Publication Details
Abstract
Background: Protein misfolding and aggregation is one of the fundamental disease-inducing processes in neurodegenerative diseases and systemic amyloidoses. Cellular protein quality control mechanisms maintain the protein folds in homeostasis and are unable to do so when disease develops. Objective: This study aimed to examine the processes of protein misfolding, the constraints of cellular quality control systems, and the efficacy of therapeutic interventions that target pathways involved in aggregation, with a focus on combination methods. Methodology: To determine the aggregation kinetics of disease related proteins (alpha-synuclein, tau, A beta 40/42) we employed a combination of biochemical assay, cell culture models comprised of several types of brain cellular models such as neurons, and molecular biology to examine the aggregating kinetics of disease related protein (using thioflavin T fluorescence, dynamic light scattering, and electron microscopy). The reactions to quality control are in the form of expression of heat shock proteins, proteasome activity and autophagy flux. Dose-response and combination therapy optimization were systematically determined to identify systematically the therapeutic agents. Results: All three proteins aggregated by a nucleation-dependent process, and could be described by various profiles: 6.2 +/- 0.8 hours lag phase of 4-synuclein, critical cofactor concentrations less than 10 muM tau, and fast kinetics (0.8 +/- 0.2 hours) with A beta 4 2. Quality control systems had very little adaptive capacity, with a 4.2-fold increase in Hsp70 in the first instance of time, but then reduced after 48 h, which is a manifestation of exhaustion. Ironically, stress inhibited proteasome activity (75 percent, 48 hours) and autophagy was curtailed by lysosomal capacity. These findings are discussed in Table 2, where not only are the benefits of chaperone activators in therapeutic efficacy, but also the benefits of autophagy regulators in therapeutic efficacy and narrow windows of treatment, and chaperone activators + autophagy modulators synergistic effects (combination indices 0.58-0.72), with sequential and simultaneous protocols displaying highly contrasting therapeutic efficacy. Conclusions: Achieving effective therapeutic intervention implies that a large number of quality control processes must be managed concurrently. Other major system constraints that have unique intervention targets include proteasome vulnerabilities and lysosomal bottlenecks. The combination therapy of chaperone activators and autophagy enhancers, as synergistic combinations, forms quantitative therapeutic paradigms to the clinical translation of chaperonal therapies to treat the protein conformational diseases.