Research Grants Awarded
Promotion Of Anti-Tumor Immune Responses By Conversion Of Tumor-Associated Macrophages To Inflammatory Phenotype
Investigator Initiated Research
Macrophages play a critical role in progression and metastasis of many types of cancer, most notably mammary carcinomas. Our studies on macrophage biology over the past 2 decades have revealed that macrophages retain functional plasticity. That is, inflammatory macrophages can readily be converted to anti-inflammatory macrophages and vice versa. This led us to test whether the immunosuppressive and tumor-promoting activities of the various tumor-associated macrophages could be reduced and/or converted to inflammatory and pro-immunogenic activities. We have demonstrated that treatment of tumor-bearing mice with liposome encapsulated IL-12 induces a very rapid (90 minutes) shift in function of macrophages within the primary tumor, within the metastases, and within distal tissues (spleen and peritoneum). The impact of this functional conversion appears to be two-fold. The shift in macrophage function (1) disrupts the symbiotic relationship between the tumor and macrophages, impairing tumor progression and metastasis and (2) promotes an adaptive immune response. Part of this functional response of macrophages to IL-12 involves the release of cytoplasmic IL-15, which we have shown to be critical for IL-12 induced leukocytic infiltration of the primary tumor and metastases and for subsequent tumor destruction by NK cells and cytotoxic T cells. We also demonstrated that the inflammatory activity of the macrophages is sustained for approximately 4 days after a single injection of IL-12, presumably due to the pro-inflammatory activities of the NK and T cells involved in the anti-tumor response.
We therefore hypothesize that the conversion of tumor-associated macrophages to an inflammatory, pro-immunogenic phenotype creates a pro-immunogenic window during which tumor progression is impaired by lack of macrophage support and an immune response is facilitated. An anti-tumor response of variable efficacy (depending on primary tumor mass) occurs without the administration of a vaccine optimized for elicitation of an anti-tumor response. Tumor vaccination has been explored by many investigators, but administration of tumor vaccines alone has been somewhat disappointing. We hypothesize that the efficacy of tumor vaccines can be increased dramatically by administering the vaccine after converting the tumor associated macrophages from an immunosuppressive to an inflammatory functional state and by sustaining that inflammatory, pro-immunogenic functional state of macrophages for as long as possible. The current proposal describes an investigation into the novel perspective of tumor associated macrophages as critical immunotherapeutic allies in the establishment of tumor immune responses rather than as ?adversaries? requiring cytotoxic elimination.
We propose to investigate mechanisms of prolonging the pro-immunogenic window initiated by IL-12 treatment and to determine if prolonging that pro-immunogenic window enhances the efficacy of tumor vaccines. We will use the 4T1 model of mammary carcinoma in Balb/c mice. The first aim will be to investigate 1a) the contribution of NK and T cells to the duration of the pro-immunogenic window and 1b) the contribution of inflammatory cytokines such as IL-15, IL-18 and IFN? to the duration of the pro-immunogenic window. The baseline for macrophage response to IL-12 will be determined in NK-depleted tumor-bearing SCID mice. Subsequently the contribution of NK, NK-T, CD4 and CD8+ T cells will be determined. Inflammatory cytokines such as IL-15 and IL-18 have been investigated for potential tumor therapeutic potential, but have displayed limited impact. Both of these cytokines contribute, directly or through their ability to elicit production of other cytokines such as IFN?, to enhancement of macrophage inflammatory activity, and/or to NK activation and cytokine production, and/or to T cell activation and survival, with expansion of the central memory compartment of T cells. We therefore propose to investigate whether split administration of liposome encapsulated IL-15 or IL18 1-5 days after administration of IL-12 extends the pro-immunogenic window as assessed by sustained inflammatory activity and minimal immunosuppressive activity of tumor-associated macrophages. The second aim will directly examine the degree to which optimal induction of the pro-immunogenic window enhances the anti-tumor response to a 4T1-dendritic cell vaccine in tumor-bearing mice. We anticipate that optimization of the inflammatory and pro-immunogenic environment of the host, and thus minimization of the immunosuppressive environment, will significantly enhance the intensity and duration of the immune response to the tumor, in particular mammary carcinomas in which the tumor-promoting and immunosuppressive activities of macrophages play such a forceful role.
Tumor vaccines as a means of initiating immune destruction of an existing tumor have received substantial research attention. However, despite their theoretical promise, tumor vaccines have had somewhat mixed and disappointing results. Many tumors, especially mammary carcinomas, recruit host macrophages to assist intumor growth, metastasis, and immunosuppression. The immunosuppressive activities of the macrophages is not limited to the tumor mass. Tumors release membrane encapsulated cytoplasmic micsrovesicles which circulate throughout the body, displaying ligands and cytokines that promote and activate immunosuppressive macrophage activities systemically. This creates a very strong immunosuppressive environment which makes it difficult to induce a tumor specific immune response in the tumor host. To date, there has been no systematic study of means to counter that immunosuppressive environment. Yet, theoretically, successful immunotherapy of mammary carcinoma will require neutralization of that immunosuppressive environment. Recognizing this, several investigators have initiated studies designed to evaluate means of cytotoxic destruction of macrophages in tumor-bearing hosts. This approach is problematic since there are no clearly defined macrophage-specific surface molecules and macrophage inflammatory activity plays a significant role in leukocytic infiltration and amplification of immune responses.
We hypothesized and subsequently demonstrated, that injection of liposome-encapsulated IL-12 into tumor-bearing mice would result in a rapid functional change in tumor-associated macrophages, marked by a termination of immunosuppressive and tumor supportive activities and increase in inflammatory activities. We have since demonstrated that this functional response of tumor-associated macrophages to IL-12 plays a critical role in initiating leukocytic infiltration into the tumor and the induction of a destructive anti-tumor response. It is therefore clear that administration of IL-12 is one method of at least transiently reducing the immunosuppressive environment of tumor hosts. In lung carcinoma models, administration of encapsulated IL-12, without administration of a tumor vaccine, results in a tumor-destructive immune response that is capable of destroying the primary tumor as well as the metastases. This provides proof of principle that it is possible to neutralize the immunosuppressive environment sustained by tumor associated macrophages and that neutralizing the immunosuppressive environment will establish a pro-immunogenic window and strongly promote the generation of an immune response against a tumor. We now propose to investigate the efficacy of establishing a pro-immunogenic window by redirecting the functional profile of tumor-associated macrophages in a mouse model of breast cancer. The study will determine if combined cytokine treatment, targeting NK cells and T cells in addition to macrophages, prolongs the pro-immunogenic window sufficiently to allow the induction, by administration of a tumor vaccine, of a sustained and effective anti-tumor cytotoxic immune response.
The perspective that tumor-associated macrophages may be recruited as allies in immunotherapeutic intervention in mammary carcinoma and the concept of redirecting the functional activities of the tumor-associated macrophages toward inflammatory and pro-immunogenic activities is extremely novel. The hypothesis that redirecting the activities of tumor-associated macrophages will enhance the efficacy of tumor vaccines has never been proposed, much less tested. Our published preliminary studies strongly indicate that the hypothesis holds substantial merit. The current proposal focuses on means of optimizing the conversion of tumor-associated macrophages to inflammatory function and directly testing whether this conversion allows successful long-term destruction of the tumor upon vaccination, using a murine model of mammary carcinoma. If successful, this research will open an exciting new avenue of tumor immunotherapy that, as yet, has never been explored and may prove to increase the efficacy of several immunotherapeutic interventions currently under investigation, in addition to tumor vaccines.