Homozygous Group Research Articles

Investigation of the Effects of Biotinidase Deficiency on Plasma Cholinesterase Activity

Purpose: Biotinidase deficiency (BD) is a rare autosomal recessive metabolic disorder that impairs the body's ability to recycle biotin, a crucial coenzyme for carboxylase enzymes involved in various metabolic processes. This study aims to evaluate the effects of biotinidase deficiency on cholinesterase activity in plasma, hypothesizing that the metabolic disruptions caused by inadequate biotin recycling may lead to alterations in cholinesterase function. Materials and methods: Plasma samples were collected from 73 individuals categorized into four genetic groups: wild type (n = 12), heterozygous (n = 30), homozygous (n = 19), and compound heterozygous (n = 12). Cholinesterase activity was measured using a colorimetric method. Results: The study discovered that the cholinesterase activity of the Heterozygous group was higher than the homozygous group (p = 0.0356). Additionally, cholinesterase activity was significantly lower in homozygous and compound heterozygous people than in wild and heterozygous groups (p = 0.0272). The statistically significant changes suggested a relationship between biotinidase deficiency and altered cholinergic activity. Conclusion: The findings indicate that biotinidase deficiency, particularly in its severe variants, may cause considerable reductions in cholinesterase activity, contributing to the neurological symptoms found in affected patients. More studies are needed to investigate the processes behind this association and develop strategies for reducing the effects of BD on cholinesterase activity and neurological health.

Deciphering metabolomics and lipidomics landscape in zebrafish hypertrophic cardiomyopathy model

To elucidate the lipidomic and metabolomic alterations associated with hypertrophic cardiomyopathy (HCM) pathogenesis, we utilized cmybpc3-/- zebrafish model. Fatty acid profiling revealed variability of 10 fatty acids profiles, with heterozygous (HT) and homozygous (HM) groups exhibiting distinct patterns. Hierarchical cluster analysis and multivariate analyses demonstrated a clear separation of HM from HT and control (CO) groups related to cardiac remodeling. Lipidomic profiling identified 257 annotated lipids, with two significantly dysregulated between CO and HT, and 59 between HM and CO. Acylcarnitines and phosphatidylcholines were identified as key contributors to group differentiation, suggesting a shift in energy source. Untargeted metabolomics revealed 110 and 53 significantly dysregulated metabolites. Pathway enrichment analysis highlighted perturbations in multiple metabolic pathways in the HM group, including nicotinate, nicotinamide, purine, glyoxylate, dicarboxylate, glycerophospholipid, pyrimidine, and amino acid metabolism. Our study provides comprehensive insights into the lipidomic and metabolomic unique signatures associated with cmybpc3-/- induced HCM in zebrafish. The identified biomarkers and dysregulated pathways shed light on the metabolic perturbations underlying HCM pathology, offering potential targets for further investigation and potential new therapeutic interventions.

The NOS1AP gene rs10494366 common genetic variant does not modify the risk of sudden cardiac death in users of digoxin.

Common genetic variations in the nitric oxide synthase-1 adaptor protein (NOS1AP) gene are associated with QT-interval prolongation. In a previous study, we observed an association between the rs10494366 variant of this gene and an increased QT-interval shortening in digoxin users. As QT-interval shortening is a risk factor for sudden cardiac death (SCD), in this study, we investigated whether the association between digoxin use and risk of SCD differs in participants with different NOS1AP rs10494366 genotypes. We included 11 377 individuals from the prospective population-based cohort of the Rotterdam Study. We used Cox proportional hazard regression analysis with digoxin as time-dependent exposure to estimate the associations between current digoxin use and the risk of SCD among different rs10494366 genotype groups in the adjusted models. We also studied whether such an association was dose-dependent, comparing high dosage (≥ 0.250 mg), moderate dosage (0.125 mg ≤ dose< 0.250 mg) and low dosage (< 0.125 mg) digoxin users with non-users. The median baseline age of the total study population was 62 (interquartile range [IQR] 58-71) years. The cumulative incidence of SCD was 4.1% (469 cases), and among them, 74 (15.7%) individuals were current digoxin users at the time of death, during a median follow-up of 11.5 (IQR 6.5-17) years. Current digoxin users had an increased risk of SCD (multivariable adjusted model hazard ratio [HR]: 3.07; 95% confidence interval [CI]: 2.38-3.98), with no significant differences between the three genotype groups. The adjusted HRs were 4.03 [95% CI: 1.98-8.21] in the minor homozygous GG, 3.46 [95% CI: 2.37-5.04] in the heterozygous TG and 2.56 [95%CI: 1.70-3.86] in the homozygous TT genotype groups. Compared to low- and moderate-dose, high-dose digoxin users with GG genotype had the highest risk of SCD (HR: 5.61 [95% CI: 1.34-23.47]). Current use of digoxin is associated with a significantly increased risk of SCD. The NOS1AP gene rs10494366 variant did not modify the digoxin-associated risk of SCD in a population of European ancestry.

Effect of FADS1 SNPs rs174546, rs174547 and rs174550 on blood fatty acid profiles and plasma free oxylipins.

Previous studies have indicated that activity of fatty acid desaturase 1 (FADS1), is involved in cardiometabolic risk. Recent experimental data have shown that FADS1 knockdown can promote lipid accumulation and lipid droplet formation in liver cells. In this study, we aimed to characterize whether different FADS1 genotypes affect liver fat content, essential fatty acid content and free oxylipin mediators in the blood. We analyzed the impact of FADS1 single-nucleotide polymorphisms (SNPs) rs174546, rs174547, and rs174550 on blood fatty acids and free oxylipins in a cohort of 85 patients from an academic metabolic medicine outpatient center. Patients were grouped based on their genotype into the homozygous major (derived) allele group, the heterozygous allele group, and the homozygous minor (ancestral) allele group. Omega-3 polyunsaturated fatty acids (n-3 PUFA) and omega-6 polyunsaturated fatty acids (n-6 PUFA) in the blood cell and plasma samples were analyzed by gas chromatography. Free Oxylipins in plasma samples were analyzed using HPLC-MS/MS. Liver fat content and fibrosis were evaluated using Fibroscan technology. Patients with the homozygous ancestral (minor) FADS1 genotype exhibited significantly lower blood levels of the n-6 PUFA arachidonic acid (AA), but no significant differences in the n-3 PUFAs eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). There were no significant differences in liver fat content or arachidonic acid-derived lipid mediators, such as thromboxane B2 (TXB2), although there was a trend toward lower levels in the homozygous ancestral genotype group. Our findings suggest that FADS1 genotypes influence the blood levels of n-6 PUFAs, while not significantly affecting the n-3 PUFAs EPA and DHA. The lack of significant differences in liver fat content and arachidonic acid-derived lipid mediators suggests that the genotype-related variations in fatty acid levels may not directly translate to differences in liver fat or inflammatory lipid mediators in this cohort. However, the trend towards lower levels of certain lipid mediators in the homozygous ancestral genotype group warrants further investigation to elucidate the underlying mechanisms of different FADS1 genotypes and potential implications for cardiometabolic risk.

A Molecular Characterization of the Allelic Expression of the BRCA1 Founder Δ9-12 Pathogenic Variant and Its Potential Clinical Relevance in Hereditary Cancer.

Hereditary breast and ovarian cancer (HBOC) syndrome is a genetic condition that increases the risk of breast cancer by 80% and that of ovarian cancer by 40%. The most common pathogenic variants (PVs) causing HBOC occur in the BRCA1 gene, with more than 3850 reported mutations in the gene sequence. The prevalence of specific PVs in BRCA1 has increased across populations due to the effect of founder mutations. Therefore, when a founder mutation is identified, it becomes key to improving cancer risk characterization and effective screening protocols. The only founder mutation described in the Mexican population is the deletion of exons 9 to 12 of BRCA1 (BRCA1Δ9-12), and its description focuses on the gene sequence, but no transcription profiles have been generated for individuals who carry this gene. In this study, we describe the transcription profiles of cancer patients and healthy individuals who were heterozygous for PV BRCA1Δ9-12 by analyzing the differential expression of both alleles compared with the homozygous BRCA1 control group using RT-qPCR, and we describe the isoforms produced by the BRCA1 wild-type and BRCA1Δ9-12 alleles using nanopore long-sequencing. Using the Kruskal-Wallis test, our results showed a similar transcript expression of the wild-type allele between the healthy heterozygous group and the homozygous BRCA1 control group. An association between the recurrence and increased expression of both alleles in HBOC patients was also observed. An analysis of the sequences indicated four wild-type isoforms with diagnostic potential for discerning individuals who carry the PV BRCA1Δ9-12 and identifying which of them has developed cancer.

Mapping the Behavioral Signatures of Shank3b Mice in Both Sexes

Autism spectrum disorders (ASD) are characterized by social and repetitive abnormalities. Although the ASD mouse model with Shank3b mutations is widely used in ASD research, the behavioral phenotype of this model has not been fully elucidated. Here, a 3D-motion capture system and linear discriminant analysis were used to comprehensively record and analyze the behavioral patterns of male and female Shank3b mutant mice. It was found that both sexes replicated the core and accompanied symptoms of ASD, with significant sex differences. Further, Shank3b heterozygous knockout mice exhibited distinct autistic behaviors, that were significantly different from those those observed in the wild type and homozygous knockout groups. Our findings provide evidence for the inclusion of both sexes and experimental approaches to efficiently characterize heterozygous transgenic models, which are more clinically relevant in autistic studies.

Are transmembrane 6 superfamily member 2 gene polymorphisms associated with steatohepatitis after pancreaticoduodenectomy?

After pancreaticoduodenectomy, 20-40% of patients develop steatotic liver disease (SLD), and steatohepatitis can be a problem. Although patatin-like phospholipase domain-containing 3 protein (PNPLA3) and transmembrane 6 superfamily member 2 (TM6SF2) polymorphisms are involved in SLD and steatohepatitis development, whether this is the case after pancreaticoduodenectomy is unclear. Forty-three patients with pancreatic cancer who underwent pancreaticoduodenectomy at our hospital between April 1, 2018, and March 31, 2021, were included. We extracted DNA from noncancerous areas of residual specimens after pancreaticoduodenectomy and determined PNPLA3 and TM6SF2 gene polymorphisms using real-time polymerase chain reaction. SLD was defined as a liver with an attenuation value of ≤40 HU or a liver-to-spleen ratio of ≤0.9 on computed tomography. We defined high hepatic fibrosis indexes (HFI) instead of steatohepatitis as a Fibrosis-4 index of ≥2.67 or nonalcoholic fatty liver disease fibrosis score of ≥0.675 in patients with SLD. The cumulative incidence of SLD (P = 0.299) and high HFI (P = 0.987) after pancreaticoduodenectomy were not significantly different between the PNPLA3 homozygous and minor allele groups. The incidences of high HFI at 1 year after pancreaticoduodenectomy were 16.8% and 27.0% in the TM6SF2 major homozygous and minor allele groups, respectively, with a significant difference in the cumulative incidence (P = 0.046). The TM6SF2 minor allele may contribute to steatohepatitis development after pancreaticoduodenectomy.

CYP1A2 Polymorphism and Drug Co-administration Affect the Blood Levels and Adverse Effects of Pirfenidone.

Mutations in metabolic enzymes and co-administration of drugs may affect the blood concentration of pirfenidone effective in pulmonary fibrosis. To provide a basis for the precise clinical use of pirfenidone, the authors analyzed the correlation between steady-state pirfenidone trough concentration and adverse drug reactions (ADRs) and examined the impact of CYP1A2*1C (rs2069514) and *1F (rs762551) variants and co-administration on pirfenidone blood concentrations and ADRs. Forty-four patients were enrolled. The blood concentration of pirfenidone was determined using high-performance liquid chromatography. CYP1A2*1C and *1F genotypes were determined using direct SNP sequencing. Additional information related to drug associations was collected to screen factors affecting drug metabolism. The highest predictive value of ADRs was observed when the steady-state trough concentration of pirfenidone was 3.18 mcg·mL-1 and the area under the receiver operating characteristic curve was 0.701 (P = 0.024). The pirfenidone concentration-to-dose ratio (C/D) in CYP1A2*1F homozygous AA mutants was lower than that in C carriers (CC+AC) (1.28 ± 0.85 vs. 2.03 ± 1.28 mcg·mL-1; P = 0.036). Adverse drug reaction (ADR) incidence in the homozygous AA mutant group (28.0%) was significantly lower than that in the C carriers (CC+AC) (63.2%; P = 0.020), and ADR incidence in the A carriers (AC+AA) was considerably lower than that in the CC group (85.7%; P = 0.039). The C/D value of the combined lansoprazole/rabeprazole group was lower than that of the noncombination group (P < 0.05). The ADR incidence was positively correlated with pirfenidone blood concentration. The CYP1A2 (rs762551) AA genotype is associated with lower pirfenidone concentrations and fewer ADRs. Lansoprazole/rabeprazole co-administration reduced pirfenidone concentrations. Randomized controlled trials should further explore personalized dosing of pirfenidone and combination therapies.

Spinal Manifestation of Sickle Cell Disease. A Report On 35 Patients with Literature Review

Background: Sickle cell disease is a hereditary hemoglobinopathy affecting the spine in a wide range of pathology. Despite the several pathological involvements, some patients remain asymptomatic even with positive imaging findings. Objectives: Theme of this study is to report the various pathological disorders involving the spine of patients with sickle cell disease in Basra “south of Iraq” Patients and method: 35 Homozygous and heterozygous sickle cell patients were included in this study. 22 Males and 13 females, their age range from 6 to 69 years they were evaluated clinically, and also by laboratory imaging studies like plain radiography, computerized tomography, magnetic resonance imaging and mineral bone density “DEXA scan “ Results: 54% of the patients were homozygous, and the remaining were heterozygous, 66% of them were asymptomatic, back pain was the complaint in 60% while spine tenderness was reported in 40%, 31% had tenderness in the sacroiliac region and only 6% had neurological claudication and focal neurological deficit. Biconcave vertebra was confirmed in 72% of the patients and mainly localized in the lumbar Spine, MRI findings of vertebral infarction were observed in 58% of the studied patients while sacroiliac joint changes appeared in 60%, 60% of those who had DEXA scan showed low mineral bone density in the lumbar spine Conclusion: Sickle cell disease involve the whole Spine in a wide range of pathological process, some show clinical symptoms, and some remain asymptomatic in spite of changes. The structural changes are more obvious in the homozygous group (55) the lumber spine is dominantly involved, disc prolapse, and neurological deficit is rare while the low mineral changes is common. We did not report a case of spondylodiscitis in this series, though it is very much expected, so routine screening of the spine in sticklers is mandatory

Should We Consider UGT1A1 Mutation Analysis in Evaluating the Prolonged Jaundice of Newborn Infants?

Purpose: Uridine diphosphate glucuronosyltransferase 1A isoform 1 (&lt;i&gt;UGT1A1&lt;/i&gt;) is a crucial enzyme in bilirubin metabolism. Mutations in this gene cause prolonged unconjugated hyperbilirubinemia in infants. This study aimed to investigate the prevalence of &lt;i&gt;UGT1A1&lt;/i&gt; mutations and their association with prolonged, unexplained, and unconjugated hyperbilirubinemia in infants.Methods: From July 2019 to March 2023, 74 infants with prolonged jaundice lasting &gt;21 days were enrolled in this study. Diagnostic evaluations, including &lt;i&gt;UGT1A1&lt;/i&gt; mutation analysis, were performed to identify the underlying causes of hyperbilirubinemia. This retrospective review evaluated the incidence and types of mutations in &lt;i&gt;UGT1A1&lt;/i&gt;. The clinical and laboratory findings were compared based on the specific mutations detected.Results: Thirty-three infants agreed to &lt;i&gt;UGT1A1&lt;/i&gt; mutation analysis, and 30 (90.9%) were positive for &lt;i&gt;UGT1A1&lt;/i&gt; mutations. Single-nucleotide variants were detected in 20 (66. %) infants. The remaining 10 (33.3%) infants had multiple variants. No significant demographic differences were observed between the group that underwent &lt;i&gt;UGT1A1&lt;/i&gt; mutation analysis and the group that did not. Among the identified genetic variants, c.211G&gt;A (46.5%) and c.-3275T&gt;G (30.2%) were the two most common variants. The other variants had the following percentages: c.1456T&gt;G, c.-64G&gt;C, and c.1091C&gt;T (4.7% each); and c.-3152G&gt;A, c.189C&gt;T, and c.-41.-40 dup (2.3% each). Among the 20 infants with the c.211G&gt;A variant, eight (40.0%) had a homozygous genotype and 12 (60.0%) had a heterozygous genotype. Infants harboring other variants exhibited heterozygous genotypes. When comparing the group with confirmed &lt;i&gt;UGT1A1&lt;/i&gt; mutations to the group without detected mutations, breastfeeding was the only significant factor (&lt;i&gt;p&lt;/i&gt;=0.027). No significant differences were found between the group with singlenucleotide variants and the group with multiple genetic variants or between the homozygous genotype group with c.211G&gt;A and the heterozygous genotype group.Conclusion: Neonates with prolonged unconjugated hyperbilirubinemia may have a higher chance of &lt;i&gt;UGT1A1&lt;/i&gt; mutation than expected. Analysis of &lt;i&gt;UGT1A1&lt;/i&gt; mutations may be beneficial in infants with prolonged unexplained jaundice.

Study based on the acetaldehyde dehydrogenase 2 gene polymorphism and acetaminophen-induced liver injury

Objective: To explore the association between aldehyde dehydrogenase 2 (ALDH2) gene polymorphisms and abnormal liver function-induced by acetaminophen (APAP) drugs. Methods: An ALDH2 gene knockout mouse model was constructed using CRISPR/Cas9 gene editing technology. The obtained heterozygous mice were mated with opposite sex of heterozygotes. Genomic DNA was extracted from the tail of the offspring mouse. The polymerase chain reaction (PCR) method was used to determine the ALDH2 genotype. APAP was further used to induce acute drug-induced liver injury models in wild-type and ALDH2 knockout mice. Blood and liver tissues of mice were collected for liver function index, HE staining, F4/80 immunohistochemistry, and other detections. The intergroup mean was compared using a one-way ANOVA. The LSD- t test was used for pairwise comparison. Results: ALDH2 knockout mice were bred successfully. The genotyping of the offspring was segregated into the wild-type (ALDH2(+/+)), heterozygous mutant (ALDH2(+/-)), and homozygous mutant (ALDH2(-/-)), respectively. Biochemical and histological results after APAP modeling showed that the level of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), and total bilirubin (TBil) was not significantly increased in the blank control group (P < 0.05), while the ALT, AST,ALP, and TBil were all elevated in the APAP experimental group. The levels of ALT (P = 0.004), AST (P = 0.002), and TBil (P = 0.012) were significantly elevated among the mutant group compared to those in the wild-type group, and the expression levels of these indicators were also significantly elevated among the homozygous mutant group compared to those in the heterozygous mutant group (P = 0.003, 0 and 0.006). In addition, the ALP levels were higher in the heterozygous mutation group than those in the homozygous mutant group (P = 0.085) and wild-type group mice, but the difference was only statistically significant compared to wild-type mice (P = 0.002). HE staining results showed that mice in the APAP experimental group had hepatocyte degeneration, necrosis, and increased inflammatory cell infiltration, which was mostly evident in mutant mice. Simultaneously, the F4/80 immunohistochemical staining results showed that brown granules were visible in the liver tissue of APAP experimental group mice, and its expression levels were significantly enhanced compared to the blank control group. Conclusion: APAP-induced liver function abnormalities were associated with the ALDH2 gene polymorphism. The liver injury symptoms were increased in ALDH2 mutant mice following APAP modeling, and the ALDH2 gene defect may alleviate, to some extent, APAP-induced liver function abnormalities.

Association between ApoE ε4 genotype and attentional function in non-demented, middle-aged, and older adults from rural India.

Several genetic factors have been associated with cognitive decline in aging. Apolipoprotein E (ApoE) ε4 has been widely studied in the risk for pathological cognitive decline, including dementia. However, the association between ApoE ε4 and cognitive functioning in the healthy aging Indian population has been understudied, and the results are ambiguous. This study aims to examine the role of the ApoE genotype with attentional function in aging adults (≥45 years) in a rural Indian population. Cross-sectional (baseline) data (n = 2100) was utilized from an ongoing longitudinal cohort study on aging (Srinivaspura Aging, Neurosenescence, and Cognition study). Participants hailed from villages of Srinivaspura in Karnataka, southern India. Participants were categorized based on ApoE-ε4 status into three categories: No ε4, heterozygous ε4, and homozygous ε4. Attentional function was assessed using the auditory and visual attention subtests from a computerized neurocognitive test battery. Linear regression was performed adjusting for age, gender, and education. In model 1 (unadjusted), we did not find an association between ApoE and attention function. In the partially adjusted model 2 (adjusting for age), ApoE ε4 with age was significantly associated with the attention function. Further, with increasing age, there was a decline in attention among homozygous ε4 individuals. Model 3 (model 2 + gender) found that ApoE ε4, age, and gender explained a significant variance in attention function. In addition, with increasing age, males had poor attention in the homozygous as compared to heterozygous group. Model 4 (model 3+ education) explained a significant variance in attention and also revealed that with increasing age, attention declined in the illiterate and low literacy groups in both homozygous and heterozygous groups among both genders. Although ApoE ε4 alone was not associated, it interacted with age, gender, and education to affect attention function in this rural Indian population. Longitudinal cognitive monitoring will yield insights into understanding whether the ApoE ε4 genotype influences the rate of cognitive decline in this rural, aging population.

Splicing Polymorphism rs12459419 Modulates Antibody-Bound Internalization of CD33 Isoforms

Background: While the standard regimen for AML has been intense chemotherapy followed by stem-cell transplantation (SCT), addition of immunotherapeutics such as gemtuzumab ozogamicin (GO) to this has shown to improve outcomes in multiple clinical trials. Since these trials, GO has been added to the National Comprehensive Cancer Network (NCCN) practice guidelines for use in AML therapy (NCCN., 2018). GO is a humanized monoclonal antibody conjugated to calicheamicin and targets the cell surface antigen CD33, observed on most AML blasts. Upon binding to GO, CD33 is internalized and the subsequent release of calicheamicin mediates cytotoxicity. CD33 contains two extracellular domains: IgV and IgC, the IgV domain is coded by CD33 exon 2 and is the binding site for GO. A splicing SNP, rs12459419 (C&amp;gt;T; A14V) in exon 2 results in a shorter isoform (CD33-D2) and has been previously associated with CD33 abundance on cell surface as well as clinical benefit from GO addition to standard chemotherapy. Patients with CC genotype (~50% of the cohort) expressed full length CD33 (CD33-FL) and benefited significantly from GO. However, heterozygous CT and homozygous TT genotype groups did not derive the same benefit. These results were perplexing as CT genotype group had intermediate levels of CD33-FL and CD33-D2 isoforms (Lamba et al., 2017). It is hypothesized that lack of IgV domain in CD33-D2 and putative CD33-FL/CD33-D2 dimerization prevents GO binding and subsequent internalization in rs12459419 CT/TT genotype cells, however this has never been tested. We have generated novel antibodies directed against the IgC domain that recognizes CD33-D2 in AML cell lines as well as primary AML cells (Gbadamosi et al., 2021) thus facilitating testing the above indicated hypothesis. Objective: In this study, we first tested the ability of antibody-bound CD33-D2 to internalize, followed by impact of the internalization status of CD33-FL and CD33-D2 in cells co-expressing both isoforms along with the concomitant impact on GO induced cytotoxicity. Methods: Using pMXs retroviral particles encoding CD33-FL or CD33-D2, we engineered CRISPR/Cas9 mediated CD33 KO in AML cell lines to overexpress either one or both CD33 isoforms to mimic cells expression varying proportions of CD33-FL/CD33-D2. Internalization of either CD33-FL or CD33-D2 over a duration of 4 hours in the presence of antibodies recognizing either of the two isoforms was performed. Cells were labeled with unconjugated mouse anti-CD33-FL P67.6 or anti-CD33-D2 HL2541 antibody on ice and incubated at 37°C for 0-4 hours, followed by staining with mouse anti-mouse IgG antibody conjugated with biotin. Surface CD33-FL or CD33-D2 was then detected using streptavidin-FITC conjugate. Percentage antibody-bound CD33-FL or CD33-D2 internalization was expressed as change in FITC MFI between each time point incubated and cells kept at 0°C post CD33 antibody labeling. Results: While P67.6-bound-CD33-FL internalized as expected in AML cell lines expressing CD33-FL only (Fig. 1A), we show for the first time that HL2541-bound-CD33-D2 also internalizes upon antibody binding in cell lines expressing CD33-D2 isoform only (Fig. 1B). In AML cell lines co-expressing both CD33-FL and CD33-D2 isoforms, a &amp;gt;2.5-3 fold decrease in this antibody-bound internalization was observed for each isoform (Figs. 1C and D). In response to GO exposure for 48 hours, cells co-expressing both CD33-FL and CD33-D2 isoforms had 7-9.5-fold higher IC50 values (233 nmol and 277 nmol) compared to cells expressing only CD33-FL (33.19 nmol and 29 nmol) (Figs. 1E and F). As expected, cells expressing CD33-D2 only did respond to GO. Discussion: Our finding that antibody-bound CD33-D2 internalizes like CD33-FL adds a novel piece of evidence in understanding the biology of this variant isoform and opens opportunities to develop antibody drug conjugate (ADC) such as GO using CD33-D2 antibody. This new CD33-D2 targeting ADC will provide a treatment option in patients with TT genotypes. Our results also provide a mechanistic insight on the previous clinical observation showing lack of benefit from GO in patients heterozygous for the splicing SNP. Our ongoing studies are focused on validating this result using live cell imaging and efforts towards design of new ADC or T cell engagers using CD33-D2 antibody. The study was supported by Leukemia Lymphoma Society, UFHealth Cancer Center, and the Frank A. Duckworth Endowment.

A NUDT15 Genetic Polymorphism Study in Adult Acute Leukemia Patients Integrating Pharmacogenomics and Clinical Practice: A Single Centre Experience from South India

Background: Severe myelosuppression secondary to thiopurines such as 6-Mercaptopurine (6-MP) in patients with acute leukemia undergoing maintenance therapy is frequently attributed to inherited genetic polymorphisms of Thiopurine Methyltransferase (TPMT); however, this is rare in the Asian population. Nucleoside diphosphate-linked moiety X-type motif 15 (NUDT15) is also responsible for encoding critical enzymes involved in 6-MP metabolism. We aimed to determine the frequency of genetic polymorphisms of NUDT15 in the South Indian population and the impact of the omission of 6-MP from the treatment protocol of these patients. Methods: We retrospectively reviewed 17 patients diagnosed with acute leukemia between May 2018 and May 2020 and analyzed the prevalence of NUDT15 genetic polymorphisms in these patients. The NUDT15 mutation study (NUDT15 415C&amp;gt;T) used allele-specific Real-Time PCR in peripheral blood. 6-Mercaptopurine (6-MP) was excluded from the treatment regimen of patients with NUDT15 genetic polymorphisms and the impact of its exclusion was also studied. Results: Among the 17 patients for whom we cared, 10 were diagnosed with B-cell Acute Lymphoblastic Leukemia (B-ALL), three with T-cell Acute Lymphoblastic Leukemia (T-ALL), one with Mixed Phenotype Acute Leukemia (MPAL), and one with Acute Promyelocytic Leukemia (APML). Treatment records could not be retrieved for 2 patients. NUDT15 mutations were present in six patients (36%) (see fig1), of whom two had homozygous (33%) polymorphisms and four had heterozygous (67%) polymorphisms. In the heterozygous group, 2 were Ph. (Philadelphia chromosome) negative B-ALL; one was treated with BFM and the other with hyper-CVAD, one Ph. positive B-ALL was treated with BFM + Ponatinib, and one High-risk APML was treated with the APML 4 protocol. In the homozygous group, treatment details could not be retrieved for one case, and the other patient had T-ALL, which was treated with hyper-CVAD. 6-MP was omitted from the treatment protocol for all NUDT15 mutation-positive patients, both homozygous and heterozygous. Of these patients, two developed grade 1 myelosuppression, one developed grade 2 myelosuppression, one developed dyselectrolytemia, and one patient did not develop any toxicity. The incidence of severe or grade 3 and 4 myelosuppression in NUDT15 polymorphisms receiving 6MP can be as high as 50% (Khaeso K et.al Front Pharmacol. 2021). In comparison, none of our patients developed grade 3 or 4 myelosuppression. Overall, treatment-related adverse events included grade 1-2 myelosuppression (n=6), grade 1-2 hepatotoxicity (n=1), deep vein thrombosis (n=2), and sepsis (n=2). Five of the 6 patients with NUDT15 mutations achieved complete remission (CR). Conclusion: The prevalence of NUDT15 polymorphisms was high in our study population. Based on a meta-analysis, the prevalence of NUDT15 in the South Indian population was 16.5%, and in patients with adverse effects, the pooled prevalence of NUDT15 polymorphisms was 49.51% (Anuraag Jena et.al. Expert Review of Clinical Pharmacology 2021). Based on our results, although limited by the small number of cases, we suggest that pre-emptive testing to detect these polymorphisms should be undertaken in all adult acute leukemia patients of South Indian origin, given their high prevalence in this population. The omission of 6-MP from treatment protocols can help prevent severe myelosuppression and treatment interruption or discontinuation, which in turn can improve treatment outcomes.

The Chinese-Han Population Carrying Wild-type Genotypes of SLCO1B1 388A&gt;G, SLCO1B1 521T&gt;C, CYP3A4 1B, CYP3A4 1G, and CYP3A5*3 Exhibits a Significant Alteration in the Pharmacokinetics of Atorvastatin Calcium

Due to the diverse genetic characteristics of metabolism and high drug plasma exposure, great inter-subject variability exists in the clinical efficacy and incidence of adverse events. This study aimed to evaluate the associations between the SLCO1B1 388A＞G (rs2306283) polymorphism and the pharmacokinetics of atorvastatin calcium (AC) in healthy volunteers who carried the wild genotypes of SLCO1B1 521T＞C (rs4149056), CYP3A4 1B (rs2740574), CYP3A4 1G (rs2242480), and CYP3A5*3 (rs776746). A FISH technique was employed to investigate the genetic polymorphisms in 187 healthy male volunteers. The pharmacokinetic study was conducted on a group of healthy male Chinese-Han volunteers with wild-type genotypes of SLCO1B1 521T&gt;C, CYP3A4 1B, CYP3A41G and CYP3A53 genes, consisting of either mutant heterozygotes (n=10) or mutant homozygotes (n=10) of SLCO1B1 388A&gt;G. The results were then compared to the pharmacokinetic parameters of AC in subjects with the wild-type genotype of SLCO1B1 388A&gt;G, as previously described. Based on the distribution of genotypes, the 187 volunteers could be divided into 28 groups. The top 10 groups accounted for nearly 85% of the total volunteers. No significant differences (P&gt;0.05) were observed in the pharmacokinetic parameters between subjects carrying homozygous and heterozygous genotypes of SLCO1B1 388A&gt;G. However, The Cmax of subjects carrying the wild-type genotype of SLCO1B1 388A＞G was about 14.75 times higher than that of the heterozygous genotype group and 10.43 times higher than that of the homozygous genotype group. The AUC0-72h of volunteers with the wild-type genotype of SLCO1B1 388A＞G was about 13.81 times higher than that of the heterozygous genotype group and 11.96 times higher than that of the homozygous genotype group. Volunteers carrying wild genotypes of SLCO1B1 388A＞G, SLCO1B1 521T＞C, CYP3A4 1B, CYP3A4 1G, and CYP3A5*3 showed significantly higher levels of Cmax and AUC (P&lt;0.01), as well as markedly decreased values of CLz/F and Vz/F (P&lt;0.01) of AC. In conclusion, patients carrying the wild genotype of SLCO1B1 388A&gt;G, SLCO1B1 521T&gt;C, CYP3A41G, CYP3A41B, and CYP3A5*3 should receive a lower dose of AC to minimize the risk of adverse effects.

The Clinical Characteristics and Prognosis of Exon 2 Mutations in Familial Mediterranean Fever

Objective: It is unclear whether exon 2 mutations are variations or a mutations that causes the disease. This study aimed to evaluate the clinical features and prognosis exon 2 mutations in Familial Mediterranean Fever. Methods: The clinical features, disease severity and prognosis of all patients with at least one exon 2 mutations were evaluated retrospectively. These data were compared seperately for homozygous (Group 1), heterozygous (Group 2), compound heterozygous (Group 3), and complex alleles (Group 4), and the data were compared by grouping patients into those with and without exon 10 mutations. Results: There were a total of 119 patients with exon 2 mutations, including 11.7% in Group 1, 36.1% in Group 2, 21.8% in Group 3, and 30.2% in Group 4 were similar in terms of demographic data, clinical characteristics, and disease course. When compared patients with exon 10 mutations (+) to those with exon 10 mutations (-), the exon 10 mutations (+) group had a higher presence of chest pain (100%, p=0.02) and a significantly higher mean Pras severity score (6.66±1.87, 6.01±1.40; p=0.02). Additionally, a higher number of patients with exon 10 mutation (-) achieved remission with treatment (76 (67.9%), 36 (32.1%); p=0.03). Conclusion: Exon 2 mutations have a milder course and higher remission rates but they should be considered as Familial Mediterranean Fever disease because of their similar clinical presentation and response to colchicine treatment with exon 10 mutations. Early treatment and close follow-up should be performed.

Effects of GLP-1 Receptor Polymorphisms on Adolescent Obesity

Obesity is becoming a concerning disease in developing countries. Like other multifactorial diseases, genetics plays a substantial role in the development of this disease. We tried to investigate genetic variations (mutation/polymorphism) of GLP-1R gene in children diagnosed with obesity and to identify their possible connections with obesity and other conditions. Genomic DNA was extracted from 162 overweight/obese patients and 100 controls. Later, full exon sequencing and association studies were carried out. Three polymorphisms and one mutation were detected in the fourth and fifth exons of the GLP-1R gene. Some variations were detected in three cases from which 1/3 had non-alcoholic fatty liver disease (NAFLD) but none showed insulin resistance (IR). There were also statistically meaningful results for ‘Odds Ratio’ among different genotypes and allele frequencies in groups with NAFLD and/or IR. In addition, there was an increase in risk for NAFLD and a decrease in risk for IR. In the homozygous group, also the prospect of IR was double declined. Patients with the A allele of this polymorphism showed a drop in risk for IR as well. GLP-1R polymorphisms could influence obesity and diabetes and thus the functional analysis of the GLP-1R polymorphisms is benevolent.

Blood glucose trends in glycogen storage disease type Ia: A cross-sectional study.

Glycogen storage disease type Ia (GSDIa) is caused by biallelic pathogenic variants in the glucose-6-phosphatase gene (G6PC) and mainly characterized by hypoglycemia, hepatomegaly, and renal insufficiency. Although its symptoms are reportedly mild in patients carrying the G6PC c.648G>T variant, the predominant variant in Japanese patients, details remain unclear. Therefore, we examined continuous glucose monitoring (CGM) data and daily nutritional intake to clarify their associations in Japanese patients with GSDIa with G6PC c.648G>T. This cross-sectional study enrolled 32 patients across 10 hospitals. CGM was performed for 14 days, and nutritional intake was recorded using electronic diaries. Patients were divided according to genotype (homozygous/compound heterozygous) and age. The durations of biochemical hypoglycemia and corresponding nutritional intake were analyzed. Multiple regression analysis was performed to identify factors associated with the duration of biochemical hypoglycemia. Data were analyzed for 30 patients. The mean daily duration of hypoglycemia (<4.0 mmol/L) in the homozygous group increased with age (2-11 years [N = 8]: 79.8 min; 12-18 years [5]: 84.8 min; ≥19 years [10]: 131.5 min). No severe hypoglycemic symptoms were recorded in the patients' diaries. The mean frequency of snack intake was approximately three times greater in patients aged 2-11 years (7.1 times/day) than in those aged 12-18 years (1.9 times/day) or ≥19 years (2.2 times/day). Total cholesterol and lactate were independently associated with the duration of biochemical hypoglycemia. Although nutritional therapy prevents severe hypoglycemia in patients with GSDIa with G6PC c.648G>T, patients often experience asymptomatic hypoglycemia.

The Validation of a Precursor Lesion of Epithelial Ovarian Cancer in Fancd2-KO Mice.

Ovarian cancer (OC) has the highest mortality rate of all gynaecological malignancies. The asymptomatic nature and limited understanding of early disease hamper research into early-stage OC. Therefore, there is an urgent need for models of early-stage OC to be characterised to improve the understanding of early neoplastic transformations. This study sought to validate a unique mouse model for early OC development. The homozygous Fanconi anaemia complementation group D2 knock-out mice (Fancd2-/-) develop multiple ovarian tumour phenotypes in a sequential manner as they age. Using immunohistochemistry, our group previously identified purported initiating precursor cells, termed 'sex cords', that are hypothesised to progress into epithelial OC in this model. To validate this hypothesis, the sex cords, tubulostromal adenomas and equivalent controls were isolated using laser capture microdissection for downstream multiplexed gene expression analyses using the Genome Lab GeXP Genetic Analysis System. Principal component analysis and unbiased hierarchical clustering of the resultant expression data from approximately 90 OC-related genes determined that cells from the sex cords and late-stage tumours clustered together, confirming the identity of the precursor lesion in this model. This study, therefore, provides a novel model for the investigation of initiating neoplastic events that can accelerate progress in understanding early OC.

Association between FOXO3a Gene Polymorphism and Susceptibility to Asthma in South Indian Population

Background: Asthma is an important cause of morbidity worldwide. The FOXO3a gene plays an important role in human immune regulation and homeostasis. Loss of function may lead to chronic inflammation and asthma. Objective: To evaluate the association between Foxo3a gene polymorphism and susceptibility to asthma. Methods: We conducted a case-control study in a tertiary care hospital. Participants answered a questionnaire that collected demographic and clinical information. Asthma was confirmed by pre and post bronchodilator spirometry. Genotyping of the FOXO3a polymorphisms was performed us-ing PCR-RFLP. Results: The study population included 41 cases and 33 controls. Among cases, the heterozygous (CT) genotype frequency was greater compared to wild homozygous (CC) and mutant homozygous (TT) variants. In controls, the wild homozygous (CC) genotype frequency was greater compared to heterozygous and mutant homozygous variants. In the general model, the mutant homozygous (TT) group had significantly higher odds of 7.8 (1.78-34.07) of having asthma compared to the wild ho-mozygous (CC) group. The mutant homozygous (TT) group had greater severity of asthma as com-pared to the wild homozygous (CC) group. The mutant homozygous group (TT) had much lower lung functions, as compared to the wild homozygous (CC) group in asthmatics. Among non-asthmatic controls, lower lung functions were seen in the mutant (TT) group as compared to the wild (CC) group. Conclusion: We found a significant association between the Foxo3a gene polymorphism and asth-ma. The T allele, a variant of the Foxo3a gene polymorphism, is associated with a higher risk of asthma, and greater asthma severity. It is also associated with lower lung functions in both asthmat-ics and apparently healthy control subjects.