Oleg N. Suchalko

Background: Personalized pharmacotherapy requires systematic consideration of genetic factors influencing drug efficacy and safety. The accumulation of large-scale whole-exome sequencing (WES) resources provides an opportunity to assess population frequencies of clinically significant pharmacogenetic variants; however, the applicability of exome-based pharmacogenomics across populations, particularly those underrepresented in existing reference datasets, requires further evaluation. Materials and methods: A retrospective analysis of 6102 anonymized sequencing datasets obtained between 2020 and 2025 was performed using the DNBSEQ-G400 (MGI) platform and Agilent SureSelect Human All Exon v6/v7/v8 enrichment kits. SNV and indel detection, CNV analysis, high-resolution HLA typing, and diplotype assignment for key pharmacogenes were conducted. Pharmacogenomic annotations were derived from ClinPGx (formerly PharmGKB) (levels of evidence 1 A–2B), CPIC, and PharmVar. Haplotype phasing (SHAPEIT5), genotype imputation (IMPUTE5), and phased linkage disequilibrium analysis were performed using a reference panel of 814 Russian whole-genome sequences generated with the same bioinformatic workflow to assess the feasibility of reconstructing clinically relevant non-coding pharmacogenetic variants not captured by WES. Results: WES reliably detected 33 of 34 Very Important Pharmacogenes (VIPs), allowing determination of allele frequencies, metabolizer statuses for 13 VIPs, and HLA diversity. The highest allelic and phenotypic variability was observed in CYP2D6 , CYP2C19 , and CYP2B6 . A total of 663 ClinPGx annotations were identified, predominantly related to drug metabolism (50.38%) and toxicity (29.56%), involving psychotropic drugs, anticoagulants, statins, opioid analgesics, antineoplastic agents, and immunosuppressants. However, at least 32 drugs require assessment of non-coding pharmacogenetic variants or accurate CYP2D6 copy number determination – both beyond standard WES capability. Phased haplotype analysis demonstrated that haplotype-based imputation was feasible for only a limited subset of clinically relevant non-coding pharmacogenetic variants, whereas the majority lacked informative exonic proxies required for accurate inference from WES data. Conclusion: These findings represent one of the largest reference resource to date of pharmacogenetically significant variant and HLA allele frequencies in the Russian population. The results confirm WES as a robust and scalable platform for population-level pharmacogenomic screening and broader clinical genomic applications, while demonstrating that haplotype-based imputation cannot fully overcome the intrinsic limitations of exome sequencing for comprehensive pharmacogenomic profiling. Selection of genomic testing strategies should balance diagnostic completeness, analytical complexity, cost, scalability, and long-term potential for reanalysis.

Adverse drug reactions (ADRs) remain a major public health issue, and genetic factors contribute importantly to interindividual variability in drug response. Pharmacogenetic testing helps reduce ADR risk by optimizing drug selection and dosage, particularly in monogenic disorders. Whole-exome sequencing of 6739 samples from the Russian population was performed on the DNBSEQ-G400 platform (MGI). Variants in 48 genes were examined, focusing on inherited arrhythmias, enzyme deficiencies (Glucose-6-Phosphate Dehydrogenase Deficiency [G6PDD], Porphyrias), Dravet Syndrome (DS) and Malignant Hyperthermia (MH). Variants reported as pathogenic (P), likely pathogenic (LP), or variants of uncertain significance (VUS) in ClinVar were manually re-evaluated using ACMG criteria. A total of 75 unique variants in 18 genes were observed in 119 individuals (1.77%), including 21 carriers and 13 women with a G6PD mutation. Of these, 44 variants were classified as P, 24 as LP, and 7 as VUS. Missense variants accounted for the largest proportion (73.33%). The most affected genes were KCNQ1 (24/119), which exhibited the highest number of unique variants (18), G6PD (20/119), SCN1A (15/119), and RYR1 (14/119). Regarding associated conditions, mutations linked to arrhythmias were found in 51 individuals, MH in 27, G6PDD in 20, DS in 15, and Porphyrias in 6. Integrating common and rare clinically actionable genetic variants with attention to penetrance and clinical validity may improve medication safety, reduce preventable ADRs, and enhance personalized pharmacotherapy.

Background: Direct oral anticoagulants (DOACs) are first-line medications for stroke prevention in non-valvular atrial fibrillation (AF). However, variability in drug response poses risks of hemorrhagic or thromboembolic events. Objectives: Although genetic influences on DOACs safety are increasingly recognized, robust evidence directly linking specific polymorphisms to bleeding risk remains limited. Design: Multi-center observational case–control study including exome-wide association analysis of 196 non-valvular AF patients treated with rivaroxaban or apixaban, comprising 97 with bleeding complications and 99 without. Methods: DOAC plasma concentrations, urinary 6-β-hydroxycortisol and cortisol levels were measured for CYP3A4 phenotyping. Sequencing was performed on the DNBSEQ G-400 platform. Single-nucleotide variant (SNV) associations with bleeding risk were assessed using logistic regression with additive, dominant, and recessive genetic models. Polygenic risk scores (PRSs) were calculated to evaluate cumulative genetic effects. Results: No SNVs reached Bonferroni-corrected significance under any model. PRSs showed weak predictive ability for bleeding with apixaban. For rivaroxaban, regression indicated that ln C _{ss min} /D + 1 index increased with PRS, age, and 6-β-hydroxycortisol/cortisol ratio, but decreased with higher 6-β-hydroxycortisol and coronary heart disease presence. No statistically significant differences were found for the PharmGKB Level 3 variants rs1045642 (rivaroxaban) and rs2231142 (apixaban). Trends toward statistical significance were observed for the rs2472304-G variant in rivaroxaban users, rs6977165-C in apixaban users, and for the CYP3A4*1/*36 diplotype. Conclusion: Residual equilibrium concentration of DOACs, including dose-adjusted, did not independently predict bleeding risk in non-valvular AF patients. Variants rs2472304 and rs6977165 may warrant further investigation as potential contributors to bleeding risk.

Auditory neuropathy spectrum disorder (ANSD) is often missed by standard hearing tests, accounting for up to 10% of hearing impairments (HI) and commonly linked to variants in 23 genes. We assessed 122 children with HI, including 102 with sensorineural hearing loss (SNHL) and 20 with ANSD. SNHL patients were genotyped for common GJB2 variants using qPCR, while ANSD patients underwent whole exome sequencing, with variants analyzed across 249 genes. Homozygous GJB2 variants were found in 54.9% of SNHL patients. In 60% of ANSD patients, variants were detected in OTOF (25%), CDH23, TMC1, COL11A1, PRPS1, TWNK, and HOMER2 genes, including eight novel variants. Transient evoked otoacoustic emissions testing revealed differences at 4000 Hz (p = 0.0084) between the ANSD and SNHL groups. The auditory steady-state response (ASSR) test showed significant differences at 500 Hz (p = 2.69 × 10⁻⁴) and 1000 Hz (p = 0.0255) compared to pure-tone audiometry (PTA) in ANSD patients. Our questionnaire shows that the parents of children with SNHL often report an improved quality of life with hearing aids or cochlear implants, while parents of children with ANSD frequently experience uncertainty about outcomes (p = 0.0026), leading to lower satisfaction.

This study reports a 14-year-old male patient with multiple congenital anomalies, including hypospadias, complete cleft palate, and recurrent pneumonia. His clinical presentation includes significant physical and intellectual developmental delays, autism-like symptoms, and spastic diplegia. Whole-exome sequencing (WES) was performed due to these complex symptoms, revealing a novel heterozygous deletion on chromosome 10q24.31-q24.33. Laboratory findings indicated agammaglobulinemia, leading to prophylactic antibiotic therapy and immunoglobulin replacement. Additional imaging studies showed cystic malformation of the middle lobe of the right lung, sliding hiatal hernia with prolapse of the gastric mucosa, and brain anomalies consistent with Joubert syndrome. This case underscores the importance of genetic analysis in understanding the etiology of congenital anomalies and neurodevelopmental disorders, demonstrating that WES can provide critical insights into the genetic basis of complex clinical presentations. The identified chromosomal deletion contributes to the existing literature on genomic imbalances associated with similar phenotypes.

The MGI (MGI Tech Co. Ltd., China) next-generation sequencing platform, including the DNBSEQ-G50, -G400, and -T7 sequencers, is being actively adopted in research. Despite its widespread adoption, challenges persist in the form of limitations associated with the manufacturer's provided barcode set for library preparation. These limitations include constraints on the number of samples that can be concurrently sequenced, compatibility issues with barcodes from diverse or incomplete sets, and restrictions on the sample ratio. Purpose: to develop a universal method that allows sequencing of up to 252 samples simultaneously on a single sequencer lane, while eliminating barcode-related limitations. We proposed a “quad method” that provides 4 or 4n+2 equilibration of barcodes. This paper also delves into its comprehensive analysis, verification procedures, seamless integration into the sequencing process and validation of the method on the DNBSEQ G-400 platform. The quad method showed efficiency and reliability, allowing sequencing of up to 252 samples simultaneously without compromising data quality. The proposed method optimizes library preparation and improves the flexibility of sequencing on the MGI platform.

Background/Objectives: Direct oral anticoagulants (DOACs) are first-line medications for stroke prevention in non-valvular atrial fibrillation (AF). However, variability in drug response poses risks of hemorrhagic or thromboembolic events. The role of genetics in determining DOAC safety is gaining attention, but evidence linking specific genetic polymorphisms to bleeding risk is lacking. Therefore, the aim of our study was to conduct an Exome-Wide Association Study with 198 non-valvular AF patients treated with rivaroxaban or apixaban, including 97 with bleeding complications and 99 without. Methods: DOAC plasma concentrations, 6-β-hydroxycortisol and cortisol levels in urine were measured for CYP3A4 phenotyping. Sequencing was performed on the DNBSEQ G-400 platform. Single nucleotide variants (SNV) associations with bleeding risk were assessed using logistic regression with additive, dominant, and recessive genetic models. Results: No SNVs reached Bonferroni-corrected significance in any genetic model. Polygenic risk scores (PRS) showed weak predictive ability for bleeding with apixaban. For rivaroxaban, regression indicated that ln Css min/D + 1 index increased with PRS, age, and 6-β-hydroxycortisol/cortisol ratio, but decreased with higher 6-β-hydroxycortisol and coronary heart disease presence. Conclusions: The findings revealed that the residual equilibrium concentration of anticoagulants, including dose-adjusted, cannot alone predict bleeding risk in non-valvular AF patients. No new genetic variants associated with bleeding risk were identified.

Background: Auditory Neuropathy Spectrum Disorder (ANSD) is often missed by common hearing screening tests, still accounting for up to 10% of hearing impairments. ANSD has an underlying genetic factor being mostly caused by pathogenic variants in 13 genes. Methods: We examined 122 children with impaired hearing, including 102 pediatric patients (mean age was 3.7±4.1 years) with sensorineural hearing loss (SNHL) of varying severity and 20 children with a clinically confirmed ANSD (mean age was 5.65±4.63 years). For children with SNHL, we genotyped the most frequent variants (c.35delG, c.167delT, c.235delC, c.313-326del14 and c.358-360delGAG) in the GJB2 gene using quantitative PCR. For children with a clinically confirmed ANSD, we performed the whole exome sequencing and studied the obtained variants in a custom panel of 248 genes. Results: Our findings show that fifty-six (54.9%) SNHL patients carried homozygous variants in GJB2 gene. In 12 (60%) ANSD patients, we detected variants in nucleotide sequences of the OTOF (25%), CDH23, TMC1, COL11A1, PRPS1 and HOMER2 genes (8 of which had not been previously described). Transient Evoked Otoacoustic Emissions testing revealed differences at 500 Hz (AS, p = 0.04181) and 4000 Hz (AD, p = 0.00126) between the ANSD and SNHL patient groups. The Auditory Steady-State Response (ASSR) test demonstrated significant differences at all frequencies (p < 0.01). When comparing the results obtained from the pure-tone audiometry and ASSR tests in ANSD patients revealed statistically significant differences at 500 (AD, AS) and 1000 (AD) Hz. Conclusions: These findings indicate that audiologists and otorhinolaryngologists should use the ASSR test in clinical practice for differential diagnosis of auditory neuropathies. According to the survey data from the parents of hearing-impaired children, rehabilitation was more successful in SNHL patients compared to ANSD patients.

Whole exome sequencing (WES) is essential for identifying genetic variants linked to diseases. This study compares available to date four exome enrichment kits: Agilent SureSelect Human All Exon v8, Roche KAPA HyperExome Probes, Vazyme VAHTS Target Capture Core Exome Panel, and Nanodigmbio NEXome Plus Panel v1. We evaluated target design, coverage statistics, and variant calling accuracy across these four different exome capture products. All kits showed high coverage completeness, with mean x10 coverage exceeding 97.5% and x20 coverage above 95%. Roche exhibited the most uniform coverage, indicated by the lowest fold-80 scores, while Nanodigmbio had more on-target reads due to fewer off-target reads. Variant calling performance, evaluated using in-lab standard E701 DNA sample, showed high recall rates for all kits, especially Agilent v8. All kits achieved an F-measure above 95.87%. Nanodigmbio had the highest precision with the fewest false positives but a slightly lower F-measure than other kits. This study also highlights the performance of new Chinese solutions from Vazyme and Nanodigmbio, which were comparable to Agilent v8 and Roche KAPA kits. These findings assist researchers and clinicians in selecting appropriate exome capture solutions.

Trio-based exome sequencing and high-resolution HLA typing were employed to analyze three patients with autoimmune adrenal insufficiency (AAI), including those with autoimmune polyglandular syndrome (APS) type 2, as well as their parents. Benign or likely benign variants of the AIRE gene were identified in all participants of the study. These variants, coupled with clinical data and the results of antibody studies to type I interferons, helped to exclude APS-1. Patients with APS-2, in contrast to patient with AAI, inherited distinct variants of unknown significance in the CLEC16A gene, which is associated with autoimmune diseases including AAI. Various risk alleles in other genes associated with autoimmunity were identified in all patients. HLA typing of second class loci revealed alleles related to APS types 2, 3, and 4. Nevertheless, the frequencies of the haplotypes identified are substantial in the healthy Russian population, precluding from regarding these alleles as genetic determinants linked to APS development. Immunological examination can detect antibody carriers and predict the risk of autoimmune disease development. In the future, to identify genetic predictors of autoimmune endocrinopathies, it is recommended to analyze the whole genome of patients and their relatives, examining clinically relevant variants in non-coding regions.