Retroviral vectors containing a variant dihydrofolate reductase gene for drug protection and in vivo selection of hematopoietic cells.
Retroviral vectors containing a variant dihydrofolate reductase gene for drug protection and in vivo selection of hematopoietic cells.
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含有变体二氢叶酸还原酶基因的逆转录病毒载体，用于药物保护和体内造血细胞选择。
DOI：
10.1002/stem.5530160827
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发表时间：
1998
期刊：
影响因子：
--
通讯作者：
Sorrentino,BP
中科院分区：
文献类型：
--
作者：
Allay,JA;Galipeau,J;Blakley,RL;Sorrentino,BP
正在研究将耐药基因转移到造血细胞作为防止与癌症化疗相关的骨髓抑制的手段，以及作为体内选择和扩增遗传修饰细胞的策略。本研究的目的是测试逆转录病毒介导的二氢叶酸还原酶（DHFR）变体（L22 Y）的基因转移是否可用于体内选择转导的骨髓细胞，并确定保护免受骨髓抑制所需的转导细胞比例。基于先前的工作表明，用抗叶酸剂进行选择也可能需要抑制核苷转运机制，移植有DHFR转导的骨髓细胞的小鼠用曲美曲沙和核苷转运抑制剂前药硝基苄基巯基嘌呤核苷磷酸进行治疗。在药物治疗后，在骨髓和循环成熟外周血细胞中观察到转导的髓系祖细胞的体内选择。这些结果表明，L22 Y-DHFR cDNA、曲美蝶呤和硝基苄基巯基嘌呤核苷磷酸的新组合可用于选择转导的骨髓细胞，并且这种方法值得在大型动物模型中进一步研究。使用含有人CD 24报告基因的双顺反子载体来确定化学保护所需的修饰细胞的数量。当超过10%的骨髓细胞表达载体时，可以看到部分保护作用，防止中性粒细胞减少，当比例超过30%时，可以获得高水平的保护作用。这些结果表明，基因转移可能对某些儿科癌症的骨髓保护有用，但将这种方法应用于成人癌症患者需要更有效的基因转移。
Transfer of drug resistance genes to hematopoietic cells is being studied as a means to protect against the myelosuppression associated with cancer chemotherapy and as a strategy for the in vivo selection and amplification of genetically modified cells. The goal of this study was to test if retroviral‐mediated gene transfer of a dihydrofolate reductase (DHFR) variant (L22Y) could be used for in vivo selection of transduced myeloid cells and to determine what proportion of transduced cells was required for protection from myelosuppression. Based on previous work suggesting that selection with antifolates may also require inhibition of nucleoside transport mechanisms, mice transplanted with DHFR‐transduced bone marrow cells were treated with trimetrexate and the nucleoside transport inhibitor prodrug nitrobenzylmercaptopurine riboside phosphate. In vivo selection of transduced myeloid progenitors was seen in the bone marrow and in circulating mature peripheral blood cells following drug treatment. These results show that the novel combination of the L22Y‐DHFR cDNA, trimetrexate and nitrobenzylmercaptopurine riboside phosphate can be used to select for transduced myeloid cells, and that this approach warrants further study in large animal models. A bicistronic vector containing a human CD24 reporter gene was used to determine the number of modified cells needed for chemoprotection. Partial protection from neutropenia was seen when greater than 10% of myeloid cells expressed the vector, and high levels of protection were obtained when the proportion exceeded 30%. These results suggest that gene transfer may be useful for myeloprotection in certain pediatric cancers, but that more efficient gene transfer will be required to apply this approach to adult cancer patients.