Limits...
An effective cytokine adjuvant vaccine induces autologous T-cell response against colon cancer in an animal model

View Article: PubMed Central - PubMed

ABSTRACT

Background: Despite recent advances in early detection and improvements in chemotherapy for colon cancer, the patients still face poor prognosis of postoperative recurrence and metastasis, the median survival for patients with metastatic colorectal cancer is approximately 22–24 months. Some immunotherapeutic approaches had been attempted in colon cancer patients to significantly increase overall survival. A vaccine based approach has shown a novel direction for colon cancer prevention and therapy.

Methods: In this study, the experiments were designed including prevention and therapeutic stages in order to attain effect against tumor recurrence in clinical settings. The anti-tumor efficacy of a novel cytokine adjuvant vaccine that contained cytokines GM-CSF and IL-2 and inactivated colon CT26.WT whole cell antigen was evaluated in BALB/c mouse tumor models by measuring tumor growth post vaccination and the survival time of tumor-bearing mice, analyzing the expression and distribution of CD4, CD8, CD11c, CD80, CD86 and CD83 positive cells in control and treated mice by flow cytometry and immunochemistry. The tumor-specific cytotoxic T cells (CTL) were analyzed by tumor proliferation and the lactic dehydrogenates (LDH) release assays. IFN-γ, IL-2 and GM-CSF secretion in serum was assayed by ELISA.

Results: Our results suggested that cytokine adjuvant vaccine significantly inhibited tumor growth and extended the survival period at least 160d. It was found that the levels of CD8 + T and the tumor-specific cytotoxicity were significantly higher in prevention and treatment group vaccinated by cytokine adjuvant vaccine. CD8 + T cells play a key role in anti-tumor response.

Conclusions: The novel GM-CSF and IL-2 based adjuvant vaccine effectively activated autologous T-cell response and represented a promising immunotherapeutic approach for patients with colon cancer.

Electronic supplementary material: The online version of this article (doi:10.1186/s12865-016-0172-x) contains supplementary material, which is available to authorized users.

No MeSH data available.


Related in: MedlinePlus

Standardization of the tumor model and its tentative mode of action The animals were vaccinated for two times before injected tumor cells during pre-experimental stage. Panel (a), shows the tumor growth after day 0, 10 and 30, as a result of subcutaneous injection of CT26.WT tumor cells into the neck of BALB/c mice. Panel (b), depicts the efficacy of cytokine adjuvant vaccine, where each group with 5 mice was vaccinated twice at interval of 7 and 3 days, before subcutaneous injection of CT26.WT cells. Panel (c), shows the comparison between male and female mice in terms of tumor uniformity. Panel (d), represents the standardization of inactivated antigen for effective anti-tumor response. M stand for million, * stand for Optimization. Panel (e), represents the effect of combination application compared to single group by observing survival period, which each group with 15 mice was vaccinated twice before subcutaneous injection of CT26.WT cells. *denotes p < 0.05, ***denotes p < 0.001. Panel (f), The experiment timeline showing the times of vaccinated. In order to establishing the tumor recurrence model stimulating the clinical status, the experiment was divided into prevention stage and therapeutic stage. The Prevention group was vaccinated vaccine for two times in prevention stage, the Treatment group was vaccinated vaccine for four times include prevention stage and therapeutic stage, and the Tumor group vaccinated inactivated antigen for control. All groups were inoculated tumor cells at the same time. Tumor inoculation was the initial point (day 0)
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC5037582&req=5

Fig1: Standardization of the tumor model and its tentative mode of action The animals were vaccinated for two times before injected tumor cells during pre-experimental stage. Panel (a), shows the tumor growth after day 0, 10 and 30, as a result of subcutaneous injection of CT26.WT tumor cells into the neck of BALB/c mice. Panel (b), depicts the efficacy of cytokine adjuvant vaccine, where each group with 5 mice was vaccinated twice at interval of 7 and 3 days, before subcutaneous injection of CT26.WT cells. Panel (c), shows the comparison between male and female mice in terms of tumor uniformity. Panel (d), represents the standardization of inactivated antigen for effective anti-tumor response. M stand for million, * stand for Optimization. Panel (e), represents the effect of combination application compared to single group by observing survival period, which each group with 15 mice was vaccinated twice before subcutaneous injection of CT26.WT cells. *denotes p < 0.05, ***denotes p < 0.001. Panel (f), The experiment timeline showing the times of vaccinated. In order to establishing the tumor recurrence model stimulating the clinical status, the experiment was divided into prevention stage and therapeutic stage. The Prevention group was vaccinated vaccine for two times in prevention stage, the Treatment group was vaccinated vaccine for four times include prevention stage and therapeutic stage, and the Tumor group vaccinated inactivated antigen for control. All groups were inoculated tumor cells at the same time. Tumor inoculation was the initial point (day 0)

Mentions: The schematic vaccine preparation and subsequent functional mechanisms for the induction of autologous T-cell anti-tumor response in a mouse colon cancer model is depicted in Additional file 1: Figure S1. Before analyzing treatment efficacy of the cytokine adjuvant vaccine, extensive standardization of different parameters to develop an ideal tumor model was applied, including the assessment of tumor formation time and volumes (Fig. 1a), specific anti-tumor effect of the vaccine (Fig. 1b), comparisons of tumor sizes between male and female mice (Fig. 1c), and optimized doses of inactivated tumor whole cell antigen in the vaccine (Fig. 1d). The results revealed that, there was no significant effect on the use of adjuvant or inactivated antigen alone, inactivated antigen obtained from 1 × 106 tumor cells in the vaccine had the most significant effect on inhibition of tumor development in the female mice that subcutaneously injected with 5 × 105 tumor cells. The tumor mass was generally established within 7–10 d post injection of the tumor cells. Based on these optimization steps, we compared the preliminary results of cytokine adjuvant vaccines between GM-CSF group, IL-2 group and GM-CSF + IL-2 group by observing the survival period. The results revealed that combined application extended the survival period and significantly inhibited the tumor growth compared to single group (Fig. 1e). Then we further examined treatment efficacy and mechanism of the cytokine adjuvant vaccination in a BALB/c mice colon cancer model. We divided the experimental procedures into the preventive and therapeutic stages, which may further strengthen anti-tumor effect during the course of tumor recurrence in clinical settings (Fig. 1f).Fig. 1


An effective cytokine adjuvant vaccine induces autologous T-cell response against colon cancer in an animal model
Standardization of the tumor model and its tentative mode of action The animals were vaccinated for two times before injected tumor cells during pre-experimental stage. Panel (a), shows the tumor growth after day 0, 10 and 30, as a result of subcutaneous injection of CT26.WT tumor cells into the neck of BALB/c mice. Panel (b), depicts the efficacy of cytokine adjuvant vaccine, where each group with 5 mice was vaccinated twice at interval of 7 and 3 days, before subcutaneous injection of CT26.WT cells. Panel (c), shows the comparison between male and female mice in terms of tumor uniformity. Panel (d), represents the standardization of inactivated antigen for effective anti-tumor response. M stand for million, * stand for Optimization. Panel (e), represents the effect of combination application compared to single group by observing survival period, which each group with 15 mice was vaccinated twice before subcutaneous injection of CT26.WT cells. *denotes p < 0.05, ***denotes p < 0.001. Panel (f), The experiment timeline showing the times of vaccinated. In order to establishing the tumor recurrence model stimulating the clinical status, the experiment was divided into prevention stage and therapeutic stage. The Prevention group was vaccinated vaccine for two times in prevention stage, the Treatment group was vaccinated vaccine for four times include prevention stage and therapeutic stage, and the Tumor group vaccinated inactivated antigen for control. All groups were inoculated tumor cells at the same time. Tumor inoculation was the initial point (day 0)
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC5037582&req=5

Fig1: Standardization of the tumor model and its tentative mode of action The animals were vaccinated for two times before injected tumor cells during pre-experimental stage. Panel (a), shows the tumor growth after day 0, 10 and 30, as a result of subcutaneous injection of CT26.WT tumor cells into the neck of BALB/c mice. Panel (b), depicts the efficacy of cytokine adjuvant vaccine, where each group with 5 mice was vaccinated twice at interval of 7 and 3 days, before subcutaneous injection of CT26.WT cells. Panel (c), shows the comparison between male and female mice in terms of tumor uniformity. Panel (d), represents the standardization of inactivated antigen for effective anti-tumor response. M stand for million, * stand for Optimization. Panel (e), represents the effect of combination application compared to single group by observing survival period, which each group with 15 mice was vaccinated twice before subcutaneous injection of CT26.WT cells. *denotes p < 0.05, ***denotes p < 0.001. Panel (f), The experiment timeline showing the times of vaccinated. In order to establishing the tumor recurrence model stimulating the clinical status, the experiment was divided into prevention stage and therapeutic stage. The Prevention group was vaccinated vaccine for two times in prevention stage, the Treatment group was vaccinated vaccine for four times include prevention stage and therapeutic stage, and the Tumor group vaccinated inactivated antigen for control. All groups were inoculated tumor cells at the same time. Tumor inoculation was the initial point (day 0)
Mentions: The schematic vaccine preparation and subsequent functional mechanisms for the induction of autologous T-cell anti-tumor response in a mouse colon cancer model is depicted in Additional file 1: Figure S1. Before analyzing treatment efficacy of the cytokine adjuvant vaccine, extensive standardization of different parameters to develop an ideal tumor model was applied, including the assessment of tumor formation time and volumes (Fig. 1a), specific anti-tumor effect of the vaccine (Fig. 1b), comparisons of tumor sizes between male and female mice (Fig. 1c), and optimized doses of inactivated tumor whole cell antigen in the vaccine (Fig. 1d). The results revealed that, there was no significant effect on the use of adjuvant or inactivated antigen alone, inactivated antigen obtained from 1 × 106 tumor cells in the vaccine had the most significant effect on inhibition of tumor development in the female mice that subcutaneously injected with 5 × 105 tumor cells. The tumor mass was generally established within 7–10 d post injection of the tumor cells. Based on these optimization steps, we compared the preliminary results of cytokine adjuvant vaccines between GM-CSF group, IL-2 group and GM-CSF + IL-2 group by observing the survival period. The results revealed that combined application extended the survival period and significantly inhibited the tumor growth compared to single group (Fig. 1e). Then we further examined treatment efficacy and mechanism of the cytokine adjuvant vaccination in a BALB/c mice colon cancer model. We divided the experimental procedures into the preventive and therapeutic stages, which may further strengthen anti-tumor effect during the course of tumor recurrence in clinical settings (Fig. 1f).Fig. 1

View Article: PubMed Central - PubMed

ABSTRACT

Background: Despite recent advances in early detection and improvements in chemotherapy for colon cancer, the patients still face poor prognosis of postoperative recurrence and metastasis, the median survival for patients with metastatic colorectal cancer is approximately 22&ndash;24 months. Some immunotherapeutic approaches had been attempted in colon cancer patients to significantly increase overall survival. A vaccine based approach has shown a novel direction for colon cancer prevention and therapy.

Methods: In this study, the experiments were designed including prevention and therapeutic stages in order to attain effect against tumor recurrence in clinical settings. The anti-tumor efficacy of a novel cytokine adjuvant vaccine that contained cytokines GM-CSF and IL-2 and inactivated colon CT26.WT whole cell antigen was evaluated in BALB/c mouse tumor models by measuring tumor growth post vaccination and the survival time of tumor-bearing mice, analyzing the expression and distribution of CD4, CD8, CD11c, CD80, CD86 and CD83 positive cells in control and treated mice by flow cytometry and immunochemistry. The tumor-specific cytotoxic T cells (CTL) were analyzed by tumor proliferation and the lactic dehydrogenates (LDH) release assays. IFN-&gamma;, IL-2 and GM-CSF secretion in serum was assayed by ELISA.

Results: Our results suggested that cytokine adjuvant vaccine significantly inhibited tumor growth and extended the survival period at least 160d. It was found that the levels of CD8&thinsp;+&thinsp;T and the tumor-specific cytotoxicity were significantly higher in prevention and treatment group vaccinated by cytokine adjuvant vaccine. CD8&thinsp;+&thinsp;T cells play a key role in anti-tumor response.

Conclusions: The novel GM-CSF and IL-2 based adjuvant vaccine effectively activated autologous T-cell response and represented a promising immunotherapeutic approach for patients with colon cancer.

Electronic supplementary material: The online version of this article (doi:10.1186/s12865-016-0172-x) contains supplementary material, which is available to authorized users.

No MeSH data available.


Related in: MedlinePlus