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Adjuvant properties of IFN-γ and GM-CSF in the scFv6.C4 DNA vaccine against CEA-expressing tumors


Tumor-associated carcinoembryonic antigen (CEA) is a natural target for vaccines against colorectal cancers. Our previous experience with a DNA vaccine with scFv6.C4, a CEA surrogate, showed a CEA-specific immune response with 40% of tumor-free mice after challenge with B16F10-CEA and 47% with MC38-CEA cells. These percentages increased to 63% after using FrC as an adjuvant. To further enhance the vaccine efficacy, we tested GM-CSF and IFNγ as adjuvants. C57BL/6J-CEA2682 mice were immunized 4 times with uP-PS/scFv6.C4, uP-PS/scFv6.C4 + uP-IFNγ, or uP-PS/scFv6.C4 + uP-GMCSF. After one week, the mice were challenged with MC38-CEA, and tumor growth was monitored over 100 days. Immunization with scFv6.C4 and scFv6.C4 + GM-CSF resulted in a gradual increase in the anti-CEA antibody titer, while scFv6.C4 + IFNγ immunization led to a rapid and sustained increase in the titer. The addition of IFNγ also induced higher CD4 + and CD8 + responses. When challenged, almost 80% of the scFv6.C4 + IFNγ-vaccinated mice did not develop tumors, while the others had a significant tumor growth delay. The probability of being tumor-free was 2700% higher using scFv6.C4 + IFNγ than scFv6.C4. The addition of GM-CSF had no additional effect on tumor protection. DNA immunization with scFv6.C4 + IFNγ, but not GM-CSF, increased the antitumor effect via readily sustained specific humoral and cytotoxic responses to CEA.

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Fig. 1: Humoral response and protection against MC38CEA tumor challenge by scFv6.C4 + IFNγ and scFv6.C4 + GM-CSF DNA immunization.
Fig. 2: Evaluation of the humoral response by immunocytochemistry using sera from the scFv6.C4 + IFNγ- or scFv6.C4 + GM-CSF-immunized mice.
Fig. 3: Cellular response of the scFv6.C4 + IFNγ-immunized animals determined by proliferation assay.
Fig. 4: CTL activity in CD8 + cells from the scFv6.C4 + INFγ- immunized animals by LDH activity assay.


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This work was supported by the São Paulo Research Foundation (FAPESP; Grant Numbers: 2012/21861-1 and # 2013/17224-9). BFZ was a recipient of a FAPESP scholarship (2012/21861-1).

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BFZ: conception and design of the study, acquisition of data, analysis and interpretation of the data, drafting the article; CPF: acquisition of data, analysis, and interpretation of data; JRCV: analysis and interpretation of the data; SWH: conception and design of the study, drafting the article, revising it critically for important intellectual content, and final approval of the version to be submitted.

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Correspondence to Sang Won Han.

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Zanetti, B.F., Ferreira, C.P., Vasconcelos, J.R.C. et al. Adjuvant properties of IFN-γ and GM-CSF in the scFv6.C4 DNA vaccine against CEA-expressing tumors. Gene Ther (2021).

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