Hb Tianshui (HBB: C.119A > G) in Compound Heterozygosity with Hb S (HBB: C.20A > T) from Odisha,India

We describe here a rare β-globin gene variant, Hb Tianshui [β39(C5)Glu→Arg; HBB: c.119A?>?G], detected during routine screening in Odisha, India. This is the second report of Hb Tianshui and the first to describe the cation exchange high performance liquid chromatography (HPLC) and DNA studies of two cases of this variant. Both cases had coinherited Hb S (HBB: c.20A?>?T) but none presented with typical symptoms of sickle cell disease. One of the cases was heterozygous for a common α-thalassemia (α-thal) allele (?α^3.7) (rightward) (NG_000006.1: g.34164_37967del3804) and marginally raised Hb F percentage, while the other Hb S/Hb Tianshui case was completely benign and healthy. An atypical Asian Indian haplotype [+???+???+] could be assigned to the Hb Tianshui variant. Hb Tianshui seems to mimic a few other Hb variants in cation exchange HPLC. However, we report two specific patterns in the chromatograms that are characteristic to Hb Tianshui. Combining an alkaline electrophoresis result with cation exchange HPLC at screening would be preferred to detect this rare variant, especially in regions with considerable frequency of Hb E [β26(B8)Glu→Lys; HBB: c.79G?>?A] or Hb S.     

Compound Heterozygote of Hb S (HBB: c.20A>T)/Hb Westdale (HBB: c.380_396delTGCAGGCTGCCTATCAG): Report of Four Cases from Odisha State,India

We report four cases of compound heterozygotes for Hb S (HBB: c.20A>T) and a rare β⁰-thalassemia (β⁰-thal) mutation, Hb Westdale (HBB: c.380_396delTGCAGGCTGCCTATCAG), characterized by a 17?bp deletion between codons 126 to 131 in exon 3 of the β-globin gene of human hemoglobin (Hb) confirmed by direct β-globin gene sequencing. All four cases were from four unrelated families belonging to the Agharia caste, an endogamous ethnic community of the Sundargarh and Jharsuguda districts of Odisha State, India. Detailed observations indicated that all four cases of Hb S/Hb Westdale were clinically severe. On family screening, six family members were found to be heterozygous for Hb Westdale and were asymptomatic. Deletional α-thalassemia (α-thal) and XmnI polymorphism were studied for all the Hb Westdale cases. The Hb S/Hb Westdale cases had an early median age at onset of symptoms and presentation, more requirement of blood transfusions, splenomegaly and hepatomegaly and were found to be clinically more severe when compared with the Hb S-β-thal with IVS-I-5 (G>C) (HBB: c.92?+?5G>C) cases. Overall, the findings indicate that this rare and hitherto unreported compound heterozygosity of Hb S/Hb Westdale is a clinically significant hemoglobinopathy and its finding in a large endogamous community of Odisha State, India will have important implication in the epidemiology and understanding of the clinical spectrum of sickle cell disease in Indian context and prenatal diagnosis.     

Pricing and availability of some essential child specific medicines in Odisha

Objectives:

Continuous availability of affordable medicines in appropriate formulations is essential to reduce morbidity and mortality in children. Odisha an eastern Indian state records very high mortality of children. The study aims at documenting the availability and prices paid for purchasing essential child-specific medicines.

Materials and Methods:

The survey of 34 essential medicines was conducted in six randomly selected districts of Odisha. Data were collected from medicine outlets of the public, private, and other sector (Nongovernmental Organization [NGO]/mission sectors) of six randomly selected districts, using WHO/Health Action International medicine price collection methodology. For each medicine surveyed, data were collected on the highest and lowest-priced formulations available in each facility.

Results:

Both public sector and other sector health facilities procure only one brand of medicines, mean percentage availability of medicines being 17% and 21.8%, respectively. In the private sector, the mean percentage availability of the high and lowest-priced medicines for a particular drug product was 10.8% and 38.5%, respectively. The public sector procurement price is 48% lower than international reference prices. In the private sector, high-priced, and low-priced products are sold at 1.83 and 1.46 times the international reference price, respectively. Substantial price variation was observed for some medicines across individual outlets. Medicines were found to cost 2.08 times their international reference price in NGO/mission sector facilities.

Conclusions:

The availability of children''s medicines in public sector facilities of Odisha state is poor. Medicines for children cost relatively high in both private and NGO sectors compared to the international reference price. The availability medicines should be improved on an urgent basis to improve access of medicines for children of Odisha.KEY WORDS: Availability, children, drug pricing, essential medicines     

A novel strategy for the manufacture of idelalisib: controlling the formation of an enantiomer

A novel and scalable synthesis of 5-fluoro-3-phenyl-2-[(1S)-1-(9H-purin-6-ylamino)propyl]-4(3H)-quinazolinone, idelalisib 1, has been developed. This strategy controls the desfluoro impurity of 13 during reduction of nitro intermediate 4, and also arrests the formation of the enantiomer during cyclisation of diamide 17, without affecting the neighbouring chiral centre. This process is demonstrated on a larger scale in the laboratory and achieved good chemical and chiral purities coupled with good yields.

A novel and scalable synthesis of 5-fluoro-3-phenyl-2-[(1S)-1-(9H-purin-6-ylamino)propyl]-4(3H)-quinazolinone, idelalisib 1, has been developed.     

Side-by-side comparison of Notch- and C83 binding to γ-secretase in a complete membrane model at physiological temperature

γ-Secretase cleaves the C99 fragment of the amyloid precursor protein, leading to formation of aggregated β-amyloid peptide central to Alzheimer''s disease, and Notch, essential for cell regulation. Recent cryogenic electron microscopy (cryo-EM) structures indicate major changes upon substrate binding, a β-sheet recognition motif, and a possible helix unwinding to expose peptide bonds towards nucleophilic attack. Here we report side-by-side comparison of the 303 K dynamics of the two proteins in realistic membranes using molecular dynamics simulations. Our ensembles agree with the cryo-EM data (full-protein Cα-RMSD = 1.62–2.19 Å) but reveal distinct presenilin helix conformation states and thermal β-strand to coil transitions of C83 and Notch100. We identify distinct 303 K hydrogen bond dynamics and water accessibility of the catalytic sites. The RKRR motif (1758–1761) contributes significantly to Notch binding and serves as a “membrane anchor” that prevents Notch displacement. Water that transiently hydrogen bonds to G1753 and V1754 probably represents the catalytic nucleophile. At 303 K, Notch and C83 binding induce different conformation states, with Notch mostly present in a closed state with shorter Asp–Asp distance. This may explain the different outcome of Notch and C99 cleavage, as the latter is more imprecise with many products. Our identified conformation states may aid efforts to develop conformation-selective drugs that target C99 and Notch cleavage differently, e.g. Notch-sparing γ-secretase modulators.

Distinct membrane dynamics and conformations of C83- and Notch-bound γ-secretase may aid the development of Notch-sparing treatments of Alzheimer''s disease.     

Retracted: Evaluating the immunomodulatory responses of LdODC-derived MHC Class-II restricted peptides against VL

In a bid to develop a novel immunoprophylactic measure against visceral leishmaniasis (VL), MHC class-II–restricted epitopes of LdODC were identified by reverse vaccinology approach. Five consensus HLA-DRB1*0101-restricted epitopes were screened. The analysis revealed that the set of epitopes was presented by at least 54 diverse MHC class-II alleles. Based on in silico screening, followed by molecular dynamics simulation, population coverage analysis, and HLA cross-presentation ability, five best epitopes were evaluated. PBMCs isolated from treated VL subjects, when stimulated with synthetic peptide alone or as a cocktail of peptides, triggered a secretory IFN-γ, but not the IL-10 level. Support in this notion came from intracellular cytokine level with a considerable up-regulated IFN-γ produced by CD4⁺ T cells. Also, the enhanced IFN-γ seemed to be augmented with the activation of macrophages with prominent IL-12 production. Therefore, it can be concluded that the screened MHC class-II–restricted epitope hotspots derived from Leishmania ODC can trigger CD4⁺ T cells, which can skew macrophage functions towards protection. However, a detailed analysis can explore its potentiality as a vaccine candidate.     

First Report of Hb Limassol [β8(A5)Lys→Asn; HBB: c.27G>C] from Odisha,India

In this short report, we describe the clinical presentation of a rare hemoglobin (Hb) variant, Hb Limassol [β8(A5)Lys→Asn; HBB: c.27G>C] with a faster electrophoretic mobility than Hb A and that elutes in the P3 window on cation exchange high performance liquid chromatography (HPLC). This sequence variation at codon 8 (AAG>AAC) of the HBB gene was found in the four heterozygous cases, all of whom were clinically asymptomatic.