LAK cell proliferation by hollow fiber bioreactor

标准治疗失败并有多处转移疾病的骨肉瘤患者似乎很难治愈。Rosenberg等人最近报道，LAK细胞和IL-2的联合治疗能够导致已建立的转移性癌症的消退。在以往的小鼠模型中，认为治疗效果与LAK细胞的数量和IL-2的用量直接相关。为了将这一过继免疫疗法应用于晚期骨肉瘤患者，我们首先研究了免疫治疗中有效的LAK细胞的体外扩增方法，采用中空纤维生物反应器系统(Cellmax-100)，观察其对LAK细胞的增殖作用，并在抗CD3抗体包被的培养瓶中培养7天。取约2~4×10个细胞接种于中空纤维生物反应器中，继续培养2周。中空纤维生物反应器中LAK细胞数量未见增加。在他们的表面标记中。Leu4a(CD3+)细胞在第1周和第2周均为98-99%。Leu2a(CD8+)细胞在1周时为55%，2周时为74%。1周时Leu3a(CD4+)细胞占48%。2周时为25%。随着治疗时间的延长，CD4/8比值逐渐降低，不能用于晚期骨肉瘤患者的过继免疫治疗。

Osteosarcoma patients who had failed standard therapy and had multiple metastatic disease seems to be intractable. Rosenberg et al, have recently reported that combined therapy with LAK cells and IL-2 was capable of causing the regression of established metastatic cancer. In prior murine model, it is said that the therapeutic effect was directly related to the number of LAK cells and amount of IL-2 adminstered. In order to apply this adoptive immunotherapy to the advanced osteosarcoma patients, at first we have studied methods for in vitro expansion of LAK cells effective in immunotherapy.A hollow fiber biorcactor system (Cellmax 100) was used and its efficacy was investigated in LAK cell proliferatioin, PBL obtained from 20 ml of peripheral blood were cultured in an anti-CD 3 antibody coated flask for 7 days. About 2-4 *10 cells were harvested.These LAK cells were transfered to the hollow fiber bioreacter and cultured for further 2 weeks. The number of LAK cells did not increase in hollow fiber bioreactor. In their surface marker. Leu 4a (CD3+) cells were 98-99% at both 1 and 2 weeks. Leu 2a (CD8+) cells were 55% at 1 week, 74% at 2 weeks. Leu 3a (CD4+) cells were 48% at 1 week. 25% at 2 weeks. The ration of CD4/8 was decreased with time.We could not apply this adoptive immunotherapy to advanced osteosarcoma patients.