The Structural Effect of FLT3 Mutations at 835th Position and Their Interaction with Acute Myeloid Leukemia Inhibitors: In Silico Approach.

FMS-like tyrosine kinase 3 (FLT3) gene mutations have been found in more than one-third of Acute Myeloid Leukemia (AML) cases. The most common point mutation in FLT3 occurs at the 835th residue (D835A/E/F/G/H/I/N/V/Y), in the activation loop region. The D835 residue is critical in maintaining FLT3 inactive conformation; these mutations might influence the interaction with clinically approved AML inhibitors used to treat the AML. The molecular mechanism of each of these mutations and their interactions with AML inhibitors at the atomic level is still unknown. In this manuscript, we have investigated the structural consequence of native and mutant FLT-3 proteins and their molecular mechanisms at the atomic level, using molecular dynamics simulations (MDS). In addition, we use the molecular docking method to investigate the binding pattern between the FLT-3 protein and AML inhibitors upon mutations. This study apparently elucidates that, due to mutations in the D835, the FLT-3 structure loses its conformation and becomes more flexible compared to the native FLT3 protein. These structural changes are suggested to contribute to the relapse and resistance responses to AML inhibitors. Identifying the effects of FLT3 at the molecular level will aid in developing a personalized therapeutic strategy for treating patients with FLT-3-associated AML.

Novel Biological-Based Strategy for Synthesis of Green Nanochitosan and Copper-Chitosan Nanocomposites: Promising Antibacterial and Hematological Agents.

Two novel samples of nanoparticles based on chitosan were greenly synthesized using pomegranate peel extract. The extract served as a nanoparticle precursor, facilitating the precipitation of nanosized chitosan through the ionic gelation method. Additionally, by mixing the green chitosan nanoparticles with copper ions, a nanoscale composite of chitosan and copper oxide was also produced. Structural and morphological investigations (FTIR, XRD, SEM, EDX, and TGA analyses) were performed for greenly synthesized chitosan nanoparticles and their copper oxide composite to determine all the significant characteristics of those nanoparticles. In addition, both samples were tested using some biological investigations, such as antimicrobial activity and hematological effects. The antimicrobial tests yielded promising results for both the green chitosan nanoparticles and the CuO composite when tested using two bacterial strains and two fungal strains. Moreover, the results showed that using a similar concentration of both green-based chitosan samples resulted in a slightly larger inhibition zone and a lower minimum inhibition concentration (MIC) for the copper oxide chitosan composite compared to the chitosan nanoparticles for all microorganisms included in the test. The mean count of blood components (RBCs and platelets), clotting time, and cholesterol levels in three different blood samples were used to indicate the hematological activity of both greenly synthesized nanoparticles. The results verified a slight reduction in blood component count after the addition of green chitosan nanoparticles, but the chitosan copper oxide composite did not have a noticeable effect on the three blood samples. The chitosan nanoparticles were able to cause a considerable reduction in clotting time and cholesterol levels for all blood samples, thus acting as procoagulants. However, the mixing of CuO with chitosan nanoparticles prolonged the rate of clotting in blood samples from hypercholesteremic individuals, and thus, the mixture acted as an anticoagulant agent.

Anti-annexin V IgG and IgM antibodies in sickle cell disease patients with vaso-occlusive crisis.

Vaso-occlusive crisis (VOC) is a significant cause of morbidity and mortality in sickle cell anemia (SCA) patients; however, its mechanisms are poorly understood. In view of their prothrombotic nature, we hypothesized that SCA-associated VOC may be due to the presence of anti-annexin V antibodies. Anti-annexin V antibodies were measured with ELISA in 177 VOC and 81 steady-state SCA patients. Anti-annexin V IgM and IgG concentrations were significantly higher in VOC patients than in steady-state patients and were associated with elevated VOC risk. After categorizing anti-annexin V antibodies, the adjusted odds ratio increased as the percentile value increased. Monovariate logistic regression analysis demonstrated a positive dose-effect relationship for anti-annexin V IgM with VOC, with increased VOC risk seen with increased antibody titers. Multivariate logistic regression analyses confirmed the association of anti-annexin V IgM, more so than IgG, as an independent VOC risk factor. Anti-annexin V IgG antibodies correlated positively with VOC type and negatively with HbF and age of VOC onset, while anti-annexin V IgM correlated positively with VOC type, duration, frequency, site, pain severity, hospitalization, and medication, and negatively with age of VOC onset and HbS levels. High levels of anti-annexin V IgM antibodies constitute a risk factor for VOC in SCA patients.

Case Report: Case report: Mixed infection of Plasmodium vivax and Plasmodium falciparum in a tertiary hospital.

Mixed infections with two or more species of Plasmodium are frequently reported due to vector factors, parasite factors (formation of hypnozoites) and host factors (residing in endemic areas, travel to endemic areas, inadequately treated previous infection, lack of compliance to therapy). Here we report a case of a 33-year-old Saudi female who had a significant travel history, and a peripheral blood smear (PBS) revealed mixed infection with P. falciparum and P. vivax. The case was successfully treated with a combination therapy of artemisinin and primaquine with follow up testing at three, seven, 14, and 28 days. Mixed malaria infections are especially reported in travelers to endemic areas. Hence, adequate diagnosis and appropriate treatment of the cases contributes majorly to preventing relapse and controlling the disease. Travel consultations should be given to all travelers before their trips to endemic countries.

Coagulopathies in novel coronavirus (SARS-CoV-2) pandemic: Emerging evidence for hematologists.

The coronavirus disease (COVID-19), which is also known as acute respiratory syndrome coronavirus-2 (SARS-CoV2) is a transmissible disease, has phenotypes varying from asymptomatic to Acute Respiratory Distress Syndrome (ARDS) or multiple organ dysfunction syndrome (MODS) and ultimately death in certain cases. Coagulation disorders are being frequently reported amongst these patients and the pathogenesis is still not completely understood. Proposed mechanisms for these coagulopathies comprise a hypercoagulable state with micro- and/or macro-thrombosis in the vessels. A number of changes have been reported or proposed in circulating prothrombotic factors in COVID-19 patients and includes elevation in both factor VIII and fibrinogen, circulating prothrombotic microparticles and hyperviscosity. The COVID-19 patients are showing varied coagulopathies and are at high risk for venous thromboembolism (VTE) which demands an early intervention. This paper reviews the evolving data regarding the evaluation and managing of coagulopathies in patients with COVID-19.

Investigation of nonsynonymous mutations in the spike protein of SARS-CoV-2 and its interaction with the ACE2 receptor by molecular docking and MM/GBSA approach.

COVID-19 is an infectious and pathogenic viral disease caused by SARS-CoV-2 that leads to septic shock, coagulation dysfunction, and acute respiratory distress syndrome. The spreading rate of SARS-CoV-2 is higher than MERS-CoV and SARS-CoV. The receptor-binding domain (RBD) of the Spike-protein (S-protein) interacts with the human cells through the host angiotensin-converting enzyme 2 (ACE2) receptor. However, the molecular mechanism of pathological mutations of S-protein is still unclear. In this perspective, we investigated the impact of mutations in the S-protein and their interaction with the ACE2 receptor for SAR-CoV-2 viral infection. We examined the stability of pathological nonsynonymous mutations in the S-protein, and the binding behavior of the ACE2 receptor with the S-protein upon nonsynonymous mutations using the molecular docking and MM_GBSA approaches. Using the extensive bioinformatics pipeline, we screened the destabilizing (L8V, L8W, L18F, Y145H, M153T, F157S, G476S, L611F, A879S, C1247F, and C1254F) and stabilizing (H49Y, S50L, N501Y, D614G, A845V, and P1143L) nonsynonymous mutations in the S-protein. The docking and binding free energy (ddG) scores revealed that the stabilizing nonsynonymous mutations show increased interaction between the S-protein and the ACE2 receptor compared to native and destabilizing S-proteins and that they may have been responsible for the virulent high level. Further, the molecular dynamics simulation (MDS) approach reveals the structural transition of mutants (N501Y and D614G) S-protein. These insights might help researchers to understand the pathological mechanisms of the S-protein and provide clues regarding mutations in viral infection and disease propagation. Further, it helps researchers to develop an efficient treatment approach against this SARS-CoV-2 pandemic.

Investigation of the antimicrobial activity and hematological pattern of nano-chitosan and its nano-copper composite.

Novel synthesized Chitosan-Copper oxide nanocomposite (Cs-CuO) was prepared using pomegranate peels extract as green precipitating agents to improve the biological activity of Cs-NP's, which was synthesized through the ionic gelation method. The characterization of biogenic nanoparticles Cs-NP's and Cs-CuO-NP's was investigated structurally, morphologically to determine all the significant characters of those nanoparticles. Antimicrobial activity was tested for both Cs-NP's and Cs-CuO-NP's via minimum inhibition concentration and zone analysis against fungus, gram-positive and gram-negative. The antimicrobial test results showed high sensitivity of Cs-CuO-NP's to all microorganisms tested in a concentration less than 20,000 mg/L, while the sensitivity of Cs-NP's against all microorganisms under the test started from a concentration of 20,000-40,000 mg/L except for the C. albicans species. The hematological activity was also tested via measuring the RBCs, platelet count, and clotting time against healthy, diabetic, and hypercholesteremia blood samples. The measurement showed a decrease in RBCs and platelet count by adding Cs-NP's or Cs-CuO-NP's to the three blood samples. Cs-NP's success in decreasing the clotting time for healthy and diabetic blood acting as a procoagulant agent while adding biogenic CuO-NP's to Cs-NP's increased clotting time considering as an anti-coagulant agent for hypercholesteremia blood samples.

Human platelet alloantigens (HPA) 1, HPA2, HPA3, HPA4, and HPA5 polymorphisms in sickle cell anemia patients with vaso-occlusive crisis.

OBJECTIVES: Vaso-occlusive crisis (VOC) is a significant cause of morbidity and mortality in sickle cell anemia (SCA) patients. Insofar as polymorphism in human platelet alloantigen (HPA) exhibit a prothrombotic nature, we hypothesized that specific HPA polymorphic variants are associated with VOC. We investigated the distribution of HPA1, HPA2, HPA3, HPA4, and HPA5 alleles genotypes among VOC and non-VOC control SCA patients. PATIENTS/METHODS: This was a case-control study. Study subjects comprised SCA patients with (VOC group; n = 127) or without (Steady-state group; n = 130) VOC events. HPA genotyping was done by PCR-SSP. RESULTS: Significantly higher frequencies of HPA-2b, HPA-3b, and HPA-5b alleles, and marked enrichment of HPA-3b/3b, HPA-5a/5b, and HPA-5b/5b genotypes, were seen in VOC than in control SCA patients. Taking homozygous wild-type genotypes as reference, univariate analysis identified HPA-3a/3b, HPA-3b/3b, and HPA-5b/5b to be associated with VOC. Multivariate analysis confirmed the independent association of only HPA-3a/3b and HPA-3b/3b genotypes with VOC. HPA-3 genotypes were significantly correlated with VOC frequency, type, and medication, and requirement for hospitalization. While both HPA 3a/3b (P = 0.002; OR = 2.94; 95% CI = 1.49-5.77) and 3b/3b (P = 0.006; OR = 3.16; 95% CI = 1.40-7.17) genotypes were associated with need for hospitalization, only HPA-3b/3b was associated with VOC frequency, type (localized vs. generalized), and medication (narcotics vs. NSAIDs). CONCLUSION: This confirms the association of HPA polymorphisms with SCA VOC, of which HPA-3 appears to be independent genetic risk factors for SCA VOC.

Polymorphisms of the human platelet alloantigens HPA-1, HPA-2, HPA-3, and HPA-4 in ischemic stroke.

Polymorphism in human platelet antigen (HPA)-1 and HPA-3 (GPIIb/IIIa), HPA-2 (GPIb/IX), HPA-4 (GPIIIa), and HPA-5 (GPIa/IIa) was investigated in 329 stroke patients and 444 matched control subjects. HPA genotyping was done by PCR-SSP method. Lower HPA-1a (P < 0.001) and higher HPA-1b (P < 0.001) allele frequencies were seen in patients than control subjects, and homozygosity for HPA-1b (P < 0.001) alleles was more prevalent in stroke cases than in controls. The allele and genotype distributions of the other HPA polymorphic variants were similar between cases and controls. Select HPA combined genotypes comprising the 2121 (Pc = 0.008) and 2221 (Pc = 0.018) genotypes, which were positively associated, and the 1111 (Pc < 0.001), which was negatively associated with stroke, thereby conferred a disease susceptibility and protective nature to these genotype combinations. Multivariate analysis confirmed the negative association of the 1111 (P < 0.001) and the positive association of the 2121 (P = 0.017) combined genotypes with stroke, after adjustment for a number of covariates. This is the first evidence demonstrating differential association of the common 4 HPA gene variants and specific HPA genotype combinations with stroke.

Association of human platelet alloantigen 1 through 5 polymorphisms with ischemic stroke.

Polymorphisms in human platelet alloantigen (HPA)-1 and HPA-3 (GPIIb/IIIa), HPA-2 (GPIb/IX), HPA-4 (GPIIIa) and HPA-5 (GPIa/IIa) were investigated in 216 stroke patients and 318 matched control subjects. HPA genotyping was done by the polymerase chain reaction method using sequence-specific primers. Higher frequencies of the HPA-1 a/b (p < 0.001) and HPA-5 a/b (p < 0.001) allele, together with HPA-1 b/b, HPA-5 a/b and HPA-5 b/b genotypes were seen in patients, which was confirmed by regression analysis after controlling for a number of confounding variables. Furthermore, HPA-1 b/b and HPA-5 b/b were significantly associated with the extent of neurological symptoms, and with the recurrence of stroke. Both susceptible (1a/ b -2a/a-3a/ b -4a/a-5a/ b ) and protective (1a/a-2a/a-3a/a-4a/a-5a/a; 1a/a-2a/a-3a/ b -4a/a-5a/a; 1a/ b -2a/a-3a/a-4a/a-5a/a; 1a/ b -2a/a-3a/ b -4a/a-5a/a) HPA genotypes were identified. This is the first evidence demonstrating differential association of the common 5 HPA gene variants with stroke, with HPA-1b and HPA-5b representing strong genetic risk factors.

Platelet function is modified by common sequence variation in megakaryocyte super enhancers.

Linking non-coding genetic variants associated with the risk of diseases or disease-relevant traits to target genes is a crucial step to realize GWAS potential in the introduction of precision medicine. Here we set out to determine the mechanisms underpinning variant association with platelet quantitative traits using cell type-matched epigenomic data and promoter long-range interactions. We identify potential regulatory functions for 423 of 565 (75%) non-coding variants associated with platelet traits and we demonstrate, through ex vivo and proof of principle genome editing validation, that variants in super enhancers play an important role in controlling archetypical platelet functions.

HLA class II haplotypes distinctly associated with vaso-occlusion in children with sickle cell disease.

We investigated the association of HLA class II alleles and haplotypes with sickle cell anemia vaso-occlusive crisis (VOC). DRB1*100101 was positively associated, while DRB1*140101, DRB1*150101, and DQB1*060101 were negatively associated, with VOC. Both susceptible (DRB1*100101-DQB1*050101) and protective (DRB1*110101-DQB1*030101 and DRB1*150101-DQB1*060101) haplotypes were identified, indicating that HLA class II haplotypes influence VOC risk.

Gene frequencies of human platelet alloantigens in Bahraini Arabs.

Human platelet antigens (HPA) are implicated in the pathophysiology of certain hematological disorders, and as varied distribution of HPA-1 alleles and genotypes were reported fordifferent countries and ethnic populations, we determined the distribution of HPA-1, -2, -3, -4, and -5 alleles, genotypes and haplotypes for 194 healthy Bahraini subjects by polymerase chain reaction with sequence specific primers. The distribution of the HPA polymorphisms was in Hardy-Weinberg equilibrium. Allele frequencies of 0.76 and 0.24 (HPA-1a and -1b), 0.77 and 0.23 (HPA-2a and -2b), 0.57 and 0.43 (HPA-3a and -3b), 0.93 and 0.07 (HPA-4a and -4b), and 0.86 and 0.13 (HPA-5a and -5b) were seen. With the exception of HPA-3a/a (30.4%), the frequencies of homozygous HPA-1a/a (56.8%), 2a/a (60.1%), 4a/a (87.2%), and 5a/a (75.7%) were higher than those of heterozygous (a/b) or homozygous (b/b) variants. Our results provide basic information for further studies of the HPA system polymorphism, which in turn will be instrumental in understanding and treating immune-mediated platelet disorders.

Association of the methylenetetrahydrofolate reductase A1298C but not the C677T single nucleotide polymorphism with sickle cell disease in Bahrain.

The association of methylenetetrahydrofolate reductase (MTHFR) gene mutations, C677T and A1298C, together with changes in homocysteine (Hcy) levels was investigated in 106 sickle cell disease patients and 156 healthy controls from Bahrain. The mutation analysis was done by restriction fragment length polymorphism-polymerase chain reaction (RFLP-PCR). While the frequencies of the mutant alleles C677T and A1298C were comparable between patients and controls, the frequency of the A1298C (C/C) (p = 0.03) but not C677T (T/T) (p = 0.67) genotype, and of the 677T/1298C haplotype were significantly higher in the patients (p = 0.05). Homocysteine levels were normal in all subjects. This suggests that the A1298C, but not C677T, mutation is associated with the genotype of sickle cell disease.