A liposome-based therapeutic vaccine against β-amyloid plaques on the pancreas of transgenic NORBA mice

Abstract

immune tolerance ͉ anti-A␤-antibodies ͉ lipid A ͉ A␤-fiber solubilization A lzheimer's disease (AD) is a progressive degenerative disorder of insidious onset characterized by memory loss, confusion, and a variety of cognitive disabilities. The major neuropathological change in the brains of AD patients is neuronal death, particularly in regions related to memory and cognition (1). One of the major pathological features of AD is the abundant presence of amyloid plaques in the brain of affected individuals (2). Intracellular bundles of paired helical filaments, composed largely of phosphorylated tau protein, accumulate in large amounts in dying neurons (3). On the neuron surfaces, insoluble aggregates of proteinaceous debris, termed amyloid, appear in the form of neuritic plaques and vascular amyloid deposits (1). The frequency and distribution of the neurofibrillar tangles and of the neuritic plaques appear to correlate well with the extent of cognitive impairment and other characteristic symptoms of AD (3).Amyloid plaques are formed by the ␤-amyloid peptide (A␤), a 39-to 43-aa-long polypeptide that is mostly coiled and slightly ␣-helical in its benign soluble form and, on conformational transition into a mainly ␤-sheet secondary structure, spontaneously aggregates into insoluble deposits. A␤ is a physiological metabolite of the much larger amyloid precursor protein (APP), 695-770 aa long, which undergoes sequential proteolysis (4). The peptide may remain in solution as a random coil or an ␣-helix (5).A mAb, raised against the sequence A␤ 1-16 of the amyloid protein, was shown in vitro to have a solubilizing effect on fibrils formed by the A␤ 1-42 amyloidogenic peptide (6). The amyloid filaments obtained were similar to those found in amyloid plaques and cerebrovascular amyloid, assembled from chemically synthesized amyloid sequences under defined experimental conditions (6).Previous studies in our laboratory had shown that palmitoylated peptide sequences of the multidrug-resistance (MDR)1 protein, reconstituted in the bilayer of liposomes, when injected into mice elicited strong immune responses, breaking the immune tolerance to this ''self'' protein (7).In the present study, we report that the palmytoilated A␤ 1-16 sequence reconstituted in liposomes-lipid A, when injected i.p. into mice, including transgenic NORBA mice, which overexpress human APP resulting in amyloid plaque deposits on their pancreases, elicited significant titers of antiamyloid antibodies displaying therapeutic as well as prophylactic action in NORBA transgenic mice. Materials and MethodsChemicals. Dichloromethane from Carlo Erba (Milan) was distilled freshly from calcium hydride under nitrogen before use. Acetic anhydride, 4-dimethylaminopyridine, and dicyclohexylcarbodiimide were purchased from Sigma-Aldrich; ␣,-dipalmitoyllysine was synthesized according to literature methods (8); ␣-fluorenylmethoxycarbonyl (Fmoc)--palmitoyllisine [FmocLys(Pal)OH], the 4-alkoxybenzyl alcohol resin for automated solid-phase synthesis, as well as A␤ 1-42 , ...

Getting into the brain: liposome-based strategies for effective drug delivery across the blood–brain barrier

“…84,85 Although the scope of this review is on liposome-strategies with the aim of facilitating BBB crossing, it is important to mention that other strategies have been developed for the use of liposomes for AD treatment. 89,[113][114][115][116] Curcumin is a natural compound extract from the plant Curcuma longa, and has been reported to be a fluorescent molecule with high affinity for the Aβ peptide and able to reduce Aβ aggregation. In this way, intracranial injection of liposomes encapsulating curcumin efficiently labeled Aβ deposits in both human and mice tissues, proving to be an effective formulation for diagnosis and treatment of AD.…”

“…113 Also, intraperitoneal injection of liposomes containing phosphatidic acid or cardiolipin was able to reduce Aβ peptides in the plasma and shifted the equilibrium that exists between brain and blood Aβ peptides, slightly affecting the plaques in the brain. 89 Lastly, different liposome-based vaccines were developed and directed toward Aβ plaques 115,116 and tau.…”

Liposomal vaccines with conformation-specific amyloid peptide antigens define immune response and efficacy in APP transgenic mice

“…Liposomes were prepared as described (19). For the synthesis of the palmitoylated amyloid 1-15 peptide in ACI-24, the orthogonally protected amino acid FmocLys(Mtt)-OH (Merck, Darmstadt, Germany) was coupled to an H 2 N-Lys(Mtt)-Wang resin.…”

“…inoculation of tetrapalmitoylated A␤1-16 reconstituted in liposomes to transgenic NORBA mice elicited significant titers of anti-A␤ antibodies, that solubilized amyloid fibers in vitro and pancreatic A␤ plaques in vivo (19). To circumvent T cell-mediated immune responses known to be causatively involved in the adverse events of meningoencephalitis of AD patients immunized with A␤1-42 (20)(21)(22), the A␤1-16 and 1-15 sequences were used for immunization of APPxPS1 double-transgenic mice (23) because strong T cell epitopes are located more toward the C-terminal region for mice (24) and humans (25).…”

Brain Drug Delivery Systems for Neurodegenerative Disorders

“…These liposomes were administered in three different strains of mice by the intra-peritoneal route [107]. A significant immune response was observed in the liposomes/Aβ(1-16) vaccinated BALB/c mice after the third inoculation.…”