Small molecule drug activity in melanoma models may be dramatically enhanced with an antibody effector

Abstract

Monoclonal antibody (mAb) 38C2 belongs to a group of catalytic antibodies that were generated by reactive immunization and contains a reactive lysine. 38C2 catalyzes aldol and retro-aldol reactions, using an enamine mechanism, and mechanistically mimics natural aldolase enzymes. In addition, mAb 38C2 can be redirected to target integrins a v b 3 and a v b 5 through the formation of a covalent bond between a b-diketone derivative of an arginine-glycine-aspartic acid (RGD) peptidomimetic and the reactive lysine residue in the antibody combining site to provide the chemically programmed mAb cp38C2. In this study, we investigated the potential of enhancing the activity of receptor-binding small molecule drug (SCS-873) through antibody conjugation. Using a M21 human melanoma xenograft model in nude mice, cp38C2 inhibited the growth of the tumor by~81%. The chemically programmed antibody was shown to be highly active at a low concentration while SCS-873 alone was ineffective even at dosages~1,000-fold higher than those used for the chemically programmed antibody. In vitro programming of the catalytic antibody was shown to be as effective as in vivo programming. In an experimental metastasis assay, treatment with mAb cp38C2 significantly prolonged overall survival of tumor-bearing severe combined immuno-deficient (SCID) mice when compared to treatment with unprogrammed mAb 38C2, SCS-873 alone or the integrin-specific monoclonal antibody LM609. In vitro, cp38C2 inhibited human and mouse endothelial and human melanoma cell adhesion, migration and invasion. Additionally, cp38C2 inhibited human and mouse endothelial cell proliferation and was active in complement-dependent cytotoxicity assays. These studies establish the potential of chemically programmed monoclonal antibodies as a novel and effective class of immunotherapeutics that combine the merits of traditional small molecule drug design with immunotherapy. ' 2006 Wiley-Liss, Inc.Key words: melanoma; angiogenesis; integrins; antibodies; tumor modelsThe discovery that tumor growth and metastasis are codependent on the formation of neovascularization 1 revealed many potential protein targets for cancer treatment. [2][3][4] The angiogenic process depends on proliferation of vascular endothelial cells, migration of tumor cells to the vascular endothelium followed by cell adhesion, and finally invasion of the endothelium. 5 A family of highly conserved adhesion molecules, known as integrins, is central to the regulation of these processes. Integrins are heterodimeric transmembrane receptor complexes composed of noncovalently associated a and b chains, and recognize the arginine-glycine-aspartic acid (RGD) sequence present in their extracellular matrix (ECM) ligands. 6 At present, 18 a and 8 b subunits are known; these form 24 different ab heterodimers with different specificity for various ECM cell-adhesive proteins. 7 Ligands for various integrins include fibronectin, collagen, laminin, von Willebrand factor, osteopontin, thrombospondin and vitronectin, all components of...

Preparation of integrin α(v)β(3)-targeting Ab 38C2 constructs

“…The conjugate formation was also established by ELISA experiments, and by flow cytometry to record the binding to cells expressing integrins a v b 3 and a v b 5 . These methods are described elsewhere [3][4][5]7,9 .…”

“…Notably, one common feature of Abs is that they are highly specific to a single target and, because of their clonal nature, a different Ab is required for each target. We have devised a new approach in which we modify the binding sites of a single Ab, 38C2, allowing it to be retargeted to more than one receptor [3][4][5][6][7] . Such Ab constructs are useful for adapter immunotherapy.…”

“…The above-mentioned cp38C2 constructs bound strongly to integrins a v b 3 -and a v b 5 -expressing cancer cells, such as human Kaposi's Sarcoma (SLK), human melanoma (M21) and human breast carcinoma (MDA-MB-231 and MDA-MB-435) [3][4][5]7 . The constructs inhibited the growth of the primary tumor and/or metastases of these human cancers as well as several others, such as human colon cancer (SW1222), which lack expression of a v b 3 .…”

“…The constructs inhibited the growth of the primary tumor and/or metastases of these human cancers as well as several others, such as human colon cancer (SW1222), which lack expression of a v b 3 . Mechanistically, the a v b 3 -and a v b 5 -integrin-targeting 38C2 constructs caused toxicity to cells not only by antagonizing the integrins or causing antiangiogenic effects, but also by instigating Ab-dependent cellular and complement-mediated cytotoxicities 5 .…”

Breaking the one antibody–one target axiom

“…The resulting conjugate of small molecule and antibody is a cpAb. Significantly, we have demonstrated that chemical programming of a catalytic antibody can occur both in vitro and in vivo to have a therapeutic effect in disease models (3,5). Key to this approach is the development of catalytic antibodies that operate using covalent reaction mechanisms (6, 7).…”