Lack of impact of the loss of constitutive folate receptor α expression, achieved by RNA interference, on the activity of the new generation antifolate pemetrexed in HeLa cells
Lack of impact of the loss of constitutive folate receptor α expression, achieved by RNA interference, on the activity of the new generation antifolate pemetrexed in HeLa cells
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DOI：
10.1158/1078-0432.ccr-04-1225
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发表时间：
2004-12-01
影响因子：
11.5
通讯作者：
Goldman, ID
中科院分区：
文献类型：
--
作者：
Chattopadhyay, S;Wang, YH;Goldman, ID
培美曲塞[PMX（Alimta）]是新一代抗叶酸剂，对多种实体瘤具有活性。它是大多数叶酸转运蛋白的优良底物，特别是还原叶酸载体（RFC）和叶酸受体（FR）-α。FR-α在PMX药理活性中的作用尚不确定。虽然高水平表达可能会增强这种代理商的活动，它是不清楚什么样的角色组成型水平的这种转运蛋白有助于PMX活动。在这项研究中，FR-α表达的组成性水平被取消的小干扰RNA诱导的沉默在HeLa细胞和RFC-null HeLa R5细胞证实的北方印迹，免疫组织化学和细胞表面结合。在不存在FR-α表达的情况下，HeLa和R5细胞中PMX生长抑制未发生变化。FR-α表达的丧失并没有减少野生型或RFC缺失HeLa细胞中PMX的净积累。同样地，野生型HeLa细胞中的叶酸池不因FR-α基因沉默而减少，并且在用5-甲酰四氢叶酸生长的RFC-无效R5亚系中受到的影响可忽略不计。FR-α在HeLa细胞中的表面结合显示出比在多种其他人实体瘤细胞系中的更大。因此，在HeLa细胞中组成型表达的FR-α在RFC功能存在或不存在的情况下对PMX活性没有贡献。这可能是许多人实体瘤细胞系的情况。
Pemetrexed [PMX (Alimta)] is a new generation antifolate with activity in a variety of solid tumors. It is an excellent substrate for most folate transporters, notably the reduced folate carrier (RFC) and folate receptor (FR)-alpha. The role of FR-alpha in PMX pharmacological activity is uncertain. Whereas high-level expression may enhance the activity of this agent, it is not clear what role constitutive levels of this transporter contribute to PMX activity. In this study, constitutive levels of FR-alpha expression were abolished by small interfering RNA-induced silencing in HeLa cells and RFC-null HeLa R5 cells as confirmed by Northern blotting, immunohistochemistry, and cell surface binding. PMX growth inhibition was unchanged in HeLa and R5 cells in the absence of FR-alpha expression. Loss of FR-alpha expression did not decrease net accumulation of PMX in either wildtype or RFC-null HeLa cells. Likewise, folate pools in wildtype HeLa cells were not decreased by FR-alpha gene silencing and were negligibly affected in the RFC-null R5 subline grown with 5-formyltetrahydrofolate. FR-alpha surface binding in HeLa cells was shown to be greater than that in a variety of other human solid tumor cell lines. Hence, constitutively expressed FR-a in HeLa cells does not contribute to PMX activity in the presence or absence of RFC function. This is likely the case in many human solid tumor cell lines.