Adoptive immunotherapy with LAK cells conjugated with epidermal growth factor receptor antibody

我们开发了一种无血清培养基，命名为RDSF，用于在RD6F培养基的基础上产生LAK细胞，RD6F培养基最初是作为骨髓瘤和杂交瘤细胞生长的无血清培养基开发的。在RDSF中生成的LAK细胞对Raji、K562和口腔癌细胞的细胞毒活性是在含有10%人ab型血清的培养基中生成的LAK细胞的3-4倍。RDSF培养基由添加转铁蛋白、2-氨基乙醇、2-巯基乙醇、亚硒酸钠和白细胞介素-2的营养混合物组成。在本研究中，我们发现胰岛素是动物细胞中最重要的多肽激素，它可以抑制外周血淋巴细胞培养的LAK细胞产生细胞毒活性。此外，我们发现转铁蛋白是LAK细胞在无血清培养中生长和产生的重要成分。这些结果表明，RDSF将用于癌症的过继免疫治疗和研究LAK细胞生长和分化的相关因素。此外，我们还研究了EGF-r的12-93单克隆抗体（MoAb）在体内对鳞状细胞癌（SCC）和唾液腺腺癌细胞（SAC）生长的影响，并在体外用12-93 MoAb偶联LAK细胞靶向免疫治疗口腔癌细胞，A 12-93 MoAb在体内抑制SCC和SAC的生长。12-93偶联LAK细胞对A431和HSG细胞的溶解作用显著增强，而对Raji细胞的溶解作用不明显。这些结果表明，12-93将是肿瘤特异性MoAb的有效替代品，12-93结合的LAK细胞可能为肿瘤的过继性免疫治疗提供有效的策略。

We have developed a serum-free medium designated RDSF for the generation of LAK cells based on RD6F medium, which was originally developed as a serum-free medium for the growth of myeloma and hybridoma cells. The cytotoxic activity of LAK cells generated in RDSF against Raji, K562 and oral cancer cells, is 3-4 times that of LAK cells generated in medium containing 10% human type-AB serum. RDSF medium consisted of nutrient mixture supplemented with transferrin, 2-aminoethanol, 2-mercaptoethanol, sodium selenite and interleukin-2. In this study, we have found that insulin, which has been found to be the most important polypeptide hormone in surum-free media for animal cells, inhibited the generation of cytotoxic activity of LAK cells cultured from pheripheral blood lymphocytes. In addition, we have found that transferrin was an essential component for the growth and generation of LAK cells in serum-free culture. Thses results suggest that RDSF will be usefel for adoptive immunotherapy of cancer and studying factors involved in the growth and differentiatiation of LAK cells.In addition, we have studied effect of monoclonal antibody (MoAb) to EGF-r designated as 12-93 on the growth of squamous cell carcinoma cells (SCC) and salivary gland adenocarcinoma cells (SAC) in vivo, and targeting immunotherapy using 12-93 MoAb-conjugated LAK cells against oral cancer cells in vitro, A 12-93 MoAb inhibited the growth og SCC and SAC in vivo. The 12-93 conjugated LAK cells showed significantly enhanced cytolysis of 12-93 reactive A431 and HSG cells but not Raji cells which dose not react with 12-93. These results indicated that 12-93 will be aviable altemative to tumor specific MoAb and 12-93-conjugated LAK cells may provide an effective strategy for targeting adoptive immunotherapy of cancer.