Anti-pancreatic cancer effect of HSP70-mutant K-ras peptide complex pulsed dendritic cells in vitro

Background: Mutant K-ras peptide cannot be well captured by dendritic cells (DC), thus the induced anti-tumor effect is limited. Aims: To investigate the anti-pancreatic cancer effect of heat shock protein 70 (HSP70)-mutant K-ras peptide complex pulsed DC in vitro. Methods: HSP70 was combined with mutant K-ras peptide, and then the formed complex was used to pulse the DC obtained by induction from human peripheral blood mononuclear cells in vitro. Levels of cytokines secreted by DC were determined by enzyme-linked immunosorbent assay (ELISA). The phenotypes of DC before and after pulsation were analyzed by flow cytometry. The proliferation of syngenic lymphocytes stimulated by pulsed DC, as well as the cytotoxicity of these activated lymphocytes on human pancreatic cancer cell lines Patu8988, PANC-1 and normal human hepatic cell line L-02 were assessed by cholecystokinin (CCK)-8 method. Results: DCs were successfully pulsed by HSP70-mutant K-ras peptide complex, the optimal dose was 0.75 μg/ml. Under this concentration, levels of interleukin (IL)-12 and tumor necrosis factor (TNF)-α secreted by DC, and expressions of CD80, CD83, CD86 and HLA-DR on surface of DC increased markedly. 0.75 μg HSP70-mutant K-ras peptide complex could pulse 1×106 DCs, inducing the proliferation of 1×107 syngenic lymphocytes, and killing specifically the Patu8988 cells, which had the same codon 12 mutant type (GGT→GTT), with a killing rate of 52.9%±5.1%; the killing effect on PANC-1 cells (mutant type GGT→GAT) and normal human hepatic cells was not obvious. Conclusions: DC pulsed with HSP70-mutant K-ras peptide complex has high capability of killing specific pancreatic cancer cells via the activation of syngenic lymphocytes.