Loss Of Function Alleles Research Articles

A study of microRNA-223 in evaluating platelet reactivity in patients with acute ischemic stroke.

To investigate the relationship between plasma microRNA-223 expression and platelet reactivity in patients with acute ischemic stroke (AIS) and to evaluate its predictive value in clopidogrel resistance or high on-treatment platelet reactivity (HTPR). A total of 120 patients with acute ischemic stroke were screened in this study, and 60 patients were included in the acute ischemic stroke group according to the inclusion criteria and platelet reactivity after clopidogrel treatment. control group was 60 non-ischemic stroke patients hospitalized. The levels of phosphorylation of vasodilator-stimulated phosphoprotein (VASP) and adenosine diphosphate-induced platelet aggregation (ADP-PAg) in platelets were detected by flow cytometry. The expression level of plasma microRNA-223 was detected before and after clopidogrel treatment using quantitative real-time polymerase chain reaction (PcR). The AIS group was then divided into the clopidogrel non-HTPR and the clopidogrel HTPR groups based on the relative inhibition rate. We found that: 1) the VASP platelet reactivity index (PRI) was positively correlated with ADP-PAg; 2) before administration, the plasma microRNA-223 expression level and VASP-PRI were higher in the AIS group than in the control group; 3) after administration, the expression level of microRNA-223 was negatively correlated with VASP-PRI; 4) before and after treatment, the plasma microRNA-223 expression level in the clopidogrel HTPR group was lower than in the non-AIS patients; 5) before treatment, there was an interaction between the expression level of microRNA-223 in the plasma and the cYP2c19 loss-of-function (LOF) allele. The study showed that decreased plasma microRNA-223 expression levels in AIS patients indicate an increased risk of clopidogrel HTPR. carrying cYP2c19 LOF alleles may result in the microRNA-223 expression being more distinct. The combined detection of plasma microRNA-223 and cYP2c19 gene polymorphisms may be effective in predicting the occurrence of clopidogrel HTPR in patients with AIS.

Effects of the CYP2C19 LoF allele on major adverse cardiovascular events associated with clopidogrel in acute coronary syndrome patients undergoing percutaneous coronary intervention: a meta-analysis.

The aggregated risk of major adverse cardiovascular events (MACE) in acute coronary syndrome (ACS) patients inheriting CYP2C19 loss-of function (LoF) alleles who underwent percutaneous coronary intervention (PCI) and were treated with clopidogrel is controversial. In the current study, we searched the literature in different databases for eligible studies. The risk ratio (RR) was measured where p<0.05 was statistically significant. The ACS patients with either one or two CYP2C19 LoF alleles who underwent PCI, treated with clopidogrel were correlated with a significantly escalated risk of MACE compared with noncarriers (RR: 1.53, 95% CI: 1.39-1.69, p < 0.00001), driven by CV death (RR: 1.88, 95% CI: 1.18-3.01, p = 0.008), MI (RR: 1.67, 95% CI: 1.21-2.31, p = 0.002) and ST (RR: 1.90, 95% CI: 1.27-2.84, p = 0.002). Patients with two CYP2C19 LoF alleles were correlated with significantly greater risk of MACE compared with noncarriers (RR: 3.91, 95% CI: 2.78-5.50, p < 0.00001). Further analysis revealed that the risk of MACE was markedly significant in Asian patients (RR: 2.02, 95% CI: 1.67-2.44, p < 0.00001) and wascomparatively low significance in western patients (RR: 1.35, 95% CI: 1.20-1.52, p < 0.00001). There was no significantly different bleeding events in patients with CYP2C19 LoF alleles compared with noncarriers (RR: 0.99, 95% CI: 0.85-1.15, p = 0.87). The ACS patients inheriting CYP2C19 LoF alleles, who underwent PCI and were treated with clopidogrel were correlated with significantly increased risk of MACE compared with noncarriers.

Transcriptome Profile of a Long-Juvenile Soybean Genotype Huaxia-3 Under Short and Long Photoperiod

The j allele delays flowering and enhances yield of long juvenile (LJ) soybean under short day (SD) condition. However, the underlying mechanism of j in flowering pathway of soybean is not fully known. The objective of the study is to profile the transcriptome of Huaxia-3 (HX3), a typical long juvenile soybean variety with a loss of function allele (j) for the J gene. This helps to identify the genes implicated in delayed flowering in the long juvenile soybean variety and the pathways involved. Phenotypic analysis revealed HX3-delayed flowering and matured later than the transgenic line with the functional J gene under SD. RNA-Seq analysis was conducted to compare the transcriptome of HX3 relative to the one overexpressing J gene under SD and long day (LD). A total of 674,800,494 clean reads were generated, of which 626,517,161 (93%) were uniquely mapped to the soybean reference genome. A total of 31 and 2311 genes were differentially expressed in the HX3 under SD and LD conditions respectively. The circadian rhythm pathway was the most significantly enriched pathway in the HX3 under SD condition. The GmELF3a (Glyma.04G050200) and FLOWERING LOCUS T (FT) genes such as GmFT2a (Glyma.16G150700) and GmFT5a (Glyma.16G044100) were downregulated, whiles GmFT4 (Glyma.08G363100) was upregulated in the HX3 under SD. Under LD FT homologs, GmFT4 (Glyma.08G363100) was downregulated and GmFT1a (Glyma.18G299000) was upregulated. Our study suggests that these FT homologs may be involved in delayed flowering of LJ soybean under SD.

Effect of Genotype-Guided Oral P2Y12 Inhibitor Selection After Percutaneous Coronary Intervention: A Systematic Review and Meta-Analysis of Randomized Clinical Trials

BackgroundClopidogrel is the most frequently used P2Y12 inhibitor as a component of the dual antiplatelet regimen in patients with acute coronary syndrome (ACS) undergoing percutaneous coronary intervention (PCI). Prior studies have shown the variable efficacy of clopidogrel due to genotypic differences in the CYP2C19 enzyme function, which converts clopidogrel to its active metabolite. The aim of this meta-analysis is to evaluate the effectiveness of genotype testing-guided P2Y12 inhibitor prescription therapy to patients after PCI for ACS compared to non-genotype guided conventional treatment. MethodsA comprehensive literature search was performed in PubMed, Embase, and Cochrane to identify relevant trials. Summary effects were calculated using a DerSimonian and Laird random-effects model as odds ratio with 95% confidence intervals for all the clinical endpoints. ResultsSeven studies with 9617 patients were included. Genotype-guided strategy arm included prasugrel or ticagrelor prescription to patients with loss of function (LOF) of CYP219 alleles (most commonly alleles being *2 and *3) and clopidogrel prescription to those without the LOF allele. The conventional arm included patients treated with clopidogrel without genotype testing. Comparison of genotype arm with conventional arm showed decreased major adverse cardiovascular events (MACE), improved cardiovascular (CV) mortality, and reduced incidence of myocardial infarction (MI) in the genotype arm, and a similar stroke incidence in the two arms. Regarding adverse events, the incidence of stent thrombosis was lower in the genotype arm than the conventional arm. ConclusionOur analysis illustrates the possible advantages of genotype-guided P2Y12 inhibitor prescription strategy compared to non-genotype-guided strategy with reductions in MACE, CV mortality, MI, and stent thrombosis. This analysis can be used as a stepping stone to conducting further trials determining the efficacy of this treatment strategy in various ACS subtypes.

A common SSD1 truncation is toxic to cells entering quiescence and promotes sporulation.

Ssd1p is an RNA binding protein in Saccharomyces cerevisiae that plays an important role in cell division, cell fate decisions, stress response and virulence. Lab strain W303 encodes the terminal truncation ssd1-2, which is typically interpreted to be a loss of function allele. We have shown that ssd1-2 is toxic to mpt5-Δ mutants and to diploids entering stationary phase and quiescence. The ssd1-Δ null shows no toxicity, indicating that ssd1-2 is disrupting an essential function that does not solely require Ssd1p. ssd1-2 cells are also more sensitive to stress than ssd1-Δ . These phenotypes are recessive to SSD1-1 . In contrast, ssd1-2 plays a dominant role in promoting sporulation.

CYP2C19 polymorphisms and clopidogrel efficacy in the secondary prevention of ischemic stroke: a retrospective observational study

The relationship between cytochrome P450 2C19 (CYP2C19) polymorphisms and clopidogrel efficacy in patients with percutaneous coronary intervention (PCI) has been widely studied. However, the relationship between CYP2C19 polymorphisms and the response to clopidogrel in patients treated for ischemic stroke (IS) remains controversial. What's more, few data address the relevance of CYP2C19 polymorphisms in patients taking clopidogrel for secondary prevention of ischemic stroke. This study investigates whether carrying CYP2C19 loss-of-function (LOF) alleles affects the risk of recurrent stroke in IS patients. One hundred twenty-two IS patients were CYP2C19 genotype screened and enrolled in the study from January 2016 to December 2017. Those with stroke recurrence, stroke sequelae, or bleeding diseases were excluded. The remaining 89 patients were divided into the following 2 groups: non-carriers of CYP2C19 LOF alleles (n=38) and carriers (n=51) of CYP2C19 LOF alleles. The variables that could influence the rate of recurrent stroke were assessed in a multivariate analysis to determine the independent risk factors. The CYP2C19*2 and *3 alleles frequencies among the 122 patients were 31.97% and 4.10%, respectively. Carriers of LOF alleles had a more significant history of hypertension compared with non-carriers [n=43/51 (86.7%) versus n=23/38 (60.5%), P=0.01]. In addition, the inclusion rate of aspirin in discharge medication was significantly higher for carriers than for non-carriers [n=19/51 (37.3%) versus n=5/38 (13.2%), P=0.01]. CYP2C19 LOF alleles were significantly associated with an increased risk of recurrent stroke [odds ratio (OR): 7.586; 95% confidence interval (CI): 1.346-42.770, P=0.022]. CYP2C19 LOF alleles may increase the risk of recurrent IS. The polymorphisms of CYP2C19 may be predictors of a poor functional outcome in patients with recurrent stroke. Instead of clopidogrel, aspirin can be prescribed as a secondary preventative measure against stroke in carriers of CYP2C19 LOF alleles.

Abstract 11621: Effect of Genotype Guided Oral P2y12 Inhibitor Selection After Percutaneous Coronary Intervention: A Systematic Review and Meta-Analysis of Randomized Clinical Trials

Introduction: Clopidogrel is the most frequently used P2Y12 inhibitor as a component of the dual antiplatelet regimen in patients with acute coronary syndrome (ACS) undergoing percutaneous coronary intervention (PCI). Prior studies have shown the variable efficacy of clopidogrel due to genotypic differences in the CYP2C19 enzyme function, which converts clopidogrel to its active metabolite. Hypothesis: The aim of this meta-analysis is to evaluate the effectiveness of genotype testing-guided P2Y12 inhibitor prescription therapy to patients after PCI for ACS compared to non-genotype guided conventional treatment. Methods: A comprehensive literature search was performed in PubMed, Embase, and Cochrane to identify relevant trials. Summary effects were calculated using a DerSimonian and Laird random-effects model as odds ratio with 95% confidence intervals for all the clinical endpoints. Results: Seven studies with 9617 patients were included. Genotype-guided strategy arm included prasugrel or ticagrelor prescription to patients with loss of function (LOF) of CYP219 alleles (most commonly alleles being *2 and *3) and clopidogrel prescription to those without the LOF allele. The conventional arm included patients treated with clopidogrel without genotype testing. Comparison of genotype arm with conventional arm showed decreased major adverse cardiovascular events (MACE) (RR 0.72, 95% CI 0.55-0.95; p=0.02, I 2 =23), improved cardiovascular (CV) mortality (RR 0.62, 95% CI 0.43-0.90; p=0.01, I 2 =0) and reduced incidence of myocardial infarction (MI) (RR 0.57, 95% CI 0.41-0.97; p&lt; 0.01), and a similar stroke incidence (RR 0.65, 95% CI 0.35-1.21; p= 0.18). Regarding adverse events, the incidence of stent thrombosis was lower in the genotype arm than the conventional arm (RR 0.52, 95% CI 0.28-0.96; p=0.04). Conclusions: Our analysis illustrates the possible advantages of genotype-guided P2Y12 inhibitor prescription strategy compared to non-genotype-guided strategy with reductions in MACE, CV mortality, MI, and stent thrombosis. This analysis can be used as a stepping stone to conducting further trials determining the efficacy of this treatment strategy in various ACS subtypes.

Functional Roles of DDX41 Mutations in the Development of Myeloid Malignancies

▪DDX41 is a newly identified leukemia predisposition gene encoding an RNA helicase, whose germline mutations are tightly associated with late-onset myeloid malignancies. Importantly, germline DDX41 mutations were also found in as many as ~7 % of sporadic cases of high-risk MDS, conferring the largest germline risk for myeloid malignancies. In typical cases, a germline loss-of-function allele (most commonly p.A500fs or p.D140fs, depending on the ethnicity) is compounded by a somatic missense mutation affecting the helicase domain in the remaining allele (p.R525H). However, little is known about the molecular mechanism by which DDX41 mutations lead to myeloid neoplasms.To clarify the role of these distinct DDX41 alleles, we generated mice models carrying either or both of conditional/constitutive Ddx41 knock-out (KO) and conditional R525H knock-in (KI) alleles. BM-specific biallelic Ddx41 deletion using Vav1-Cre resulted in embryonic lethality, suggesting that Ddx41 is indispensable for normal hematopoietic development. Next, by crossing these mice and further breeding with Rosa26-CreERT2 transgenic mice, we engineered mice that were wild-type for Ddx41 (Ddx41+/+), heterozygous Ddx41 KO (Ddx41+/-), homozygous Ddx41 KO (Ddx41-/-), heterozygous for the Ddx41 R525H mutation (Ddx41R525H/+), or hemizygous for the Ddx41 R525H mutation (Ddx41R525H/-), in which expression of the mutant allele was induced by tamoxifen administration.First, we assessed cell intrinsic effects of these Ddx41 alleles, using noncompetitive transplantation experiments. Shortly after tamoxifen administration, most of the recipient mice that were reconstituted with BM from Ddx41-/- or Ddx41R525H/- mice died within a month after CreERT2 induction due to severe BM failure (BMF). None of the mice transplanted with BM from Ddx41+/+, Ddx41+/- or Ddx41R525H/+ mice developed myeloid neoplasms.We also assessed the reconstitution capacity of whole BM cells from different Ddx41 mutant mice in competitive transplantation experiments. The donor chimerism of Ddx41-/- or Ddx41R525H/- mice-derived cells in PB was markedly reduced compared to that of cells derived from Ddx41+/+ mice. In contrast, Ddx41+/- or Ddx41R525H/+ mice-derived cells showed no significant changes in competitive bone marrow reconstitution compared to Ddx41+/+ mice-derived cells. Notably, about half of the recipient mice died due to BMF when Ddx41R525H/--derived BM cells were co-transplanted with Ddx41+/--derived BM cells but not with wild-type BM cells, suggesting some non-cell autonomous effect of Ddx41R525H/- cells on Ddx41+/- cells.Transcriptome analysis of stem cells (Kit +Sca-1 -Lin low cells) from different Ddx41 mutant mice revealed significant changes in gene expression and splicing patterns in many genes in stem cells from all the mutant mice, with larger changes for Ddx41R525H/- than Ddx41+/- or Ddx41R525H/+ cells. Notably, Ddx41R525H/- -derived stem cells exhibited a significant upregulation of genes involved in innate immunity, including an upregulation of cGAS-STING innate immunity pathways, as well as an enhanced Trp53 pathway, whereas there was a downregulation of genes related to RNA metabolism and ribosome biogenesis. Proteomics analysis confirmed the significant downregulation of ribosomal proteins in hematopoietic cells derived from Ddx41R525H/- mice.In summary, our results revealed an essential role of Ddx41 in normal hematopoiesis. While both heterozygous Ddx41 KO and heterozygous R525H knock-in did not develop myeloid neoplasm, compound biallelic loss-of function and R525 alleles led to a compromised function of hematopoietic stem cells, which was evident from reduced competitive repopulation capacity and impaired hematopoietic differentiation, where activated innate immunity and impaired ribosome functions may play important roles. Their roles in myeloid neoplasms need further evaluation. DisclosuresNakagawa: Sumitomo Dainippon Pharma Oncology, Inc.: Research Funding. Kataoka: Celgene: Honoraria; Eisai: Honoraria; Astellas Pharma: Honoraria; Novartis: Honoraria; Chugai Pharmaceutical: Honoraria; AstraZeneca: Honoraria; Sumitomo Dainippon Pharma: Honoraria; Kyowa Kirin: Honoraria; Janssen Pharmaceutical: Honoraria; MSD: Honoraria; Takeda Pharmaceutical: Honoraria; Otsuka Pharmaceutical: Honoraria; Asahi Genomics: Current equity holder in publicly-traded company; Otsuka Pharmaceutical: Research Funding; Chordia Therapeutics: Research Funding; Chugai Pharmaceutical: Research Funding; Takeda Pharmaceutical: Research Funding; Bristol-Myers Squibb: Research Funding; Eisai: Other: Scholarship; Otsuka Pharmaceutical: Other: Scholarship; Ono Pharmaceutical: Other: Scholarship; Kyowa Kirin: Other: Scholarship; Shionogi: Other: Scholarship; Takeda Pharmaceutical: Other: Scholarship; Summitomo Dainippon Pharma: Other: Scholarship; Chugai Pharmaceutical: Other: Scholarship; Teijn Pharma: Other: Scholarship; Japan Blood Products Organization: Other: Scholarship; Mochida Pharmaceutical: Other: Scholarship; JCR Pharmaceuticals: Other: Scholarship; Genetic Alterations: Patents & Royalties: PD-L1 abnormalties . Ogawa: Ashahi Genomics: Current holder of individual stocks in a privately-held company; Otsuka Pharmaceutical Co., Ltd.: Research Funding; Eisai Co., Ltd.: Research Funding; Kan Research Laboratory, Inc.: Consultancy, Research Funding; Dainippon-Sumitomo Pharmaceutical, Inc.: Research Funding; ChordiaTherapeutics, Inc.: Consultancy, Research Funding.

Assessing Prevalence and Carrier Frequency of Succinic Semialdehyde Dehydrogenase Deficiency.

Pathogenic variants in ALDH5A1 cause succinic semialdehyde dehydrogenase (SSADH) deficiency, with >180 cases reported worldwide. However, a nonspecific neurologic presentation and inconsistent variant nomenclature have limited diagnoses. In this study, pathogenic variants in ALDH5A1 were curated and variant prevalence assessed in the Genome Aggregation Database (gnomAD) to determine a minimum carrier frequency and to estimate disease prevalence. Stringent population variant analysis, including 98 reported disease-associated ALDH5A1 variants, indicates a pan-ethnic carrier frequency of ∼1/340, supporting a prevalence of SSADH deficiency of ∼1/460 000 worldwide, with highest carrier frequencies observed in East Asian and South Asian populations. Because heterozygous loss of function alleles are rare in gnomAD and >60% of reported disease-causing variants were missense changes that were not present in gnomAD, the pan-ethnic carrier frequency for SSADH deficiency is likely not fully represented in this study. Additional analyses to investigate the potential impact of more common ALDH5A1 variants with reduced but not deficient enzyme activity, including analysis in diverse populations, are needed to fully assess the prevalence of this ultra-rare disease.

Clinical Outcomes After Percutaneous Coronary Intervention Over Time on the Basis of CYP2C19 Polymorphisms.

The aim of this study was to investigate the association between CYP2C19 gene polymorphisms and the risk of cardiovascular events in the early stage and subsequent period after percutaneous coronary intervention (PCI) among patients who received clopidogrel. Between October 2015 and January 2017, CYP2C19 genotyped patients who were treated with clopidogrel after PCI were enrolled in this study. Included patients were categorized as non-loss-of-function metabolizers, intermediate metabolizers, and poor metabolizers based on CYP2C19 genotype. The primary outcome was a composite of any-cause mortality, nonfatal myocardial infarction, nonfatal ischemic stroke, and stent thrombosis occurring during exposure to clopidogrel. The rates of clinical outcome events were compared between CYP2C19 phenotypes. Landmark analyses were processed at 90 days and 1 year post-PCI. Of 1341 patients, 161 (12.0%) had 2 copies of loss-of-function (LOF) alleles, 621(46.3%) had one LOF allele, and 559 (41.7%) had no LOF allele. At the 3-month follow-up, the primary outcome events were more frequent in carriers of 2 LOF alleles (5.6%) than in noncarriers (1.8%) [adjusted hazard ratio (HR) 2.944, 95% confidence interval, 1.184-7.321, P = 0.020). A similar finding was observed among in patients with acute coronary syndrome indications at the index PCI (adjusted HR 3.046, 95% confidence interval, 1.237-7.501, P = 0.015). These differences did not persist within the subsequent 9 months of follow-up, among either all comers or subjects with acute coronary syndrome. In conclusion, these data demonstrate a higher risk for ischemic events in patients with 2 CYP2C19 LOF alleles who are prescribed clopidogrel, seen at 3 months after PCI, that is not sustained for 12 months.

Leptin system loss of function in the absence of obesity in zebrafish.

The leptin system plays a crucial role in the regulation of appetite and energy homeostasis in vertebrates. While the phenotype of morbid obesity due to leptin (Lep) or leptin receptor (LEPR) loss of function is well established in mammals, evidence in fish is controversial, questioning the role of leptin as the vertebrate adipostat. Here we report on three (Lepr) loss of function (LOF) and one leptin loss of function alleles in zebrafish. In order to demonstrate that the Lepr LOF alleles cannot transduce a leptin signal, we measured socs3a transcription after i.p. leptin which is abolished by Lepr LOF. None of the Lepr/Lepa LOF alleles leads to obesity/a body growth phenotype. We explore possible reasons leading to the difference in published results and find that even slight changes in background genetics such as inbreeding siblings and cousins can lead to significant variance in growth.

Clopidogrel versus Ticagrelor in CYP2C19 Loss-of-Function Allele Noncarriers: A Real-World Study in China

This article compares the clinical outcomes of clopidogrel and ticagrelor in patients with acute coronary syndrome (ACS) without cytochrome P450 (CYP)2C19 loss-of-function (LOF) alleles and investigates whether clopidogrel could be an alternative P2Y12 inhibitor without increasing the risk of ischemic events. Patients were divided into the clopidogrel-treated group and the ticagrelor-treated group. Inverse probability of treatment weighting (IPTW) calculated by propensity scores was used to adjust confounding covariates. The primary outcome was major adverse cardiovascular or cerebrovascular events (MACCEs) within 12 months. The secondary outcomes were MACCEs plus unstable angina, and clinically significant bleeding events. Finally, 2,199 patients were included. Of them, 1,606 were treated with clopidogrel, and 593 were treated with ticagrelor. The mean age of the original cohort was 59.92 ± 9.81 years. During the 12-month follow-up period, MACCEs occurred in 89 patients (4.0%). No significant differences were observed in MACCEs (IPTW-adjusted hazard ratio [HR], 0.87; 95% confidence interval [CI], 0.65-1.18), MACCEs plus unstable angina (IPTW-adjusted HR, 1.20; 95% CI, 0.91-1.59), or clinically significant bleeding events (IPTW-adjusted HR, 0.81; 95% CI, 0.53-1.23) between the clopidogrel- and ticagrelor-treated groups. In patients with ACS without CYP2C19 LOF alleles, clopidogrel was not associated with a higher risk of MACCEs when compared with ticagrelor. The main findings of this study support use of clopidogrel in CYP2C19 LOF noncarriers as an alternative P2Y12 inhibitor, which may reduce medical expenses and adverse reactions caused by more potent P2Y12 inhibitors in these patients.

Allele mining in diverse accessions of tropical grasses to improve forage quality and reduce environmental impact.

The C4Urochloa species (syn. Brachiaria) and Megathyrsus maximus (syn. Panicum maximum) are used as pasture for cattle across vast areas in tropical agriculture systems in Africa and South America. A key target for variety improvement is forage quality: enhanced digestibility could decrease the amount of land required per unit production, and enhanced lipid content could decrease methane emissions from cattle. For these traits, loss-of-function (LOF) alleles in known gene targets are predicted to improve them, making a reverse genetics approach of allele mining feasible. We therefore set out to look for such alleles in diverse accessions of Urochloa species and Megathyrsus maximus from the genebank collection held at the CIAT. We studied allelic diversity of 20 target genes (11 for digestibility, nine for lipid content) in 104 accessions selected to represent genetic diversity and ploidy levels of U. brizantha, U. decumbens, U. humidicola, U. ruziziensis and M. maximum. We used RNA sequencing and then bait capture DNA sequencing to improve gene models in a U. ruziziensis reference genome to assign polymorphisms with high confidence. We found 953 non-synonymous polymorphisms across all genes and accessions; within these, we identified seven putative LOF alleles with high confidence, including those in the non-redundant SDP1 and BAHD01 genes present in diploid and tetraploid accessions. These LOF alleles could respectively confer increased lipid content and digestibility if incorporated into a breeding programme. We demonstrated a novel, effective approach to allele discovery in diverse accessions using a draft reference genome from a single species. We used this to find gene variants in a collection of tropical grasses that could help reduce the environmental impact of cattle production.

B-PO02-157 INCIDENCE OF DEVICE-RELATED THROMBOSIS IN WATCHMAN PATIENTS UNDERGOING A GENOTYPE-GUIDED ANTITHROMBOTIC STRATEGY

Loss of Function (LOF) polymorphisms of cytochrome P450 2C19 (CYP2C19) are associated with reduced hepatic bioactivation of clopidogrel. To report the incidence of device-related thrombosis (DRT) and thromboembolic (TE) events when an alternative to clopidogrel is prescribed in patients with a LOF allele of CYP2C19 gene. Among 1002 Watchman patients, 645 underwent CYP2C19 genetic testing; among patients with clopidogrel resistance, clopidogrel was replaced by either prasugrel (pilot cohort) or half dose novel oral anticoagulant (NOAC; Group 1) both in combination with aspirin. We compared the incidence of DRT/TE events among genotyped patients and a control group which received standard dual antiplatelet therapy (DAPT; Group 2; n=357). All reported events occurred during a timeframe between 45- and 180-day follow-up TEEs, when the genotype-guided or standard DAPT strategies were adopted. In the pilot cohort (n=244), bleeding events occurred in 10.2% of patients receiving prasugrel, leading to discontinuation of the prasugrel-based protocol. Among Group 1 patients (n=401), 25.7% were reduced metabolizers and clopidogrel was replaced with half dose NOAC. DRT was documented in 1 (0.2%) patient of Group 1 and 7 (1.96%) of Group 2 (log-rank p-value=0.021). The composite endpoint of DRT/TE events was significantly lower among patients receiving a genotype-guided strategy (0.75% vs. 3.1%; log-rank p-value=0.017). In Watchman patients, a genotype-based antithrombotic strategy with aspirin plus half dose NOAC in reduced clopidogrel metabolizers was superior to standard DAPT with respect to DRT/TE events.

Risk of stroke in CYP2C19 LoF polymorphism carrier coronary artery disease patients undergoing clopidogrel therapy: An ethnicity-based updated meta-analysis

BackgroundAntiplatelet agent clopidogrel has been widely used for stroke management for many years, although resistance to clopidogrel may increase the chance of stroke recurrence. CYP2C19 loss-of-function (LoF) polymorphism is assumed to be responsible for the poor metabolism of clopidogrel that ultimately turns to resistance. Previous publications could not provide firm evidence due to highly conflicting and heterogeneous outcomes. AimTo get clear evidence from an updated meta-analysis on CYP2C19 LoF polymorphism association with stroke risk in clopidogrel treated patients, this study has been performed. MethodsWe conducted a meta-analysis with 72 selected studies from authentic databases, including 40,035 coronary artery disease patients treated with clopidogrel. ResultsThis analysis showed that the worldwide carrier of one or more CYP2C19 LoF alleles had a significantly higher risk of stroke and composite events than the non-LoF carriers (RR=1.78, 95% CI=1.52-2.07, p<0.00001 and RR=1.39, 95% CI=1.26-1.54, p<0.00001, respectively). Besides, subgroup analysis showed that Asian CYP2C19 LoF carriers had a significantly increased risk of stroke (RR=1.91, 95% CI=1.60-2.28, p<0.00001) while the risk of composite events was significantly higher in all ethnic populations (Asian: RR=1.58, 95% CI=1.32-1.89, p<0.00001; Caucasian: RR=1.27, 95% CI=1.08-1.50, p=0.003; Hispanic and others: RR=1.21, 95% CI=1.09-1.34, p=0.0003). ConclusionOur meta-analysis confirmed that the presence of CYP2C19 LoF alleles increases the risk of stroke and composite events recurrence in the worldwide population, especially in Asians undergoing clopidogrel treatment. Alternative antiplatelet therapy should be investigated thoroughly for the intermediate and poor metabolizers.

Clopidogrel with aspirin in High-risk patients with Acute Non-disabling Cerebrovascular Events II (CHANCE-2): rationale and design of a multicentre randomised trial

BackgroundIn patients with a minor ischaemic stroke or transient ischaemic attack (TIA), separate trials have shown that dual antiplatelet therapy with clopidogrel plus aspirin (clopidogrel–aspirin) or ticagrelor plus aspirin (ticagrelor–aspirin)...

Implementation and clinical outcomes of pharmacogenetic CYP2C19 testing for clopidogrel therapy in real-world clinical practice.

Abstract Funding Acknowledgements Type of funding sources: None. BACKGROUNDCYP2C19 loss-of-function (LOF) polymorphisms are associated with adverse ischaemic events after PCI. The use of a point-of-care assay (POC) to routinely genotype patients immediately post PCI could rapidly identify patients at risk of adverse cardiac outcomes. PURPOSE To investigate the incidence of CYP2C19 polymorphisms (*2, *17) and 30-day MACE in patients presenting to catheter laboratory for PCI (See table 1).METHODS We performed a single centre prospective analysis of patients presenting to a cardiac catheterisation laboratory for percutaneous coronary intervention. Participants underwent prospective rapid point-of-care genotyping of CYP2C19 major alleles (2*,17*), using the SpartanRx PCR device via buccal swab sample. All patients provided written consent. RESULTS:A total of 120 tests were performed, 51 patients were normal allele carriers (*1), 31 patients were carriers of LOF alleles (*2) and 38 patients were carriers of gain of function alleles (*17). All tests results returned in one hour. Rate of dyslipidaemia was significantly different between three groups (55% vs. 63% vs. 36%; p = 0.050). A numerically higher proportion of LOF allele carriers received clopidogrel prior to undergoing pharmacogenetic testing but this was not statistically significant (52% vs. 35% vs. 34%; p = 0.09). Two cases of MACE at 30 day follow up occurred in the loss-of-function group. Both cases received clopidogrel.CONCLUSIONSWe have demonstrated that a rapid POC of CYP2C19 testing can take place in a real-world setting. Our incidence rate of LOF carriers is concordant with international published literature. We found 52% of LOF carriers were commenced on clopidogrel therapy prior to genetic analysis. Comparison of CPY2C19 Metabolisers genotype Loss of function normal Gain of function p values baseline characteristics age in years, median (range) 65 (43-82) 64 (43-85) 65 (42-89) 0.717 Male, N (%) 21 (68%) 43 (64%) 27 (71%) 0.198 Hypertensive, N (%) 16 (52%) 29 (57%) 24 (50%) 0.623 Dyslipidaemia. N (%) 17 (55%) 32 (63%) 14 (36%) 0.050 Indication, N (%) St-Elevation MI 12 (39%) 18 (35%) 11 (29%) 0.558 NSTEMI 5 (16%) 15 (29%) 14 (37%) 0.142 Unstable Angina 5 (16%) 7 (14%) 3 (8%) 0.518 Stable CAD 9 (29%) 11 (22%) 10 (26%) 0.731 Antiplatelet, N (%) Ticagrelor 15 (48%) 33 (65%) 25 (66%) 0.09 Clopidogrel 16 (52%) 18 (35%) 13 (34%) Complication, N (%) 30-day MACE 2 (6.5%) 0 0 0.01

Rare deleterious de novo missense variants in Rnf2/Ring2 are associated with a neurodevelopmental disorder with unique clinical features.

The Polycomb group (PcG) gene RNF2 (RING2) encodes a catalytic subunit of the Polycomb repressive complex 1 (PRC1), an evolutionarily conserved machinery that post-translationally modifies chromatin to maintain epigenetic transcriptional repressive states of target genes including Hox genes. Here, we describe two individuals, each with rare de novo missense variants in RNF2. Their phenotypes include intrauterine growth retardation, severe intellectual disabilities, behavioral problems, seizures, feeding difficulties and dysmorphic features. Population genomics data suggest that RNF2 is highly constrained for loss-of-function (LoF) and missense variants, and both p.R70H and p.S82R variants have not been reported to date. Structural analyses of the two alleles indicate that these changes likely impact the interaction between RNF2 and BMI1, another PRC1 subunit or its substrate Histone H2A, respectively. Finally, we provide functional data in Drosophila that these two missense variants behave as LoF alleles in vivo. The evidence provide support for deleterious alleles in RNF2 being associated with a new and recognizable genetic disorder. This tentative gene-disease association in addition to the 12 previously identified disorders caused by PcG genes attests to the importance of these chromatin regulators in Mendelian disorders.

Cost-effectiveness of CYP2C19 genotyping to guide antiplatelet therapy for acute minor stroke and high-risk transient ischemic attack

Dual antiplatelet therapy (DAPT) with clopidogrel plus aspirin within 48 h of acute minor strokes and transient ischemic attacks (TIAs) has been indicated to effectively reduce the rate of recurrent strokes. However, the efficacy of clopidogrel has been shown to be affected by cytochrome P450 2C19 (CYP2C19) polymorphisms. Patients carrying loss-of-function alleles (LoFAs) at a low risk of recurrence (ESRS < 3) cannot benefit from clopidogrel plus aspirin at all and may have an increased bleeding risk. In order to optimize antiplatelet therapy for these patients and avoid the waste of medical resources, it is important to identify the subgroups that genuinely benefit from DAPT with clopidogrel plus aspirin through CYP2C19 genotyping. This study sought to assess the cost-effectiveness of CYP2C19 genotyping to guide drug therapy for acute minor strokes or high-risk TIAs in China. A decision tree and Markov model were constructed to evaluate the cost-effectiveness of CYP2C19 genotyping. We used a healthcare payer perspective, and the primary outcomes included quality-adjusted life years (QALYs), costs and the incremental cost-effectiveness ratio (ICER). Sensitivity analyses were performed to evaluate the robustness of the results. CYP2C19 genotyping resulted in a lifetime gain of 0.031 QALYs at an additional cost of CNY 420.13 (US$ 59.85), yielding an ICER of CNY 13,552.74 (US$ 1930.59) per QALY gained. Probabilistic sensitivity analysis showed that genetic testing was more cost-effective in 95.7% of the simulations at the willingness-to-pay threshold of CNY 72,100 (GDP per capita, US$ 10,300) per QALY. Therefore, CYP2C19 genotyping to guide antiplatelet therapy for acute minor strokes and high-risk TIAs is highly cost-effective in China.

Field-based Screening and Haplotyping of J Locus for Long Juvenile Trait in Tropical Soybean Genotypes

Background: The long juvenile trait influences flowering time of soybean under tropical conditions. The trait ensures sufficient vegetative growth prior to flowering. The present study aimed at identifying tropical soybean genotypes with the long juvenile trait and harboring the loss-of-function alleles for the J gene and verifying the effect of loss-of-function alleles on the long juvenile trait. Methods: A total of 159 soybean genotypes were evaluated on the field for two years in tropical city of Sanya, Hainan Islands, China for days to beginning bloom (VE-R1), days to physiological maturity (VE-R7), nodes/plant, pods/plant and plant height at maturity. The full length of the J gene was cloned in the 159 soybean genotypes. The sequence data was subjected to haplotype analysis to determine the genotypes with the functional and loss of function alleles. Result: Significant differences (p less than 0.05) were observed among genotypes for all phenotypic traits. 53 genotypes were identified with delayed flowering based on the phenotypic data. Sequence comparison of the 159 genotypes identified 10 polymorphisms comprising 7 SNPs and three deletions in the coding sequences, the three deletions resulted in the loss of function of the J gene. Six genotypes with delayed flowering had the loss-of-function alleles of the J gene.