Antibody-Based Targeting of Interferon-Beta-1a Mutein in HER2-Positive Cancer Enhances Antitumor Effects Through Immune Responses and Direct Cell Killing

Abstract

Type I interferon (IFN) has been approved as an anticancer agent to treat some malignancies. However, IFNs have a short in vivo half-life, systemic toxicity, and poor biophysical properties, which prevent it from being widely used for cancer therapy. This study aimed to construct recombinant IFN-beta-1a mutein immunocytokines that comprise a human epidermal growth factor receptor 2 (HER2)-targeting antibody and IFN-beta muteins with an additional glycosylation, which can overcome the limitation of the cytokine itself. Hence, the molecular design aims to 1) enhance productivity and biophysical properties by adding secondary glycosylation in IFN-beta, 2) increase the therapeutic index of IFN-beta therapy by preferential retention at the tumor by possessing high affinity for HER2-expressing cancer cells, and 3) improve the pharmacokinetics and, thus, the convenience of IFN-beta administration. The yield of trastuzumab-IFN-beta mutein was higher than that of trastuzumab-wild-type IFN-beta in the mammalian cell culture system. Trastuzumab-IFN-beta mutein showed similar IFN activity and HER2-targeting ability equivalent to that of IFN-beta mutein and trastuzumab, respectively. Trastuzumab-IFN-beta mutein directly inhibited the growth of HER2-positive gastric cancer cell lines and was more effective than trastuzumab or IFN-beta mutein alone. Trastuzumab-IFN-beta mutein and IFN-beta mutein displayed enhanced immune cell-mediated cytotoxicity. Collectively, trastuzumab-IFN-beta mutein may have indirect immune cell-mediated antitumor effects and direct cell growth inhibitory effects. Tumor-targeting effect of trastuzumab-IFN-beta mutein was analyzed using in vivo fluorescence imaging. The accumulation of trastuzumab-IFN-beta mutein was observed in HER2-positive tumors rather than other tissues except the liver. To evaluate the both direct tumor growth inhibition effect and indirect immune cell-mediated antitumor effect, we tested the effect of trastuzumab-IFN-beta mutein in HER2-positive cancer xenograft models using nude mice or humanized mice. Trastuzumab-IFN-beta mutein could significantly enhance tumor regression when compared with trastuzumab or IFN-beta mutein. In addition, an increase in tumor-infiltrating lymphocytes was observed in the trastuzumab-IFN-beta mutein-treated group, implying that the tumor-targeting IFN-beta may have an enhanced antitumor effect through increased immune response. Therefore, targeting IFN-beta with an anti-HER2 monoclonal antibody makes the immunocytokine more potent than either agent alone. These novel findings suggest that trastuzumab-IFN-beta mutein merits clinical evaluation as a new candidate of anticancer therapeutics.