Inflammatory risk contributes to post-COVID endothelial dysfunction through anti-ACKR1 autoantibody.

Subclinical vascular impairment can be exacerbated in individuals who experience sustained inflammation after COVID-19 infection. Our study explores the prevalence and impact of autoantibodies on vascular dysfunction in healthy COVID-19 survivors, an area that remains inadequately investigated. Focusing on autoantibodies against the atypical chemokine receptor 1 (ACKR1), COVID-19 survivors demonstrated significantly elevated anti-ACKR1 autoantibodies, correlating with systemic cytokines, circulating damaged endothelial cells, and endothelial dysfunction. An independent cohort linked these autoantibodies to increased vascular disease outcomes during a median 6.7-yr follow-up. We analyzed a single-cell transcriptome atlas of endothelial cells from diverse mouse tissues, identifying enriched Ackr1 expressions in venous regions of the brain and soleus muscle vasculatures, which holds intriguing implications for tissue-specific venous thromboembolism manifestations reported in COVID-19. Functionally, purified immunoglobulin G (IgG) extracted from patient plasma did not trigger cell apoptosis or increase barrier permeability in human vein endothelial cells. Instead, plasma IgG enhanced antibody-dependent cellular cytotoxicity mediated by patient PBMCs, a phenomenon alleviated by blocking peptide or liposome ACKR1 recombinant protein. The blocking peptide uncovered that purified IgG from COVID-19 survivors possessed potential epitopes in the N-terminal extracellular domain of ACKR1, which effectively averted antibody-dependent cellular cytotoxicity. Our findings offer insights into therapeutic development to mitigate autoantibody reactivity in blood vessels in chronic inflammation.

A five-safes approach to a secure and scalable genomics data repository.

Genomic researchers increasingly utilize commercial cloud service providers (CSPs) to manage data and analytics needs. CSPs allow researchers to grow Information Technology (IT) infrastructure on demand to overcome bottlenecks when combining large datasets. However, without adequate security controls, the risk of unauthorized access may be higher for data stored on the cloud. Additionally, regulators are mandating data access patterns and specific security protocols for the storage and use of genomic data. While CSP provides tools for security and regulatory compliance, building the necessary controls required for cloud solutions is not trivial. Research Assets Provisioning and Tracking Online Repository (RAPTOR) by the Genome Institute of Singapore is a cloud-native genomics data repository and analytics platform that implements a "five-safes" framework to provide security and governance controls to data contributors and users, leveraging CSP for sharing and analysis of genomic datasets without the risk of security breaches or running afoul of regulations.

The Singapore National Precision Medicine Strategy.

Precision medicine promises to transform healthcare for groups and individuals through early disease detection, refining diagnoses and tailoring treatments. Analysis of large-scale genomic-phenotypic databases is a critical enabler of precision medicine. Although Asia is home to 60% of the world's population, many Asian ancestries are under-represented in existing databases, leading to missed opportunities for new discoveries, particularly for diseases most relevant for these populations. The Singapore National Precision Medicine initiative is a whole-of-government 10-year initiative aiming to generate precision medicine data of up to one million individuals, integrating genomic, lifestyle, health, social and environmental data. Beyond technologies, routine adoption of precision medicine in clinical practice requires social, ethical, legal and regulatory barriers to be addressed. Identifying driver use cases in which precision medicine results in standardized changes to clinical workflows or improvements in population health, coupled with health economic analysis to demonstrate value-based healthcare, is a vital prerequisite for responsible health system adoption.

Analysis of clinically relevant variants from ancestrally diverse Asian genomes.

Asian populations are under-represented in human genomics research. Here, we characterize clinically significant genetic variation in 9051 genomes representing East Asian, South Asian, and severely under-represented Austronesian-speaking Southeast Asian ancestries. We observe disparate genetic risk burden attributable to ancestry-specific recurrent variants and identify individuals with variants specific to ancestries discordant to their self-reported ethnicity, mostly due to cryptic admixture. About 27% of severe recessive disorder genes with appreciable carrier frequencies in Asians are missed by carrier screening panels, and we estimate 0.5% Asian couples at-risk of having an affected child. Prevalence of medically-actionable variant carriers is 3.4% and a further 1.6% harbour variants with potential for pathogenic classification upon additional clinical/experimental evidence. We profile 23 pharmacogenes with high-confidence gene-drug associations and find 22.4% of Asians at-risk of Centers for Disease Control and Prevention Tier 1 genetic conditions concurrently harbour pharmacogenetic variants with actionable phenotypes, highlighting the benefits of pre-emptive pharmacogenomics. Our findings illuminate the diversity in genetic disease epidemiology and opportunities for precision medicine for a large, diverse Asian population.

TPMT and NUDT15 testing for thiopurine therapy: A major tertiary hospital experience and lessons learned.

Variants in thiopurine methyltransferase (TPMT) and nudix hydrolase 15 (NUDT15) are associated with an accumulation of cytotoxic metabolites leading to increased risk of drug-related toxicity with standard doses of thiopurine drugs. We established TPMT and NUDT15 genetic testing for clinical use and evaluated the utilization, service outcomes and potential value of multi-gene PGx testing for 210 patients that underwent pharmacogenetics (PGx) testing for thiopurine therapy with the aim to optimize service delivery for future prescribing. The test was most commonly ordered for Gastroenterology (40.0%) and Neurology (31.4%), with an average turnaround time of 2 days. Following testing, 24.3% patients were identified as intermediate or poor metabolizers, resulting in 51 recommendations for a drug or dose change in thiopurine therapy, which were implemented in 28 (54.9%) patients. In the remaining patients, 14 were not adjusted and 9 had no data available. Focusing on drug gene interactions available for testing in our laboratory, multi-gene PGx results would present opportunities for treatment optimization for at least 33.8% of these patients who were on 2 or more concurrent medications with actionable PGx guidance. However, the use of PGx panel testing in clinical practice will require the development of guidelines and education as revealed by a survey with the test providers. The evaluation demonstrated successful implementation of single gene PGx testing and this experience guides the transition to a pre-emptive multi-gene testing approach that provides the opportunity to improve clinical care.

Self-Reported Frailty Screening Tools: Comparing Construct Validity of the Frailty Phenotype Questionnaire and FRAIL.

OBJECTIVES: We examined the construct validity of 2 self-reported frailty questionnaires, the Frailty Phenotype Questionnaire (FPQ) and FRAIL, against the Cardiovascular Health Study frailty phenotype (CHS-FP). DESIGN: Cross-sectional data analysis of longitudinal prospective cohort study. SETTINGS AND PARTICIPANTS: We included data from 230 older adults (mean age: 67.2 ± 7.4 years) from the "Longitudinal Assessment of Biomarkers for characterization of early Sarcopenia and Osteosarcopenic Obesity in predicting frailty and functional decline in community-dwelling Asian older adults Study" (GeriLABS 2) recruited between December 2017 and March 2019. METHODS: We compared area under receiver operating characteristic curves (AUC), agreement, correlation, and predictive validity against outcome measures [Short Physical Performance Battery, 5 times repeat chair stand (RCS-5), Frenchay activities index, International Physical Activity Questionnaire, life-space assessment, Social Functioning Scale 8 (SFS-8), EuroQol-5 dimensions (utility value)] using logistic regression adjusted for age, gender, and vascular risk factors. We examined concurrent validity across robust versus prefrail/frail for inflammatory blood biomarkers [tumor necrosis factor receptor 1 and C-reactive protein (CRP)] and dual-energy x-ray absorptiometry body composition [bone mineral density (BMD); appendicular lean mass index (ALMI), and fat mass index (FMI)]. RESULTS: Prevalence of prefrail/frail was 25.7%, 14.8%, and 48.3% for FPQ, FRAIL, and CHS-FP, respectively. Compared with FRAIL, FPQ had better diagnostic performance (AUC = 0.617 vs 0.531, P = .002; sensitivity = 37.8% vs 18.0%; specificity = 85.6% vs 88.2%) and agreement (AC1-Stat = 0.303 vs 0.197). FPQ showed good predictive validity [RCS-5: odds ratio (OR) 2.38; 95% CI: 1.17-4.86; International Physical Activity Questionnaire: OR 3.62; 95% CI:1.78-7.34; SFS-8: OR 2.11; 95% CI: 1.64-5.89 vs FRAIL: all P > .05]. Only FRAIL showed concurrent validity for CRP, compared with both FPQ and FRAIL for TNF-R1. FRAIL showed better concurrent validity for BMD, FMI, and possibly ALMI, unlike FPQ (all P > .05). CONCLUSIONS AND IMPLICATIONS: Our results support complementary validity of FPQ and FRAIL in independent community-dwelling older adults. FPQ has increased case detection sensitivity with good predictive validity, whereas FRAIL demonstrates concurrent validity for inflammation and body composition. With better diagnostic performance and validity for blood biomarkers and clinical outcomes, FPQ has utility for early frailty detection in the community setting.

Compound Heterozygous Factor VII Deficiency c.1025G>A p.(Arg342Gln) With Novel Missense Variant c.194C>G p.(Ala65Gly).

Factor VII (FVII) deficiency manifests as prolonged prothrombin time (PT) and reduced FVII activity. We report a case of an asymptomatic 60-year-old gentleman with discrepancies in PT and FVII coagulant activity levels (FVII:C) on three different thromboplastin reagents used. Further sequence analysis on genomic DNA showed double heterozygosity for c.1025G>A p.Arg342Gln and c.194C>G p.Ala65Gly in the F7 gene. To date, p.Ala65Gly in exon 2 of the F7 gene represents a novel variant in patients with FVII deficiency and is classified as likely pathogenic. Computational prediction tools support a deleterious effect on the gene. The genotype-phenotype association and the clinical significance of this exon 2 missense variant is proposed in this case report.

Sex Modifies the Association of Fibroblast Growth Factor 21 With Subclinical Carotid Atherosclerosis.

Background and Aims: Fibroblast growth factor 21 (FGF21), an emerging metabolic hepatokine, is associated with atherosclerosis. An interaction with sex has been described in various populations. We aimed to study whether sex modulates the relationship between FGF21 and subclinical carotid atherosclerosis in a diabetes-enriched multiethnic population of Singapore. We explore differences in intermediary mechanisms, in terms of hypertension, lipids, and inflammation, between FGF21 and atherosclerosis. Methods: We recruited 425 individuals from a single diabetes center in Singapore, and demographics, anthropometry, metabolic profile, FGF21, and carotid ultrasonography were performed. Multivariable logistic regression models were used to study the association between subclinical atherosclerosis and FGF21 adjusting for age, ethnicity, body mass index (BMI), hemoglobin A1c (HbA1c), systolic and diastolic blood pressures, and low-density lipoprotein (LDL)-cholesterol separately for males and females as two groups after an interaction test. Results: An interaction test assessing interaction by sex on the relationship between subclinical atherosclerosis and FGF21 showed a significant interaction with sex (Pinteraction = 0.033). In the female subgroup, significant independent associations of standardized lnFGF21 with subclinical atherosclerosis were seen, with 1 SD increment in lnFGF21 being associated with 1.48-fold (95% CI: 1.03, 2.12; p = 0.036) increase in risk. In the male subgroup, the association of subclinical atherosclerosis with standardized lnFGF21 was not significant [odds ratio (OR) (95% CI): 0.90 (0.63, 1.28); p = 0.553]. We found sex interactions with pulse pressure being significantly associated in females only and triglycerides and C-reactive protein being associated with males only. Conclusion: FGF21 is positively associated with subclinical carotid atherosclerosis in women, but not in men. The sex-racial patterns in the mechanisms by which FGF21 causes subclinical atherosclerosis needs to be explored in larger population-based studies and mechanistically studied in greater detail.

Clinical implementation of plasma EGFR T790M testing using droplet digital PCR in TKI-resistant NSCLC patients.

BACKGROUND: Majority of non-small cell lung cancer (NSCLC) patients progressed on epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) due to acquired T790M mutation. Blood sample is increasingly used in clinical setting for EGFR T790M detection and our laboratory employed the droplet digital PCR (ddPCR) methodology for testing. This study investigated the positive rate, specimen type for rebiopsy and clinical impact of blood-based EGFR T790M testing. METHODS: We retrospectively evaluated clinical samples that underwent plasma EGFR T790M testing in TTSH Molecular Diagnostic Laboratory from August 2017 to September 2019. Data on diagnosis, EGFR activating and T790M mutations, and treatment strategies were recorded. RESULTS: A total of 104 progressive NSCLC cases were included in this study. Overall, 46 patients (44.2%) were tested T790M positive, and 47.8% of these tested positive had low levels (defined as ≤3% fractional abundance and <50 copies/mL plasma), which may be missed by the conventional methods with lower sensitivity. Of these tested with low T790M abundance, 77.3% subsequently received osimertinib. Activating mutations were not detected in 42 (40.4%) cases, indicating that the tumors were not actively shedding ctDNA. Among these, 24 patients underwent repeat testing with tissue or blood specimens. Thirteen patients were subsequently tested T790M positive and 12 of them switched treatment to osimertinib. The recommendation to repeat testing with a different biopsy or after a suitable interval increased the overall positive rate to 56.7% (59/104). CONCLUSION: The use of a highly sensitive platform such as ddPCR for the detection of low abundance T790M, and the approach of repeat testing in cases with insufficient ctDNA increased the positive rate. This in turn identified more patients who are eligible for targeted therapy.

Validation of a next generation sequencing assay for BRCA1, BRCA2, CHEK2 and PALB2 genetic testing.

BRCA1, BRCA2, CHEK2 and PALB2 genes are associated with hereditary breast and ovarian cancer syndrome. Genetic testing of these genes is of increasing importance to guide therapeutic and management decisions. In this study, we evaluated the performance of a next generation sequencing (NGS) assay for the complete analysis of BRCA1, BRCA2, CHEK2 and PALB2 genes using Agilent's SureMASTR BRCA Screen that enabled the detection of single nucleotide variants (SNVs), small insertions/deletions (indels) and copy number variations (CNVs) in a single-tube PCR based library preparation. The results showed 100% sensitivity and specificity on a set of 52 known samples from de-identified patients and external quality assessment program. A concordance rate of 87.5% was achieved in the comparison of variant classification with the external laboratories. The high accuracy of the assay supports the use of SureMASTR BRCA Screen in clinical diagnostic laboratories (SureMASTR BRCA Screen is for research use only, not for use in diagnostic procedures).

Impact of Vitamin E supplementation on vascular function in haptoglobin genotype stratified diabetes patients (EVAS Trial): a randomised controlled trial.

AIMS: Vitamin E (Vit-E) may preferentially improve cardiovascular risk in haptoglobin 2-2 (Hp2-2) genotype diabetes individuals. We studied the impact of Vit-E supplementation on vascular function in diabetes individuals stratified by haptoglobin genotype in Singapore. METHODS: In this 24-week, double blind, placebo-controlled RCT, we recruited 187 subjects (101 Hp2-2, 86 non-Hp2-2). INTERVENTION: alpha-tocopherol-400 IU. PRIMARY OUTCOME: Change in EndoPAT-derived reactive-hyperaemia index (RHI) and augmentation index (AIx); Secondary Outcomes: Pulse-Wave velocity (Sphygmocor-PWV), carotid intima media thickness (CIMT), inflammation (hsCRP), derivatives of reactive-oxygen metabolites (dROMs), biological antioxidant-potential (BAPs), HbA1c, LDL-C, HDL-C and oxidised LDL-C (ox-LDL). RESULTS: Overall, with Vit-E supplementation no significant change in RHI, PWV, CIMT, hsCRP, dROMS, BAPs, HDL-C and HbA1c was observed (p > 0.05); an increase in LDL-C with concomitant decrease in ox-LDL, and incidentally increase in eGFR was observed (p < 0.05). No interaction effect with haptoglobin genotype was seen for all outcomes (p > 0.05). Subgroup analysis: In the non-Hp-2-2 group, Vit-E supplementation led to a higher EndoPAT-derived AIx, accompanied by higher LDL and ox-LDL concentrations (p < 0.05); Hp2-2 group: Vit-E supplementation led to higher eGFR when compared to the non-Hp2-2 group (exploratory) (p < 0.05). We observed an interaction effect for baseline haptoglobin concentration (threshold > 119 mg/dl) with intervention in terms of increased EndoPAT-derived AIx in the Hp > 119 mg/dl group whereas no change in the group with Hp ≤ 119 mg/dl. CONCLUSION: Vit-E supplementation did not show any preferential benefit or deleterious effect on vascular function in Hp2-2 diabetes subjects in Singapore. A possible deleterious effect of an increase in arterial stiffness in individuals with Hp > 119 mg/dl was observed. Future studies should consider personalisation based on baseline Hp concentrations in patients with T2DM rather than just Hp2-2 genotype to evaluate impact on the detailed lipid pathways, cardiac and renal physiology. The impact of ethnic differences needs to be explored in greater details.

CYP2C19 phenotype in South-East Asian Acute Coronary Syndrome patients and impact on major adverse cardiovascular events.

WHAT IS KNOWN AND OBJECTIVE: Several Caucasian cohort studies have associated at least one loss-of-function CYP2C19 on Clopidogrel (LoF-Clopidogrel) with major adverse cardiovascular events (MACE), and only a couple have used Clinical Pharmacogenetics Implementation Consortium (CPIC® ) phenotype grouping to study the associations. We primarily aimed to study the impact of use of platelet reactivity testing to escalate antiplatelet therapy and secondarily to study the association of CPIC phenotype with MACE outcomes in South-East Asian Acute Coronary Syndrome (ACS) subjects. METHOD: A retrospective genotype study was performed on 238 percutaneous coronary intervention subjects, originally planned for escalation of antiplatelets using platelet reactivity testing. RESULTS AND DISCUSSION: There was no difference in MACE between the switched and unswitched groups; however, 'all bleeds' and 'clinically significant bleeds' (CSB) were statistically higher in the patients who were switched to prasugrel. The subgroup of patients who remained on clopidogrel (n = 199) were analysed using phenotype categories and MACE. Eleven percent (11.4%) of CYP2C19 poor metabolizers (PM) suffered MACE, compared with 1.3% of extensive metabolizers (EM). LoF-Clopidogrel patients are significantly more likely to experience MACE compared with non-LoF subjects (8.0% vs 5.4%, P: .041). WHAT IS NEW AND CONCLUSION: In our multivariate analysis, LoF-Clopidogrel, malay ethnicity, diabetics and use of proton pump inhibitors were independent predictors of MACE. There were numerically more bleeds in LoF subjects who were on prasugrel compared with Clopidogrel (23.5% vs 11%, P = .082). Our data corroborate with current findings on platelet reactivity testing, suggesting that the assay would not be sensitive enough to pick up sufficient 'at-risk' subjects as compared to the use of CYP2C19 genotyping.

Large-Scale Whole-Genome Sequencing of Three Diverse Asian Populations in Singapore.

Underrepresentation of Asian genomes has hindered population and medical genetics research on Asians, leading to population disparities in precision medicine. By whole-genome sequencing of 4,810 Singapore Chinese, Malays, and Indians, we found 98.3 million SNPs and small insertions or deletions, over half of which are novel. Population structure analysis demonstrated great representation of Asian genetic diversity by three ethnicities in Singapore and revealed a Malay-related novel ancestry component. Furthermore, demographic inference suggested that Malays split from Chinese ∼24,800 years ago and experienced significant admixture with East Asians ∼1,700 years ago, coinciding with the Austronesian expansion. Additionally, we identified 20 candidate loci for natural selection, 14 of which harbored robust associations with complex traits and diseases. Finally, we show that our data can substantially improve genotype imputation in diverse Asian and Oceanian populations. These results highlight the value of our data as a resource to empower human genetics discovery across broad geographic regions.

Non-invasive detection of actionable mutations in advanced non-small-cell lung cancer using targeted sequencing of circulating tumor DNA.

OBJECTIVE: To evaluate the feasibility of detecting actionable gene mutations in circulating tumor DNA (ctDNA) in patients with advanced non-small-cell lung cancer (NSCLC) using targeted next-generation sequencing (NGS). MATERIALS AND METHODS: In total 50 plasma samples from patients newly diagnosed with advanced NSCLC or resistant to first-line tyrosine kinase inhibitors (TKIs) were subjected to deep sequencing on a seven-gene panel (BRAF, EGFR, ERBB2, KRAS, NRAS, PIK3CA, PTEN) incorporated with molecular barcodes to improve accuracy in variant detection. When possible, results were compared with those from matched tissue samples. RESULTS: At least one alteration in the ctDNA was detected in 44 out of 50 patients (88%); EGFR was the most frequently mutated gene. Half the total number of patients (50%, 25 of 50) had at least one actionable genetic alteration with targeted therapies available for treatment. Our results showed a high concordance rate of 81% in detection of EGFR mutation between 26 matched tissue and plasma samples. For progressive patients, from whom tissue is mostly unavailable, the resistant EGFR T790 M mutation was validated using the droplet digital polymerase chain reaction (ddPCR), yielding a concordance of 92% between alternative platforms. CONCLUSION: Our study demonstrated that therapeutically actionable mutations can be detected with high accuracy in ctDNA using NGS. This promising approach offers alternative and non-invasive diagnostic methods for treatment guidance and clinical monitoring.

Patient with multiple acyl-CoA dehydrogenase deficiency disease and ETFDH mutations benefits from riboflavin therapy: a case report.

BACKGROUND: Lipid storage myopathy (LSM) is a diverse group of lipid metabolic disorders with great variations in the clinical phenotype and age of onset. Classical multiple acyl-CoA dehydrogenase deficiency (MADD) is known to occur secondary to mutations in electron transfer flavoprotein dehydrogenase (ETFDH) gene. Whole exome sequencing (WES) with clinical correlations can be useful in identifying genomic alterations for targeted therapy. CASE PRESENTATION: We report a patient presented with severe muscle weakness and exercise intolerance, suggestive of LSM. Diagnostic testing demonstrated lipid accumulation in muscle fibres and elevated plasma acyl carnitine levels. Exome sequencing of the proband and two of his unaffected siblings revealed compound heterozygous mutations, c.250G > A (p.Ala84Thr) and c.770A > G (p.Tyr257Cys) in the ETFDH gene as the probable causative mutations. In addition, a previously unreported variant c.1042C > T (p.Arg348Trp) in ACOT11 gene was found. This missense variant was predicted to be deleterious but its association with lipid storage in muscle is unclear. The diagnosis of MADD was established and the patient was treated with riboflavin which resulted in rapid clinical and biochemical improvement. CONCLUSIONS: Our findings support the role of WES as an effective tool in the diagnosis of highly heterogeneous disease and this has important implications in the therapeutic strategy of LSM treatment.

Endothelial cell apoptosis correlates with low haptoglobin concentrations in diabetes.

OBJECTIVE: The haptoglobin 2-2 genotype is associated with lower haptoglobin concentrations and atherosclerosis in diabetes. Endothelial cell apoptosis contributes significantly to atherosclerosis. We studied endothelial cell apoptosis in diabetes patients with haptoglobin 2-2 and non-haptoglobin 2-2 genotype. Approach and results: We pooled plasma from 10 patients with haptoglobin 2-2 and non-haptoglobin 2-2 genotype and quantified endothelial cell apoptosis using a hemodynamic lab-on-chip system. Then, we conducted similar experiments on individual diabetes plasma samples with the haptoglobin 2-2 ( n = 20) and non-haptoglobin 2-2 genotype ( n = 20). Haptoglobin beta concentrations were measured by Western blot analysis. We looked for association with demographic, metabolic variables, inflammation and oxidative stress. In pooled plasma, endothelial cell apoptosis was higher in haptoglobin 2-2 group (haptoglobin 2-2: 23.18% vs non-haptoglobin 2-2:15.32%). In individual samples, univariate analysis showed that endothelial cell apoptosis correlated with haptoglobin beta concentration [ ß = -10.29 (95% confidence interval: -13.44, -7.14), p < 0.001] and total haptoglobin concentration [ ß = -0.03 (95% confidence interval: -0.05, -0.002), p = 0.03]. After multivariable analysis, only haptoglobin beta concentrations remained significant [ ß = -9.24 (95% confidence interval: -13.10, -5.37), p < 0.001]. The interaction term between haptoglobin genotypes and haptoglobin beta was not significant ( p > 0.05). CONCLUSION: These results show that regardless of the haptoglobin genotype, haptoglobin is associated with prevention of endothelial cell apoptosis in diabetes.

NLRP1, PTPN22 and PADI4 gene polymorphisms and rheumatoid arthritis in ACPA-positive Singaporean Chinese.

Studies have shown that the genetic risk factors for rheumatoid arthritis (RA) differ substantially between Asian and Caucasian populations. Even among Asian populations, the genetic contributions of NLRP1, PTPN22 and PADI4 have been controversial. Consequently, we sought to address these separate findings and determine whether any of these proposed risk variants are associated with RA susceptibility, onset, DAS activity and erosion in a Singaporean Chinese cohort. We genotyped five SNPs within NLRP1 (rs878329 and rs6502867), PTPN22 (rs2488457 and rs6665194), and PADI4 (rs2240340) in 500 anti-cyclic citrullinated peptide antibody-positive (ACPA) patients with RA and 500 healthy controls using TaqMan assays. The CC genotype of NLRP1 rs878329 and TT genotype of PADI4 rs2240340 were associated with RA susceptibility. The risk association of the T allele of PADI4 rs2240340 with RA was confirmed through a meta-analysis based on previous reports in Asian populations. The GG genotype of PTPN22 rs6665194 (-3508A>G) was associated with significantly reduced risk of RA. No significant association was found for NLRP1 rs6502867 T/C and PTPN22 rs2488457 G/C polymorphisms. None of the five SNPs was associated with RA's clinical features. This work supports the association of the T allele of PADI4 rs2240340 with RA in Asians. The roles of NLRP1 rs878329 G/C and PTPN22 rs6665194 A/G polymorphisms were demonstrated for the first time. We also propose rs6665194 to be a promising candidate for RA risk evaluation between ethnicities.

Analysis of Genetic Variation in CYP450 Genes for Clinical Implementation.

BACKGROUND: Genetic determinants of drug response remain stable throughout life and offer great promise to patient-tailored drug therapy. The adoption of pharmacogenetic (PGx) testing in patient care requires accurate, cost effective and rapid genotyping with clear guidance on the use of the results. Hence, we evaluated a 32 SNPs panel for implementing PGx testing in clinical laboratories. METHODS: We designed a 32-SNP panel for PGx testing in clinical laboratories. The variants were selected using the clinical annotations of the Pharmacogenomics Knowledgebase (PharmGKB) and include polymorphisms of CYP2C9, CYP2C19, CYP2D6, CYP3A5 and VKORC1 genes. The CYP2D6 gene allele quantification was determined simultaneously with TaqMan copy number assays targeting intron 2 and exon 9 regions. The genotyping results showed high call rate accuracy according to concordance with genotypes identified by independent analyses on Sequenome massarray and droplet digital PCR. Furthermore, 506 genomic samples across three major ethnic groups of Singapore (Malay, Indian and Chinese) were analysed on our workflow. RESULTS: We found that 98% of our study subjects carry one or more CPIC actionable variants. The major alleles detected include CYP2C9*3, CYP2C19*2, CYP2D6*10, CYP2D6*36, CYP2D6*41, CYP3A5*3 and VKORC1*2. These translate into a high percentage of intermediate (IM) and poor metabolizer (PM) phenotypes for these genes in our population. CONCLUSION: Genotyping may be useful to identify patients who are prone to drug toxicity with standard doses of drug therapy in our population. The simplicity and robustness of this PGx panel is highly suitable for use in a clinical laboratory.