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Abstract: TH-PO206

Small Molecule Screening Identifies TW-37 as a KIM-1 Inhibitor and Potential Anti-Fibrotic Molecule

Session Information

Category: Diabetic Kidney Disease

  • 601 Diabetic Kidney Disease: Basic

Authors

  • Ajay, Amrendra Kumar, Brigham and Women's Hospital Department of Medicine, Boston, Massachusetts, United States
  • Mori, Yutaro, Brigham and Women's Hospital Department of Medicine, Boston, Massachusetts, United States
  • Sabbisetti, Venkata, Brigham and Women's Hospital Department of Medicine, Boston, Massachusetts, United States
  • Bonventre, Joseph V., Brigham and Women's Hospital Department of Medicine, Boston, Massachusetts, United States
Background

Kidney Injury Molecule-1 (KIM-1) is a glycosylated protein upregulated following proximal tubular injury. With acute and chronic injury, KIM-1 mediates the uptake of apoptotic cells, oxidized lipids, advanced glycation end products (AGEs), and albumin bound to long chain fatty acids. Overexpression of KIM-1 causes chronic kidney disease (CKD) in mice.

Methods

We developed a high throughput cell-based functional assay for KIM-1 mediated uptake of ox-LDL, and screened 14,414 unique small molecules. After setting up a histological score based on the potential to inhibit cellular ox-LDL uptake for each compound, 240 potential hits were cherry-picked from the primary screening. Raman spectroscopy was employed to investigate the binding of KIM-1 to TW-37. The effectiveness of the selected compound was evaluated in a mouse model of kidney fibrosis.

Results

TW-37, a second-generation benzenesulfonyl derivative of gossypol, had the greatest inhibitory effect on ox-LDL uptake. TW-37 is known to have Bcl2 inhibitory activity; however, Bcl-2 blockade with another specific Bcl-2 inhibitor, ABT-263, did not inhibit KIM-1 dependent ox-LDL uptake. TW-37 is not toxic to cells at concentrations up to 11µM. TW-37 neither cleaves KIM-1 nor quenches the fluorophores in our assay systems. Our in silico docking revealed a putative TW-37 binding pocket which spans from residues 37 to 52 of KIM-1. TW-37 specifically binds to recombinant KIM-1 and not to the BSA as determined by Raman spectroscopy. We observed a drop in the loops and turn content (44 to 34.5 percent) and an enhanced beta sheet content (42 to 53.5 percent) in the KIM-1+TW-37 sample compared to native KIM-1 spectrum. TW-37 significantly inhibits the uptake of BODIPY-labeled palmitic acid bound to albumin. TW-37 reduces kidney fibrosis induced by palmitic acid-albumin (0.5 mmol/kg PA and 2.5 g/kg BSA) administration daily from days 7 to 14 after aristolochic acid (5 mg/kg) administration.

Conclusion

We have identified and characterized TW-37 as an inhibitor of KIM-1 binding. TW-37 protects mice from kidney fibrosis. Thus, TW-37 has potential use as a therapeutic for the treatment of kidney disease where chronic KIM-1 mediated uptake of luminal contents into the proximal tubule contributes to chronic injury and maladaptive repair.

Funding

  • NIDDK Support