Pharmaceutical Sciences
Permanent URI for this collectionhttps://hdl.handle.net/20.500.12503/30821
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Browsing Pharmaceutical Sciences by Author "Qunies, Alshaima'a"
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Item Design and discovery of new tool compounds for studying the role of Slack potassium channels in malignant migrating partial seizure of infancy(2022) Qunies, Alshaima'a; Spitznagel, Brittany; Du, Yu; Weaver, C.; Emmitte, KylePurpose: Slack channels are sodium-activated potassium channels encoded by the KCNT1 gene, and are key regulators of electrical activity in the central nervous system. Slack channels belong to the Slo family of potassium channels (Slo2.2). The development of malignant migrating partial seizure of infancy (MMPSI), a type of severe infantile epilepsy, has been linked to KCNT1 gain of function mutations. The aim of this project is to design and synthesize Slack channel inhibitors for use as in vivo probes via an iterative hit optimization approach. Methods: 110K-member library was screened in a cell-based assay against wild-type Slack and three MMPSI-associated KCNT1 mutants. The hit compound VU0531245 (VU245) was selected for the development of structure-activity relationship (SAR) studies in order to optimize its potency and drug metabolism and pharmacokinetic (DMPK) properties. A thallium flux assay in HEK-293 cell lines stably expressing Slack channels was utilized to evaluate Slack inhibitory activity of the resulting compounds. Results: Compound libraries were designed around VU245 through the systematic scanning of the chemical space and incorporating various bioisosteric replacements. Our data suggest that modifications at the phenyl ring A lead to mode switching from inhibition to activation. However, fluorinated, alicyclic, and deuterated alkoxy groups at 2-position of the phenyl ring maintained Slack inhibitory activity; moreover, they improved some DMPK properties. We have demonstrated that piperidine replacement at ring B was tolerated. In addition, an ethylene linker in place of the 1,2,4-oxadiazole ring at position C maintained the Slack activity. A 4-fluorophenyl ring at position D was tolerated, improved the metabolic stability, and was used as the basis for further SAR exploration. Finally, a sulfonamide linker was optimal for Slack inhibitory activity. Collectively, the tested analogs within this series demonstrated good passive permeability with no evidence of P-gp-mediated efflux; however, high protein binding (fu ~ 0.01 - 0.04) was observed. Conclusions: SAR for Slack activity and DMPK properties was identified around VU245. Further optimization is required to develop suitable Slack in vivo probes with improved potency and DMPK properties.Item Library synthesis of Slack potassium channel activators based on a high-throughput screening hit(2022) Nguyen, Dalena; Qunies, Alshaima'a; Du, Yu; Weaver, C.; Emmitte, KyleIntroduction: Fragile X syndrome (FXS) is an X-linked disorder that is associated with cognitive disabilities. Previous studies have shown an association between a mutation in the FMR1 gene and FXS. The mutation is an overexpansion of the promoter region, resulting in hypermethylation and silencing of fragile X mental retardation protein 1 (FMRP).1 FMRRP is necessary for activating Slack proteins, which are important for normal neuronal activity.2, 3 Objective: To synthesize a library of small molecules in two distinct regions of an HTS hit chemotype made of sulfonamides and heteroaryl amines for functional testing versus Slack channels. Methods: Solution-phase and microwave-assisted organic chemistry were utilized to synthesis small molecules. Purification of compounds involved using automated liquid chromatography. Final compounds were characterized through NMR and HRMS data obtained from a Bruker Fourier 300HD and Agilent 6230 time-of-flight LC/MS, respectively. Activity of new compounds versus Slack was measured utilizing a Thallium flux assay in HEK293 cells stably expressing WT Slack channels. Results: SAR studies conducted around hit compound VU0521448 have thus far discovered analogs showing only weak activity against Slack proteins. Data for SAR studies developed around hit compound VU0521398 are in the process of being collected. Conclusion: The sulfonamide library prepared using different sulfonyl chloride substituents gave some analogs with weak activity in activating Slack proteins. Additional SAR studies will be carried out to examine other aspects of the chemotype, such as modifying the piperidine ring to pyrroline or pyrrolidine. FUNDING STATEMENT Research reported in this publication was supported by the National Heart, Lung, and Blood Institute (R25HL125447) and the National Institute of Mental Health (R21MH125257), both of the National Institutes of Health. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institute of Health.