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Human interleukin‐17 (hIL‐17) gene without the signal sequence was isolated from activated peripheral blood lymphocytes by RT‐PCR, then highly expressed in the yeast Pichia pastoris in the form of the glycosylated monomer. The monomer of rhIL‐17 stimulated mouse fibroblast 3T3 cells to secrete IL‐6 and was specifically bound to its receptors on 3T3 cells.

Endostatin is a 20 kDa COOH-terminal fragment of collagen XVIII that inhibits angiogenesis and tumor growth. The cDNA coding for human endostatin in human fetal liver has been cloned into the secreting expression organism Pichia pastoris, and the high level expression of human endostatin has been achieved (about 200 mg of endostatin in 1 l of culture). The recombinant human endostatin was purified to homogeneity by heparin-affinity column, and showed antiproliferative effect on rat brain micro-vascular endothelia cells.

The pleiotropic cytokine interleukin‐6(IL‐6) triggers the formation of a high affinity receptor complex constituted by the ligand‐binding subunit IL‐6 receptor α(IL‐6Rα) and the signal transducer gpl30. To construct the antagonist of IL‐6, a DNA segment encoding the soluble human IL‐6Rα(shlL‐6Rα) mutant with two substitutions C277D/H280I was generated by overlap extension polymerase chain reaction. The DNA segment was then subcloned into vector pET‐3b and expressed in E.coli at a high level. The refolded and purified recombinant protein, which was designated as DM650, exhibited an increased IL‐6‐binding capability by 8‐fold. DM650 antagonized IL‐6 perfectly, resulting in the growth‐inhibition of human myeloma AF‐10 cells. DNA fragmentation assay proved that the growth of AF‐10 cells was inhibited through the induction of apoptosis suppressed by IL‐6.

Internal deletion K128‐Q175 of the human interleukin‐6 (hIL‐6) has been generated at the cDNA level. With pBV220 as expressing vector, the recombinant pBV*‐DIL‐6 encoding the deletion mutant (12kD) of hIL‐6 has been constructed. The resulted recombinant plasmids were then used to transform E. coli strain HB101, and the expression in the PLPR promoter system, which is temperature‐regulatable, was achieved. After purification and renaturation, the biological activity of the expressed product, designated as DM120, was measured by MTT method in an IL‐6‐dependent cell line 7TD1. The results show that the amino acid residues of IL‐6 128 to 175 are crucial for IL‐6 activity. Receptor binding assay in vitro indicates that the entire region is not involved in forming the receptor binding surface.