Enhancing the immunotherapeutic Trastuzumab for selective activity in the low pH tumor microenvironment

Abstract

Abstract Highly selective killing of tumor cells is a key requirement for a successful and tolerable intervention in cancer treatment. While immunotherapies have been revolutionary in reducing the systemic impacts of chemotherapy, dramatic and sometimes life-threatening side effects can occur when selectivity is not stringent enough. For example, the popular HER2+ breast cancer therapeutic Trastuzumab has well-documented cardiotoxic effects due to expression of HER2 on cardiomyocytes. To enhance selectivity of immunotherapeutics for tumor cells, we can take advantage of the unique tumor microenvironment and its low pH relative to healthy tissues to bias the activity of antibodies toward tumor tissues. We have engineered antibodies for enhanced binding and efficacy at low pH, which may alleviate the requirement for binding an antigen that is only present on cancer cells while simultaneously reducing side effects due to binding non-tumor cells. Trastuzumab CDRs were mutagenized and screened for binding of HER2 antigen at a tumor pH of 6.5 and reduced binding at the normal tissue pH of 7.4. In a more generalizeable approach, the Fc region of Trastuzumab was also mutagenized and screened for similar pH dependent binding to FcγrIIIA, the interaction responsible for recruitment of key anti-tumor natural killer cells. All library screening took place on the surface of CHO cells, ensuring selection of clones that express well and contain the glycosylation necessary for Fc structure and function. Improved Trastuzumab variants identified through this work have the potential to dramatically improve this popular breast cancer therapy by reducing the cardiotoxic side effects, making the therapy more tolerable and available to more patients.