Investigating the effect of cGRP78 vaccine against different cancer cells and its role in reducing melanoma metastasis

Abstract

Background and purpose: Treatment of malignancies with chemotherapy and surgery is often associated with disease recurrence and metastasis. Immunotherapy improves cancer treatment by creating an active response against tumor antigens. Various cancer cells express a large amount of glucose-regulated protein 78 (GRP78) protein on their surface. Stimulating the immune system against this antigen can expose cancer cells to the immune system. Herein, we investigated the effectiveness of a cGRP78-based vaccine against different cancer cells.

Experimental approach: BALB/c mice were immunized with the cGRP78. The humoral immune response against different cancer cells was assessed by Cell-ELISA. The cellular immunity response was determined by splenocyte proliferation assay with different cancer antigens. The effect of vaccination on metastasis was investigated in vaccinated mice by injecting melanoma cancer cells into the tail of mice.

Findings/results: These results indicated that the cGRP78 has acceptable antigenicity and stimulates the immune system to produce antibodies. After three injections, the amount of produced antibody was significantly different from the control group. Compared to the other three cell types, Hela and HepG2 showed the highest reaction to the serum of vaccinated mice. Cellular immunity against the B16F10 cell line had the best results compared to other cells. The metastasis results showed that after 30 days, the growth of B16F10 melanoma cancer cells was not noticeable in the lung tissue of vaccinated mice.

Conclusion and implications: Considering the resistance of vaccinated mice to metastasis, this vaccine offers a promising prospect for cancer treatment by inhibiting the spread of cancer cells.

Keywords: Immunotherapy.; Melanoma; Metastasis; cGRP78 vaccine.

Fig. 1.

The SDS-PAGE and western blotting of cGRP78 protein. (A) Expression and purification of cGRP78 recombinant protein. Line 1: soluble fraction from protein expression in E. coli before purification; lines 2 and 3: recombinant protein cGRP78 after purification by Ni-NTA; line 4: PageRuler™ plus Prestained Protein Ladder. (B) Western blotting of cGRP78. Line 1: purified protein; line 2: Prestained Protein Ladder. SDS-PAGE, Sodium dodecyl-sulfate polyacrylamide gel electrophoresis; GRP, glucose-regulated protein 78; Ni-NTA, nickel-nitrilotriacetic acid.

Fig. 2.

Anti-cGRP78 IgG titer after mouse immunization. OD represents the amount of produced antibody. ^**P < 0.01 and ^***P < 0.001 indicate significant differences in comparison with the control group. GRP, glucose-regulated protein 78; OD, optical density.

Fig. 3.

Humoral immune status of immune mice against different types of cancer cells. Two human cancer cells (A549 and Hela) and three mouse cancer cells (4T1, B16F10, and HepG2) were used to examine the humoral immunity in vaccinated mice. Hela and HepG2 cells reacted the most with the serum of vaccinated mice. ^*P < 0.05 and ^**P < 0.01 indicate significant differences compared to the respective control group. OD, Optical density.

Fig. 4.

Cellular immune status of vaccinated mice against different types of cancer cells in the “vaccinated + antigen group” (vaccinated mouse splenocyte along with cancer antigens) and “vaccinated -antigen group” (vaccinated mouse splenocyte in the absence of cancer antigens). The B16F10 cells' antigens more than other cancer cell antigens increased the proliferation of splenocytes in vaccinated mice. ^*P < 0.05, ^**P < 0.01, ^***P < 0.001 indicates significant differences compared to the respective control and ^###P < 0.001 indicates the differences between vaccinated groups. OD, Optical density.

Fig. 5.

Investigating lung metastasis of melanoma cells in vaccinated mice. The black nodules in the lungs of control group mice indicate the metastasis of cancer cells in the lungs.