© 2004 The Japanese Biochemical Society
MOLECULAR BIOLOGY |
Soluble Expression of Recombinant Human Secondary Lymphoid Chemokine (SLC) in E. coli and Research on Its In Vitro and In Vivo Bioactivity
1 The Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, 100101, Beijing, China; and 2 Beijing ABT Genetic Engineering Technology Co. Ltd., 102206, Beijing, China
Secondary lymphoid tissue chemokine (SLC) is a CC chemokine that plays an important role in leukocytes homing to lymphoid tissues. The ability of SLC to co-localize both T cells and dendritic cells formed the rationale to evaluate its utility in cancer immunotherapy. The in vivo antitumor effect of murine SLC (mSLC) has been well documented, but little is known about that of human SLC (hSLC). To investigate the antitumor efficiency in vivo of hSLC, the hSLC gene was artificially synthesized and induced to express as a soluble form in Escherichia coli. After purification, the purity of the recombinant human SLC (rhSLC) protein was above 95% by SDS-PAGE analysis. The Kd of rhSLC binding to peripheral blood lymphocytes (PBLs) was 0.2186 ± 0.02675 µM as assessed by FACS, and the maximal chemotactic index of rhSLC was 9.49 at 100 nM as assessed by in vitro chemotaxis assay. Then genomic sequences of hSLC and mSLC, and of human CCR7 (hCCR7) and murine CCR7 (mCCR7), the receptor for SLC, were aligned. It was found that hSLC and mSLC share 70.72% identity and hCCR7 and mCCR7share 86.77% identity. Furthermore, we found that rhSLC could chemoattract murine peripheral blood mononuclear cells (PBMCs) in vitro. On the basis of these facts, immune competent mice inoculated with S180 sarcoma cells were chosen as an in vivo model. Intratumoral injections of rhSLC inhibited tumor growth and increased survival. These findings suggest that, despite its incapability to bind to either human or murine CXCR3, which is related to angiostasis, rhSLC can induce an antitumor response in vivo by another route. This report proves that rhSLC has a potent tumor-inhibition ability that makes it a promising candidate agent in cancer immunotherapy.
* To whom correspondence should be addressed at: Beijing ABT Genetic Engineering Technology Co. Ltd., ZGC Life Science Park, No 55, Beiqing Road, Beijing, 102206, China. Fax: +86-10-80726906, E-mail: hlhuang{at}genetics.ac.cn