Proteolytic degradation patterns of the receptor for advanced glycation end products peptide fragments correlate with their neuroprotective activity in Alzheimer's disease models.

The receptor for advanced glycation end products (RAGE) plays an essential role in Alzheimer's disease (AD). We previously demonstrated that a fragment (60-76) of RAGE improved the memory of olfactory bulbectomized (OBX) and Tg 5 × FAD mice - animal models of AD. The peptide analog (60-76) with protected N- and C-terminal groups was more active than the free peptide in Tg 5 × FAD mice. This study investigated proteolytic cleavage of the RAGE fragment (60-76) and its C- and N-terminally modified analog by blood serum using HPLC and mass spectrometry. The modified peptide was proteolyzed slower than the free peptide. Degrading the protected analog resulted in shortened fragments with memory-enhancing effects, whereas the free peptide yielded inactive fragments. After administering the different peptides to OBX mice, their performance in a spatial memory task revealed that the effective dose of the modified peptide was five times lower than that of the free peptide. HPLC and mass spectrometry analysis of the proteolytic products allowed us to clarify the differences in the neuroprotective activity conferred by administering these two peptides to AD animal models. The current study suggests that the modified RAGE fragment is more promising for the development of anti-AD therapy than its free analog.

Synthetic Fragment of Receptor for Advanced Glycation End Products Prevents Memory Loss and Protects Brain Neurons in Olfactory Bulbectomized Mice.

Activation of receptor for advanced glycation end products (RAGE) plays an essential role in the development of Alzheimer's disease (AD). It is known that the soluble isoform of the receptor binds to ligands and prevents negative effects of the receptor activation. We proposed that peptide fragments from RAGE prevent negative effects of the receptor activation during AD neurodegeneration. We have synthesized peptide fragments from surface-exposed regions of RAGE. Peptides were intranasally administrated into olfactory bulbectomized (OBX) mice, which developed some characteristics similar to AD neurodegeneration. We have found that only insertion of fragment (60-76) prevents the memory of OBX mice. Immunization of OBX mice with peptides showed that again only (60-76) peptide protected the memory of animals. Both intranasal insertion and immunization decreased the amyloid-ß (Aß) level in the brain. Activity of shortened fragments of (60-76) peptide was tested and showed only the (60-70) peptide is responsible for manifestation of activity. Intranasal administration of (60-76) peptide shows most protective effect on morpho-functional characteristics of neurons in the cortex and hippocampal areas. Using Flu-(60-76) peptide, we revealed its penetration in the brain of OBX mice as well as colocalization of Flu-labeled peptide with Aß in the brain regions in transgenic mice. Flu-(60-76) peptide complex with trimer of Aß was detected by SDS-PAGE. These data indicate that Aß can be one of the molecular target of (60-70) peptide. These findings provide a new peptide molecule for design of anti-AD drug and for investigation of RAGE activation ways in progression of AD neurodegeneration.

Vaccination with peptide 173-193 of acetylcholine receptor α7-subunit prevents memory loss in olfactory bulbectomized mice.

We studied the ability of four non-conjugated alpha7-subunit fragments of the nicotinic acetylcholine receptor to induce an immune response and to protect memory in olfactory bulbectomized mice which demonstrate abnormalities similar to Alzheimer's disease (AD). Vaccination only with the alpha7-subunit fragment 173-193 was shown to rescue spatial memory, to restore the level of alpha7 acetylcholine receptors in the cortex, and to prevent an increase in the amyloid-beta (Abeta) level in brain tissue in these animals. Antibodies against the peptide 173-193 were revealed in blood serum and cerebrospinal liquid in the bulbectomized mice. Passive immunization with mouse blood sera containing antibodies to the peptide 173-193 also restored memory in bulbectomized animals. The observed positive effect of both active and passive immunization with the fragment of alpha7-subunit on memory of bulbectomized mice provides a new insight into an anti-AD drug design.

Antibodies to a nonconjugated prion protein peptide 95-123 interfere with PrP( Sc ) propagation in prion-infected cells.

1. Vaccination-induced anti-prion protein antibodies are presently regarded as a promising approach toward treatment of prion diseases. Here, we investigated the ability of five peptides corresponding to three different regions of the bovine prion protein (PrP) to elicit antibodies interfering with PrP(Sc) propagation in prion-infected cells.2. Rabbits were immunized with free nonconjugated peptides. Obtained immune sera were tested in enzyme-linked immunosorbent assay (ELISA) and immunoblot for their binding to recombinant PrP and cell-derived pathogenic isoform (PrP(Sc)) and normal prion protein (PrP(c)), respectively. Sera positive in all tests were chosen for PrP(Sc) inhibition studies in cell culture.3. All peptides induced anti-peptide antibodies, most of them reacting with recombinant PrP. Moreover, addition of the serum specific to peptide 95-123 led to a transient reduction of PrP(Sc) levels in persistently prion-infected cells.4. Thus, anti-PrP antibodies interfering with PrP(Sc) propagation were induced with a prion protein peptide nonconjugated to a protein carrier. These results point to the potential application of the nonconjugated peptide 95-123 for the treatment of prion diseases.