Exciting applications of single chain trimers of MHC-I molecules

Dear Editor Oved et al. reported their findings utilizing singlechain versions of MHC class I, wherein all three components of the class I heterotrimer were covalently attached with flexible linkers in the following order: antigenic peptide-linker-b2m-linker-heavy-chain [1]. It should be noted that this format, which we have termed single chain trimer, was previously reported by us and others in papers not cited. In their report, Oved et al. demonstrated that HLA-A2-containing versions of SCTs could be produced in recombinant form and used successfully for tetramer production for enumeration and stimulation of cognate CTLs. They speculate that their SCTs could have many important applications, largely due to the expected enhancement of the stability of single-chain molecules relative to native peptide/ MHC class I complexes. Indeed, the authors write: ‘‘We hypothesize that the covalent attachment of the peptide will make the complex more stable by its ability to continuously and efficiently rebind the covalently attached peptide’’ [1]. We heartily agree. In fact, we previously published extensive experimental evidence demonstrating that the superior stability of SCTs is ‘‘rendered by their ability to continuously and efficiently rebind the covalent peptide’’ [2]. Furthermore, in the original SCT paper that appeared in the Journal of Immunology in 2002, we demonstrated that SCTs are expressed efficiently, are recognized by specific antibodies and T cells, are unusually stable at the cell surface, and can stimulate antigen-specific T and B cells ([3], and highlighted in Nature Reviews in Immunology 2, 306 (2002) for its vaccine potential). Our follow-up report in 2003 explored the mechanism underlying the unique properties of SCTs, with special emphasis on understanding how the peptide-binding groove is able to accommodate the linker extending from the C-terminus of the peptide. Further, we demonstrated that recombinant forms of the SCTs could be produced in bacteria, refolded in vitro for tetramerization, and used to stain cognate CTLs [2]. It is also important to note that another group published an independent report, using a format very similar to SCTs, showing that HLAA2 versions were recognized by T cells and that these SCT-like molecules could be used as specific T cellstaining reagents [4]. In addition, a similar format was described for the surface display of MHC class I on bacteriophage [5]. There is a substantial body of published work on the engineering of class I and b2m to present single peptides through covalent tethering [6–13 for example]. However, it appears that placing all three components of class I together in the specific order found with SCTs leads to the production of high quality molecules, and this technology seems to be broadly applicable. The unique ability of SCTs to present single peptides with relatively high efficiency has led us and others to explore a variety of applications that exploit this technology. One of the most exciting applications of this technology is to prime CTL responses. In three different systems, DNA vaccination has now been used to successfully prime specific CTL responses in mice [14–16]. In one of these cases, Gene Gun immunization was used to compare the efficiency of CTL priming against tumors by an SCT versus expression of the entire protein from which the peptide used in SCT construction was derived [14]. The results indicated that the preassembled nature of the SCT offered a dramatic advantage in terms of generating CTL responses for tumor rejection. The ability to engineer class I to efficiently present a single peptide has also proven useful in more basic research in immunology. We have generated transgenic Ted. H. Hansen (&) Department of Pathology and Immunology, Washington University School of Medicine, St Louis, MO 63110, USA E-mail: hansen@immunology.wustl.edu