Early diagnosis of transthyretin amyloidosis by detection of monomers in plasma microsamples using a protein crystal-based assay

Abstract

Amyloid diseases are frequently associated with the appearance of an aberrant form of a protein, whose detection enables early diagnosis. In the case of transthyretin amyloidosis, the aberrant protein, the monomers, constitute the smallest species of the amyloid cascade, which creates engineering opportunities for sensing that remain virtually unexplored. Here, a two-step assay is devised, combining molecular sieving and immunodetection, for quantification of circulating monomeric transthyretin in the plasma. It is shown that mesoporous crystals built from biomolecules can selectively uptake transthyretin monomers up to measurable quantities. Furthermore, it was found that the use of endogenous molecules to produce the host framework drastically reduces unspecific adsorption of plasma proteins at the crystal surface, a feature that was observed with metal-organic frameworks. The assay was used to analyse plasma microsamples of patients and healthy controls. It shows a significant increase in the levels of monomeric transthyretin in the patients, proving its usefulness to establish the monomers as soluble and non-invasive marker of the disease. In addition, the assay can evaluate transthyretin stabilizers, an emergent strategy that broadened the treatment approach to the disease. Sensing the initial event of the transthyretin amyloid cascade with the proposed assay can make the difference for early diagnosis and eliminate the currently adopted invasive biopsies modalities for detection of the final products of the aggregation pathway.