Chemically linked phage idiotype vaccination in the murine B cell lymphoma 1 model

Abstract

Background: B cell malignancies are characterized by clonal expansion of B cells expressing tumor-specific idiotypes on their surface. These idiotypes are ideal target antigens for an individualized immunotherapy. However, previous idiotype vaccines mostly lacked efficiency due to a low immunogenicity of the idiotype. The objective of the present study was the determination of the feasibility, safety and immunogenicity of a novel chemically linked phage idiotype vaccine.

Methods: In the murine B cell lymphoma 1 model, tumor idiotypes were chemically linked to phage particles used as immunological carriers. For comparison, the idiotype was genetically expressed on the major phage coat protein g8 or linked to keyhole limpet hemocynanin. After intradermal immunizations with idiotype vaccines, tolerability and humoral immune responses were assessed.

Results: Feasibility and tolerability of the chemically linked phage idiotype vaccine was demonstrated. Vaccination with B cell lymphoma 1 idiotype expressing phage resulted in a significant survival benefit in the murine B cell lymphoma 1 protection model (60.2±23.8 days vs. 41.8±1.6 days and 39.8±3.8 days after vaccination with wild type phage or phosphate buffered saline, respectively). Superior immunogenicity of the chemically linked phage idiotype vaccine compared to the genetically engineered phage idiotype and keyhole limpet hemocynanin-coupled idiotype vaccine was demonstrated by significantly higher B cell lymphoma 1 idiotype-specific IgG levels after vaccination with chemically linked phage idiotype.

Conclusion: We present a novel, simple, time- and cost-efficient phage idiotype vaccination strategy, which represents a safe and feasible therapy and may produce a superior immune response compared to previously employed idiotype vaccination strategies.

Figure 1

Determination of the phage vaccine dosage. The humoral immune response after vaccination of mice with phage was determined by ELISA (antibody dilution: 1/1000). (A) Total IgG response at day 28 was measured after the last vaccination with 10¹⁰, 10¹¹ or 5 × 10¹¹ phage particles weekly for 4 weeks (n = 6). (B) Anti-phage specific IgG isotype levels before and at day 28 after 4 weekly vaccinations with 10¹⁰, 10¹¹ or 5 × 10¹¹ phage particles (n = 6). * p > 0.05.

Figure 2

Tumor protection procured by Id-phage vaccines. Kaplan-Meier plot of the survival of the mice vaccinated weekly with 5 × 10¹¹ bacteriophages (BCL1-g8 or PBS or wild type phage as control). Seven days after the last vaccination, 10⁵ BCL1 cells were injected intraperitoneally; n = 12. p < 0.05 (BCL1-g8 vs. wild type phage and vs. PBS).

Figure 3

Anti-phage IgG response. The humoral immune response after vaccination of mice with phage was determined by ELISA (antibody dilution: 1/1000). (A) Total IgG response of single mice 4 weeks (day 28) after the last of 4 weekly vaccinations with PBS, BCL1-KLH, BCL1-g8, BCL1-WT or wild type phage (n = 6). (B) Sustainability of the humoral immune response to phage vaccines: Anti-phage IgG levels before (day 0) and at day 14, 28 and 120 after the last of 4 weekly vaccinations with BCL1-g8, BCL1-WT or wild type phage (5 × 10¹¹ each; n = 6). * p > 0.05, *** p < 0.001.

Figure 4

Anti-phage IgM response and IgG response with or without addition of GM-CSF. (A) IgM responses at day 28 were measured after the last vaccination with 10¹⁰, 10¹¹ or 5 × 10¹¹ phage particles weekly for 4 weeks (n = 6; antibody dilution: 1/200). (B) Anti-phage specific IgG isotype levels at day 28 after 4 weekly vaccinations with 5 × 10¹¹ phage particles with or without addition of 20 μg GM-CSF (n = 6; antibody dilution: 1/1000).

Figure 5

Anti-phage IgG response. The anti-phage humoral immune response after vaccination of mice with phage was determined by ELISA (antibody dilution: 1/1000) (A) Anti-BCL1-specific IgG response before (day 0) and at day 14, 28 and 120 after vaccination with PBS, wild type phage, BCL1-KLH, BCL1-g8 or BCL1-WT (5 × 10¹¹ each; n = 6). (B) Anti-BCL1-specific IgG isotypes at day 14 and 120 after vaccination. * p > 0.05, ** p < 0.01; *** p < 0.001.