Hofmann A, Blau H M
Department of Molecular Pharmacology, Stanford University School of Medicine, California 94305, USA.
Somat Cell Mol Genet. 1997 Jul;23(4):249-57. doi: 10.1007/BF02674416.
Anticancer therapy for solid tumors suffers from inadequate methods for the localized administration of cytotoxic agents. Fas ligand (FasL) has been reported to be cytotoxic to a variety of cells, including certain tumor cell lines. We therefore postulated that myoblasts could serve as non-transformed gene therapy vehicles for the continuous localized delivery of cytotoxic anticancer agents such as FasL. However, contrary to previous reports, fluorescence activated cell sorting (FACS) analyses revealed that both primary mouse and human myoblasts express Fas, the receptor for FasL. To avoid self-destruction and test the cytotoxic potential of myoblasts, the cells were isolated from mice deficient in Fas (lpr/lpr), the mouse counterpart of human autoimmune lymphoproliferative syndrome (ALPS). These primary mouse myoblasts were transduced with a retroviral vector encoding mouse FasL and expression of a biologically active and soluble form of the molecule was confirmed by the apoptotic demise of cocultured Fas-expressing Jurkat cells, the standard in the field. To test whether the lpr myoblasts expressing FasL could be used in anticancer therapy, human rhabdomyosarcoma derived cell lines were assayed for Fas and then tested in the apoptosis coculture assay. The majority of Fas-expressing muscle tumor cells were rapidly killed. Moreover, FasL expressing myoblasts were remarkably potent; indeed well characterized cytotoxic antibodies to Fas were only 20% as efficient at killing rhabdomyosarcoma cells as FasL expressing myoblasts. These findings together with previous findings suggest that primary myoblasts, defective in Fas but genetically engineered to express FasL, could function as potent anticancer agents for use in the localized destruction of solid tumors in vivo by three synergistic mechanisms: (1) directly via Fas/FasL mediated apoptosis, (2) indirectly via neutrophil infiltration and immunodestruction, and (3) as allogeneic inducers of a bystander effect via B and T cells.
实体瘤的抗癌治疗存在细胞毒性药物局部给药方法不足的问题。据报道，Fas配体（FasL）对包括某些肿瘤细胞系在内的多种细胞具有细胞毒性。因此，我们推测成肌细胞可作为未转化的基因治疗载体，用于持续局部递送细胞毒性抗癌药物，如FasL。然而，与之前的报道相反，荧光激活细胞分选（FACS）分析显示，原代小鼠和人类成肌细胞均表达FasL的受体Fas。为避免自我破坏并测试成肌细胞的细胞毒性潜力，从缺乏Fas的小鼠（lpr/lpr）中分离细胞，lpr/lpr是人类自身免疫性淋巴细胞增生综合征（ALPS）的小鼠对应物。这些原代小鼠成肌细胞用编码小鼠FasL的逆转录病毒载体进行转导，通过共培养的表达Fas的Jurkat细胞（该领域的标准）的凋亡死亡证实了该分子生物活性和可溶性形式的表达。为了测试表达FasL的lpr成肌细胞是否可用于抗癌治疗，对源自人横纹肌肉瘤的细胞系进行Fas检测，然后在凋亡共培养试验中进行测试。大多数表达Fas的肌肉肿瘤细胞迅速死亡。此外，表达FasL的成肌细胞非常有效；事实上，特征明确的抗Fas细胞毒性抗体在杀死横纹肌肉瘤细胞方面的效率仅为表达FasL的成肌细胞的20%。这些发现与之前的发现共同表明，Fas有缺陷但经过基因工程改造以表达FasL的原代成肌细胞可作为有效的抗癌药物，通过三种协同机制在体内局部破坏实体瘤：（1）直接通过Fas/FasL介导的凋亡，（2）间接通过中性粒细胞浸润和免疫破坏，以及（3）作为通过B细胞和T细胞产生旁观者效应的同种异体诱导剂。
