Cancer Res Treat.  2015 Jul;47(3):518-526. 10.4143/crt.2013.241.

Improved Efficacy of a Dendritic Cell-Based Vaccine against a Murine Model of Colon Cancer: The Helper Protein Effect

• Zarnani AH
• Torabi-Rahvar M
• Bozorgmehr M
• Zareie M
• Mojtabavi

• KMID: 2148503
• DOI: http://doi.org/10.4143/crt.2013.241

Abstract

PURPOSE
Targeted immunotherapy using dendritic cells (DCs) has been employed in numerous investigations aiming at combating neoplasms. We previously showed that copulsing of an antigen with a helper protein could considerably enhance antigen presenting capacity of ex vivo-generated DCs. In this study, we attempted to administer an effective treatment in a murine model of colon cancer with DCs pulsed with the mixture of a tumor-specific gp70-derived peptide (AH1) and a helper protein, ovalbumin (OVA).
MATERIALS AND METHODS
First, the presence of gp70 in CT26 tumor cells and tumor tissues was verified using immunofluorescence and Western blot analyses. Next, DCs were purified from normal mice, loaded ex vivowith AH1 and OVA (DC-Pep-OVA), and injected into tumor-bearing mice. Tumor volume, in vitro antigen (Ag)-specific proliferation of splenic cells, and survival rate were measured to determine the efficacy of DC-Pep-OVA. As the control groups, tumor-bearing mice were vaccinated with DC-Pep, unpulsed DC, and DCs loaded with a mixture of OVA and an irrelevant peptide (P15), or were not vaccinated at all.
RESULTS
DC-Pep-OVA showed superior efficacy over other groups, as indicated by smaller tumor volume, higher Ag-specific proliferation rate of splenic cells, and prolonged survival.
CONCLUSION
Overall, in the present study we showed for the first time that DCs copulsed with AH1 (tumor Ag) and OVA (helper molecule) could be considered as potentially robust weapons for use in future antitumor immunotherapies.

Keyword

Neoplasms; Dendritic cells; Vaccination; Helper protein; Immunotherapy

MeSH Terms

Animals
Blotting, Western
Colonic Neoplasms*
Dendritic Cells
Fluorescent Antibody Technique
Immunotherapy
Mice
Ovalbumin
Ovum
Survival Rate
Tumor Burden
Vaccination
Ovalbumin

Figure

• 

  Fig. 1. Verification of gp70 expression in CT26 cells and tumors. (A-D) Immunofluorescence staining. CT26 cells showed positive staining with rabbit-anti-mouse AH1 antibody (green cytoplasm). (A) The nuclei were stained using 7-AAD (red). (B) Murine 4T1 cell line served as the negative cell control. (C) CT26 cells were stained with peptide-adsorbed Ab. (D) CT26 cells were stained with rabbit IgG as the negative reagent control (A-D, ×40). (E) Western blot analysis of gp70 expression. 1, CT26 cell lysate; 2 and 5, CT26-derived tumor lysate; 3, murine 4T1 cell lysate (as the negative cell control); 4, normal murine muscle tissue lysate (as the negative tissue control); 6, CT26 cell lysate treated with rabbit IgG (as the negative reagent control); 7, CT26 cell-derived tumor lysate treated with rabbit IgG (as the negative reagent control). The 90 kDa band corresponds to gp90, the precursor of gp70.

• 

  Fig. 2. Effect of various DC-based vaccination protocols on tumor size. Mice were inoculated subcutaneously with CT26 tumor cells (day 0). On days 3 and 10, mice were injected with DCs loaded with different Ag mixtures, shown in the figure. The tumor volume in all vaccination groups was measured from day 0 at 3-day intervals for 42 days. DC, dendritic cell; Ag, antigen; OVA, ovalbumin. a)As of day 18, the average tumor volume in tumor-bearing mice vaccinated with either DC-Pep or DC-Pep-OVA was significantly smaller than in those treated with DCP15-OVA, with naïve DC (DC-alone), or not treated at all (tumor), b)From day 36 and beyond, a significant difference in terms of tumor size was observed between the DC-Pep and DC-Pep-OVA groups. Data are shown as the average tumor volume of five mice per group on each day.

• 

  Fig. 3. Proliferation of splenic cells obtained from tumor-bearing mice vaccinated with DCs. On days 3 and 10 following tumor induction, groups of five mice were vaccinated with DC pulsed with either tumor peptide or OVA or their mixture. Some mice were vaccinated with DC-P15-OVA, and DC-alone or not vaccinated at all (tumor). On day 42, mice were sacrificed and their splenic cells were stimulated in vitro with either tumor peptide (AH1) or OVA and the proliferation rate was measured using 3H-thymidine incorporation. As the negative control, splenic cells of all groups were treated with PBS in some wells. DC, dendritic cell; OVA, ovalbumin; PBS, phosphate buffered saline. Data are presented as mean±SD; **p ≤ 0.01.

• 

  Fig. 4. The therapeutic impact of DC-based vaccination on the survival rate of tumor-bearing mice. Groups of 10 mice were inoculated subcutaneously with CT26 tumor cells (day 0). On days 3 and 10, mice were injected with DCs loaded with different Ag mixtures, shown in the figure. The survival rate of all groups was determined weekly for 10 weeks and expressed as the percentage of live mice per treatment group. Significantly higher survival rate was observed in DC-Pep-OVA-treated mice compared to those vaccinated with DC-Pep. DC, dendritic cell; Ag, antigen; OVA, ovalbumin.

Reference

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