Anti-tumor Immunotherapy With Anti-HER2 Antibody/Flt3L Bispecific Fusion Protein

Tumor immunotherapy involves the induction of tumor regression by modulation of natural host defense mechanisms or by manipulation with a immunological agent. Immunotherapy is a recognized therapeutic modality for the treatment of malignancies along with the traditional modalities of surgical resection, radiotherapy and chemotherapy. In fact, immunotherapy is sometimes used as "complementary therapy" for the more common therapies such as surgery and radiation. The impetus for such combination therapy lies in the shortcomings in traditional modalities. More importantly, even if the modality of surgical resection is available to tumor patients, the problem of distant, undetected micrometastases remains untreated by such therapy. Likewise, the traditional therapies of radiotherapy and chemotherapy also have significant limitations, most prominently the systemic inhibition of the hematopoietic and immune system. Thus, the toxic effects of radiotherapy and chemotherapy limit efficacy of these therapies in the cases where radical treatment is most desired in the patient with significant tumor burden at the time of diagnosis. Therefore, it is desirable to find novel effective strategies that will complement traditional therapies.Immunotherapy of tumors can be effected through the administration of antibodies specific for tumor antigens. While antibodies typically have been used as delivery agents for toxic moieties, recent studies indicated that the monoclonal antibodies(mAbs) against certain cell surface molecules, e.g., FAS, EGFR, and HER2, directly induced tumor cell death through the triggering of apoptotic pathways. This suggests that the modulation of particular signaling pathways, particularly those resulting in tumor cell death, may provide a successful strategy for antibody-mediated tumor immunotherapy. At least one antibody employing this strategy has been successful during clinical trials. Herceptin, a monoclonal antibody specific for human HER2, induces apoptosis in Her2~+ tumor cells and has been used successfully for the in vivo treatment of breast cancer. However, one of the recognized limitations of such antibody therapy is the likelihood that distant metastases may still escape such therapy or that antigen-negative variants will develop, leading to a later relapse with metastatic disease.Immunotherapy can also be effected through the elicitation of an active anti-tumor immune response from the patient following the administration of a tumor vaccine. Ideally, the tumor vaccine delivers immunogenic tumor antigens to suitable antigen presenting cells, resulting in the generation of an effective and long-lasting anti-tumor immune response.Studies have demonstrated that the dendritic cell (DC), a type of antigen presenting cell, plays a crucial role in an effective anti-tumor immune response. DCs stimulate the differentiation of naive CD4+ and CD8+ T cells to T helper cells (Th) and cytotoxic T lymphocytes (CTLs), respectively. DCs can express high levels of both class I and class II major histocompatibility complex (MHC) antigens, costimulatory molecules, adhesion molecules and secrete high levels of IL-12, a potent cytokine in CTL differentiation and activation. As the CTL-mediated anti-tumor response is believed to generate long term protection against tumor regrowth, DCs appear to be the antigen presenting cell of choice for tumor immunotherapy.While tumor vaccines clearly confer long term protection against tumor metastatic outgrowth and even subsequent tumor challenges, the clinical application of this knowledge has proved to be difficult. First, it has proven difficult to reliably expand functional DCs in ex vivo expansion protocols. Because the immune is necessarily MHC-restricted, any ex vivo DCs employed in an immunotherapy strategy must be the DCs of the patient being treated. Second, reproducible activation of DCs in vivo has not yet been achieved. Third, no clear protocol has been established that permits the activation and antigen loading of the desired DC population, i.e., those capable of eliciting an anti-tumor response. In sum, the expansion of activated DCs selectively located at tumor site that present immunogenic tumor antigens is a problem that remains unsolved.Therefore, while it is clear that immune molecules, e.g., tumor-specific antibodies, and vaccines eliciting immune responses can effect tumor growth, a unified approach that permits the simultaneous reduction of tumor growth and the generation of lasting protective immune response is still lacking.Provided herein is a chimeric protein that permits the simultaneous eradication of tumor cells and the stimulation of an effective anti-tumor immune response. Specifically, the chimeric protein comprises two components. The first component is a biologically active fragment of Flt3. Flt3 is a potent chemotactic molecule and activator for DCs and other anti-tumor effectors such as NK cells. The second component is a tumoricidal agent-anti-HER2 antibody- that incude cell death through the direct initiation of the apoptotic cascade. Thus, the chimeric protein reduces tumor burden by directly inducing the apoptosis of tumor cells while also targeting and activating DCs, and other antitumor effectors, e.g., NK cells, to infiltrate the tumor tissues. Tumor antigens released by the dying tumor cells then can be processed and presented by Flt3-activated DCs, that then effectively serve as antigen-presenting cells for a specific anti-tumor immune response. Therefore, this...