Treatment of rat C6 meningeal dissemination model by intrathecal injection o frecomvinant soluble FasL

Fas(CD95)配体(FasL)通过与Fas结合，具有诱导Fas表达的胶质瘤细胞凋亡的能力：有几种分子被设计为治疗用的可溶性Fas配体。大多数只具有Fas配体胞外区的Fas配体(可溶性FasL)由于生物活性不足或生产效率降低而不能实际使用。我们。成功构建了可溶性嵌合体。通过将用于Se IF-齐聚的异亮氨酸拉链基序和标志序列融合到人Fas配体的胞外区(FIZ-shFasL)。FIZ-shFasL对Jurkat细胞的杀伤作用与膜结合型FasL相当，比激动型抗Fas抗体(CH-11)强10倍左右。用流式细胞仪检测细胞Fas和Bcl2的表达，并与FIZ-shFasL的敏感性进行比较。流式细胞仪分析表明，Fas在人脑肿瘤细胞系中的差异表达与FIZ-shFasL诱导的细胞凋亡水平部分相关。静脉注射FIZ-shFasL 1.0 mg/kg可致大鼠死亡。鞘内注射FIZ-shFasL可成功治疗Fas阳性的C6胶质瘤细胞所致的大鼠脑膜扩散模型，且无明显不良反应。这些结果提示鞘内注射FIZ-shFasL可能在不久的将来应用于软脑膜胶质瘤病的治疗。

Fas (CD95) ligand (FasL) has the ability to induce apoptosis in Fas-expressing glioma cells by binding to with Fas: Several molecular species have been designed to be soluble Fas ligands for therapeutic purposes. Most of them possessing just only extracellular domain of Fas ligand (soluble FasL) cannot be used practically due to either insufficient biological activity or reduced productivity. We. successfully constructed a chimeric soluble. FasL by fusing a isoleucine zipper motif for se If-oligomerization and a FLAG sequence to the extracellular domain of the human Fas ligand (FIZ-shFasL). The cytotoxic effect of FIZ-shFasL on Jurkat cells was equivalent to that of membrane-bound FasL and approximately ten-fold stronger than that of agonistic anti-Fas antibody (CH-11). Expression of Fas and Bcl-2 on cell lines was determined using flow cytometry and compared with sensitivity to FIZ-shFasL. Flowcytometric analysis demonstrated that the differential Fas expression of human brain tumor cell lines partially correlated with levels of apoptosis through FIZ-shFasL. The intravenous administration of 1.0mg/kg of FIZ -shFasL resulted in a lethal effect in rats. A rat meningeal dissemination model with rat Fas-expressing C6 glioma cells can be treated successfully by intrathecal FIZ-shFasL administration without any apparent adverse effects. These results suggest that intrathecal administration of FIZ-shFasL maybe applied to the treatment of patients with leptomeningeal gliomatosis in the near future.