Enhanced and long term immunogenicity of a Her-2/neu multi-epitope vaccine conjugated to the carrier CRM197 in conjunction with the adjuvant Montanide

Abstract

BackgroundWe previously identified three short single peptides (P4, P6 and P7) representing different B-cell epitopes on the extracellular domain of Her-2/neu for a vaccine that was tested in a phase-I clinical trial. Here we describe the improvement of the multi peptide vaccine by fusing the single peptides to a hybrid peptide P467.MethodsAfter coupling to either virosomes or to diphtheria toxoid CRM197 (CRM), the hybrid peptide was tested in different concentrations in combination with either Montanide or Aluminium hydroxide (Alum) in preclinical studies.ResultsAlready low amount (10 μg) of P467 conjugated to CRM led to faster onset of high antibody levels compared to the P467-virosome. The formulation P467-CRM-Montanide induced higher serum IgG antibody titers, compared with P467-CRM-Alum, as examined by ELISA using recombinant Her-2/neu or Her-2/neu natively expressed on the tumor cell line SK-BR-3. Compared to P467-CRM-Alum, higher in vitro production of IL-2 and IFNγ in the Montanide-immunized mice was induced after re-stimulation of splenocytes with CRM but also with P467, indicating a clear Th1-biased response. In contrast to the single B cell peptides, the hybrid peptide led to T cell proliferation and cytokine production as CD4 T cell epitopes were generated in the fusion region of the single peptides P4 and P6 or P6 and P7. Additionally, a significantly higher proportion IFNγ-producing CD8+ T cells was found in the P467-CRM-Montanide immunized mice, probably by Montanide-driven bystander activation. Importantly, anti-P467 IgG antibodies exhibited anti-tumor properties and the combination of anti-P467 specific IgG with Herceptin® was found to inhibit the proliferation of Her-2/neu-overexpressing cell line SK-BR-3 in a significantly higher capacity than Herceptin® alone.ConclusionsFusion of the B cell peptides has led to additional generation of CD4 T cell epitopes, and this P467-multi epitope vaccine was found to induce polyclonal antibody responses with anti-proliferative capacity against Her-2/neu. The hybrid vaccine together with Montanide induced higher and long-lasting antibody levels, Th1-biased cellular responses being superior to vaccination with the single B cell peptides. This vaccine formulation is now planned to be evaluated in a phase Ib/II study in Her-2/neu overexpressing cancer patients.

Designing a multi-epitope based vaccine to combat Kaposi Sarcoma utilizing immunoinformatics approach

“…Thus, the in silico cloning was performed in the present study to ensure the efficient expression and translation of the multi-epitope vaccine construct in an expression vector: pET-28a (+). The similar kind of strategy of designing a multi-epitope vaccine have been applied recently by several research groups against multiple myeloma, oncoprotein Her-2/neu, avian leukosis virus, Chikungunya, Hepatitis C virus, Dengue, Visceral leishmaniasis etc 12,28–33 . The proposed mechanism of action of the final vaccine construct was also predicted (Fig.…”

B-Cell Epitope Peptide Cancer Vaccines: A New Paradigm for Combination Immunotherapies with Novel Checkpoint Peptide Vaccine

“…The HER-2 multipeptide vaccine was safe, well tolerated and effective in overcoming immunological tolerance to HER-2. More recently, Wiedermann group developed a new vaccine: three short single peptides (P4, P6 and P7) representing different HER-2 ECD B-cell epitopes fused as a single hybrid peptide P467 on either virosomes or to diphtheria toxoid CRM197 (CRM) [ 97 ]. The formulation P467-CRM-Montanide induced higher serum IgG antibody titers, compared with P467-CRM-Alum.…”

Application of built-in adjuvants for epitope-based vaccines

“…Moreover, CRM can rapidly activate CD4 + T cells by generating a multitude of Th1 and Th2 cytokines, thereby promoting the proliferation of B cells and regulating the level of antibody production (Kamboj et al, 2001). Several short B cell epitopes (P4, P6, and P7) on the Her-2/neu protein were combined with the polyepitope peptide, P467, using CRM197 as a carrier protein to conjugate with this complex epitope, demonstrated a strong anti-tumor response (Tobias et al, 2017). The function of DTT is to assist the enzyme active region (C-domain) in passing through the endocytosis membrane, and there are no associated risks when the transmembrane domain of DTT is used as the protein carrier for exogenous antigen (Ladokhin, 2013; Malito et al, 2012).…”