Photodynamic therapy-generated cancer vaccine elicits acute phase and hormonal response in treated mice

Abstract

Photodynamic therapy (PDT)-generated cancer vaccines have shown promising results in preclinical studies and are being introduced in the clinics. Using an SCCVII mouse squamous cell carcinoma-based whole-cell autologous PDT vaccine model developed in our previous work, we have examined systemic effects in vaccinated mice that could be related to the induction of acute phase response. The upregulation of gene encoding serum amyloid P component (prototypic mouse acute phase reactant) was detected in the liver and to a lesser degree in the tumor of vaccinated mice at 24 h post-PDT vaccine treatment. A strong upregulation of gene for heat shock protein 70 was found in both the liver and tumor of mice at 4 h after their PDT vaccine treatment. Changes in the expression of genes for glucocorticoid-induced leucine zipper and serum- and glucocorticoid-regulated kinase 1 that are highly responsive to glucocorticoid modulation were uncovered in both the tumor and liver of vaccinated mice. A rise in the levels of serum corticosterone was detected in mice at 24 h after PDT vaccine treatment. The results indicate that a sudden appearance of a large number of PDT vaccine cells elicits host responses for securing their optimized clearance, which in addition to producing seminal acute phase reactants includes the engagement of glucocorticoid hormones. It is becoming increasingly clear that a consummate execution of this process of PDT vaccine cell removal is critical for tumor antigen recognition and the attainment of potent antitumor immune response.

Fig. 1

Expression of SAP gene in the tumor and liver of mice treated with PDT vaccine. Mice bearing SCCVII tumors were treated with either PDT vaccine or control vaccine using SCCVII cells exposed only to X-rays (60 Gy). The vaccine was prepared by incubating SCCVII cells with ce6 (0.5 μg/ml) for 30 min followed by exposure to 665 ± 10 nm light (1 J/cm²) and then 16-h post-incubation at 37°C. The cells were then collected, exposed to X-rays (60 Gy), and injected into SCCVII tumor-bearing mice (2 × 10⁷ cells per mouse peritumorally). The mice were killed at 24 h after vaccination and their tumor and liver tissues collected for quantitative RT-PCR-based analysis of the expression of SAP gene. The results are presented as GAPDH-normalized SAP gene expression relative to that in the same tissue of untreated tumor-bearing mice. Each treatment group consisted of 4 mice; bars represent standard deviations. *Statistically significant difference in response (P < 0.05) compared to the unvaccinated control group; **statistically significant difference in response (P < 0.05) compared to control vaccine group

Fig. 2

Expression of Hsp70 gene in the tumor and liver of mice treated with PDT vaccine. Mice bearing SCCVII tumors were treated with either PDT vaccine or control vaccine as described for Fig. 1. The mice were killed either 4 or 24 h after vaccination and their tumor and liver tissues collected for quantitative RT-PCR-based analysis of the expression of Hsp70 gene. The results are presented as GAPDH-normalized Hsp70 gene expression relative to that in the same tissue of untreated tumor-bearing mice. Each treatment group consisted of 4 mice; bars represent standard deviations. *Statistically significant difference in response (P < 0.05) compared to the unvaccinated control group; **statistically significant difference in response (P < 0.05) compared to control vaccine group

Fig. 3

Effect of inhibition of heat shock protein synthesis induction in PDT vaccine cells on therapy outcome. Mice bearing SCCVII tumors were treated with PDT vaccine or control vaccine as described for Fig. 1. The 16 h post-incubation in this protocol was carried out either in the presence or absence of KNK437 (25 μg/ml). The vaccine response was monitored by tumor size measurement and is shown as percentage of growth-inhibited tumors smaller than the means minus twofold SD of unvaccinated control group. Each treatment group consisted of 6 mice. *Statistically significant difference in response (P < 0.05) compared to the unvaccinated control group; **statistically significant difference in response (P < 0.05) compared to PDT vaccine alone group

Fig. 4

Expression of GILZ and SGK1 genes in the tumor and liver of mice treated with PDT vaccine. Mice bearing SCCVII tumors were treated with either PDT vaccine or control vaccine as described for Fig. 1. The mice were killed either 4 or 24 h after vaccination and their tumor and liver tissues collected for quantitative RT-PCR-based analysis of the expression of a GILZ gene and b SGK1 gene. The results are presented as GAPDH-normalized GILZ or SGK1 gene expression relative to that in the same tissue of untreated tumor-bearing mice. Each treatment group consisted of 4 mice; bars represent standard deviations. *Statistically significant difference in response (P < 0.05) compared to the unvaccinated control group; **statistically significant difference in response (P < 0.05) compared to control vaccine group

Fig. 5

Effect of glucocorticoid activity inhibition on the therapeutic effect of PDT vaccine. Mice bearing SCCVII tumors were treated with PDT vaccine as described for Fig. 1. At 30 min before vaccination, the mice received intraperitoneal injection of either a mifepristone (50 mg/kg) or b metyrapone (100 mg kg). Tumor responses to vaccine treatment are presented as percentage of growth-inhibited tumors (same as in Fig. 1). Each treatment group consisted of 6 mice. *Statistically significant difference in response (P < 0.05) compared to the unvaccinated control group; **Statistically significant difference in response (P < 0.05) compared to PDT vaccine alone group

Fig. 6

Serum corticosterone levels in mice treated by PDT vaccine. Mice bearing SCCVII tumors were treated with either PDT vaccine or control vaccine as described for Fig. 1. Their blood was collected at indicated post-vaccination time intervals for the ELISA-based determination of serum corticosterone. Each treatment group consisted of 4 mice; bars represent standard deviations. *Statistically significant difference in response (P < 0.05) compared to the unvaccinated control group; **statistically significant difference in response (P < 0.05) compared to PDT vaccine alone group