Belldegrun A, Tso C L, Sakata T, Duckett T, Brunda M J, Barsky S H, Chai J, Kaboo R, Lavey R S, McBride W H
Department of Surgery, UCLA School of Medicine 90024.
J Natl Cancer Inst. 1993 Feb 3;85(3):207-16. doi: 10.1093/jnci/85.3.207.
Combination therapy with systemically administered interleukin-2 (IL-2) and interferon alpha (IFN-alpha) has resulted in long-term objective remissions in 30% of patients with metastatic renal cell carcinoma (RCC), but toxic effects are clinically significant.
We have thus investigated an alternative therapeutic approach--continuous intratumoral production of IL-2 and/or IFN-alpha by a cytokine-transfected human RCC tumor cell line.
Plasmid vectors were used to transfect the R11 RCC line with the genes for human IL-2 and/or IFN-alpha by the calcium phosphate precipitation method. Biologic characteristics of the cytokine-transfected tumor cells were determined by assays of thymidine incorporation and cytotoxicity, fluorescence-activated cell-sorter analysis, Northern blotting, and in vivo studies in C3Hf/Sed/Kam mice rendered T-cell deficient.
The transfected cell lines produced the following amounts of cytokine per 10(6) cells per day: R11-IL-2 (220 U), R11-IFN-alpha (10,240 U), and R11-IL-2 + IFN-alpha (95 U + 1270 U, respectively). Gamma irradiation did not eliminate cytokine secretion. Morphology and growth rates were identical to those for the parental R11 cell line, except for IFN-alpha-producing clones, which showed significant growth inhibition. All cytokine-producing cells demonstrated increased susceptibility to cell killing by peripheral blood leukocytes (PBL). IFN-alpha producers exhibited enhanced HLA antigen expression and suppressed c-myc messenger RNA expression; when cocultured in vitro, they induced similar changes in parental R11 cells. IL-2 producers could stimulate growth and cytotoxicity of naive (i.e., freshly isolated, uncultured) and activated PBL. All cytokine-producing cells lost their tumorigenicity, as evidenced by failure to grow in the T-cell-depleted mice. When co-injected at a local site but not at a distant site, these cells prevented growth of parental R11 cells. Histologic examination of the injection sites revealed a substantial influx of macrophages. Intraperitoneal administration of IL-2 and/or IFN-alpha could not, however, prevent growth of the parental R11 tumors.
Local production of high concentrations of IL-2 and IFN-alpha at the tumor site is more effective in preventing tumor growth than systemic administration.
Continuous local delivery of cytokines via transfer of cytokine genes into tumor cells for use as live cancer vaccines is a novel strategy for manipulation of host-mediated antitumor immune response in patients with advanced RCC.
全身给予白细胞介素-2（IL-2）和干扰素α（IFN-α）的联合疗法使30%的转移性肾细胞癌（RCC）患者获得了长期客观缓解，但毒副作用在临床上较为显著。
因此，我们研究了一种替代治疗方法——通过细胞因子转染的人RCC肿瘤细胞系在肿瘤内持续产生IL-2和/或IFN-α。
采用磷酸钙沉淀法，用质粒载体将人IL-2和/或IFN-α基因转染R11 RCC细胞系。通过胸苷掺入和细胞毒性测定、荧光激活细胞分选分析、Northern印迹以及在T细胞缺陷的C3Hf/Sed/Kam小鼠体内进行的研究，确定细胞因子转染肿瘤细胞的生物学特性。
转染后的细胞系每10⁶个细胞每天产生的细胞因子量如下：R11-IL-2（220 U）、R11-IFN-α（10240 U）和R11-IL-2 + IFN-α（分别为95 U + 1270 U）。γ射线照射并未消除细胞因子的分泌。除了产生IFN-α的克隆显示出显著的生长抑制外，其形态和生长速率与亲代R11细胞系相同。所有产生细胞因子的细胞对外周血白细胞（PBL）杀伤细胞的敏感性均增加。产生IFN-α的细胞表现出HLA抗原表达增强和c-myc信使RNA表达受抑制；在体外共培养时，它们可诱导亲代R11细胞发生类似变化。产生IL-2的细胞可刺激未致敏（即新鲜分离、未培养）和活化的PBL的生长和细胞毒性。所有产生细胞因子的细胞均失去了致瘤性，这在T细胞缺陷小鼠中未能生长得到证实。当在局部部位而非远处部位共同注射时，这些细胞可阻止亲代R11细胞的生长。注射部位的组织学检查显示巨噬细胞大量浸润。然而，腹腔内给予IL-2和/或IFN-α并不能阻止亲代R11肿瘤的生长。
在肿瘤部位局部产生高浓度的IL-2和IFN-α在预防肿瘤生长方面比全身给药更有效。
通过将细胞因子基因转移到肿瘤细胞中以持续局部递送细胞因子用作活癌症疫苗，是晚期RCC患者中操纵宿主介导的抗肿瘤免疫反应的一种新策略。
