Amyloid fibril formation in microwell plates for screening of inhibitors

Fibril formation is the basis of amyloid production in a number of disease states, such as Alzheimer's disease, diabetes and immunocytic dyscrasias. C ompounds that inhibit fibril formation could be directly relevant to the tr eatment of amyloid diseases, and may also provide a foundation for the deve lopment of interventions in other molecular condensation diseases ranging f rom sickle cell anemia to atherosclerosis. We developed an economical and convenient high-throughput method for screen ing compounds against fibril formation in microwell plates. Chalcones, flav onoids and biflavonoids were screened against fibril formation by a recombi nant antibody variable domain (V-L). Chalcones 6 and 14 were found to demon strate inhibition at 0.1 PM in 79 muM 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 muM using ThT as a monitoring agent. Molec ular modeling studies indicated that both compounds could be individually d ocked into a binding site at the monomer-monomer interface of the V-L prote in dimer. These studies suggested that these compounds could potentially st abilize the VL dimer and therefore reduce its tendency to form fibrils. The se findings may provide the basis for a new therapeutic approach to prevent or treat amyloid diseases.