters remains poorly understood. Here, we use massively parallel reporter assays to directly measure the transcriptional outputs of thousands of individual regulatory elements in embryonic stem cells and measure cis and trans effects between human and mouse.
Our approach reveals that cis effects are widespread across transcribed regulatory elements, and the strongest cis effects are associated with the disruption of motifs recognized by strong transcriptional activators. Conversely, we find that trans effects are rare but stronger in enhancers than promoters and are associated with a subset of transcription factors that are differentially expressed between human and mouse. While we find that cis-trans compensation is common within promoters, we do not see evidence of widespread cis-trans compensation at enhancers. Cis-trans compensation is inversely correlated with enhancer redundancy, suggesting that such compensation may often occur across multiple enhancers.
Our results highlight differences in the mode of evolution between promoters and enhancers in complex mammalian genomes and indicate that studying the evolution of individual regulatory elements is pivotal to understand the tempo and mode of gene expression evolution.
Understanding the consequences of individual transcriptome variation is fundamental to deciphering human biology and disease. We implement a statistical framework to quantify the contributions of 21 ...individual traits as drivers of gene expression and alternative splicing variation across 46 human tissues and 781 individuals from the Genotype-Tissue Expression project. We demonstrate that ancestry, sex, age, and BMI make additive and tissue-specific contributions to expression variability, whereas interactions are rare. Variation in splicing is dominated by ancestry and is under genetic control in most tissues, with ribosomal proteins showing a strong enrichment of tissue-shared splicing events. Our analyses reveal a systemic contribution of types 1 and 2 diabetes to tissue transcriptome variation with the strongest signal in the nerve, where histopathology image analysis identifies novel genes related to diabetic neuropathy. Our multi-tissue and multi-trait approach provides an extensive characterization of the main drivers of human transcriptome variation in health and disease.
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•Ancestry, sex, age, and BMI make tissue-specific contributions to expression variation•Contributions to expression variation are mostly additive, and interactions are rare•Ribosomal proteins have widespread splicing differences between populations•Systemic analysis of diabetes reveals genes associated with neuropathy in the tibial nerve
García-Pérez et al. perform a multi-tissue analysis of the association between demographic and clinical traits and human transcriptome variation. Ancestry, sex, age, and BMI and certain diseases make large tissue-specific contributions to gene expression variation, whereas alternative splicing variation is mostly driven by ancestry and under genetic control.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Abstract
Three inhibitors of CDK4/6 kinases have been recently approved for use in combination with endocrine therapy. These inhibitors considerably increase the progression-free survival of patients ...with advanced Estrogen receptor (ER)-positive breast cancer in first-line treatment. As the resistance to CDK4/6 inhibitors is currently a major problem faced in pre-clinical and clinical drug development, the identification of novel treatment combinations and predictive markers is of high importance. BET proteins, specifically BRD2/4, have important roles in cell cycle regulation and proliferation. Thus, BET inhibitors (BETi)have the potential to overcome some pathways of resistance, especially for those mediated by upregulation of specific transcription factors.
We have investigated the response to BET inhibition in patient-derived xenographs (PDXs) from patients with advanced ER+ breast cancer ex vivo and in vivo. Specifically, 14 breast cancer PDXs (10 ER+/HER2- and 4 ER+/HER2+) that were resistant to CDK4/6i in vivo were treated with ZEN-3694, a selective BET inhibitor currently in clinical development. Nine of the PDXs were sensitive to BET inhibition in monotherapy (65%). The non-responder PDXs showed an improved response when they were treated with ZEN-3694 in combination with fulvestrant (a selective ER degrader) or with a CDK4/6 inhibitor (ribociclib or abemaciclib). RNA-seq and computational analyses comparing responder versus non-responder PDXs revealed 493 differential expressed genes for responders, and 252 for non-responders. From those, 142 genes were differentially expressed in both conditions (untreated and treated). Furthermore, phosphorylation of BRD4 was observed in the ZEN-3694 non-responder PDX group, a mechanism of resistance previously observed in triple-negative breast cancer cell lines. This phosphorylation was abrogated by adding a CDK4/6 inhibitor. In the PDXs, resistance to single agent CDK4/6 and BET inhibitors might be overcome using a combination of both inhibitors. Finally, the molecular mechanism involved in phosphorylation of BRD4 was characterized in a MCF-7 cell line resistant to CDK4/6 inhibitors. Our results show that CDK2 could be one of the kinases involved in the phosphorylation of BRD4.
Conclusion: BET inhibition has an impact in advanced ER+ breast cancer cells, either used alone or in combination with endocrine therapy and CDK4/6 inhibitors. Detection of phosphorylated BRD4 (as a mechanism of resistance to BETi) has high potential for use as a biomarker for selecting patients who could benefit from a combined treatment of BET and CDK4/6 inhibitors.
Citation Format: Marc Cosín, Tian Tian, Winona Oliveros, Olesya A. Kharenko, Edward H. van der Horst, Reena G. Patel, Cyrus Calosing, Eric Campeau, Ravi Jahagirdar, Sanjay Lakhotia, Marta Melé, Marta Palafox, Joaquín Arribas, Mafalda Oliveira, Cristina Saura, Violeta Serra, Sandra Peiro. Overcoming resistance to inhibitors of CDK4/6 and BET in estrogen receptor-positive breast cancer abstract. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1282.
Cholangiocarcinoma (CCA) is usually diagnosed at advanced stages, with limited therapeutic options. Preclinical models focused on unresectable metastatic CCA are necessary to develop rational ...treatments. Pathogenic mutations in IDH1/2, ARID1A/B, BAP1, and BRCA1/2 have been identified in 30%-50% of patients with CCA. Several types of tumor cells harboring these mutations exhibit homologous recombination deficiency (HRD) phenotype with enhanced sensitivity to PARP inhibitors (PARPi). However, PARPi treatment has not yet been tested for effectiveness in patient-derived models of advanced CCA.
We have established a collection of patient-derived xenografts from patients with unresectable metastatic CCA (CCA_PDX). The CCA_PDXs were characterized at both histopathologic and genomic levels. We optimized a protocol to generate CCA tumoroids from CCA_PDXs. We tested the effects of PARPis in both CCA tumoroids and CCA_PDXs. Finally, we used the RAD51 assay to evaluate the HRD status of CCA tissues.
This collection of CCA_PDXs recapitulates the histopathologic and molecular features of their original tumors. PARPi treatments inhibited the growth of CCA tumoroids and CCA_PDXs with pathogenic mutations of BRCA2, but not those with mutations of IDH1, ARID1A, or BAP1. In line with these findings, only CCA_PDX and CCA patient biopsy samples with mutations of BRCA2 showed RAD51 scores compatible with HRD.
Our results suggest that patients with advanced CCA with pathogenic mutations of BRCA2, but not those with mutations of IDH1, ARID1A, or BAP1, are likely to benefit from PARPi therapy. This collection of CCA_PDXs provides new opportunities for evaluating drug response and prioritizing clinical trials.
High blood pressure (BP) is the major risk factor for cardiovascular disease. Genome-wide association studies have identified genetic variants for BP, but functional insights into causality and ...related molecular mechanisms lag behind. We functionally characterize 4,608 genetic variants in linkage with 135 BP loci in vascular smooth muscle cells and cardiomyocytes by massively parallel reporter assays. High densities of regulatory variants at BP loci (i.e., ULK4, MAP4, CFDP1, PDE5A) indicate that multiple variants drive genetic association. Regulatory variants are enriched in repeats, alter cardiovascular-related transcription factor motifs, and spatially converge with genes controlling specific cardiovascular pathways. Using heuristic scoring, we define likely causal variants, and CRISPR prime editing finally determines causal variants for KCNK9, SFXN2, and PCGF6, which are candidates for developing high BP. Our systems-level approach provides a catalog of functionally relevant variants and their genomic architecture in two trait-relevant cell lines for a better understanding of BP gene regulation.
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•Functional testing of 4,608 genetic variants associated with blood pressure•High densities of regulatory variants spatially converge to influence target genes•Heuristic scoring using epigenomic properties defines likely causal variants•CRISPR prime editing determines gene-regulatory effects of causal variants
Functional testing of 4,608 genetic variants associated with blood pressure revealed high densities of regulatory variants at candidate genes. Epigenomic properties defined likely causal variants, which were ultimately tested for their gene-regulatory effects by CRISPR prime editing.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Cardiomyopathy (CMP) is a heritable disorder. Over 50% of cases are gene-elusive on clinical gene panel testing. The contribution of variants in non-coding DNA elements that result in cryptic ...splicing and regulate gene expression has not been explored. We analyzed whole-genome sequencing (WGS) data in a discovery cohort of 209 pediatric CMP patients and 1953 independent replication genomes and exomes. We searched for protein-coding variants, and non-coding variants predicted to affect the function or expression of genes. Thirty-nine percent of cases harbored pathogenic coding variants in known CMP genes, and 5% harbored high-risk loss-of-function (LoF) variants in additional candidate CMP genes. Fifteen percent harbored high-risk regulatory variants in promoters and enhancers of CMP genes (odds ratio 2.25, p = 6.70 × 10
versus controls). Genes involved in α-dystroglycan glycosylation (FKTN, DTNA) and desmosomal signaling (DSC2, DSG2) were most highly enriched for regulatory variants (odds ratio 6.7-58.1). Functional effects were confirmed in patient myocardium and reporter assays in human cardiomyocytes, and in zebrafish CRISPR knockouts. We provide strong evidence for the genomic contribution of functionally active variants in new genes and in regulatory elements of known CMP genes to early onset CMP.
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