Enhancement of therapeutic DNA vaccine potency by melatonin through inhibiting VEGF expression and induction of antitumor immunity mediated by CD8+ T cells
To be effective, therapeutic cancer vaccines should stimulate both an effective cell-mediated and a robust cytotoxic CD8+ T-cell response against human papillomavirus (HPV)-infected cells to treat the pre-existing tumors and prevent potential future tumors. In this study, the therapeutic experiments were designed in order to evaluate antitumor effect against the syngeneic TC-1 tumor model. The anti-tumor efficacy of a HPV-16 E7 DNA vaccine adjuvanted with melatonin (MLT) was evaluated in a C57BL/6 mouse tumor model by measuring tumor growth post vaccination and the survival rate of tumor-bearing mice, analyzing the specific lymphocyte proliferation responses in control and vaccinated mice by MTT assay. The E7-specific cytotoxic T cells (CTL) were analyzed by lymphocyte proliferation and lactate dehydrogenates (LDH) release assays. IFN-γ, IL-4 and TNF-α secretion in splenocyte cultures as well as vascular endothelial growth factor (VEGF) and IL-10 in the tumor microenvironment were assayed by ELISA. Our results demonstrated that subcutaneous administration of C57BL/6 mice with a DNA vaccine adjuvanted with MLT dose-dependently and significantly induced strong HPV16 E7-specific CD8+ cytotoxicity and IFN-γ and TNF-α responses capable of reducing HPV-16 E7-expressing tumor volume. A significantly higher level of E7-specific T-cell proliferation was also found in the adjuvanted vaccine group. Furthermore, tumor growth was significantly inhibited when the DNA vaccine was combined with MLT and the survival time of TC-1 tumor bearing mice was also significantly prolonged. In vivo studies further demonstrated that MLT decreased the accumulation of IL-10 and VEGF in the tumor microenvironment of vaccinated mice. These data indicate that melatonin as an adjuvant augmented the cancer vaccine efficiency against HPV-associated tumors in a dose dependent manner.
Human papilloma virus
Cytolytic T lymphocyte
- IFN- γ
Tumor necrosis factor
Polyvinylidene difluoride membranes
Cytotoxic T cells
3[4,5-Dimethylthiazol-2-ll]-2,5-diphenyltetrazolium bromide, thiazolyl-blue
Fetal bovine serum
1640 Roswell Park Memorial Institute (name of the medium)
Food and Drug Administration
T helper, vascular endothelial growth factor (VEGF)
Enzyme-linked immunosorbent assay
The authors would like to acknowledge Institute Pasteur of Iran and Golestan Medical University for the financial support. This project was extracted from a MSC thesis.
Compliance with ethical standards
This study was supported by Institute Pasteur of Iran and Research Deputy at Golestan Medical University through the Grant project number (35/21332).
Conflict of interest
The authors declare that they have no conflict of interest.
The animal protocol used in this study was approved by the local animal ethics council of Golestan Ethic Committee of Golestan University of Medical Sciences (ethics number: et- 35/21332). All experimental procedures involving mice were performed in accordance with the national experimental guidelines.
- 1.Anisimov VN, Egormin PA, Piskunova TS, Popovich IG, Tyndyk ML, Yurova MN, Zabezhinski MA, Anikin IV, Karkach AS, Romanyukha AA (2010) Metformin extends life span of HER-2/neu transgenic mice and in combination with melatonin inhibits growth of transplantable tumors in vivo. Cell Cycle 9:188–197CrossRefPubMedGoogle Scholar
- 6.Bush SH, Lacaze-Masmonteil N, McNamara-Kilian MT, MacDonald AR, Tierney S, Momoli F, Agar M, Currow DC, Lawlor PG (2016) The preventative role of exogenous melatonin administration to patients with advanced cancer who are at risk of delirium: study protocol for a randomized controlled trial. Trials 17:399CrossRefPubMedPubMedCentralGoogle Scholar
- 28.Moeini S, Saeidi M, Fotouhi F, Mondanizadeh M, Shirian S, Mohebi A, Gorji A, Ghaemi A (2017) Synergistic effect of programmed cell death protein 1 blockade and secondary lymphoid tissue chemokine in the induction of anti-tumor immunity by a therapeutic cancer vaccine. Arch Virol 162:333–346CrossRefPubMedGoogle Scholar
- 32.Rondanelli M, Faliva MA, Perna S, Antoniello N (2013) Update on the role of melatonin in the prevention of cancer tumorigenesis and in the management of cancer correlates, such as sleep-wake and mood disturbances: review and remarks. Aging Clin Exp Res 25:499–510CrossRefPubMedPubMedCentralGoogle Scholar