Frequency Of Single Nucleotide Polymorphisms Research Articles

NEW MULTIPLEX SNP GENOTYPING ASSAY TO SIMUTANEOUSLY SCREEN FOR EIGHT VOLTAGE-GATED SODIUM CHANNEL MUTATIONS IN AEDES AEGYPTI

Aedes aegypti has been implicated as the vector responsible for transmission of dengue, chikungunya, and Zika viruses during disease outbreaks in Florida within the past 15 years. Recently, locally acquired dengue cases have increased dramatically, with more than 450 cases documented in Florida since 2019. This mosquito is known to be resistant to pyrethroid-based insecticides in Florida. Resistance of insects to pyrethroids due to knockdown resistance (kdr) is the result of single nucleotide polymorphisms (SNPs) in the voltage-gated sodium channel gene (vgsc). Recently, two novel SNPs, F174I and E478K, and four known SNPs, V410L, S723T, D1763Y, and Q1853R were reported circulating in Floridian Ae. aegypti populations for the first time. The present study provides a more comprehensive estimate of these SNP frequencies through the screening of a larger number of Floridian Ae. aegypti samples using a new custom multiplex SNP assay we developed using the Agena Biosciences iPLEX Assay platform to facilitate the rapid screening of multiple SNPs at an affordable cost. Our assay was successful in screening 162 Ae. aegypti mosquitoes for 8 SNPs from 4 counties in Southern Florida (Broward, Collier, Palm Beach, and Monroe Counties). This new assay can be used for studies examining the association between genetic mutations and pyrethroid-resistant phenotypes in Ae. aegypti populations such as increased time of survival after insecticide exposure.

Sirtuin-1 level and gene polymorphisms in multiple sclerosis

BackgroundSirtuin-1 (SIRT1) may affect multiple sclerosis (MS) disease. This study aimed to investigate the level of serum SIRT1, mRNA expression and genetic polymorphisms in Egyptian MS sufferers. Also, to assess its role as a possible biomarker in predicting the risk of MS and to evaluate the association between its levels and disability of MS. Measurement of SIRT1, serum level, mRNA expression level and genotyping for sirtuin-1 gene polymorphisms in 240 Egyptian subjects; 120 MS sufferers and 120 healthy control subjects.ResultsThere was a significant diminishment of level of serum sirtuin-1, and sirtuin-1 mRNA expression in MS sufferers compared to control subjects. Different sirtuin-1 single nucleotide polymorphism frequencies were statistically significant in MS sufferers compared to the control subjects. Moreover, a negative correlation of serum level of sirtuin-1 in MS sufferers with MS disease duration, disability according to Expanded Disability Status Scale (EDSS) score, cholesterol, and triglyceride serum levels. Regarding the sirtuin-1 gene polymorphisms in MS sufferers, the rs7895833 GG genotype had significant higher cholesterol, and low-density lipoprotein (LDL) levels than the GA and AA genotypes and that the rs7069102 GG genotype had a higher LDL level than the CG and CC genotypes while the rs2273773 TT genotype was significantly associated with cholesterol, and LDL levels than the TC and CC genotypes. No significant difference was detected in EDSS score comparing different sirtuin-1 genotypes among MS sufferers. In MS sufferers, rs7895833 G allele can be independently associated with cholesterol, triglycerides, and LDL levels. rs7069102 C allele can be independently associated with LDL level. With regard to rs2273773, T allele, it can be independently associated with cholesterol and LDL levels.ConclusionThere was a significant association between different sirtuin-1 gene polymorphisms and dyslipidemia which may modulate the course of MS disease. Furthermore, serum sirtuin-1 level can be considered as a possible predictor of disability in multiple sclerosis sufferers.

Clinical Presentation of Blastomycosis is Associated With Infecting Species, Not Host Genotype.

Objective: To determine if host genetics may be a risk factor for severe blastomycosis.Design: A cohort of patients who had contracted blastomycosis underwent targeted SNP (single nucleotide polymorphism) genotyping. The genetics of these patients were compared to a set of age and gender-matched controls and between patients with severe versus mild to moderate blastomycosis.Setting: The Marshfield Clinic Health System in central and northern WisconsinParticipants: Patients with a diagnosis of blastomycosis prior to 2017 were contacted for enrollment in this study. A phone hotline was also set up to allow interested participants from outside the Marshfield Clinic Health System to request enrollment.Methods: SNP frequency was assessed for significant differences between the patient cohort and controls and between patients with severe versus mild to moderate blastomycosis. We also tested the effect of Blastomyces species identified in clinical isolates on disease symptoms and severity.Results: No significant differences were found in SNP frequency between cases and controls or between those with severe or mild to moderate blastomycosis. We did detect significant differences in symptom frequency and disease severity by Blastomyces species.Conclusions: Our study did not identify any genetic risk factors for blastomycosis. Instead, the species of Blastomyces causing the infection had a significant effect on disease severity.

Autophagy-Related Gene ATG7 Polymorphism Could Potentially Serve as a Biomarker of the Progression of Atrophic Gastritis.

Cytotoxin-associated gene A (CagA) is an oncoprotein that H. pylori injects into the host's gastric epithelial cells and that induces proinflammatory cytokines, such as interleukin (IL)-18 and IL-1β. As a result, it leads to atrophic gastritis (AG), a precancerous lesion of gastric cancer. On the other hand, host cells degrade CagA using autophagy systems. However, few studies exist about the single nucleotide polymorphisms (SNPs) in MAP1LC3A, MAP1LC3B, ATG4A, ATG4B, ATG4C, ATG7, and ATG13, which belong to the autophagy-related genes concerning AG. This study aimed to detect biomarkers associated with AG. Herein, H. pylori-positive subjects (n = 200) were divided into the AG (n = 94) and non-AG (n = 106) groups. Thirty tag SNPs were selected from the above seven candidate genes. The SNP frequency between the two groups was analyzed. The frequency of the C/T or T/T genotype at rs4683787 of ATG7 was significantly lower in the AG group than in the non-AG group (p = 0.034, odds ratio = 0.535). Based on multivariate analysis, the C/C genotype of rs4684787 and age were independently associated with gastric mucosal atrophy. This finding helps stratify the patients needing timely endoscopic screening or early eradication of H. pylori.

Defining genetic diversity of rhesus macaque Fcγ receptors with long-read RNA sequencing.

Fcγ receptors (FcγRs) are membrane-bound glycoproteins that bind to the fragment crystallizable (Fc) constant regions of IgG antibodies. Interactions between IgG immune complexes and FcγRs can initiate signal transduction that mediates important components of the immune response including activation of immune cells for clearance of opsonized pathogens or infected host cells. In humans, many studies have identified associations between FcγR gene polymorphisms and risk of infection, or progression of disease, suggesting a gene-level impact on FcγR-dependent immune responses. Rhesus macaques are an important translational model for most human health interventions, yet little is known about the breadth of rhesus macaque FcγR genetic diversity. This lack of knowledge prevents evaluation of the impact of FcγR polymorphisms on outcomes of preclinical studies performed in rhesus macaques. In this study we used long-read RNA sequencing to define the genetic diversity of FcγRs in 206 Indian-origin Rhesus macaques, Macaca mulatta. We describe the frequency of single nucleotide polymorphisms, insertions, deletions, frame-shift mutations, and isoforms. We also index the identified diversity using predicted and known rhesus macaque FcγR and Fc-FcγR structures. Future studies that define the functional significance of this genetic diversity will facilitate a better understanding of the correlation between human and macaque FcγR biology that is needed for effective translation of studies with antibody-mediated outcomes performed in rhesus macaques.

UV-C LED-induced cyclobutane pyrimidine dimer formation, lesion repair and mutagenesis in the biofilm-forming diatom, Navicula incerta

The use of ultraviolet-C (UV-C) irradiation in marine biofouling control is a relatively new and potentially disruptive technology. This study examined effects of UV-C exposure on the biofilm-forming diatom, Navicula incerta. UV-C-induced mutations were identified via Illumina HiSeq. A de novo genome was assembled from control sequences and reads from UV-C-exposed treatments were mapped to this genome, with a quantitative estimate of mutagenesis then derived from the frequency of single nucleotide polymorphisms. UV-C exposure increased cyclobutane pyrimidine dimer (CPD) abundance with a direct correlation between lesion formation and fluency. Cellular repair mechanisms gradually reduced CPDs over time, with the highest UV-C fluence treatments having the fastest repair rates. Mutation abundances were, however, negatively correlated with CPD abundance suggesting that UV-C exposure may influence lesion repair. The threshold fluence for CPD formation exceeding CPD repair was >1.27 J cm−2. Fluences >2.54 J cm−2 were predicted to inhibit repair mechanisms. While UV-C holds considerable promise for marine antifouling, diatoms are just one, albeit an important, component of marine biofouling communities. Determining fluence thresholds for other representative taxa, highlighting the most resistant, would allow UV-C treatments to be specifically tuned to target biofouling organisms, whilst limiting environmental effects and the power requirement.

LDL-receptor gene polymorphism as a predictor of coronary artery disease: an Egyptian pilot study: relation to lipid profile and angiographic findings.

Coronary artery disease (CAD) is the main cause of death in Egypt. Many LDL-R gene locus single nucleotide polymorphisms (SNP) are found to be associated with the risk of CAD. This research aimed to assess the allelic and genotypic frequencies of rs1122608 SNP and their association with the extent of vessel affection and lipid profile in a population of Egyptians.100 CAD patients and 100 healthy controls of Egyptians were included. PCR-RFLP was used to genotype rs1122608 SNPs. Significantly higher proportion of 'T' allele among patient (risk allele). This association is of low strength (ϕ lies between 0.1 and 0.3). A participant with 'T' allele has 1.95 times higher odds to exhibit CAD versus a participant with 'G' allele. Significantly higher proportion of 'T/T' genotype among cases versus control (risk genotype). This association is of low strength (Cramer's V lies between 0.1 and 0.3). A participant with 'T/T' genotype has 4.5 times higher odds to exhibit CAD versus a participant with 'G/G'. Gensini score showed no significant association with rs1122608 genotypes (p = 0.863). The mutant GT and TT genotypes and minor T allele of rs1122608 were positively correlated with CAD and considered as independent risk factors for CAD.

Drug resistance markers in Plasmodium vivax isolates from a Kanchanaburi province, Thailand between January to May 2023.

Plasmodium vivax has become the predominant species in the border regions of Thailand. The emergence and spread of antimalarial drug resistance in P. vivax is one of the significant challenges for malaria control. Continuous surveillance of drug resistance is therefore necessary for monitoring the development of drug resistance in the region. This study aims to investigate the prevalence of the mutation in the P. vivax multidrug resistant 1 (Pvmdr1), dihydrofolate reductase (Pvdhfr), and dihydropteroate synthetase (Pvdhps) genes conferred resistance to chloroquine (CQ), pyrimethamine (P) and sulfadoxine (S), respectively. 100 P. vivax isolates were obtained between January to May 2023 from a Kanchanaburi province, western Thailand. Nucleotide sequences of Pvmdr1, Pvdhfr, and Pvdhps genes were amplified and sequenced. The frequency of single nucleotide polymorphisms (SNPs)-haplotypes of drug-resistant alleles was assessed. The linkage disequilibrium (LD) tests were also analyzed. In Pvmdr1, T958M, Y976F, and F1076L, mutations were detected in 100%, 21%, and 23% of the isolates, respectively. In Pvdhfr, the quadruple mutant allele (I57R58M61T117) prevailed in 84% of the samples, followed by (L57R58M61T117) in 11%. For Pvdhps, the double mutant allele (G383G553) was detected (48%), followed by the triple mutant allele (G383M512G553) (47%) of the isolates. The most prevalent combination of Pvdhfr (I57R58M61T117) and Pvdhps (G383G553) alleles was sextuple mutated haplotypes (48%). For LD analysis, the association in the SNPs pairs was found between the intragenic and intergenic regions of the Pvdhfr and Pvdhps genes. The study has recently updated the high prevalence of three gene mutations associated with CQ and SP resistance. Genetic monitoring is therefore important to intensify in the regions to further assess the spread of drug resistant. Our data also provide evidence on the distribution of drug resistance for the early warning system, thereby threatening P. vivax malaria treatment policy decisions at the national level.

Genetic Association Between Polycystic Ovary Syndrome and the APOA5 rs662799 and PLIN1 rs894160 Metabolic Variants in the Western Saudi Population: A Case-Control Study.

Polycystic ovary syndrome (PCOS) is a common endocrinological condition affecting women of reproductive age, associated with insulin resistance and obesity. PCOS pathogenesis is complex and multifactorial, involving genetic and environmental factors. This study aimed to determine and compare genotype and allele frequencies of single nucleotide polymorphisms (SNPs) in the apolipoprotein A5 (APOA5; rs662799) and perilipin 1 (PLIN1; rs894160, rs1052700 and rs6496589) genes in Western Saudi women to investigate their association with PCOS and its clinical characteristics. This was a case-control study conducted on women with (n = 104) and without (n = 87) PCOS. The SNPs were genotyped using TaqMan genotyping assays. Significant and direct associations were detected between PCOS susceptibility and APOA5 SNP rs662799 and PLIN1 SNP rs894160 (P < .001). For APOA5 SNP rs662799, women with the A allele were more likely to have PCOS (relative risk [RR] = 1.348, odds ratio [OR] = 2.313, P < .001) and hypertriglyceridaemia (OR = 17.0, P = .5) than women with the G allele. For PLIN1 SNP rs894160, women with the T allele were more likely to have PCOS than women with the C allele (RR = 8.043, OR = 7.427, P < .001). For PLIN1 SNP rs1052700, women with the TT genotype were more likely to have hyperandrogenism (OR = 29.75, P = .02) and an irregular period (OR = 0.07, P = .040) than women with the AT genotype. We identified novel alleles and genotypes contributing to the genetic risk of PCOS in the Western Saudi population.

Alcohol Intubation and Synergistic Effect of Folic Acid and Egg Yolk Powder Solution on Alcohol Dehydrogenase [ADH] Class I Gene Expression and Single Nucleotide Polymorphism Frequency in Parent and Prenatal Alcohol Exposed Wistar Rats

Alcohol Intubation and Synergistic Effect of Folic Acid and Egg Yolk Powder Solution on Alcohol Dehydrogenase [ADH] Class I Gene Expression and Single Nucleotide Polymorphism Frequency in Parent and Prenatal Alcohol Exposed Wistar Rats

Common Genetic Variants in SIRT1 Gene Promoter and Type 2 Diabetes Mellitus in Saudi Arabia.

Type 2 diabetes mellitus (T2DM) is common in Saudi Arabia and represents a major health concern. Silent information regulator of transcription-1 (SIRT1) positively influences insulin sensitivity and might contribute to the pathogenesis of T2DM. This study aimed to investigate the frequency of two common functional single nucleotide polymorphisms (SNPs) in the promoter region of SIRT1; rs12778366 (T>C) and rs3758391 (T>C) in Saudi Arabian population and examine any association with T2DM. A total of 445 volunteers were divided into 224 healthy controls and 221 patients previously diagnosed with T2DM. Genomic DNA was extracted from all samples and genotyped for SIRT1 rs12778366 and rs3758391 SNPs by TaqMan RT-PCR allelic discrimination assay. Logistic regression analysis was used to establish any relationship between these polymorphisms and T2DM. In the total study population, rs12778366 genotype frequencies were TT (89.2%), TC (10.3%), and CC (0.45%) and for the rs3758391 they were TT (16.4%), TC (44.5%), and CC (39.1%). The distribution of these genotypes, in both polymorphisms, were similar among T2DM and controls. Logistic regression analysis confirmed the lack of association between the presence of CC or CT variants and T2DM for rs12778366 and rs3758391 SNP (OR = 0.98 [CI]: 0.55 - 1.75; p = 0.999 and OR= 0.75; [CI]: 0.45 - 1.24; p = 0.313), respectively. This study revealed the frequency of SIRT1 rs12778366 and rs3758391 SNPs in our population and reported no association between these polymorphisms and the risk for T2DM. This finding might add to the growing body of literature exploring the genetics of T2DM.

The association of CYP11A1 gene polymorphisms with the polycystic ovary syndrome patients.

The objective of this study was to investigate the allele frequencies of polymorphisms in genes CYP11A1 rs4886595 and CYP11A1 rs4887139 that are responsible for the steroidogenesis mechanism in polycystic ovary syndrome patients and control females. Samples were obtained from the Department of Obstetrics and Gynecology in the Near East University Hospital from September 2019 to December 2019. Only the nonobese patients between the ages of 18-40 years were included in this study following informed consent. Obese patients and patients more than 40 years of age were excluded from the study. Nonobese women and normal ovulation were included in the control group. DNA was isolated from blood samples. Real-time polymerase chain reaction (PCR) was used to analyze single nucleotide polymorphisms (SNPs) in various genes linked to polycystic ovary syndrome. The studies were carried out using the samples obtained from 120 women, of whom 55 were nonobese and had normal ovulation, and 65 were polycystic ovary syndrome patients. The allelic frequencies of SNPs in genes linked to polycystic ovary syndrome were calculated using real-time PCR outcomes. The variation of the CYP11A1 rs4887139 G>A did not show any significance, while the variation of CYP11A1 rs4886595 C>A showed significant differences between the patient and the control groups (p=0.01), respectively. Future research ought to focus on elucidating the susceptible causes of polycystic ovary syndrome with a wide range of SNPs and more sample size. The genome-wide association studies in polycystic ovary syndrome patients of different origin will be important to identify candidate genes as well as proteins that are implied in polycystic ovary syndrome risk.

Association of HLA rs3135388 and rs9271366 Single-Nucleotide Polymorphisms With Multiple Sclerosis in the Chaharmahal and Bakhtiari Province, Iran

Background and aims: Genetics and environment are synergistic in multiple sclerosis (MS). Human leukocyte antigen (HLA) class II has a strong genetic association with MS. The aim of this study was to determine the association of HLA rs3135388 and rs9271366 single-nucleotide polymorphisms (SNPs) with MS. Methods: The rs3135388, rs9271366, and rs422951 SNPs were genotyped in 173 Iranian relapsing-remitting MS (RRMS) patients and 200 matched healthy controls, using the polymerase chain reaction-restriction fragment length polymorphism method. Odds ratios (ORs) and 95% confidence intervals (CIs) were calculated using statistical software to estimate the risk factor for disease. The allele and genotype frequencies of SNPs were determined based on data analysis using SPSS, version 20. Results: Our results identified a strong association in the allele distribution for both rs3135388 and rs9271366 SNP, such that the A allele of rs3135388 (OR=1.765; 95% CI: 1.071–2.909) and the G allele of rs9271366 were found to be more frequent in MS patients than in healthy controls (OR=1.861; 95% CI: 1.025–3.3378). The mutated G allele for rs422951 SNP was relatively frequent (OR=1.128; 95% CI: 0.745–1.707). Conclusion: Our findings revealed the critical role of the rs3135388 and rs9271366 SNPs in MS disease progression. Genotyping MS patients could facilitate personalized medical management.

Association of Gene Polymorphisms with Breast Cancer Risk in the Kazakh Population.

The research aim is analyzing and identify reliable genetic markers of breast cancer risk in the Kazakh population. The databases were analyzed with the selection of polymorphisms associated with the development of breast cancer and further genotypic study of a group of women with a confirmed diagnosis of breast adenocarcinoma (group No. 1) and a group of relatively healthy women (group No. 2). The research presents the results of a study on the frequency of certain single-nucleotide polymorphisms in patients with breast cancer in the Republic of Kazakhstan. The frequency of single-nucleotide polymorphisms rs4646, rs1065852, rs4244285, rs67376798, rs6504950, rs2229774, rs1800056, rs16942, rs4987047 is statistically significant compared to the control group of patients. These polymorphisms in the Kazakh population have a direct association with an increased risk of breast cancer in women and may be used as cancer indicators during the genetic screening of patients with a complicated family history. Single-nucleotide polymorphisms such as rs55886062, rs3918290, rs12721655, rs4987117, rs2229774, rs11203289, rs137852576, rs11571833, rs80359062 and rs11571746 were found in more than 40. Zero percent of patients with breast cancer may be used as markers for detecting patients at increased risk of breast malignancy in the Kazakh population without a history of poor family history. The usage of the data obtained in a set of state programs for early screening of patients will improve the rates of early breast tumor detection, form groups of patients with a high risk of disease development and improve the quality and expectancy of life.

Abstract PR013: Association of a polymorphic variant in JMJD1C with response to chemoradiotherapy in rectal cancer and head and neck cancer: Implications for a diverse population

Abstract Neoadjuvant chemoradiotherapy (nCRT) is the standard treatment for locally advanced rectal cancer (LARC). Although nCRT has improved outcomes for many patients, ~70-80% of patients show poor or no response, and CRT is toxic. No biomarker exists that can predict which patients may benefit from nCRT. CRT efficacy arises from unrepaired double-strand breaks (DSBs). Recent work has suggested that dynamic changes in lysine modification of chromatin regulate DSB repair. This study investigated the association of germline single nucleotide polymorphisms (SNPs) in lysine-modifying genes with DSB repair and CRT outcomes in LARC. The LARC patients (n=84) were segregated into poor (PoR) and complete (CR) responders using the Neoadjuvant Rectal score. Blood DNA was sequenced to identified SNPs, matched LARC patient-derived cells were tested for sensitivity to DNA damage, and results correlated. Significant SNP-response correlations were confirmed in an independent cohort of CRT-treated head and neck squamous cell carcinoma (HNSCC) patients who had received chemoradiation (n=90). The HNSCC patients were segregated into 3 groups based on response: disease-free (n=30), recurrent disease (n=28), and never-disease free (n=32). Isogenic HNSCC lines with either the wildtype (WT) or SNP were generated using CRISPR/Cas9 technology, and bulk RNA sequencing and immunofluorescence-based studies were performed to assess pathways affected. LARC CRs were significantly enriched versus PoRs for a germline SNP in the lysine H3K9 demethylase, JMJD1C, resulting in a missense variant (discovery (n=30, p&amp;lt;0.00001), validation (n=54, p=0.001); Fisher’s Exact). In the HNSCC cohort, the JMJD1C SNP was significantly associated with no recurrence (p=0.0003, Fisher’s Exact). LARC patient-derived cell lines carrying the SNP were significantly more sensitive to DSB damage versus those carrying the WT (n=12, p&amp;lt;0.001, Mann-Whitney). Cells bearing the SNP were significantly enriched for altered mRNA levels of genes associated with DNA repair compared with WT cells (q-value&amp;lt;0.05). On irradiation, the SNP cells showed robust activation of DSB repair (γH2AX, BRCA1 foci, JMJD1C nuclear localization) versus the WT cells. Finally, analysis of the JMJD1C SNP frequency in large population datasets found that while it was abundant in European Americans (EAs), it is relatively rare in African Americans (AAs) (EA= 0.3139 vs. AA=0.072; gnomAD population). This study provides novel insight into a SNP in a lysine-modifying gene affecting DSB repair that may serve as a predictive biomarker for CRT response in 2 different cancer types. Our data suggest that host genetics impact DSB repair and warrant further investigation to better understand disparities in response to CRT. Future research will further explore the racial/ethnic disparities in tumor recurrence and confirm results in in vitro and in vivo models of cancer, and in large patient datasets. Citation Format: Adria Hasan, Elena Demidova, Philip Czyzewicz, Shreya Shah, Karthik Devarajan, Thomas J. Galloway, Margret B. Einarson, Barbara Burtness, Erica A. Golemis, Joshua E. Meyer, Sanjeevani Arora. Association of a polymorphic variant in JMJD1C with response to chemoradiotherapy in rectal cancer and head and neck cancer: Implications for a diverse population [abstract]. In: Proceedings of the 16th AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2023 Sep 29-Oct 2;Orlando, FL. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2023;32(12 Suppl):Abstract nr PR013.

The Impacts of Genetic Polymorphisms on the Von Willebrand Factor Level in Type 1 Von Willebrand Disease

Background: von Willebrand disease (VWD) is the most common inherited bleeding disease, which results from deficiency or defect of von Willebrand factor (VWF). In type 1 VWD, there are only 53-81% patient whose mutations of VWF gene can be identified. Some studies reported that variants outside the VWF gene may also contribute to the VWF level. There are 7 genetic variants of single nucleotide polymorphism ( SNP ) reported to be associated with the variation of VWF levels, including CLEC4M, STXBP5, SCARA5, ABO, VWF, STAB2 and UFM1. The aims of this study were to explore the frequency of SNPs of the 7 genes among type 1 VWD patients and the effects of the 7 genetic variants on the VWF levels in type 1 VWD patients. Patients and methods: There were 62 patients ( the ratio of M/F: 1 ) with type 1 VWD enrolled in this study, including 35 (56.5%) patients with found mutations of VWF gene. Each patient's bleeding symptoms were assessed by the MCMDM-1 VWD bleeding score. The results of VWF levels: VWF:Ag and VWF activity ( VWF:ACL or VWF:RCo ) and FVIII: C at the VWD diagnosis were retrospectively collected, as shown in Table 1. Each enrolled patient's genomic DNA was extracted and the SNPs of the 7 gene were targeted by PCR amplification followed by direct sequencing. Results: There was high correlations between VWF:Ag and VWF activity ( r = 0.73, p &amp;lt; 0.0001 ) and between VWF:Ag and FVIII:C ( r = 0.60, p &amp;lt; 0.0001 ), respectively. The patients without variant ( A/A) of UFM1 has significantly lower median VWF:Ag of 33% than 44.7% of patients with heterozygous variant ( A/C ) type ( p = 0.033 ). Patients without variant (A/A ) of STXBP gene had both significantly lower median value of VWF:Ag ( 31.9% vs 46.1%, p = 0.017 ) and VWF activity (26.55% vs 30.4%, p = 0.0467 ) than those patients with heterozygous variant ( A/G ). Four alleles substitution of SNP of UFM1( C&amp;gt;A ) and STXBP genes ( G&amp;gt;A ) variant has remarkably lowest median VWF antigen level of 27.4%, which was significantly lower than 43.05% of the three alleles substitution ( p = 0.0014 ) and the 48.6% of the two alleles substitution ( p = 0.0279 ), as shown in Figure 1. Conclusions: Our study demonstrates that both C to A variant of UFM1 gene and G to A variant of STXBP5 gene were associated with lower VWF: Ag level than those without variant, respectively. The combination of four allelic variant of UFM1 and STXBP5 genes had the most reducing effect on VWF: Ag level.

2163. Prevalence and geographic distribution of mutations in TonB-dependent receptors in the Prospective Observational Pseudomonas aeruginosa (PA) Study

Abstract Background Cefiderocol (FDC) is a last line treatment for carbapenem-resistant Pseudomonas aeruginosa (CR-PA). FDC retains activity against CR-PA by inhibition of penicillin-binding proteins facilitated by active uptake of drug via TonB-dependent receptors (TBDR). Prior, we have shown an association between mutations in TBDR genes and loss of FDC susceptibility, particularly in association with changes at the PA-derived cephalosporinase (PDC/AmpC) active site. Here, we outline geographic variation and prevalence of TBDR mutations in the Prospective Observational Pseudomonas (POP) study.Figure 1.SNP frequency in the pirR, pirS, pirA, and piuA/piuD genes across geographic location.Figure 2.Genotypic variation of PirRS and PiuA/D genes in the 972 P. aeruginosa isolates of the POP cohort. Black versus gray differentiates presence of either PiuA versus PiuD homologous proteins. Color legend corresponds to presence of TBDR gene variants/SNPs per gene pirR, pirS, pirA, piuA/piuD, and ampC. Methods The genomes of 972 CR-PA isolates collected across 10 countries in North America, South America, Asia, and Oceania were screened for single nucleotide polymorphisms (SNP) resulting in missense, insertion/deletion (indel), frameshift, and early stop codon mutations in the TBDR genes pirR, pirS, pirA, piuA/piuD, and ampC using PAO1 as a reference. SNPs were defined as variant mutations occurring in ≤10% of isolates. A core genome maximum likelihood phylogenetic tree was used to assess distribution and prevalence of genotypic variants. Neighbor-joining trees of the variants in each gene were used to analyze clusters across lineages. Results Across all regions except Oceania, SNPs were most frequently seen in pirS and pirA (Fig 1). Gene variants comprised from 8% up to 50% of each geographic population. On phylogenetic mapping, isolates within the same lineage tended to have clustering of shared mutations across the TBDR genes pirS, pirR, and pirA, and the majority of piuA/piuD mutations clustered within two distinct lineages (Fig 2). Major mutations, including indel, frameshift, and early stop codon mutations occurred in 16 isolates (1.6%), predominantly in isolates from the United States (n=14). Variations in PDC were frequently observed, with 57 different variants across 35.8% of the population. Seven of the PDC variants (n=10 isolates) have been associated with decreased susceptibility to ceftolozane/tazobactam, but these variants did not co-occur with major TBDR mutations. Conclusion TBDR mutations occur in PA isolates worldwide. Mutations previously associated with reduced susceptibility to FDC occurred most frequently in isolates from the United States. Disclosures Michael J. Satlin, MD, AbbVie: IDMC member|Biomerieux: Grant/Research Support|Merck: Grant/Research Support|Shionogi: Advisor/Consultant|SNIPRBiome: Grant/Research Support William R. Miller, M.D., Merck: Grant/Research Support|UpToDate: Honoraria

The Role of Coagulation Gene Polymorphisms in Sars-CoV2 Infection in the Bergamo Area

Introduction: Given the great variability of the individual response to SARS-CoV2 infection, it is important to understand whether and how biological and genetic factors might predispose to the infection and to the degree of severity, including the development of systemic coagulopathy and thrombosis. In this study, we evaluated the frequency of prothrombotic allelic variants (AVs) in coagulation genes in a population of COVID-19 patients, to verify whether these polymorphisms, alone or in combination, are associated to an increased susceptibility to develop COVID-19 and/or a greater severity. Methods: A cohort of 358 patients (257M/101F; median age: 55 years) from the Bergamo area with two different degrees of COVID-19 severity, i.e., severe disease (hospitalized, n=212) or mild disease (non-hospitalized, n=146), was enrolled from April 2020 to June 2021 and followed-up for disease outcome and thrombosis. A cohort of 377 healthy subjects (177M/200F; median age: 49 years) from the same area without COVID-19 acted as a control group. DNA was analyzed for a panel of prothrombotic/inflammatory single nucleotide polymorphisms (SNPs) including FII rs1799963, FV rs6025, FV rs118203907, FXIIIA1 rs5985, FGB rs1800790, MTHFR rs1801131 and MTHFR rs1801133 as well as two SNPs located in the coding region of angiotensin-converting enzyme (ACE2) gene, namely ACE2 rs140312271 and ACE2 rs41303171. Statistical analysis was performed using SPSS statistical software. Results: Among the AVs analyzed, the rs6025 polymorphism of FV gene (FV Leiden) was more frequent in COVID-19 patients than in the control group. By a logistic regression corrected for age and gender, FV Leiden was associated with a 3-fold risk of developing COVID-19 (OR: 3.2 [95% CI 1.02-10]). In contrast, the rs41303171 polymorphism of the ACE2 gene was more frequent in the control group and was a protective factor for COVID-19 (OR: 0.12 [95% CI 0.02-0.99]) even after correction for age and gender. No significant differences appeared in the frequency of the other SNPs alone or in combination. Within the COVID-19 patient cohort, the rs1801131 SNP of MTHFR gene was associated with less severe COVID-19 disease. In the hospitalized severe cohort, 17 (8%) patients developed an overt venous thromboembolic event, and 7 COVID-19-related deaths (4%) were recorded. No significant associations were found between any of the AVs analyzed and the outcomes of thrombosis and death. However, the analyses of circulating biomarkers of clotting (i.e. D-dimer, FVIII, fibrinogen, prothrombin fragment 1+2 [F1+2], and FXIII), fibrinolysis (i.e. tissue plasminogen activator [tPA], and plasminogen activator inhibitor-1 [PAI-1]) and endothelial cell activation (i.e. thrombomodulin [TM], and von Willebrand Factor [vWF]), and the neutrophil-to-lymphocyte ratio (NLR) as an inflammatory biomarker revealed thathaving fibrinogen (OR: 4.9 [95% CI 1.04-23.1]), vWF:Ag (OR: 11.8 [95% CI 2.1-66.9]) and NLR (OR: 26.1 [95% CI 2.9-237] levels above the 75th percentile was significantly associated with death. Of interest, in the same hospitalized patient group, but not in the control group, the presence of FXIII SNPs was significantly associated with high levels of F1+2. Conclusions: Our study shows that the FV Leiden polymorphism predisposes to COVID-19 infection, while the rs41303171 ACE2 polymorphism has a protective role. In addition, the rs1801131 SNP of gene MTHFR correlates with a milder COVID-19 disease. In patients with severe COVID-19, the inflammatory marker NLR, and the circulating hemostatic proteins vWF and fibrinogen are associated with mortality.

Impact of MTHFR Polymorphisms C677T and A1298C in Acute Lymphoblastic Leukemia and Burkitt Lymphoma Patients Exposed to Methotrexate and Asparaginase

Background: Asparaginase (Asp) and high-dose methotrexate (HDMTX) are pivotal drugs in the treatment of aggressive hematological malignancies as acute lymphoblastic leukemia (ALL) and Burkitt lymphoma (BL). Asp and MTX present specific drug-related toxicities; for Asp, prothrombotic alteration of the coagulation profile, hepatobiliary and pancreatic toxicities, for MTX, renal, hepatobiliary, hematologic, gastro-intestinal and neuro- toxicity. The 5,10-methylenetetrahydrofolate reductase (MTHFR) is the enzyme involved in conversion of homocysteine into methionine. MTHFR most frequent single nucleotide polymorphisms C677T and A1298C are associated with reduced enzymatic activity up to 60%, that implies hyperhomocysteinemia and increased toxicity of MTX. Numerous studies have explored the relationship between C677T and A1298C polymorphisms of MTHFR and MTX toxicity and/or outcome in children ALL, but results are controversial. Nevertheless, it is common practice to reduce MTX dosage in adult patients in homozygosis. Aims: To explore the relationship between the MTHFR C677T and A1298C polymorphisms in homozygosis and development of MTX toxicity in a cohort of adult patients; furthermore, to evaluate the relationship with thrombotic risk in patients exposed to Asp. Materials and Methods: Data from consecutive adult onco-hematologic patients for whom MTHFR C677T and A1298C polymorphism analysis was available, treated with chemotherapeutic regimens containing HDMTX and/or Asp was retrospectively collected at a single center Italian institution. Adverse events (AEs) were graded according to CTCAE 5.0. Statistical analysis was performed with R language. The impact of MTHFR polymorphisms was evaluated by mean of a multivariate analysis adjusting Odds Ratio (OR) estimates for drug dosage and disease status. Results: Data from 78 patients, 57 affected by ALL, 3 by mixed phenotype leukemia (MPAL) and 18 by BL, was retrospectively collected. Overall, 13 (16.7%) of patients were 677TT and 8 (10.3%) were 1298CC. Patients with ALL/MPAL received mostly pediatric-inspired chemotherapy regimens (containing MTX 2.5 - 5 g/m 2 and Asp 1000-2000 UI/m 2), patients with BL received modified Rituxumab (R)-BFM protocol (MTX 1.5 g/m2); age/comorbidities-adjusted dose reduction was performed according to institutional guidelines. Median age for ALL and BL patients was 37.5 (IQR 27.0-54.2) and 33.5 (26.0-49.7) years, respectively; 54/78 (69.2%) of patients were male. Patients received a median of 3 (IQR 2-6) cycles containing HDMTX and/or Asp, for a total number of 299 cycles (94 containing Asp, 184 HDMTX and 24 both); 142 (48.0%) cycles were administered for active disease (AD), at diagnosis/relapse, and 154 (52.0%) in complete remission (CR). The impact of different dosages of Asp &amp;lt; 1000 UI/m 2 and ≥ 1000 UI/m² and of HDMTX &amp;lt;1.5 g/m 2 and ≥ 1.5 g/m 2 as well as the impact of disease status (AD vs CR) on AEs was explored. Concerning Asp therapy and thrombotic risk, 13 deep vein thrombosis (DVT) were observed, but no association between 677TT nor 1298CC was appreciated. AD status was strongly related to DVT (21.4% vs 1.8%, p 0.003). Coherently with the MTHFR-independent drug's mechanism of action, the polymorphisms had no impact on the development of Asp-related toxicities. Diversely, concerning HDMTX, in multivariate analysis, patients with 677TT were more likely to experience AST increase grade ≥3 (OR 10.55, 95%CI 1.34-83.09) while a protective role versus any grade GGT increase was observed (OR 0.09, 95%CI 0.01-0.71) (Figure 1A). Patients with 1298CC exhibited a superior incidence any grade AST/ALT increase, not confirmed for grade ≥3 (Figure 1B). No significant superior toxicity was observed in terms of renal, gastro-intestinal, hematologic AEs. The impact of a MTX dose reduction &amp;gt;25% of expected dose was then explored in a homogeneous population of 45 front-line pediatric-regimen treated ALL patients (35 received MTX dose ≥75% and 10 &amp;lt; 75%) and no impact in terms of OS and RFS was appreciated. Conclusions: The analysis ofMTHFR polymorphisms in adult patients is useful in identifying patients at higher risk for transaminitis, providing a biological parameter to integrate with hepatosteatosis, liver stiffness evaluation and BMI, for a comprehensive hepatic baseline evaluation. Patients at higher risk for hepatic complications may benefit from MTX dose reduction with no impact on disease outcome.

Analysis of the Association between the Tgfb1 Gene Haplotype and Liver Diseases in Children.

Transforming growth factor-β1 (TGF-β1), a cytokine with immunosuppressive and pro-fibrogenic activity, is a potential marker of infection, liver transplant rejection, and fibrosis. Its levels in the blood and tissues depend on many factors; however, the role of gene polymorphism is still unclear. In this work, the distribution frequency of three single nucleotide polymorphism (SNP) variants of the Tgfb1 gene, namely rs1800469, rs1800470, and rs1800471, was studied in children with end-stage liver disease (ESLD). The study included 225 pediatric liver recipients aged 1 month to 16 years (median, 8 months), including 100 boys and 125 girls, and 198 healthy individuals aged 32.7 ± 9.6 years, including 78 men and 120 women. The indication for liver transplantation in children was ESLD, which was mostly caused by congenital and inherited liver diseases. SNPs were detected by real-time polymerase chain reaction using TaqMan probes and DNA isolated from peripheral blood. SNP frequency distribution was in Hardy-Weinberg equilibrium and did not differ between children with liver diseases and the healthy ones. Analysis of the SNPs frequency based on allelic interaction models did not reveal any differences between patients and the healthy individuals. Evaluation of linkage disequilibrium for Tgfb1 polymorphic variant pairs revealed a statistically significant linkage between all studied variants. Seven haplotypes, which are variants of SNP combinations, were observed in the studied groups of patients and healthy individuals. A total of 80% of the group had three haplotypes, whose frequencies did not differ between patients and the healthy individuals. Significant differences were found in the frequency of the haplotypes A-A-C, G-G-C, and G-A-G (at rs1800469, rs1800470, and rs1800471, respectively), which were observed up to 11 times more often in recipients compared to the healthy individuals. It is possible that these haplotypes are ESLD-predisposing variants, which may also contribute to the development of complications after liver transplantation in children.