Development of Culture system for LAK cells and its application to adoptive immunotheraphy for lung cancer.

用于癌症的外科辅助治疗。我们用淋巴激活杀伤细胞(LAK)和重组白介素2(IL-2)设计了一种独特的过继免疫治疗方案。该方案的特点是长期(连续6天)皮下注射低剂量的IL-2，并从手术时获得的区域淋巴结淋巴细胞中转移体外产生的LAK细胞。按照这个养生法。19例病理分期为1的肺癌患者在肺叶切除后约2周接受免疫治疗。临床毒性包括发热。疲惫。&lt；5%的体重增加。所有毒副作用均在治疗结束后4天内逆转。13例。治疗后24小时获取的外周血淋巴细胞具有体外LAK活性。因此，就患者的术后状态而言，该方案被认为具有良好的耐受性和免疫活性，以改善LAK细胞的常规生成和扩增性能。我们开发了一种新的培养系统(LIFROC充液旋转培养：正式命名为JCC设备)。LIFROC系统基本上是基于透析培养的。透析膜将培养容器与基础培养基外循环分开。通过旋转培养容器使细胞得到适当的分散，并通过硅管向培养容器提供氧气。细胞密度最高可达2.7×10~(-7)个/毫升，细胞存活率大于90%，LAK活性较高。与传统的分批培养系统相比，采用组织培养皿。LIFROC系统能够减少基础培养基用量。IL-2。从培养液中回收LAK细胞的培养容器、容器和工具都是一次性的。该培养体系可用于LAK细胞的安全培养和过继免疫治疗。

For application with surgical adjuvant therapy of cancer. we have designed a unique regimen of adopuve immunotherapy with lymphokineacuvated killer (LAK) cells and recombinant interleukin-2 (IL-2). The regimen features the prolonged (6 consecutive days) subcutaneous administration of a low-dose IL-2 and the transfer of ex vivo generated LAK cells from regional lymph node lymphocytes, obtained at the time of operations. According to this regimen. 19 patients with pathological stage-1 lung cancer received immunotheraph about 2 weeks after pulmonary lobectomy. Clinical toxicities included fever. fatigue. <5% weight gain. and edema.All toxicities reversed within 4 days after the complection of therapy. In 13 patients. peripheral blood lymphecytes obtained 24h after therapy exhibited in vitro LAK activity. Thus the regimen is considered to be well-tolerable and immunologically active, in regard to the postoperative state of the patients.To improve the routine performance of the generation and expansion of LAK cells. we have developed a new culture system (LIFROC Liquid-Filled Rotary Culture : formaly designated JCC device). The LIFROC system is essentially based on dialysis culture. A dialysis membrane divides the culture vessel from the outside circulation of basal medium. The culture vessel is rotated for the proper dispersion of cells and supplied with oxygen through a silicon tube.With this LIFROC system. the cell density reached a maximum 2.7x10^7 cells/ml with greater than 90% viability and a substantial level of LAK activity. In comparison with a convenstonal batchwise culture system suing tissue culture dishes. the LIFROC system was able to reduce the consumption of basal medium. IL-2. and serum by 20%, 84%, and96%, respectively.Culture vessel, reservoir, and tools for the recovery of LAK cells from culture medium were devised to be disposable. This culture system is apphed with safe to culture LAK cells for adoptive immunotheraphy.