Journal Article Ppublish 1 citation
PubMed: 36399908 DOI: 10.1016/j.jpba.2022.115152
Quantitative assessment of the in-vitro binding kinetics of antisickling aromatic aldehydes with hemoglobin A: A universal HPLC-UV/Vis method to quantitate Schiff-base adduct formation.
J Pharm Biomed Anal 223:115152
Authors: Xiaomeng Xu, Mohini S Ghatge, Boshi Huang, Ahmed Alghamdi, Huiqun Wang, B Daniel Pierce, Osheiza Abdulmalik, Yan Zhang, Martin K Safo, Jürgen Venitz
Affiliations
- Division of Cardiometabolic and Endocrine Pharmacology, Office of Clinical Pharmacology, Center of Drug Evaluation and Research, US FDA, Silver spring, MD 20993, USA; Department of Pharmaceutics, School of Pharmacy, Virginia Commonwealth University, Richmond, VA 23298, USA.
- Department of Medicinal Chemistry and The Institute for Structural Biology, Drug Discovery and Development, School of Pharmacy, Virginia Commonwealth University, Richmond, VA 23298, USA.
- Department of Biology, University of Richmond, Richmond, VA 23173, USA.
- Division of Hematology, The Children's Hospital of Philadelphia, PA 19104, USA.
- Department of Medicinal Chemistry and The Institute for Structural Biology, Drug Discovery and Development, School of Pharmacy, Virginia Commonwealth University, Richmond, VA 23298, USA.
- Department of Pharmaceutics, School of Pharmacy, Virginia Commonwealth University, Richmond, VA 23298, USA.
Abstract
Aromatic aldehydes act as allosteric effectors of hemoglobin (AEH), forming Schiff-base adducts with the protein to increase its oxygen (O 2) affinity; a desirable property in sickle cell disease (SCD) treatment, as the high-O 2 affinity hemoglobin (Hb) does not polymerize and subsequently prevents erythrocytes sickling. This study reports the development, validation, and application of a weak cation-exchange HPLC assay - quantifying the appearance of Hb-AEH adduct - as a "universal" method, allowing for the prioritization of AEH candidates through an understanding of their Hb binding affinity and kinetics. Concentration- and time-dependent Hb binding profiles of ten AEHs were determined with HPLC, followed by the appropriate non-linear modeling to characterize their steady-state binding affinity (K D ss), and binding kinetics second-order association (k on) and first-order dissociation (k off) rate constants. Vanillin-derived AEHs exhibited enhanced binding affinity to Hb, primarily due to their faster k on. Across AEH, k on and k off values are strongly correlated (r = 0.993, n = 7), suggesting that modifications of the AEH scaffold enhanced their interactions with Hb as intended, but inadvertently increased their Hb-AEH adduct dissociation. To our knowledge, the present study is the first to provide valuable insight into Hb binding kinetics of antisickling aromatic aldehydes, and the assay will be a useful platform in screening/prioritizing drug candidates for SCD treatment.
MeSH
Hemoglobins, Hemoglobin A, Chemistry, Pharmaceutical, Aldehydes, Schiff Bases, Therapeutics