Amyloid fibril formation in microwell plates for screening of inhibitors

Abstract

Fibril formation is the basis of amyloid production in a number of disease states, such as Alzheimer's disease, diabetes and immunocytic dyscrasias. Compounds that inhibit fibril formation could be directly relevant to the treatment of amyloid diseases, and may also provide a foundation for the development of interventions in other molecular condensation diseases ranging from sickle cell anemia to atherosclerosis.

We developed an economical and convenient high-throughput method for screening compounds against fibril formation in microwell plates. Chalcones, Jlavonoids and bijluvonoids were screened against fibril formation by a recombinant antibody variable domain (V_L). Chalcones 6 and 14 were found to demonstrate inhibition at 0.1 M in 79 M of protein solution in both test tube and microwell plate assays. The concentration of protein in the microwell plate assay could be as low as 5 M using ThTas a monitoring agent. Molecular modeling studies indicated that both compounds could be individually docked into a binding site at the monomer-monomer interface of the V_Lprotein dimer. These studies suggested that these compounds could potentialty stabilize the V_L dimer and therefore reduce its fendency to form fibrils. These findings may provide the basis for a new therapeutic approach to prevent or treat amyloid diseases.

Key Words:

• fibril formation
• microwell plate assay
• LEN V_l inhibitor