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Abstract: FR-PO0775

Conformation-Dependent Antibody Variants Targeting Integrin α3β1 Restore Podocyte Function and Reduce Injury in FSGS Mouse Models

Session Information

Category: Glomerular Diseases

  • 1401 Glomerular Diseases: Mechanisms, including Podocyte Biology

Authors

  • Youssef, Mohamed A., The University of Texas Medical Branch at Galveston Department of Internal Medicine, Galveston, Texas, United States
  • Balza Pineda, Santiago, Allosite Therapeutics, Miami, Florida, United States
  • Machineni, Prathyushasai, The University of Texas Medical Branch at Galveston Department of Internal Medicine, Galveston, Texas, United States
  • Lopez-Rodriguez, Darlah M., Allosite Therapeutics, Miami, Florida, United States
  • Gonzalez-Lerma, Paola, Allosite Therapeutics, Miami, Florida, United States
  • Helmuth, Richard, Rush University Medical Center, Chicago, Illinois, United States
  • Gurusamy Kamaraj, Sowmya, The University of Texas Medical Branch at Galveston Department of Internal Medicine, Galveston, Texas, United States
  • Brackman, Sheri, The University of Texas Medical Branch at Galveston Department of Internal Medicine, Galveston, Texas, United States
  • Barbosa, Antonio, Allosite Therapeutics, Miami, Florida, United States
  • Gupta, Vineet, The University of Texas Medical Branch at Galveston Department of Internal Medicine, Galveston, Texas, United States
Background

Healthy podocytes preserve glomerular integrity by adhering to the basement membrane via integrin α3β1. Disruption of this adhesion, due to inflammation, mutations in α3β1 or CD151, or cytoskeletal defects, leads to podocyte detachment and proteinuria. Loss of adhesion undermines the filtration barrier and drives progressive diseases such as focal segmental glomerulosclerosis (FSGS). Strategies that reinforce integrin-mediated adhesion, such as a conformational-dependent antibody, may prevent podocyte loss and slow FSGS progression.

Methods

Cell damage and agonist-mediated protection were quantified in PAN-treated podocytes by high-content screening. α3β1-activating antibodies were isolated from a naïve human phage display library and screened on recombinant integrin, K562 cells expressing α3β1, differentiated podocytes, and SK-OV3 cells. Antibody effects on adhesion and morphology were quantified using image analysis. Candidate antibodies were characterized via ELISA on recombinant α3β1, flow cytometry with wild-type or chimeric α3β1-expressing K562 cells, bio-layer interferometry kinetic assays, and functional cell adhesion and scratch wound healing assays. Albuminuria was measured in an LPS-induced and anti-GBM nephritis FSGS mouse models.

Results

PAN injury reduced F-actin fibers, focal adhesions and active integrin in podocytes. Known β1 agonists (Ab9EG7, Pyrintegrin) and novel α3 antibodies reversed these changes, enhanced adhesion and inhibited migration. Epitope mapping revealed α3 head-domain binding. In FSGS mice models, treatment with an α3 antibody significantly reduced albuminuria, attenuated glomerular injury, decreased podocyte foot process effacement and reduced glomerular deposition of IgG and complement component C3.

Conclusion

Integrin α3β1 activation strengthens podocyte adhesion and reduces injury. Ongoing in vitro studies include epitope mapping, functional assays, and signaling pathway evaluation via Western blotting and in vivo studies will assess optimal dosing and mechanism of protection.

Funding

  • Other U.S. Government Support

Digital Object Identifier (DOI)