Identification of a Small Molecule that Increases Hemoglobin Oxygen Affinity and Reduces SS Erythrocyte Sickling

Nakagawa, Akito; Lui, Francine E.; Wassaf, Dina; Yefidoff-Freedman, Revital; Casalena, Dominick; Palmer, Michelle; Meadows, Jacqueline; Mozzarelli, Andrea; Ronda, Luca; Abdulmalik, Osheiza; Bloch, Kenneth D.; Safo, Martin K.; Zapol, Warren M.

doi:10.1021/cb500230b

Cited by 48 publications

(82 citation statements)

References 46 publications

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“…The β chains of Hb from four crystal structures: 2DN1, 68 4NI0, 69 1YZI, 70 and 1GZX 71 were examined to identify a suitable overall hemoglobin structure into which to dock HONH-ANL, HONH-ABP, HONH-AαC, HONH-PhIP, and HONH-MeIQx. All four β chains were close to identical in configuration, including the proximity of their heme pockets; RMSD values between the four B chains were 0.77–0.79Å.…”

Section: Resultsmentioning

confidence: 99%

Methemoglobin Formation and Characterization of Hemoglobin Adducts of Carcinogenic Aromatic Amines and Heterocyclic Aromatic Amines

Pathak

Chiu

Amin

et al. 2016

Chem. Res. Toxicol.

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Arylamines (AA) and heterocyclic aromatic amines (HAA) are structurally related carcinogens formed during combustion of tobacco or cooking of meat. They undergo cytochrome P450 mediated N-hydroxylation to form metabolites which bind to DNA and lead to mutations. The N-hydroxylated metabolites of many AA also can undergo a co-oxidation reaction with oxy-hemolgobin (HbO2) to form methemoglobin (met-Hb) and the arylnitroso intermediates, which react with the β-Cys93 chain of Hb to form Hb-arylsulfinamide adducts. The biochemistry of arylamine metabolism has been exploited to biomonitor certain AAs through their Hb arylsulfinamide adducts in humans. We examined the reactivity of HbO2 with the N-hydroxylated metabolites of 4-aminobiphenyl (ABP, HONH-ABP), aniline (ANL, HONH-ANL), and the HAA 2-amino-9H-pyrido[2,3-b]indole (AαC, HONH-AαC), 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP, HONH-PhIP) and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx, HONH-MeIQx). HONH-ABP, HO-ANL, and HONH-AαC induced methemoglobinemia and formed Hb sulfinamide adducts. However, HONH-MeIQx and HONH-PhIP did not react with the oxy-heme complex, and met-Hb formation and chemical modification of β-Cys93 residue was negligible. Molecular modeling studies showed that the distances between the H-ON-AA or H-ON-HAA substrates and the oxy-heme complex of HbO2 were too far away to induce methemoglobinemia. Different conformational changes in flexible helical and loop regions around the heme pocket induced by the H-ON-AA or H-ON-HAAs may explain the different proclivities of these chemicals to induce methemoglobinemia. Hb-Cys93β sulfinamide and sulfonamide adducts of ABP, ANL, and AαC were identified, by Orbitrap MS, following proteolysis of Hb with trypsin, Glu-C, or Lys-C. Hb sulfinamide and sulfonamide adducts of ABP were identified in blood of mice exposed to ABP, by Orbitrap MS. This is the first report of the identification of intact Hb sulfinamide adducts of carcinogenic AAs in vivo. The high reactivity of HONH-AαC with HbO2 suggests that the Hb sulfinamide adduct of AαC may be a promising biomarker of exposure to this HAA in humans.

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Section: Resultsmentioning

confidence: 99%

Methemoglobin Formation and Characterization of Hemoglobin Adducts of Carcinogenic Aromatic Amines and Heterocyclic Aromatic Amines

Pathak

Chiu

Amin

et al. 2016

Chem. Res. Toxicol.

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“…Highthroughput screens have been developed for the induction of HbF (42)(43)(44)(45)(46). The only high-throughput screen for antipolymerization compounds up until now tests for increasing oxygen affinity on isolated hemoglobin solutions, but it does not address the question of cell permeability (47).…”

Section: Discussionmentioning

confidence: 99%

Kinetic assay shows that increasing red cell volume could be a treatment for sickle cell disease

Henry

Hofrichter

et al. 2017

Proc. Natl. Acad. Sci. U.S.A.

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Although it has been known for more than 60 years that the cause of sickle cell disease is polymerization of a hemoglobin mutant, hydroxyurea is the only drug approved for treatment by the US Food and Drug Administration. This drug, however, is only partially successful, and the discovery of additional drugs that inhibit fiber formation has been hampered by the lack of a sensitive and quantitative cellular assay. Here, we describe such a method in a 96-well plate format that is based on laser-induced polymerization in sickle trait cells and robust, automated image analysis to detect the precise time at which fibers distort ("sickle") the cells. With this kinetic method, we show that small increases in cell volume to reduce the hemoglobin concentration can result in therapeutic increases in the delay time prior to fiber formation. We also show that, of the two drugs (AES103 and GBT440) in clinical trials that inhibit polymerization by increasing oxygen affinity, one of them (GBT440) also inhibits sickling in the absence of oxygen by two additional mechanisms.sickle cell | drugs | hemoglobin S | treatment | screening assay

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“…10 This has prompted several studies to find alternate covalent modifiers of Hb, such as isothiocynates and thiols to treat the disease. 23,24 …”

Section: Introductionmentioning

confidence: 99%

Identification of a novel class of covalent modifiers of hemoglobin as potential antisickling agents

et al. 2015

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Aromatic aldehydes and ethacrynic acid (ECA) exhibit antipolymerization properties that are beneficial for sickle cell disease therapy. Based on ECA pharmacophore and its atomic interaction with hemoglobin, we designed and synthesized several compounds--designated as KAUS (imidazolylacryloyl derivatives)--that we hypothesized would bind covalently to βCys93 of hemoglobin and inhibit sickling. The compounds surprisingly showed weak allosteric and antisickling properties. X-ray studies of hemoglobin in complex with representative KAUS compounds revealed an unanticipated mode of Michael addition reaction between the β-unsaturated carbon and the N-terminal αVal1 nitrogen at the α-cleft of hemoglobin, with no observable interaction with βCys93. Interestingly, the compounds exhibited almost no reactivity with the free amino acids, L-Val, L-His and L-Lys, however showed some reactivity with both glutathione and L-Cys. Our findings provide a molecular level explanation to the compounds biological activities and an important framework for targeted modifications that would yield novel potent antisickling agents.

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Identification of a Small Molecule that Increases Hemoglobin Oxygen Affinity and Reduces SS Erythrocyte Sickling

Cited by 48 publications

References 46 publications

Methemoglobin Formation and Characterization of Hemoglobin Adducts of Carcinogenic Aromatic Amines and Heterocyclic Aromatic Amines

Methemoglobin Formation and Characterization of Hemoglobin Adducts of Carcinogenic Aromatic Amines and Heterocyclic Aromatic Amines

Kinetic assay shows that increasing red cell volume could be a treatment for sickle cell disease

Identification of a novel class of covalent modifiers of hemoglobin as potential antisickling agents

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