Differential whole-genome doubling and homologous recombination deficiencies across breast cancer subtypes from the Taiwanese population.

Whole-genome doubling (WGD) is an early macro-evolutionary event in tumorigenesis, involving the doubling of an entire chromosome complement. However, its impact on breast cancer subtypes remains unclear. Here, we performed a comprehensive and quantitative analysis of WGD and its influence on breast cancer subtypes in patients from Taiwan and consequently highlight the genomic association between WGD and homologous recombination deficiency (HRD). A higher manifestation of WGD was reported in triple-negative breast cancer, conferring high chromosomal instability (CIN), while HER2 + tumors exhibited early WGD events, with widely varied CIN levels, compared to luminal-type tumors. An association of higher activity of de novo indel signature 2 with WGD and HRD in Taiwanese breast cancer patients was reported. A control test between WGD and pseudo non-WGD samples was further employed to support this finding. The study provides a better comprehension of tumorigenesis in breast cancer subtypes, thus assisting in personalized treatment.

Evolutionary Trajectories and Genomic Divergence in Localized Breast Cancers after Ipsilateral Breast Tumor Recurrence.

The evolutionary trajectories that drive clinical and therapeutic consequences in localized breast cancers (BCs) with ipsilateral breast tumor relapse (IBTR) remain largely unknown. Analyses of longitudinal paired whole-exome sequencing data from 10 localized BC patients with IBTR reveal that, compared to primary breast tumors, homologous recombination (HR) deficiency, inactivation of the HR pathway, chromosomal instability, and somatic driver mutations are more frequent. Furthermore, three major models of evolution in IBTR are summarized, through which relative contributions of mutational signatures shift, and the subclonal diversity expansions are shown. Optimal treatment regimens are suggested by the clinically relevant molecular features, such as HR deficiency (20%) or specific alterations (30%) with sensitivity to available FDA-approved drugs. Finally, a rationale for the development of the therapeutic management framework is provided. This study sheds light on the complicated evolution patterns in IBTR and has significant clinical implications for future improvement of treatment decisions.

Prevalence of sudden arrhythmic death syndrome-related genetic mutations in an Asian cohort of whole genome sequence.

AIMS: Recently, the spectrum of background mutation in the genes implicated in sudden arrhythmic death syndrome (SADS), has been elucidated in the Caucasian populations. However, this information is largely unknown in the Asian populations. METHODS AND RESULTS: We assessed the background rare variants (minor allele frequency < 0.01) of major SADS genes in whole genome sequence data of 1514 healthy Taiwanese subjects from the Taiwan Biobank. We found up to 45% of healthy subjects have a rare variant in at least one of the major SADS genes. Around 3.44% of healthy subjects had multiple mutations in one or multiple genes. The background mutation rates in long QT syndrome, catecholaminergic polymorphic ventricular tachycardia, and arrhythmogenic right ventricular cardiomyopathy genes were similar, but those in Brugada syndrome (BrS) (SCN5A) and hypertrophic cardiomyopathy (HCM) genes (MYBPC3, MYH7, and TNNT2) were higher, compared to those reported in the Caucasian populations. Furthermore, the rate of incidental pathogenic variant was highest in MYBPC3 gene. Finally, the number of variant was proportional to the exon length of the gene (R2 = 0.486, P = 0.0056) but not related to its functional or evolutionary importance (degree of evolutionary conservation) (R2 = 0.0008, P = 0.9218), suggesting that the mutation was random. The ratio of variant number over exon nucleotide length was highest in MYBPC3, MYH7, and TNNT2 genes. CONCLUSION: Unique features of background SADS gene mutation in the Asian populations include higher prevalence of incidental variant in HCM, BrS, and long QT 3 (SCN5A) genes. HCM genes have the highest variant number per exon length.

Genome-Wide Copy Number Variation Association Study of Atrial Fibrillation Related Thromboembolic Stroke.

Atrial fibrillation (AF) is a common cardiac arrhythmia and is one of the major causes of ischemic stroke. In addition to the clinical factors such as CHADS2 or CHADS2-VASC score, the impact of genetic factors on the risk of thromboembolic stroke in patients with AF has been largely unknown. Single-nucleotide polymorphisms in several genomic regions have been found to be associated with AF. However, these loci do not contribute to all the genetic risks of AF or AF related thromboembolic risks, suggesting that there are other genetic factors or variants not yet discovered. In the human genome, copy number variations (CNVs) could also contribute to disease susceptibility. In the present study, we sought to identify CNVs determining the AF-related thromboembolic risk. Using a genome-wide approach in 109 patients with AF and thromboembolic stroke and 14,666 controls from the Taiwanese general population (Taiwan Biobank), we first identified deletions in chromosomal regions 1p36.32-1p36.33, 5p15.33, 8q24.3 and 19p13.3 and amplifications in 14q11.2 that were significantly associated with AF-related stroke in the Taiwanese population. In these regions, 148 genes were involved, including several microRNAs and long non-recoding RNAs. Using a pathway analysis, we found deletions in GNB1, PRKCZ, and GNG7 genes related to the alpha-adrenergic receptor signaling pathway that play a major role in determining the risk of an AF-related stroke. In conclusion, CNVs may be genetic predictors of a risk of a thromboembolic stroke for patients with AF, possibly pointing to an impaired alpha-adrenergic signaling pathway in the mechanism of AF-related thromboembolism.

Global Expression Profiling Identifies a Novel Hyaluronan Synthases 2 Gene in the Pathogenesis of Lower Extremity Varicose Veins.

Lower extremities varicose veins (VV) are among the most easily recognized venous abnormalities. The genetic mechanism of VV is largely unknown. In this study, we sought to explore the global expressional change of VV and identify novel genes that might play a role in VV. We used next-generation ribonucleic acid (RNA) sequence (RNA seq) technology to study the global messenger RNA expressional change in the venous samples of five diseased and five control patients. We identified several differentially expressed genes, which were further confirmed by conventional reverse transcription polymerase chain reaction (RT-PCR). Using these significant genes we performed in silico pathway analyses and found distinct transcriptional networks, such as angiogenesis, cell adhesion, vascular injury, and carbohydrate metabolisms that might be involved in the mechanism of VV. Among these significant genes, we also found hyaluronan synthases 2 gene (HAS2) played a pivotal role and governed all these pathways. We further confirmed that HAS2 expression was decreased in the venous samples of patients with VV. Finally, we used a zebrafish model with fluorescence emitting vasculature and red blood cells to see the morphological changes of the venous system and blood flow. We found that HAS2 knockdown in zebrafish resulted in dilated venous structural with static venous flow. HAS2 may modulate the transcriptional networks of angiogenesis, cell adhesion, vascular injury, and carbohydrate metabolisms in venous tissues and downregulation of HAS2 may underlie the mechanism of VV.

Genome-wide screening identifies a KCNIP1 copy number variant as a genetic predictor for atrial fibrillation.

Atrial fibrillation (AF) is the most common sustained cardiac arrhythmia. Previous genome-wide association studies had identified single-nucleotide polymorphisms in several genomic regions to be associated with AF. In human genome, copy number variations (CNVs) are known to contribute to disease susceptibility. Using a genome-wide multistage approach to identify AF susceptibility CNVs, we here show a common 4,470-bp diallelic CNV in the first intron of potassium interacting channel 1 gene (KCNIP1) is strongly associated with AF in Taiwanese populations (odds ratio=2.27 for insertion allele; P=6.23 × 10(-24)). KCNIP1 insertion is associated with higher KCNIP1 mRNA expression. KCNIP1-encoded protein potassium interacting channel 1 (KCHIP1) is physically associated with potassium Kv channels and modulates atrial transient outward current in cardiac myocytes. Overexpression of KCNIP1 results in inducible AF in zebrafish. In conclusions, a common CNV in KCNIP1 gene is a genetic predictor of AF risk possibly pointing to a functional pathway.

Next-generation sequencing of nine atrial fibrillation candidate genes identified novel de novo mutations in patients with extreme trait of atrial fibrillation.

BACKGROUND: Atrial fibrillation (AF) is the most common sustained cardiac arrhythmia. Genome-wide association studies (GWAS) have identified common variants in nine genomic regions associated with AF (KCNN3, PRRX1, PITX2, WNT8A, CAV1, C9orf3, SYNE2, HCN4 and ZFHX3 genes); however, the genetic variability of these risk variants does not explain the entire genetic susceptibility to AF. Rare variants missed by GWAS may also contribute to genetic risk of AF. METHODS: We used an extreme trait design to sequence carefully selected probands with extreme phenotypes and their unaffected parents to identify rare de novo variants or mutations. Based on the hypothesis that common and rare variants may colocate in the same disease susceptibility gene, we used next-generation sequencing to sequence these nine published AF susceptibility genes identified by GWAS (a total of 179 exons) in 20 trios, 200 unrelated patients with AF and 200 non-AF controls. RESULTS: We identified a novel mutation in the 5' untranslated region of the PITX2 gene, which localised in the transcriptionally active enhancer region. We also identified one missense exon mutation in KCNN3, two in ZFHX3 and one in SYNE2. None of these mutations were present in other unrelated patients with AF, healthy controls, unaffected parents and are thus novel and de novo (p<10(-4)). Functional study showed that the mutation in the 5' untranslated region of the PITX2 gene significantly downregulated PITX2 expression in atrial myocytes, either in basal condition or during rapid pacing. In silico analysis showed that the missense mutation in ZFHX3 results in damage of the ZFHX3 protein structure. CONCLUSIONS: The genetic architecture of subjects with extreme phenotypes of AF is similar to that of rare or Mendelian diseases, and mutations may be the underlying cause.

Next-Generation Sequencing in the Genetics of Human Atrial Fibrillation.

UNLABELLED: The International Human Genome Sequencing Consortium published the first draft of the human genome in the journal Nature in February 2001, providing the sequence of the entire genome's three billion base pairs. The Human Genome Project involves a concerted effort to better understand the human DNA sequence through identification of all the genes. The knowledge that can be derived from the genome could result in the development of novel diagnostic assays, targeted therapies and the improved ability to predict the onset, severity and progression of diseases. This has been made possible by many parallelized, high-throughput technologies such as next-generation sequencing. In this review, we discuss the possible application of next-generation sequencing in finding the susceptibility gene(s) or disease mechanism of an important human arrhythmia called atrial fibrillation. KEY WORDS: Arrhythmia; Atrial fibrillation; Genetics, Next-generation sequencing.

Angiotensin II does not influence expression of sarcoplasmic reticulum Ca2 + ATPase in atrial myocytes.

INTRODUCTION: The sarcoplasmic reticulum Ca(2+) ATPase (SERCA) is essential for the regulation of the intracellular calcium level in cardiomyocytes. Previous studies have found that angiotensin II (Ang II) decreased SERCA2 gene expression in ventricular myocytes. Alteration of SERCA activity is important in the mechanism of atrial fibrillation. The present study was undertaken to examine Ang II effects on atrial myocytes. MATERIALS AND METHODS: An approximately 1.75-kb promoter region of SERCA2 gene was cloned with the pGL3 luciferase vector. The direct effects of Ang II on SERCA2 gene expression in HL-1 atrial myocytes were examined by promoter activity assay, followed by Western blot analysis for protein levels and quantitative real-time reverse transcription polymerase chain reaction for mRNA amounts. RESULTS: Ang II did not increase the promoter activity of the 1,754-bp promoter-receptor construct of the SERCA2 gene. The levels of SERCA2 protein and mRNA were also unchanged at different time points after Ang II treatment. CONCLUSIONS: Although Ang II had prominent effects on SERCA2 in ventricular myocytes, it did not alter SERCA2 gene expression and protein levels in atrial myocytes. We provide a model for further investigation of the regulation of SERCA2 gene expression in atrial myocytes.

Angiotensin II activates signal transducer and activators of transcription 3 via Rac1 in atrial myocytes and fibroblasts: implication for the therapeutic effect of statin in atrial structural remodeling.

BACKGROUND: Recently, activation of the local renin-angiotensin system and mitogen-activated protein kinase pathways in atrial myocardium has been found to play an important role in atrial structural remodeling related to atrial fibrillation. Another important mediator of the angiotensin II (Ang II) effect is the Janus kinase/signal transducers and activators of transcription (STAT) pathway, which has never been characterized in the atrium. METHODS AND RESULTS: In cultured atrial myocytes and fibroblasts, Ang II induced tyrosine phosphorylation of STAT3 through a Rac1-dependent mechanism, which was inhibited by dominant-negative Rac1, losartan, and simvastatin. In atrial myocytes, activation of STAT3 by Rac1 was mediated by direct association of Rac1 with STAT3; however, in atrial fibroblasts, it was mediated by an indirect paracrine effect. Constitutively active STAT3 increased protein synthesis, and dominant-negative STAT3 abrogated Ang II-induced protein synthesis in atrial myocytes and fibroblasts. Rats infused long term with Ang II exhibited higher levels of activated Rac1, phospho-STAT3, collagen synthesis, and atrial fibrosis in the atria, all of which were attenuated by oral losartan and simvastatin. In human atrial tissues from patients with atrial fibrillation, Ang II and phospho-STAT3 levels were also elevated. CONCLUSIONS: The Ang II/Rac1/STAT3 pathway is an important signaling pathway in the atrial myocardium to mediate atrial structural remodeling, and losartan and statin may be able to reverse Ang II-induced atrial structural remodeling in atrial fibrillation.

Angiotensin II increases expression of alpha1C subunit of L-type calcium channel through a reactive oxygen species and cAMP response element-binding protein-dependent pathway in HL-1 myocytes.

Angiotensin II (Ang II) is involved in the pathogenesis of atrial fibrillation (AF). L-type calcium channel (LCC) expression is altered in AF remodeling. We investigated whether Ang II modulates LCC current through transcriptional regulation, by using murine atrial HL-1 cells, which have a spontaneous calcium transient, and an in vivo rat model. Ang II increased LCC alpha1C subunit mRNA and protein levels and LCC current density, which resulted in an augmented calcium transient in atrial myocytes. An approximately 2-kb promoter region of LCC alpha1C subunit gene was cloned to the pGL3 luciferase vector. Ang II significantly increased promoter activity in a concentration- and time-dependent manner. Truncation and mutational analysis of the LCC alpha1C subunit gene promoter showed that cAMP response element (CRE) (-1853 to -1845) was an important cis element in Ang II-induced LCC alpha1C subunit gene expression. Transfection of dominant-negative CRE binding protein (CREB) (pCMV-CREBS133A) abolished the Ang II effect. Ang II (1 micromol/L, 2 hours) induced serine 133 phosphorylation of CREB and binding of CREB to CRE and increased LCC alpha1C subunit gene promoter activity through a protein kinase C/NADPH oxidase/reactive oxygen species pathway, which was blocked by the Ang II type 1 receptor blocker losartan and the antioxidant simvastatin. In the rat model, Ang II infusion increased LCC alpha1C subunit expression and serine 133 phosphorylation of CREB, which were attenuated by oral losartan and simvastatin. In summary, Ang II induced LCC alpha1C subunit expression via a protein kinase C-, reactive oxygen species-, and CREB-dependent pathway and was blocked by losartan and simvastatin.