Controlled release of neurosin enzyme for alternative treatment of Parkinson's Disease

Parkinson's Disease (PD) is a common neurodegenerative disorder that shows the motor and non-motor symptoms. Misfolded alpha-synuclein aggregates (Lewy bodies) are found in the brain pathology of Parkinson's patients. As a result of studies, it has been determined that these alpha-synuclein aggregates were directly related to the degrees of neurodegeneration. Although these pathological aggregates are found inside the cell, they are able to pass to healthy neurons by a prion-like mechanism. In previous studies, neurosin (Kallikrein 6, KLK6) has been shown to destroy the alpha-synuclein by cleavage at the NAC (non Amyloid-β component) region of the enzyme. In controlled drug release systems, nanoparticles are preferred because of their nano size, allowing ease of uptake into cells. Human serum albumin (HSA), the most abundant protein in the body, is an important candidate for nanoparticle synthesis due to its easy purification, biocompatibility, biodegradability, and having many functional groups. The most common method in the synthesis of nanoparticles from HSA is the desolvation technique. In this method, generally glutaraldehyde, which has a toxic effect on cells, is used as a cross-linker. This study, it is aimed to encapsulate neurosin, which is a therapeutic agent candidate, into HSA nanoparticles by desolvation technique. Additionally, it is aimed to reduce the toxic effect by using natural crosslinker genipin instead of glutaraldehyde in the production of HSA nanoparticles. First of all, the optimum synthesis condition for genipin-crosslinked HSA nanoparticles was determined. Zetasizer, FTIR, SEM analyzes were performed for the characterization of the synthesized nanoparticles. Variables affecting nanoparticles production were estimated as HSA concentration, crosslinker concentration, crosslinking temperature, crosslinking time and speed of magnetic stirrer. According to the parameters examined, the optimum condition of nanoparticle synthesis was determined as 20 mg/ml HSA concentration and 0.2% w/w genipin per mg of HSA concentration. The condition of crosslinking was determined as 37°C for 2 hours at 600 rpm. Then, neurosin enzyme was loaded into HSA nanoparticles which were produced under optimum condition. Encapsulation efficiency was calculated using the ELISA kit. In vitro release study of neurosin loaded HSA nanoparticles was performed in PBS buffer using the dialysis membrane. As a result of the studies, genipin cross-linked HSA nanoparticles with desired properties were produced. In the release studies of neurosin loaded nanoparticles, it was observed that neurosin was released with burst effect in the first 30 minutes. Afterward, it was observed that the drug release continues slowly according to the samples taken for 7 days. It is predicted that neurosin loaded genipin-crosslinked HSA nanoparticles will contribute to the drug delivery systems and literature as an alternative drug system in Parkinson's disease.