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本文根据已发表的肿瘤坏死因子(TNF)氨基酸序列和核苷酸序列,用大肠杆菌偏性密码人工合成全基因,并在大肠杆菌中表达,表达量占总菌体蛋白的22～33%,均为可溶性TNF·菌体裂解液经抗TNF 单抗亲和柱纯化后得到SDS—PAGE 上分子量为17KD 的单带蛋白,比活性1～4×10~7u/mg。体外细胞毒试验表明:部分人肿瘤细胞系例如HL-60、Molt-4、OVCAR-3等对TNF 很敏感;部分肿瘤细胞例如KG_1、H7402等对TNF 呈中等敏感;也有部分肿瘤细胞对TNF 不敏感。从白血病患者周血来的原代白血病细胞对TNF 也呈现不同敏感性。TNF 与各种化疗药物,如阿霉素(AD)、更生霉素(KSM)等,或某些细胞因子,如γ-干扰素(IFN-γ)等联合应用时,对肿瘤细胞的杀伤有协同作用。裸鼠移植瘤模型上也观察到不论瘤内局部注射还是腹腔注射,均能明显抑制肿瘤生长,如果TNF 与化疗药物合用则抑瘤效果更好。
Based on the published amino acid sequence and nucleotide sequence of tumor necrosis factor (TNF), the full-length gene was synthesized by Escherichia coli biased codon and expressed in E.coli. The expression level accounted for 22-33% All soluble TNF · bacterial lysate purified by anti-TNF monoclonal antibody affinity column SDS-PAGE molecular weight of 17KD single band protein, specific activity 1 ~ 4 × 10 ~ 7u / mg. In vitro cytotoxicity test showed that some human tumor cell lines such as HL-60, Molt-4 and OVCAR-3 are very sensitive to TNF. Some tumor cells such as KG_1 and H7402 are moderately sensitive to TNF. Some tumor cells have no effect on TNF sensitive. Primary leukemia cells from leukemic patients are also differentially expressed on TNF. Tumor necrosis factor (TNF) is associated with various chemotherapeutic drugs such as adriamycin (AD), dactinomycin (KSM), etc., or certain cytokines such as interferon-γ (IFN- γ) Synergy. Nude mice transplantation model was also observed regardless of intratumoral injection or intraperitoneal injection, can significantly inhibit tumor growth, if the combination of TNF and chemotherapy drugs antitumor effect is better.