(M0930-11-74) Mannose Functionalized Chitosan Coated PLGA Nanoparticles for Cannabidiol Delivery to the Brain for the Treatment of Alzheimer’s disease

Purpose: Alzheimer's disease (AD) is a debilitating neurodegenerative disease accounting for 60% of cases of dementia. Accumulation of amyloid-b, tau hyperphosphorylation, neuroinflammation, and oxidative stress play an important role in AD progression. Nowadays, cannabidiol (CBD) is drawing great attention because of its antioxidant, anti-inflammatory, and neuroprotective properties. Moreover, CBD has been found to reduce the hyperphosphorylation of tau protein, inhibit acetylcholinesterase activity, and reduced deposition as well as expression of beta-amyloid. These properties suggest that CBD can be used to treat many pathologies typically found in AD. The major limitation of drug delivery to the brain is the selective permeability of the blood-brain barrier (BBB) which restricts the entry of drugs into the brain. In recent years, nanoparticles are being used to solve the unmet problem of enhancing drug transport across the BBB. The purpose of this study is to develop and evaluate a brain-targeted PLGA nanoparticles-based drug delivery system to deliver CBD across the BBB to the brain in a controlled way.

Methods: Chitosan (CHT) was functionalized with mannose (MAN) using a reductive amination reaction. Briefly, the aldehyde (-CHO) functionality of D-mannose reacts with the amine (-NH₂) functionality of chitosan in the presence of sodium triacetoxyborohydride [NaBH(OAc)₃], a reducing agent. Further, Mannose-functionalized chitosan (CHT-MAN) was coated on PLGA nanoparticles followed by loading with CBD. The developed CHT-MAN-PLGA-CBD nanoparticles were subjected to physical characterization including size, Zeta potential, and polydispersity index. The CBD released pattern from CHT-MAN-PLGA-CBD was established in PBS (pH~7.4) containing 0.5% Tween-80 at 37 ^o C. Cytotoxicity and hemotoxicity of the developed CHT-MAN-PLGA-CBD nanoparticles was investigated in bEND.3 cells and RBCs, respectively. The in-vitro cellular uptake potential of the developed nanoparticles was evaluated in bEND.3 cells on different time points. Chitosan-coated PLGA nanoparticles loaded with CBD were noted as CHT-PLGA-CBD nanoparticles whereas mannose-functionalized chitosan-coated PLGA nanoparticles loaded with CBD were noted as CHT-MAN-PLGA-CBD nanoparticles.

Results: The size of the developed CHT-PLGA-CBD and CHT-MAN-PLGA-CBD nanoparticles were 294.2 ± 7.52 nm and 306 ± 8.12 nm, respectively. The Zeta potential of CHT-PLGA-CBD was 33.4 ± 1. 32 mV, whereas CHT-MAN-PLGA-CBD nanoparticles has a zeta potential of 31.7 ± 1.53 mV. The encapsulation efficiency of CHT-PLGA-CBD and CHT-MAN-PLGA-CBD was found to be 71 ± 5.34 % and 65 ± 6.54%, respectively. In vitro, drug release studies unveiled that the developed CHT-PLGA-CBD and CHT-MAN-PLGA-CBD nanoparticles were able to release the drug for a longer time in a sustained way. It has been observed from the release profile that CHT-PLGA-CBD and CHT-MAN-PLGA-CBD can release CBD up to 90 % and 67 % at 22 days (Figure 1). The developed nanoparticles were nontoxic even at the higher concentration of 10 mg/ml (Figure 2(A)) and did not exhibit any potential hemotoxicity (Figure 2(B)).

Conclusion: CBD-loaded mannose functionalized chitosan-coated PLGA nanoparticles were successfully developed with higher drug encapsulation efficiency. The developed nanoparticles release the drug in a controlled manner for a longer time and have the potential to be used as a drug delivery vehicle. Further, the in vivo therapeutic efficacy of drug-loaded nanoparticles will be evaluated in the AD mouse model.

Acknowledgements: This research was supported by National Institutes of Health Grants R01 AG051574, RF1 AG068034 and 3RF1AG068034-01A1S1.

Cummulative percent release of CBD from mannose functionalized chitosan-coated PLGA nanoparticles and chitosan-coated PLGA nanoparticles for 22 days. The data represent mean ± SD (n=4)

(A) Graphical representation of percent relative cell viability at different concentrations of indicated samples. Cytotoxicity was evaluated in bEND.3 cell line through MTT assay. (B) Graphical representation of percent hemolysis in rat red blood cells upon incubation with indicated samples at different concentrations. Data represent the mean ± SD (n=4).