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Active immunization against complement factor C5a: a new therapeutic approach for Alzheimer's disease.

Landlinger C, Oberleitner L, Gruber P, Noiges B, Yatsyk K, Santic R, Mandler M, Staffler G - J Neuroinflammation (2015)

Bottom Line: Especially, complement factor C5a and its receptor have been found to be up-regulated in microglia in the immediate surroundings of cerebral amyloid plaques and blocking of C5aR resulted in a reduction of pathological markers in a model of AD.Both C5a-targeting vaccines were highly immunogenic and induced sustained antibody titers against C5a.C5a-peptide vaccines represent a safe and well-tolerated immunotherapy, which is able to induce a strong and specific immune response against the pro-inflammatory molecule C5a.

View Article: PubMed Central - PubMed

Affiliation: AFFiRiS AG, Karl-Farkas-Gasse 22, Vienna, 1030, Austria. christine.landlinger@affiris.com.

ABSTRACT

Background: Alzheimer's disease (AD) is the most common neurodegenerative disease characterized by neuronal loss due to amyloid beta aggregations, neurofibrillary tangles, and prominent neuroinflammation. Recently, interference with neuroinflammation as a new therapeutic approach for AD treatment gained great interest. Microglia cells, one of the major contributors in neuroinflammation, are activated in response to misfolded proteins such as amyloid β and cell debris leading to a sustained release of pro-inflammatory mediators. Especially, complement factor C5a and its receptor have been found to be up-regulated in microglia in the immediate surroundings of cerebral amyloid plaques and blocking of C5aR resulted in a reduction of pathological markers in a model of AD. Here, we investigate the effect of active vaccination against the complement factor C5a to interfere with neuroinflammation and neuropathologic alterations in a mouse model of AD.

Methods: Short antigenic peptides AFF1 and AFF2, which mimic a C-terminal epitope of C5a, were selected and formulated to vaccines. These vaccines are able to induce a highly specific antibody response to the target protein C5a. Tg2576 mice, a common model of AD, were immunized with these two C5a-peptide vaccines and the induced immune response toward C5a was analyzed by ELISA and Western blot analysis. The influence on memory retention was assessed by a contextual fear conditioning test. Microglia activation and amyloid plaque deposition in the brain was visualized by immunohistochemistry.

Results: Both C5a-targeting vaccines were highly immunogenic and induced sustained antibody titers against C5a. Tg2576 mice vaccinated at early stages of the disease showed significantly improved contextual memory accompanied by the reduction of microglia activation in the hippocampus and cerebral amyloid plaque load compared to control mice. Late-stage immunization also showed a decrease in the number of activated microglia, and improved memory function, however, had no influence on the amyloid β load.

Conclusion: C5a-peptide vaccines represent a safe and well-tolerated immunotherapy, which is able to induce a strong and specific immune response against the pro-inflammatory molecule C5a. In a mouse model of AD, C5a-peptide vaccines reduce microglia activation and thus neuroinflammation, which is supposed to lead to reduced neuronal dysfunction and AD symptomatic decline.

No MeSH data available.


Related in: MedlinePlus

C5a-peptide vaccines induce target-specific antibody response in Tg2576 mice. a Tg2576 mice were immunized with AFF1- (n = 12) and AFF2- (n = 13) containing vaccines 4 times in biweekly intervals (week 0, 2, 4, and 6) followed by 3 boosts in a monthly interval (week 11, 15, and 19). The immunizations are indicated by an asterisk. Plasma samples were collected at the indicated time points (week 2–28). All samples were analyzed by ELISA and the mean IgG antibody titers (ODmax/2) against the injected peptide AFF1 and AFF2 are presented. b The titers against both forms of the target protein, C5a desARG and C5a ARG, were analyzed in the plasma obtained after the last immunization in W28. Also control immunized mice (n = 11) were tested against C5a proteins (c). Western blot analyses where recombinant C5a ARG and C5a desARG were loaded and detected by AFF1 (second panel), AFF2 induced immune plasma (third panel), as well as plasma obtained from control immunized mice (fourth panel), and untreated wt mice (fifth panel) as a negative control. Rabbit anti-mouse C5a antibody was used as a positive control (first panel). The Precision Plus Protein™ Dual Color Standards (Bio-Rad) was used as a marker. d CSF was obtained from five randomly selected AFF1 and AFF2 immunized mice and tested for AFFITOPE®-specific antibodies in week 28. All data points represent the group means ± SEM of n animals
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Fig3: C5a-peptide vaccines induce target-specific antibody response in Tg2576 mice. a Tg2576 mice were immunized with AFF1- (n = 12) and AFF2- (n = 13) containing vaccines 4 times in biweekly intervals (week 0, 2, 4, and 6) followed by 3 boosts in a monthly interval (week 11, 15, and 19). The immunizations are indicated by an asterisk. Plasma samples were collected at the indicated time points (week 2–28). All samples were analyzed by ELISA and the mean IgG antibody titers (ODmax/2) against the injected peptide AFF1 and AFF2 are presented. b The titers against both forms of the target protein, C5a desARG and C5a ARG, were analyzed in the plasma obtained after the last immunization in W28. Also control immunized mice (n = 11) were tested against C5a proteins (c). Western blot analyses where recombinant C5a ARG and C5a desARG were loaded and detected by AFF1 (second panel), AFF2 induced immune plasma (third panel), as well as plasma obtained from control immunized mice (fourth panel), and untreated wt mice (fifth panel) as a negative control. Rabbit anti-mouse C5a antibody was used as a positive control (first panel). The Precision Plus Protein™ Dual Color Standards (Bio-Rad) was used as a marker. d CSF was obtained from five randomly selected AFF1 and AFF2 immunized mice and tested for AFFITOPE®-specific antibodies in week 28. All data points represent the group means ± SEM of n animals

Mentions: Human APP transgenic Tg2576 mice on a 129S6 genetic background were used as a model of AD-like disease. In a first study, starting at the age of 8 months, Tg2576 mice were either immunized with AFF1- or AFF2-formulated anti-C5a vaccines. Two weeks after the second immunization (W4), titers against the immunizing peptides AFF1 and AFF2 approached approximately 1/20.000 and 1/45.000, respectively, and remained at this level until week 28 (W28) corresponding to 15 months of age (Fig. 3a). Compared to the results obtained from immunized wt mice, AFF2-containing vaccines induced similar high titers against the antigenic peptide AFF2, whereas AFF1-containing vaccines elicited a clearly lower titer against the peptide moiety (Figs. 1a and 3a). ELISA for the proteins C5a desARG and C5a ARG revealed that AFF1-containing vaccine mounted higher antibody titers against C5a desARG whereas AFF2-containing vaccine showed a much higher immune response against C5a ARG (Fig. 3b), similar to those results obtained from AFF1 and AFF2 immunized wt mice (Fig. 1b). As expected, control immunized mice did not show any immune response against the C5a proteins (Fig. 3b). Western blot analyses using recombinant C5a ARG and C5a desARG confirmed this reactivity pattern of AFF1- and AFF2-induced antibodies. Again, AFF1-elicited antibodies showed a better recognition for C5a desARG, whereas AFF2-induced immune plasma revealed a stronger band for C5a ARG. Control immunized mice as well as plasma obtained from untreated wt littermates at the age of 15 months did not show any signal toward both forms of C5a (Fig. 3c).Fig. 3


Active immunization against complement factor C5a: a new therapeutic approach for Alzheimer's disease.

Landlinger C, Oberleitner L, Gruber P, Noiges B, Yatsyk K, Santic R, Mandler M, Staffler G - J Neuroinflammation (2015)

C5a-peptide vaccines induce target-specific antibody response in Tg2576 mice. a Tg2576 mice were immunized with AFF1- (n = 12) and AFF2- (n = 13) containing vaccines 4 times in biweekly intervals (week 0, 2, 4, and 6) followed by 3 boosts in a monthly interval (week 11, 15, and 19). The immunizations are indicated by an asterisk. Plasma samples were collected at the indicated time points (week 2–28). All samples were analyzed by ELISA and the mean IgG antibody titers (ODmax/2) against the injected peptide AFF1 and AFF2 are presented. b The titers against both forms of the target protein, C5a desARG and C5a ARG, were analyzed in the plasma obtained after the last immunization in W28. Also control immunized mice (n = 11) were tested against C5a proteins (c). Western blot analyses where recombinant C5a ARG and C5a desARG were loaded and detected by AFF1 (second panel), AFF2 induced immune plasma (third panel), as well as plasma obtained from control immunized mice (fourth panel), and untreated wt mice (fifth panel) as a negative control. Rabbit anti-mouse C5a antibody was used as a positive control (first panel). The Precision Plus Protein™ Dual Color Standards (Bio-Rad) was used as a marker. d CSF was obtained from five randomly selected AFF1 and AFF2 immunized mice and tested for AFFITOPE®-specific antibodies in week 28. All data points represent the group means ± SEM of n animals
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4537556&req=5

Fig3: C5a-peptide vaccines induce target-specific antibody response in Tg2576 mice. a Tg2576 mice were immunized with AFF1- (n = 12) and AFF2- (n = 13) containing vaccines 4 times in biweekly intervals (week 0, 2, 4, and 6) followed by 3 boosts in a monthly interval (week 11, 15, and 19). The immunizations are indicated by an asterisk. Plasma samples were collected at the indicated time points (week 2–28). All samples were analyzed by ELISA and the mean IgG antibody titers (ODmax/2) against the injected peptide AFF1 and AFF2 are presented. b The titers against both forms of the target protein, C5a desARG and C5a ARG, were analyzed in the plasma obtained after the last immunization in W28. Also control immunized mice (n = 11) were tested against C5a proteins (c). Western blot analyses where recombinant C5a ARG and C5a desARG were loaded and detected by AFF1 (second panel), AFF2 induced immune plasma (third panel), as well as plasma obtained from control immunized mice (fourth panel), and untreated wt mice (fifth panel) as a negative control. Rabbit anti-mouse C5a antibody was used as a positive control (first panel). The Precision Plus Protein™ Dual Color Standards (Bio-Rad) was used as a marker. d CSF was obtained from five randomly selected AFF1 and AFF2 immunized mice and tested for AFFITOPE®-specific antibodies in week 28. All data points represent the group means ± SEM of n animals
Mentions: Human APP transgenic Tg2576 mice on a 129S6 genetic background were used as a model of AD-like disease. In a first study, starting at the age of 8 months, Tg2576 mice were either immunized with AFF1- or AFF2-formulated anti-C5a vaccines. Two weeks after the second immunization (W4), titers against the immunizing peptides AFF1 and AFF2 approached approximately 1/20.000 and 1/45.000, respectively, and remained at this level until week 28 (W28) corresponding to 15 months of age (Fig. 3a). Compared to the results obtained from immunized wt mice, AFF2-containing vaccines induced similar high titers against the antigenic peptide AFF2, whereas AFF1-containing vaccines elicited a clearly lower titer against the peptide moiety (Figs. 1a and 3a). ELISA for the proteins C5a desARG and C5a ARG revealed that AFF1-containing vaccine mounted higher antibody titers against C5a desARG whereas AFF2-containing vaccine showed a much higher immune response against C5a ARG (Fig. 3b), similar to those results obtained from AFF1 and AFF2 immunized wt mice (Fig. 1b). As expected, control immunized mice did not show any immune response against the C5a proteins (Fig. 3b). Western blot analyses using recombinant C5a ARG and C5a desARG confirmed this reactivity pattern of AFF1- and AFF2-induced antibodies. Again, AFF1-elicited antibodies showed a better recognition for C5a desARG, whereas AFF2-induced immune plasma revealed a stronger band for C5a ARG. Control immunized mice as well as plasma obtained from untreated wt littermates at the age of 15 months did not show any signal toward both forms of C5a (Fig. 3c).Fig. 3

Bottom Line: Especially, complement factor C5a and its receptor have been found to be up-regulated in microglia in the immediate surroundings of cerebral amyloid plaques and blocking of C5aR resulted in a reduction of pathological markers in a model of AD.Both C5a-targeting vaccines were highly immunogenic and induced sustained antibody titers against C5a.C5a-peptide vaccines represent a safe and well-tolerated immunotherapy, which is able to induce a strong and specific immune response against the pro-inflammatory molecule C5a.

View Article: PubMed Central - PubMed

Affiliation: AFFiRiS AG, Karl-Farkas-Gasse 22, Vienna, 1030, Austria. christine.landlinger@affiris.com.

ABSTRACT

Background: Alzheimer's disease (AD) is the most common neurodegenerative disease characterized by neuronal loss due to amyloid beta aggregations, neurofibrillary tangles, and prominent neuroinflammation. Recently, interference with neuroinflammation as a new therapeutic approach for AD treatment gained great interest. Microglia cells, one of the major contributors in neuroinflammation, are activated in response to misfolded proteins such as amyloid β and cell debris leading to a sustained release of pro-inflammatory mediators. Especially, complement factor C5a and its receptor have been found to be up-regulated in microglia in the immediate surroundings of cerebral amyloid plaques and blocking of C5aR resulted in a reduction of pathological markers in a model of AD. Here, we investigate the effect of active vaccination against the complement factor C5a to interfere with neuroinflammation and neuropathologic alterations in a mouse model of AD.

Methods: Short antigenic peptides AFF1 and AFF2, which mimic a C-terminal epitope of C5a, were selected and formulated to vaccines. These vaccines are able to induce a highly specific antibody response to the target protein C5a. Tg2576 mice, a common model of AD, were immunized with these two C5a-peptide vaccines and the induced immune response toward C5a was analyzed by ELISA and Western blot analysis. The influence on memory retention was assessed by a contextual fear conditioning test. Microglia activation and amyloid plaque deposition in the brain was visualized by immunohistochemistry.

Results: Both C5a-targeting vaccines were highly immunogenic and induced sustained antibody titers against C5a. Tg2576 mice vaccinated at early stages of the disease showed significantly improved contextual memory accompanied by the reduction of microglia activation in the hippocampus and cerebral amyloid plaque load compared to control mice. Late-stage immunization also showed a decrease in the number of activated microglia, and improved memory function, however, had no influence on the amyloid β load.

Conclusion: C5a-peptide vaccines represent a safe and well-tolerated immunotherapy, which is able to induce a strong and specific immune response against the pro-inflammatory molecule C5a. In a mouse model of AD, C5a-peptide vaccines reduce microglia activation and thus neuroinflammation, which is supposed to lead to reduced neuronal dysfunction and AD symptomatic decline.

No MeSH data available.


Related in: MedlinePlus