Boosted Memory and Improved Brain Bioavailability of Rivastigmine: Targeting Effort to the Brain Using Covalently Tethered Lower Generation PAMAM Dendrimers with Lactoferrin.

Abstract

Currently, there is no treatment strategy which can reverse the process of neuro-degeneration in progression of Alzheimer's disease (AD). Practically, it is desired to achieve and maintain high therapeutic doses in the brain, but it is hard due to selective permeability of the blood-brain barrier (BBB). In the present study, lactoferrin (Lf) was conjugated to polyamidoamine generation 3.0 (PAMAM G3.0) dendrimers for the effective delivery of rivastigmine (RIV) to the brain. Conjugation of PAMAM G3.0 with lactoferrin was confirmed by FT-IR, 1H NMR, and 2D-NMR spectroscopy as well as AFM techniques. Further, RIV was loaded into PAMAM G3.0 and PAMAM-Lf conjugates. RP-HPLC was used to quantify the drug loading and release as well. Spectroscopic analysis confirmed PAMAM-Lf conjugation, the size of the conjugate was 100.04 ± 3.1 nm, and after RIV loading, the size was increased up to 216.13 ± 2.3 nm. Atomic force microscopic results revealed that the root-mean-square roughness ( Rq) and surface roughness ( Ra) were 6.31 and 5.27 nm, respectively, along with other parameters, Skewness and Kurtosis, which were 0.522 and 2.50, respectively. In vitro drug release from the PAMAM-Lf-RIV conjugate was sustained up to more than 100 h, and that of naive RIV was quite rapid (approxmately 99% release was observed in 8 h). Ex vivo hemotoxicity of the PAMAM-Lf-RIV conjugate was almost 9.8-fold lesser than the PAMAM G3.0 ( p < 0.0001), 7.77 times that of PAMAM-enc-RIV and 5 times that of naïve RIV ( p < 0.0001), respectively. The in vivo targeting potency of the conjugate was investigated in a rat model. Bioavailability of the RIV was enhanced 7.87 times compared to RIV along with improved pharmacokinetic parameters. Brain uptake of the drug was improved when treated with PAMAM-Lf-RIV over the RIV and PAMAM-enc-RIV, almost 8 and 4.2 times, respectively, after 4 h of the administration. Additionally, the behavioral studies revealed that PAMAM-Lf-RIV significantly enhanced the overall locomotor activity with higher ambulations over the pure drug and PAMAM-enc-RIV formulation. The outcome of the novel object recognition test was an indirect evidence of memory improvement. Conclusively, the development and characterization of PAMAM-Lf-RIV resulted in improved brain uptake and brain bioavailability with boosted memory, which can be beneficial in the treatment of Alzheimer's.