Catapano C V, Dayton J S, Mitchell B S, Fernandes D J
Department of Experimental Oncology, Hollings Cancer Center, Medical University of South Carolina, Charleston 29425, USA.
Mol Pharmacol. 1995 May;47(5):948-55.
Inhibitors of IMP dehydrogenase (EC 1.2.1.14), including mizoribine (Bredinin) and mycophenolic acid, have significant antitumor and immunosuppressive activities. Studies were aimed at determining the mechanism by which intracellular GTP depletion induced by these agents results in inhibition of DNA synthesis. Incubation of human CEM leukemia cells for 2 hr with IC50 concentrations of either mizoribine (4 microM) or mycophenolic acid (0.5 microM) reduced cellular GTP levels an average of 68% or 58%, respectively, compared with the levels in control cells. Under similar conditions, mizoribine and mycophenolic acid decreased the amount of [3H]adenosine incorporated into primer RNA by 75% and 70%, respectively, relative to the untreated controls, but had no significant effect on total RNA synthesis. Repletion of the guanine nucleotide pools by coincubation of CEM cells with guanosine plus 8-aminoguanosine prevented both the inhibition of primer RNA synthesis and the inhibition of tumor cell growth induced by these agents. Additional studies demonstrated that GTP depletion alone was capable of directly inducing inhibition of primer RNA synthesis. Primer RNA synthesis was inhibited an average of 84% in whole-cell lysates that lacked GTP but contained all remaining ribo- and deoxyribonucleoside triphosphates. On an M13 DNA template, RNA-primed DNA synthesis catalyzed by the purified complex of DNA primase (EC 2.7.7.6) and DNA polymerase alpha (EC 2.7.7.7) was decreased an average of 70% in the absence of GTP, compared with synthesis in the presence of 0.5 mM GTP. These results provide evidence that mizoribine and mycophenolic acid inhibit DNA replication by inducing GTP depletion, which suppresses the synthesis of RNA-primed DNA intermediates.
肌苷5'-单磷酸脱氢酶（EC 1.2.1.14）抑制剂，包括咪唑立宾（布累迪宁）和霉酚酸，具有显著的抗肿瘤和免疫抑制活性。研究旨在确定这些药物诱导的细胞内GTP耗竭导致DNA合成抑制的机制。用IC50浓度的咪唑立宾（4 microM）或霉酚酸（0.5 microM）将人CEM白血病细胞孵育2小时，与对照细胞相比，细胞内GTP水平分别平均降低了68%或58%。在类似条件下，相对于未处理的对照，咪唑立宾和霉酚酸分别使掺入引物RNA中的[3H]腺苷量减少了75%和70%，但对总RNA合成没有显著影响。通过将CEM细胞与鸟苷加8-氨基鸟苷共同孵育来补充鸟嘌呤核苷酸库，可防止这些药物诱导的引物RNA合成抑制和肿瘤细胞生长抑制。进一步的研究表明，仅GTP耗竭就能直接诱导引物RNA合成的抑制。在缺乏GTP但含有所有其余核糖和脱氧核糖核苷三磷酸的全细胞裂解物中，引物RNA合成平均被抑制了84%。在M13 DNA模板上，与存在0.5 mM GTP时的合成相比，在没有GTP的情况下，由纯化的DNA引发酶（EC 2.7.7.6）和DNA聚合酶α（EC 2.7.7.7）复合物催化的RNA引发的DNA合成平均降低了70%。这些结果证明，咪唑立宾和霉酚酸通过诱导GTP耗竭来抑制DNA复制，从而抑制RNA引发的DNA中间体的合成。
