Abstract

Anti-HER2/CD3, a T-cell-dependent bispecific antibody (TDB) construct, induces T-cell-mediated cell death in cancer cells expressing HER2 by cross-linking tumor HER2 with CD3 on cytotoxic T cells, thereby creating a functional cytolytic synapse. TDB design is a very challenging process that requires consideration of multiple parameters. Although therapeutic antibody design strategy is commonly driven by striving for the highest attainable antigen-binding affinity, little is known about how the affinity of each TDB arm can affect the targeting ability of the other arm and the consequent distribution and efficacy. To our knowledge, no distribution studies have been published using preclinical models wherein the T-cell-targeting arm of the TDB is actively bound to T cells. We used a combined approach involving radiochemistry, invasive biodistribution, and noninvasive single-photon emission tomographic (SPECT) imaging to measure TDB distribution and catabolism in transgenic mice with human CD3ε expression on T cells. Using CD3 affinity variants, we assessed the impact of CD3 affinity on short-term pharmacokinetics, tissue distribution, and cellular uptake. Our experimental approach determined the relative effects of (i) CD3 targeting to normal tissues, (ii) HER2 targeting to HER2-expressing tumors, and (iii) relative HER2/CD3 affinity, all as critical drivers for TDB distribution. We observed a strong correlation between CD3 affinity and distribution to T-cell-rich tissues, with higher CD3 affinity reducing systemic exposure and shifting TDB distribution away from tumor to T-cell-containing tissues. These observations have important implications for clinical translation of bispecific antibodies for cancer immunotherapy. Mol Cancer Ther; 17(4); 776-85. ©2018 AACR.

Highlights

  • Breast cancer is the leading cause of death for women from 20 to 59 years of age diagnosed with any cancer, with approximately 705 new cases diagnosed daily [1]

  • Cellular binding of T-cell–dependent bispecific antibody (TDB) Binding of glycoprotein D (gD)/CD3 and HER2/CD3 to HER2-expressing SKBR3 cells was determined by flow cytometry

  • Jurkat cell–binding profiles increased with CD3 affinity; the replacement of gD with the HER2 arm had no effect on CD3 binding (Supplementary Fig. S1B)

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Summary

Introduction

Breast cancer is the leading cause of death for women from 20 to 59 years of age diagnosed with any cancer, with approximately 705 new cases diagnosed daily [1]. About 15% to 20% of breast cancer overexpress the HER2 and are more likely to invade axillary lymph nodes than other subtypes [2]. Approved antibody therapeutics that target HER2 include the monoclonal antibodies trastuzumab and pertuzumab and the antibody–drug conjugate ado-trastuzumab emtansine. Low response rates and the emergence of resistance limit their efficacies [3]. Note: Supplementary data for this article are available at Molecular Cancer Therapeutics Online (http://mct.aacrjournals.org/).

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