Parkinson’s Disease (PD) is a debilitating neurodegenerative disorder. It is characterized by a substantial and irreversible loss of neurons in the substantia nigra in the brain, which leads to the loss of motor function, and eventually, death. PD is the most common movement-based neurodegenerative disorder and the second-most common cause of dementia in individuals over the age of 65. The diagnosis of PD relies entirely on the observation of the associated motor symptoms, and as such makes a timely diagnosis exceptionally challenging. This leads to a lengthy diagnosis process with the average diagnosis occurring 10 years after the onset of symptoms. By this time, the patient’s quality of life is drastically reduced. There is a need to develop early-stage diagnostics to enable more rapid treatment to improve this quality of life, while also prolonging life expectancy after diagnosis.
However, currently approved therapeutics only transiently alleviate motor symptoms. There is a need for developing therapeutic treatment modalities that effectively slow and/or halt disease progression. Among the underlying mechanisms that contribute to disease progression, the buildup of misfolded and aggregated protein in the form of Lewy Bodies (LB) is the leading contributor. As the largest component of LB, aggregated α-synuclein (αSynagg) represents a promising biomarker for both diagnostic and therapeutic thrusts to PD. Accordingly, a large sector of PD research is dedicated towards finding ways to detect αSynagg, and thereby breakdown the aggregates and halt disease progression. Most of these efforts focus on immunological approaches, utilizing antibodies that are specific to only αSynagg, and not the physiological form which is important for neuronal homeostasis. While great strides have been made with these approaches, there is yet to be a fully reliable diagnostic assay, or treatment for that matter, due to the complex nature of aggregates in vivo and the inability to achieve high levels of specificity for the pathogenic form of the protein.
In addition to the need for better therapeutics, there is a need to minimize systemic toxicity associated with delivering such disease-slowing therapeutics, while also optimizing brain bioavailability. This can be accomplished by using targeted nano-carriers which encapsulate the therapeutic. Polyanhydride nanoparticles are an attractive option in this field, due to their biocompatibility, sustained release properties, high levels of cellular internalization and ability to easily conjugate targeting ligands in multiple different ways.
In this work, a set of early-stage diagnostics is presented. The high levels of specificity for αSynagg using the singularly specific monoclonal antibodies (ssMAbs) in an enzyme-linked immunosorbent assay (ELISA) has led to high levels of discrimination between PD and age-matched control samples, including in an early-age group. Further ELISA optimization is underway to progress to clinical accuracy. Additionally, the ability of these ssMAbs to break down αSynagg in vitro suggests their use as a disease-slowing therapeutic, however there are several shortcomings to the use of full antibody therapeutics, including unfavorable immunogenicity and poor absorption properties. To overcome these issues, recombinant single chain fragment variable antibodies (scFv) based on the chemistry of ssMAb 3A8 (i.e. scFv 3A8) was proposed for therapeutic use to treat PD. The therapeutic potential for this scFv to breakdown αSynagg was observed, and to a greater degree than the ssMAbs themselves.
To enhance brain delivery of therapeutics like this scFv, we proposed the use of a polyanhydride nanoparticle delivery platform. After testing select chemistries encapsulating a fluorescent dye and monitoring fluorescence over the course of seven days in biodistribution studies, it was found that the fluorescent dye encapsulated by these nanoparticles was detected in the brain after intravenous delivery. Notably, non-functionalized nanoparticles provided the most sustained release of dye over the course of the study, compared to both a soluble dose and a functionalized polyanhydride chemistry incorporating a peptide of the BBB-targeting transferrin protein. The enhanced brain bioavailability provided by the nanoparticles was also evidenced by more favorable pharmacokinetic parameters.
Lastly, how brain bioavailability of this polyanhydride nanoparticle platform is correlated with the ability to enhance therapeutic efficacy was tested in an αSyn overexpression model. Soluble scFv and scFv-encapsulated nanoparticles were administered as treatments and it was found that this scFV-based treatment regimen recovered motor function, which was correlated with higher levels of dopaminergic activity by immunochemistry analysis. Collectively, these data suggest further optimization of this therapeutic platform so that it can be ultimately used as a disease-slowing therapeutic for PD patients
