Basic research of the OK-432-conjugated tumor vaccine

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(1)Preparation of the OK-432-conjugated tumor vaccinesOK-432 was mixed with tumor cells (B16 melanoma cells or Sq1979 cells), and conjugated with the addition of GA to increase the antigenicity of tumor cells. To examine the safety of the OK-432-conjugated tumor vaccines, a trypan blue staining was performed. Over 80 % of the conjugated vaccines were stained with trypan blue, indicating that the majority of OK-432-conjugated tumor vaccines were not viable. When tumor cells treated with various concentrations of GA (0.2-0.0002 %) were immunized, conjugation under condition of high concentrations of over 0.02 % GA inhibited strongly viability of B16 melanoma cells, so that tumor was not observed up to 180 days. The surface of OK-432-conjugated tumor vaccines was strongly stained with FITC emission of anti-OK-432 antibodies, while untreated tumor cells were not stained. Three immunizations with the B16-vaccines showed the strongest suppression of the incidence of B16 melanoma and associat … More ed mortality. The only immunizations with the Sq1979-vaccines induce anti-Sq1979 specific effects, and the B16-vaccines elicit no cross-reactive antitumor effects.(2) Preparation of KLN-205 tongue cancer modelTo analyze the mechanisms of antitumor effects by the OK-432-conjugated tumor vaccines I developed the experimental murine tongue cancer model. Two kinds of squamous cell carcinoma (KLN-205 and Sq1979 cells) were examined. In the DBA/2 mice tongue cancer model, the oncogenetic rate was 100 % in the groups of 2×10^5 and 5×10^5 cells of KLN-205. On the other hand, the oncogenetic rate of the 5×10^5 cell group was 100 % in the Sq-1979 cell model, but tumor regression was observed. Therefore, inoculation of 2×10^5 cells / 40 μl KLN-205 cells was selected as an appropriate experimental tongue cancer model.(3) Analysis of antitumor effects induced by the KLN205-vaccinesTo analyze the changes of the immunized mice after inoculation of KLN-205 cells, I carried out histological examination. Infiltrating lymphocytes were markedly observed around the tumor cells with histological section at 24h after tumor inoculation in the KLN205-vaccine group. On the other hand, scattering lymphocytes were shown and no infiltrating lymphocytes could be observed in the control group. These results indicate that immunized mice obtained immunological memory for KLN-205 cells.(4) The KLN205-vaccines elicit cytolytic activityIn the cytolytic assay, splenocytes from mice immunized with the KLN205-vaccines indicate a greater cytolytic response than that from mice immunized with the others. But KLN-205 treated with GA, OK-432 treated with GA and OK-432 alone showed almost the same level of precent specific lysis as control. These findings suggest the conjugation of both (tumor cells and OK-432) is of importance concerning cancer immunotherapy using a cell-based vaccine. Less