The HER2/neu protooncogene is overexpressed in approximately 30 percent of human breast cancers. Therapy with a recombinant humanized monoclonal antibody to HER2/neu (Herceptin) has shown efficacy in large clinical trials, however, only 15-20 percent of patients with HER2-overexpressing tumors will respond to this treatment, and responses are limited to those tumors with 3+ expression of HER2. Attempts to enhance the actions of Herceptin have been hampered by our limited knowledge of its mechanism of action. We have recently demonstrated that co-stimulation of natural killer cells with Herceptin-coated breast cancer cells and interleukin-12 (IL-12) results in the production of significant quantities of IFN-g and other immunomodulatory cytokines with anti-tumor actions. Based upon these findings, we have initiated an NCI-sponsored Phase I trial in which Herceptin is administered in combination with IL-12 to patients with HER2-overexpressing malignancies. We hypothesize that administration of Herceptin followed by interleukin-12 will stimulate the production of IFN-g and other cytokines by immune cells bearing receptors for immunoglobulin (FcR). Furthermore, we propose that the anti-tumor actions of Herceptin will be significantly enhanced via the local production of factors with effects on tumor proliferation, angiogenesis, and antigen presentation.
In Aim 1 we propose to characterize the effects of IL-12 and Herceptin administration on the patient's immune system. Serum cytokine levels will be evaluated via ELISA and intracellular staining will be used to identify the specific cellular compartments responsible for cytokine production. Real-Time PCR will be used to quantify cytokine transcripts in peripheral blood mononuclear cells. We will also use flow cytometric techniques and antibody-dependent cytotoxicity assays to evaluate the effects of this treatment on the numbers, activation state, and cytotoxic capacity of FcR-bearing cells.
In Aim 2, we will perform immunohistochemical analyses on paraffin-embedded tumor samples that have been obtained over the course of the clinical trial in an effort to characterize the effects of IL-12 and Herceptin administration on the intratumoral immune cell infiltrate. The extent, phenotype, and activation status of this infiltrate will be carefully graded. Cytokine production by infiltrating immune cells will also be analyzed via immunohistochemical techniques. We will obtain all paraffin-embedded tumor samples derived from previous operations in order that we may have a pre-therapy baseline with which to compare our results. The effects of infiltrating immune cells on tumor cell proliferation and apoptosis will be systematically evaluated. These results will be correlated with those obtained in Aim 1 in an attempt to determine the effects of Herceptin/IL-12 administration on the tumor immune microenvironment. These studies will provide information on the actions of Herceptin and IL-12, enhance our ability to monitor the effects of antibody-based therapies, and permit the identification of specific markers of responsiveness.
| Martin del Campo, Sara E; Levine, Kala M; Mundy-Bosse, Bethany L et al. (2015) The Raf Kinase Inhibitor Sorafenib Inhibits JAK-STAT Signal Transduction in Human Immune Cells. J Immunol 195:1995-2005 |
