A Novel Method for Producing Custom-Made Idiotype Vaccines for Lymphoma Immunotherapy Using a Cell-Free Expression System.

Idiotype vaccines have been used as a customized immunotherapy for lymphoma. This strategy takes advantage of a unique antigen (either immunoglobulin or T cell receptor) found on the lymphoma cells of each patient. To make this personalized vaccine feasible for widespread use a production method, which is rapid and relatively inexpensive will be required. In an E. coli-based cell-free protein expression system we were able to make ScFv proteins derived from the immunoglobulins of a mouse B cell lymphoma, 38C13, in high yields. We have used this cell-free system to make 38C13 ScFv proteins fused to either the cytokine mGM-CSF or a 9 amino acid peptide derived from hIL-1b. The 9 amino acid fragment of hIL-1b was chosen because it is known to retain immunostimulatory function while lacking pyrogenicity. We were able consistently to produce these proteins in high yield and to purify them using a histadine tag at the terminus of the molecule. The ScFv portion of the molecule is folded properly, as assayed by its binding to an anti-idiotype Ab. Similarly, the mGM-CSF fused to the ScFv can be recognized by an anti-mGM-CSF Ab. The mGM-CSF portion is biologically active as determined by its ability to support the growth a mGM-CSF dependant cell line. We immunized C3H mice with either the ScFc-hIL-1b or the ScFv-mGM-CSF fusion protein and we were able to induce an idiotype-specific Ab response, which recognizes the native idiotype protein in ELISA assays as well as the 38C13 tumor as detected by flow cytometry. Most importantly, in the highly aggressive 38C13 lymphoma model system, three bi-weekly vaccinations with either fusion protein protected mice from tumor challenge with the syngeneic 38C13 tumor. These data establish that a cell-free protein expression system can work for producing individualized idiotype vaccines for lymphoma. (This work was supported by NIH grant CA34233)