Ole Henrik Brekke

Therapeutic antibodies for human diseases at the dawn of the twenty-first century.

Antibodies are highly specific, naturally evolved molecules that recognize and eliminate pathogenic and disease antigens. The past 30 years of antibody research have hinted at the promise of new versatile therapeutic agents to fight cancer, autoimmune diseases and infection. Technology development and the testing of new generations of antibody reagents have altered our view of how they might be used for prophylactic and therapeutic purposes. The therapeutic antibodies of today are genetically engineered molecules that are designed to ensure high specificity and functionality. Some antibodies are loaded with toxic modules, whereas others are designed to function naturally, depending on the therapeutic application. In this review, we discuss various aspects of antibodies that are relevant to their use as as therapeutic agents.

The structural requirements for complement activation by IgG: does it hinge on the hinge?

The flexibility of antibody molecules principally derives from the structure of the hinge region. It has generally been accepted that the flexibility of the IgG hinge is necessary for complement activation to occur; however, recent studies dispute this premise. As described here by Ole Henrik Brekke, Terje Michaelsen and Inger Sandlie, it now appears that the only requirement of the hinge region for complement activation is the presence of inter-heavy-chain disulfide bond(s). Furthermore, the structural basis for the differences between IgG subclasses with respect to effector functions appear to be located within the CH2 domain of the immunoglobulin molecule.

Identification and Characterisation of C1q-Binding Phage Displayed Peptides

Five phage displayed peptide libraries were screened for binders to C1q, the recognition subunit of the classical complement pathway. Two rounds of panning resulted in the isolation and characterisation of several different phage displayed C1q-binding peptides from all five libraries. Two groups of the characterised peptides show sequence similarity with part of the metal ion dependent adhesion site (MIDAS) of integrin A-domains, and the site 187LRNPCPNKEKECQPPF of CD18 (integrin beta2), respectively. These results support binding of complement receptor 3 (CR3, CD11b/CD18, Mac1) to C1q and further suggest C1q binding sites in CR3. We also discuss sequence matches between the characterised peptides and proteins known to interact with C1q, as well as other proteins listed in the SwissProt databank. These findings are of interest for the study of the complement system and may lead to the development of peptides, fusion products or peptido-mimetics with C1q modulating potential.

A mutant human IgG molecule with only one C1q binding site can activate complement and induce lysis of target cells

There are potentially two binding sites for C1q on IgG, one on each CH2 domain of the gamma heavy chains, close to the lower hinge region. It is not clear whether the presence and involvement of both the C1q binding sites is necessary to induce the activation signal of human IgG. In order to clarify this issue, we made a hybrid mutant IgG1/IgG3 molecule where the IgG1 half of the molecule was made unable to activate complement through the introduction of a P329A mutation. The IgG3 half of the molecule was mutated to harbor a hinge region identical to that of IgG1, and for detection a peptide tag derived from p21ras was introduced into the FG loop of the CH1 domain. The hybrid IgG1P329A/IgG3h1‐ras molecules were isolated by Protein A affinity chromatography and shown to activate complement and induce complement‐mediated lysis at the same levels as wild‐type IgG1 and IgG3h1‐ras molecules. Thus, one C1q binding site per IgG is sufficient to induce activation. Wild‐type human IgG molecules might also normally expose only one C1q binding site as already shown for interaction with FcγR, were IgG expose one binding site per molecule.