Intranasal rifampicin for Alzheimer's disease prevention

Abstract

Introduction: Oral rifampicin has been shown to significantly reduce amyloid β (Aβ) and tau pathologies in mice. However, it shows occasional adverse effects such as liver injury in humans, making its use difficult for a long period.

Methods: To explore safer rifampicin treatment, APP_OSK mice, a model of Alzheimer's disease, were treated with rifampicin for 1 month via oral, intranasal, and subcutaneous administration, and its therapeutic efficacy and safety were compared.

Results: Intranasal or subcutaneous administration of rifampicin improved memory more effectively than oral administration. The improvement of memory was accompanied with the reduction of neuropathologies, including Aβ oligomer accumulation, tau abnormal phosphorylation, and synapse loss. Serum levels of a liver enzyme significantly rose only by oral administration. Pharmacokinetic study revealed that the level of rifampicin in the brain was highest with intranasal administration.

Discussion: Considering its easiness and noninvasiveness, intranasal administration would be the best way for long-term dosing of rifampicin.

Keywords: Alzheimer's disease; Aβ oligomers; Hepatotoxicity; Intranasal; Pharmacokinetics; Prevention; Rifampicin.

Fig. 1

Effects of differently administered rifampicin on memory in APP_OSK mice. (A and B) Rifampicin (RFP) was administered to APP_OSK mice orally (p.o.), intranasally (i.n.) or subcutaneously (s.c.) at 0.25 or 0.05 mg/day for 1 month. As controls, carboxymethylcellulose (CMC) solution was administered to age-matched Tg and non-Tg littermates. (A) Spatial reference memory in mice was assessed at 12 months of age using the Morris water maze. Each point represents the mean of five trials per day ± SEM (sample number for each group is shown). The differences between groups were evaluated using two-factor repeated measures analysis of variance followed by Fisher's PLSD test. *P = .0027 vs. Non-Tg, P = .0038 vs. RFP (i.n.) (0.25 mg), P = .0059 vs. RFP (i.n.) (0.05 mg), P = .0029 vs. RFP s.c. (0.25 mg). (B) Retention of memory was assessed by a probe trial on day 5. Data are given as time occupancy in the target quadrant. (C) Locomotor activity of APP_OSK mice in the light and dark was measured at 18 months of age by an open field test. Spontaneous locomotion of mice was estimated by the number of their beam crossing. Each point represents the mean ± SEM (n = 10 for each group). Abbreviation: SEM, standard error of the mean.

Fig. 2

Effects of differently administered rifampicin on Aβ accumulation in APP_OSK mice. After the behavioral test, the mice were divided into two groups: one for immunohistochemical analysis and the other for biochemical analysis. (A) Brain sections were stained with Aβ oligomer-specific 11A1 antibody. The photographs were taken from the hippocampal CA3 region. Scale bar = 30 μm. (B) The staining intensity in a constant area was quantified. Each bar represents the mean ± SEM (n = 6 for each group). AU, arbitrary unit. (C) Brain homogenates were separated into TBS-, SDS-, and FA-soluble fractions and subjected to Aβ40, Aβ42, and Aβ oligomer sandwich ELISA. Aβ oligomers were also measured by direct ELISA with 11A1 antibody. Each bar represents the mean ± SEM (n = 5 for CMC and subcutaneous RFP groups, and n = 6 for oral and intranasal RFP groups). *P < .05 vs. CMC treatment by Tukey-Kramer test. Abbreviations: Aβ, amyloid β; CMC, carboxymethylcellulose; ELISA, enzyme-linked immunosorbent assay; FA, formic acid; SDS, sodium dodecyl sulfate; SEM, standard error of the mean; TBS, Tris-buffered saline.

Fig. 3

Effects of differently administered rifampicin on tau phosphorylation and synapse loss in APP_OSK mice. Rifampicin-treated mice were further analyzed. (A) Brain sections were stained with PHF-1 (anti-pSer396/404-tau) antibody. The photographs were taken from the hippocampal CA2/3 region. Scale bar = 30 μm. (B) The staining intensity in a constant area was quantified. Each bar represents the mean ± SEM (n = 6 for each group). (C) Brain sections were stained with anti-synaptophysin antibody. The photographs were taken from the hippocampal CA2/3 region. Scale bar = 30 μm. (D) The staining intensity in a constant area was quantified. Each bar represents the mean ± SEM (n = 6 for each group). (E) Hippocampal homogenates were subjected to Western blot for synaptophysin and actin. The intensity of each signal was quantified. Each bar represents the mean ± SEM (n = 6 for non-Tg, oral and intranasal RFP groups, and n = 5 for CMC and subcutaneous RFP groups). Abbreviations: CMC, carboxymethylcellulose; SEM, standard error of the mean.

Fig. 4

Pharmacokinetics of differently administered rifampicin in normal mice. A mixture of unlabeled and radiolabeled rifampicin was single administered to wild-type mice orally (p.o.), intranasally (i.n.), or subcutaneously (s.c.) at a dose of 2.5 μg/mouse. At the indicated time (15, 30, 60, 120, or 180 min) after application, blood and whole brains were collected. Plasma (A) and brain homogenates (B) were prepared, and radioactivity in the samples was determined with a liquid scintillation counter. Each bar represents the mean ± SEM (n = 3 for each point except for n = 4 at 120 min after intranasal application and at 15 min after subcutaneous application). ^#P < .05 vs. p.o., ^##P < .01 vs. p.o., ^∗∗P < .01 vs. s.c. by Dunnett's test. Abbreviation: SEM, standard error of the mean.