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Abstract Human cancers employ a number of mechanisms to evade host immune responses against novel antigens generated from aberrant over-expression, mutations and/or epigenetic alterations. Humoral immunity utilizes antibodies and immune-effector cells as well as molecular immune complexes involving the complement system to mediate the killing of dysregulated cancer cells. We refer to these anti-cancer mechanisms as Humoral Immuno-Oncology (HIO). HIO suppression is mediated by tumor-produced proteins called HIO factors. One such factor is CA125, which was previously shown to bind IgG-type antibodies and inhibit their immune-effector activities, including antibody dependent (ADCC) and complement dependent (CDC) cellular cytotoxicity. Using a combination of experimental screening and literature searches, a second protein produced by tumors and associated with a variety of cancer indications was discovered and found to be soluble ICAM-1 (sICAM-1). Through various functional studies we reported that sICAM-1 and membrane ICAM-1 (mICAM-1) are capable of binding IgG1-type antibodies that in turn inhibits their immune-effector activity. Through a combination of truncation and substitution mutagenesis, we identified a four amino acid motif within the CH3 domain of IgG1 essential for ICAM-1 binding, resulting in the inhibition of ADCC activity. While traditional glycine and alanine substitutions in this region abrogated ICAM-1 binding, these modifications caused tertiary structural changes in the Fc domain that resulted in loss of ADCC activity. Through a combinatorial amino acid substitution approach, we identified a four amino acid combination within the 407YSKL410 to 407FARV410 motif that resulted in antibodies refractory to sICAM-1 binding with robust ADCC activity. Additionally, isogenic wildtype and ICAM-1 knockdown target cell lines showed mICAM-1 inhibited target cell killing of a saporin antibody drug conjugate (ADC) and this was the direct result of mICAM-1 reducing antibody internalization, a requisite for maximal ADC target cell killing. The inhibitory activity of mICAM-1 could be overcome employing antibodies containing the 407FARV410 substitution. These findings highlight yet another mechanism by which tumors can suppress the host’s immune system for survival and offer new concepts for developing antibody-based therapies that can aid in the treatment of various cancer indications, especially those over-expressing ICAM-1. Moreover, the findings here offer diagnostic and therapeutic clinical design opportunities to improve upon existing approved immune-mediated therapies for which this factor is present. Citation Format: Brad Kline, Luigi Grasso, Nicolas Nicolaides. Engineering of ICAM-1 refractory antibodies for the development of therapeutic antibodies and antibody drug conjugates (ADCs) in ICAM-1 over-expressing cancers abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts) ; 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84 (6Suppl): Abstract nr 6344.
Kline et al. (Fri,) studied this question.