Harvill E T, Morrison S L
Department of Microbiology and Molecular Genetics, University of California, Los Angeles 90024-1489, USA.
Immunotechnology. 1995 Aug;1(2):95-105. doi: 10.1016/1380-2933(95)00009-7.
The therapeutic value of Interleukin 2 (IL-2) is limited by its short half life and systemic toxicity. One approach to overcoming these problems is to fuse this protein to an antibody, a protein with a long half life and the ability to target a unique antigen within the body. To examine the biochemical properties of such a molecule a fusion protein was constructed linking the N-terminus of human IL-2 to the C-terminus of IgG3. A similar fusion between IgG1 and IL-2 has previously been shown to bind antigen, generate antibody-dependent cellular cytotoxicity (ADCC) and stimulate T cell proliferation and cytotoxicity. We now extend these studies and show that the fusion protein, termed IgG3-IL2, is appropriately N-glycosylated within the IgG3 CH2 domain, binds the human high affinity Fc receptor (Fc gamma RI) with an affinity slightly lower than that of IgG3, and is able to activate complement via the classical pathway to lyse antigen coated sheep red blood cells (SRBC). When used to stimulate the proliferation of the IL-2 dependent cell line CTLL-2, IgG3-IL2 has a specific activity slightly lower than that of human recombinant IL-2 (hrIL-2). In marked contrast, when comparable unit concentrations, as defined by the standard CTLL-2 proliferation assay, are used to stimulate human peripheral blood lymphocytes (PBL), IgG3-IL2 generates significantly greater lymphokine activated killer (LAK) cell cytotoxicity than does hrIL-2. Competition studies show that IgG3-IL2 binds the intermediate affinity form of the IL-2 receptor (IL-2R), consisting of the beta and gamma subunits, with an affinity slightly less than that of hrIL-2. In contrast, IgG3-IL2 shows a greater affinity than hrIL-2 for the high affinity IL-2R, consisting of alpha, beta and gamma subunits. Our studies show that the IgG3-IL2 fusion protein possesses a combination of the biological properties of IgG3 and IL-2 including antigen binding, complement activation, Fc gamma RI binding, IL-2R binding and stimulation of both proliferation and LAK activity. This combination of activities may allow IgG3-IL2 to target humoral and cell-mediated immune activation to the site of an antigen of interest or target an antigen to IL-2R bearing cells or organs.
白细胞介素2（IL-2）的治疗价值受到其半衰期短和全身毒性的限制。克服这些问题的一种方法是将这种蛋白质与抗体融合，抗体是一种半衰期长且能够靶向体内独特抗原的蛋白质。为了研究这种分子的生化特性，构建了一种融合蛋白，将人IL-2的N端与IgG3的C端相连。先前已证明IgG1和IL-2之间的类似融合可结合抗原、产生抗体依赖性细胞毒性（ADCC）并刺激T细胞增殖和细胞毒性。我们现在扩展这些研究，并表明称为IgG3-IL2的融合蛋白在IgG3 CH2结构域内进行了适当的N-糖基化，以略低于IgG3的亲和力与人高亲和力Fc受体（FcγRI）结合，并能够通过经典途径激活补体以裂解包被抗原的绵羊红细胞（SRBC）。当用于刺激IL-2依赖性细胞系CTLL-2的增殖时，IgG3-IL2的比活性略低于人重组IL-2（hrIL-2）。与之形成鲜明对比的是，当使用标准CTLL-2增殖试验定义的可比单位浓度来刺激人外周血淋巴细胞（PBL）时，IgG
