Process for the production of a DNA vaccine for cancer immunotherapy

A method of treating a human patient against cancer comprising administering to the patient a DNA vaccine produced by a method comprising at least the following steps:

• a) transforming an attenuated strain of Salmonella with at least one DNA molecule comprising at least one eukaryotic expression cassette encoding at least one antigen comprising a neoantigen, wherein the neoantigen is a tumor-specific antigen that arises as a consequence of a tumor-specific mutation;
• b) characterizing at least one transformed cell clone obtained in step (a);
• c) selecting at least one of the transformed cell clone(s) characterized in step (b) and further characterizing said at least one selected transformed cell clone;
• wherein step (b) comprises at least one of the following substeps:
• bi) assessing the cell growth of at least one transformed cell clone obtained in step (a) over time;
• bii) assessing the stability of the at least one DNA molecule comprising at least one expression cassette encoding at least one antigen in the at least one transformed cell clone obtained in step (a);
• biii) isolating the at least one DNA molecule comprising at least one expression cassette encoding at least one antigen from at least one transformed cell clone obtained in step (a) and characterizing the at least one isolated DNA molecule by restriction analysis and/or sequencing;
• biv) isolating the at least one DNA molecule comprising at least one expression cassette encoding at least one from at least one transformed cell clone obtained in step (a), transfecting the at least one isolated DNA molecule into at least one eukaryotic cell and assessing the expression of the at least one antigen in said at least one eukaryotic cell;
• wherein step (c) comprises at least one of the following substeps:
• ci) assessing the number of viable cells per ml cell suspension of the at least one transformed cell clone selected in step (c);
• cii) assessing the stability of the at least one DNA molecule comprising at least one expression cassette encoding at least one antigen in the at least one transformed cell clone selected in step (c);
• ciii) isolating the at least one DNA molecule comprising at least one expression cassette encoding at least one antigen from the at least one transformed cell clone selected in step (c) and characterizing the at least one isolated DNA molecule by restriction analysis and/or sequencing;
• civ) isolating the at least one DNA molecule comprising at least one expression cassette encoding at least one antigen from the at least one transformed cell clone selected in step (c), transfecting the at least one isolated DNA molecule into at least one eukaryotic cell and assessing the expression of the at least one antigen in said at least one eukaryotic cell;
• cv) testing for the presence of bacterial, fungal and/or viral contaminants in at the least one transformed cell clone selected in step (c);
• cvi) verifying the bacterial strain identity of the at least one transformed cell clone selected in step (c);
• and
• wherein the attenuated strain of Salmonella is Salmonella typhi Ty21a.