Can dendritic cells improve whole cancer cell vaccines based on immunogenically killed cancer cells?

Abstract

Immunogenic cell death (ICD) offers interesting opportunities in cancer cell (CC) vaccine manufacture, as it increases the immunogenicity of the dead CC. Furthermore, fusion of CCs with dendritic cells (DCs) is considered a superior method for generating whole CC vaccines. Therefore, in this work, we determined in naive mice whether immunogenically killed CCs per se (CC vaccine) elicit an antitumoral immune response different from the response observed when immunogenically killed CCs are associated with DCs through fusion (fusion vaccine) or through co-incubation (co-incubation vaccine). After tumor inoculation, the type of immune response in the prophylactically vaccinated mice differed between the groups. In more detail, fusion vaccines elicited a humoral anticancer response, whereas the co-incubation and CC vaccine mainly induced a cellular response. Despite these differences, all three approaches offered a prophylactic protection against tumor development in the murine mammary carcinoma model. In summary, it can be concluded that whole CC vaccines based on immunogenically killed CCs may not necessarily require association with DCs to elicit a protective anticancer immune response. If this finding can be endorsed in other cancer models, the manufacture of CC vaccines would greatly benefit from this new insight, as production of DC-based vaccines is laborious, time-consuming and expensive.

Keywords: co-incubation; fusion; immunogenic cell death; whole cancer cell vaccine.

Figure 1.

Formation and characterization of CC-DC hybrids. (A) Phagocytosis of MTX-treated (dashed line) EO771 cells and untreated (solid line) EO771 cells by DCs during 12 h of co-incubation (n = 4, error bars ± 1 SD). (B) Hybrid formation between MTX-treated EO771 cells fused to DCs (dashed line) or co-incubated with DCs (solid line) (n = 4, error bars ±1 SD). (C) Influence of fusion or co-incubation of EO771 cells with DCs on functional DC maturation (IL-12 production) (n = 3, error bars ± 1 SD). (D) Influence of fusion or co-incubation of EO771 cells with DCs on phenotypical DC maturation (CD86 surface expression) (n = 3, error bars ±1 SD). *, P < 0.05 between groups.

Figure 2.

In vivo experimental setup. All mice received on days 0 and 35 in the flanks (left and right flank, respectively) a subcutaneous injection of phosphate buffered saline (control group) or 1 of the 3 vaccines (cancer cell vaccine, co-incubation vaccine or fusion vaccine). Three mice in each group were euthanized on day 40 and spleen and serum were collected to characterize the cellular and humoral response (tumor-specific cytotoxicity, activation of Th1 or Th2 effector cells, production of tumor-specific antibodies). Tumor inoculation was performed on day 42. Eight mice in each group were euthanized on day 142 (or earlier if the tumor would reach 1 cm²) and spleen and serum were collected for the characterization of the memory cellular and humoral response (tumor-specific cytotoxicity, activation of Th1 or Th2 memory cells, production of tumor-specific antibodies).

Figure 3.

In vivo evaluation of vaccines based on immunogenically killed EO771 cells. (A) Activation of Th2 effector cells (IL-4⁺ CD4⁺ and CD8⁺ splenocytes) 5 d after the second vaccination (n = 3, ± 1 SD). (B) Activation of Th1 and CTL effector cells (IFNγ⁺ CD4⁺ and CD8⁺ splenocytes) 102 d after the second vaccination (n = 8, ± 1 SD). (C) The cellular response for tumor-specific cytotoxicity (in vitro cancer cell killing capacity of isolated splenocytes). In this cytotoxicity assay, the percentage of live EO771 cells inversely correlates with the efficacy of the vaccine/group (Pre n = 3, Post n = 8, ± 1 SD). (D) The humoral response (presence of antibodies against EO771 cells) (Pre n = 3, Post n = 8, ± 1 SD). Abbreviations: CCV, cancer cell vaccine group; CIV, co-incubation vaccine group; Control, control group; FV, fusion vaccine group; Post, 100 d post tumor inoculation or 102 d after the second vaccination; Pre, 2 d prior to tumor inoculation or 5 d after the second vaccination. *, P < 0.05 between groups.

Figure 4.

Kaplan–Meier survival curve after tumor challenge of prophylactically vaccinated mice (n= 8/group). Abbreviations: CCV, cancer cell vaccine group; CIV, co-incubation vaccine group; Control, control group; FV, fusion vaccine group.