Purpose: The overall objective of this work is to demonstrate granisetron as a potential therapeutic drug against Alzheimer's disease (AD).
Methods: Recently, we developed a high throughput-screening assay to screen for compounds that enhance a cell-based BBB model integrity, which identified multiple hits among which is granisetron, an FDA approved drug. Here, we evaluated the therapeutic potential of granisetron against AD. Animals: Wild type C57Bl/6 male mice at age of 4 months, representing the young group, and 18 months, representing the aged group, and TgSwDI transgenic mice (a CAA/AD mouse model) were used. Animals’ treatment: : young and aged wild type mice were divided into 2 groups each, control and treatment groups (n=7 mice/group. For TgSwDI mice experiments, mice were divided into two groups (n=5 mice/group). The control group in both models received sterile water as a vehicle administered intraperitonially (IP),while the treatment group that received 3 mg/kg/day granisetron administered IP for 28 days. Florescent tagged-Dextran to monitor BBB permeability in wild type young and aged mice: florescent tagged dextran; 10 kDa Texas-red dextran and 40 kDa FITC-dextran were used to measure BBB exravasation. Western Blot analysis: To measure effect on tight junction proteins, synaptic markers, amyloid precursor protein processing and calcium signaling pathway Immunohistochemical analysis: IgG exravasation as a BBB marker, GFAP as marker for astrocytes activation and Aβ load was measured with 6e10 and thioflavin-s for Aβ plaques. ELISA to measure Aβ40 and Aβ42 brain l Analysis of brain acetylcholine levels: was determined indirectly by measuring acetylcholine (ACh) levels. Behavioral testing: The Morris water maze (MWM) test was performed for TgSwDI mice to assess spatial learning and memory performance at the end of the treatment. proteomic analysis: was also performed to examine the proteomic response to granisetron. Statistical analysis: Data were expressed as mean ± SEM for n=7 mice/group for wild type mice and n=5 mice/group for TgSwDI mice. The experimental results were statistically analyzed for significant difference using Student’s t-test for two groups and one-way ANOVA for more than two-group analysis. For behavior studies, two-way ANOVA with Bonferroni post-hoc analysis was used. Values of P < 0.05 were considered statistically significant.
Results: Consistent with our in vitro-HTS findings, our data demonstrated granisetron to restore the BBB integrity and function, reduce Aβ accumulation and neuroinflammation, decrease sustained calcium flux and apoptosis in addition to enhance cognitive function in TgSwDI. In addition, we showed for the first time that granisetron effect is mediated by CREB pathway that is perturbed by aging and AD. This reduction in Aβ load was concomitant with a significant increase in the expression of P-gp, Aβ major transport protein across the BBB, in isolated microvessels, and Aβ degrading enzymes NEP and IDE in brain homogenates.
Conclusion: Granisetron significantly enhanced BBB integrity and function as measured by reduced IgG extravasation and increased expression of tight junction proteins, which is consistent with our previously reported in vitro finding in the CAA-cell based BBB model and reduced brain Aβ accumulation. Collectively, our findings demonstrate granisetron rectifies Aβ related pathology through modulating CREB signaling pathways, and support its repurposing to slow, hold the progression and/or treat AD and related disorders.
Amal Kaddoumi– Auburn University, Auburn, Alabama