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A Semi-High-Throughput Adaptation of the NADH-Coupled ATPase Assay for Screening Small Molecule Inhibitors
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Functional analysis of a species-specific inhibitor selective for human Na+-coupled citrate transporter

Kei Higuchi1, Jonathan J Kopel1, Sathish Sivaprakasam1

  • 1Department of Cell Biology and Biochemistry, Texas Tech University Health Sciences Center, Lubbock 79430, Texas, U.S.A.

The Biochemical Journal
|October 20, 2020
PubMed

Insights

BI01383298 is the first irreversible inhibitor of the human Na+-coupled citrate transporter (NaCT), showing high potency and selectivity. This discovery offers potential therapeutic strategies for metabolic diseases and cancer by blocking citrate uptake.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Pharmacology

Background:

  • The Na+-coupled citrate transporter (NaCT/SLC13A5/mINDY) facilitates citrate uptake into hepatocytes, impacting key metabolic pathways.
  • NaCT dysfunction is implicated in obesity, diabetes, metabolic syndrome, and hepatocellular carcinoma, highlighting its therapeutic relevance.
  • Selective NaCT inhibitors are sought for treating these conditions.

Purpose of the Study:

  • To characterize the novel inhibitor BI01383298 targeting the human Na+-coupled citrate transporter (NaCT).
  • To elucidate the mechanism of action and species-specific effects of BI01383298.
  • To provide structural insights into NaCT inhibition for drug development.

Main Methods:

  • Inhibition assays using HepG2 and HEK293 cells expressing human NaCT.
  • Determination of IC50 values and inhibitor kinetics (competitive/non-competitive).
  • Molecular modeling and docking studies of human and mouse NaCT structures.

Main Results:

  • BI01383298 is a potent (IC50 ~100 nM), irreversible, and non-competitive inhibitor of human NaCT.
  • The compound exhibits high selectivity for human NaCT, with no observed effect on mouse NaCT.
  • Molecular modeling identified key amino acid residues responsible for differential species affinity and inhibitor binding.

Conclusions:

  • BI01383298 represents the first irreversible inhibitor of human NaCT, offering a novel therapeutic agent.
  • Its species-specific action and high potency provide a valuable tool for studying NaCT function and developing treatments for metabolic disorders.
  • Structural insights guide the design of future NaCT inhibitors with improved efficacy and selectivity.
Keywords:
BI01383298NACT/SLC13a5/mINDYcell growth arrestirreversible inhibitionmolecular modelingspecies-specificity

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