The formation of amyloid fibrils due to protein aggregation represents a central step in the pathogenesis of many age-related diseases for which there are no current cures. At the core of this step is the misfolding of one or more key proteins, which is converted from a natively folded or unfolded state to an aggregation-prone state that is rich in b-sheet structure. While there have been intensive efforts to develop therapeutic approaches to inhibit amyloid fiber formation during its initial stages, such strategies have met with only limited success. As such, there remains a need for alternative strategies to mediate even earlier states of amyloid oligomerization by re- folding and thus rescuing misfolded proteins prior to aggregating. The overall aim of this proposal is to develop nanomaterials with chaperone-like activity to promote re-folding of misfolded proteins and thus have therapeutic potential to inhibit amyloid formation. Specifically, in this approach, we propose to rationally modify nanoparticles with novel coatings that stabilize the native structure of amyloid-associated proteins and can be delivered in vivo. Of particular interest will be investigating the extent to which chemically heterogeneous polymer brushes and mixed lipid bilayers promote re-folding of the protein amyloid-b (Ab42), which is a precursor to amyloid fibers that are formed in Alzheimer?s disease. In support of this approach, we have previously shown that random co- polymer brushes composed of poly(ethylene glycol) and poly(sulfobetaine) as well as mixed supported lipid bilayers compared of varying ratios of 1,2-dioleoyl-sn-glycero-3-phosphocholine and 1,2-dioleoyl-sn-glycero-3- phospho-(1'-rac-glycerol) both stabilized and promoted the re-folding of model proteins, including fibronectin and nitroreductase. Using coatings made of such materials, we will test the central hypothesis that nanoparticles coated with dynamic and heterogeneous layers can reduce the conversion of Ab42 to its aggregation-prone state and prevent amyloid formation via a chaperone-like mechanism. In line with this hypothesis, the specific aims of this proposal are to: (1) elucidate the mechanism and optimize the composition of heterogeneous coatings for Ab42 stabilization by nanoparticles via a chaperone-like mechanism (Aim 1) and (2) correlate Ab42 stabilization by chemically heterogeneous nanoparticles with the inhibition amyloid formation (Aim 2). The chaperone-like activity of the nanoparticle coatings will be characterized quantitatively using single-molecule biophysical methods that are uniquely sensitive to monitoring protein structure and dynamics (e.g., re-folding) on surfaces. To correlate the stabilization of Ab42 with the inhibition of amyloid formation, the impact of the coatings on the formation of amyloid fibrils will be measured in the presence of coated and uncoated nanoparticles while varying coating composition. Additionally, the utility of the coated nanoparticles in inhibiting the aggregation of Ab42 and Ab40 as well as Ab42 with mutations associated with Alzheimer?s disease (i.e., E22G) will also be measured. These studies will provide a complete molecular picture of this approach, as well as a general understanding that may be applied to the rescue of proteins implicated in other protein misfolding diseases, including cancer.
The overarching aim of this proposal is to develop a novel therapeutic approach to treat protein misfolding diseases using chemically-modified nanoparticles with chaperone-like activity. This approach involves understanding how nanoparticles with dynamic and heterogeneous layers stabilize and promote re-folding of Ab42 and correlating the stabilization of Ab42 with the anti-amyloid activity of the nanoparticles. Ultimately, this work will provide a complete molecular picture of the proposed approach, as well as a general understanding that may be applied to the rescue of proteins implicated in other protein misfolding diseases linked to aging, including cancer.
