of the individual’s tumor—it would also be possible to vaccinate with higher doses.
Antigen-Specific Tumor Vaccines. Researchers also hope to use activated T-cells to identify the relevant antigens for antigen-specific vaccines. One group has been pursuing this concept in the mouse model using CT26, an NMU-induced colon cancer, in order to identify the repertoire of antigens that are being recognized by the CD8 arm of the immune response following vaccination with GM-CSF-transduced tumor cells. Their technique involves taking bulk T-cells from the draining lymph nodes (rather than tumor-specific CTLs), eluting peptides from Class I molecules, fractionating them with reverse-phase HPLC, and using surrogate targets to assay peptide fractions for bioactivity. The results indicated that there was only one bioactive fraction, suggesting that the majority of CD8-positive immune response was focused on a single peptide among the many presented by the CT26 tumor. These results have been repeated in 40 separate experiments.
This peptide, called AH1, had a molecular weight of 1,128 and was doubly charged. It sensitized surrogate target cells down to a concentration of 5 X 10-12 molar. Upon sequencing, it proved to be a peptide derived from an endogenous murine MuLV gene that is normally completely silent in the BALB/c genome but is reactivated by altered methylation, much like MAGE-1 (see above). (The latter finding has led to new interest in endogenous human retroviruses, which also seem to be reactivated in human tumors.)
AH1 is not expressed in normal tissues from BALB/c mice, including normal colon and small intestine epithelium, but it is turned on in a number of different tumors. Interestingly, if T-cells from vaccinated animals are stimulated for two rounds with AH1 plus IL-2 and then adoptively transferred back into animals with CT26 tumors, most of the animals are cured. However, there is no significant response when tumor-bearing animals are vaccinated with AH1-pulsed dendritic cells or other approaches.
Researchers believe that they should examine several other approaches for introducing these gene products in vivo, such as viruses that target them to both the Class I and Class III MHC pathways. In addition, there are some interesting recombinant viral and bacterial approaches that should be compared head-to-head with endogenous tumor antigen models before deciding which approaches will be taken to clinical trials, with their tremendous investment of time, effort, and expense.
In response to questions from the audience, Dr. Pardoll added the following:
Although GM-CSF up-regulates B7 in macrophages, it does not do so in tumor cells and, hence, this cannot explain their increased immunogenicity. In fact, the major effect of transducing B7 into tumors is to provide a target molecule for NK cells to more actively lyse the tumor cells. A B7-transduced tumor cell isn’t nearly as good an APC as a bone marrow progenitor that differentiates into a dendritic cell in the presence of GM-CSF.