Interferon-alpha-2 (IFNa2) is indicated for the therapy of a variety of hematopoietic and solid tumors. However, its therapeutic potential has not been fully realized primarily due to its short circulating half-life and systemic toxicity. Fusion of IFNa2 to a monoclonal antibody (MAb) markedly increases circulating half-life. The immediate clinical benefit is the requirement for less frequent and lower doses in order to achieve prolonged therapeutic concentrations. Additionally, targeting of IFNa2 to tumor sites using a tumor-associated antigen- directed MAb can significantly increase the local concentration and tumor retention of IFN12 while limiting its systemic concentration, thereby potentially increasing the therapeutic index. Increased tumor concentrations of IFNa2 can augment its direct anti-proliferative, apoptotic and anti-angiogenic activity, as well as prime and focus an antitumor immune response. This project will evaluate the feasibility of developing novel MAb-IFN1 conjugates for improved therapy of cancer. MAb-IFNa conjugates comprising 4 IFNa2b groups site-specifically tethered to an IgG have been produced using the Dock-and-Lock (DNL) method. The prototype MAb-IFNa, 20-2b, uses the humanized anti-CD20 MAb, veltuzumab, for targeted delivery of IFNa2b for therapy of B-cell lymphoma. Preliminary studies indicate that this conjugate may be highly potent and effective for NHL therapy. Evaluation of this prototype will be invaluable for the development of additional MAb-IFNa for therapy of a variety of hematopoeitic and solid tumors. The first specific aim for this Phase I project is to further characterize the biochemical and biological properties of 20-2b. This will include the identification and characterization of potential impurities that will be isolated by IEC. The pharmacokinetics and biodistribution will be assessed in normal mice. The stability in formulation buffer, blood and in vivo will be also be evaluated. For the second aim, suitable host cell lines will be developed for production of clinical material in small-scale bioreactors by screening of 150-300 transgenic myeloma clones and subsequent amplification with methotrexate.
The third aim will evaluate the therapeutic efficacy of 20-2b in a mouse model, as compared to veltuzumab and an irrelevant MAb-IFNa construct, with up to 5 human lymphoma cell lines having variable densities of CD20 expression.
The final aims will use FACS analysis to evaluate the anti-lymphoma therapeutic efficacy of 20-2b, as well as its affect on normal immune cells, in an ex vivo setting using blood from healthy human volunteers mixed with NHL cell lines and blood samples from human B-cell lymphoma and chronic lymphoid leukemia patients. The primary milestones for the Phase I project would show that this innovative construct can be suitable for clinical evaluation in terms of manufacturing, stability, safety and efficacy. Since both IFNa2b and veltuzumab have been studied in patients, we believe it will be relatively straightforward to complete the required preclinical studies as we upscale production of this cytokine-antibody conjugate.
The prototype construct evaluated in this project, 20-2b, may ultimately be developed into a therapeutic biologic agent useful for the treatment of B-cell malignancies including NHL, CLL and certain autoimmune disorders. This lessons learned from this project may eventually lead to the development of additional MAb-IFNa for therapy of a variety of hematopoeitic and solid tumors.
