3-AP (Triapine) is a novel inhibitor of the M2 subunit of ribonucleotide reductase (RR).

Triapine是一种强效的α-杂环羧醛硫脲(HCT)衍生物，能够抑制hRRM2与p53R2亚型的M2亚单位[1]。Triapine被认为是通过其形成的铁螯合物而非直接从活性位点移除铁来抑制核糖核苷酸还原酶。在topoisomerase IIα较少的细胞中，会产生较少的DNA链断裂，因此在K/VP.5细胞系中，topoisomerase II毒性将较低。Dp44mT对K562和K/VP.5细胞生长抑制的IC50分别为48±9 nM和60±12 nM。Triapine对K562和K/VP.5细胞生长抑制的IC50分别为476±39 nM和661±69 nM[2]。PKIH与DpT Fe螯合剂对一系列肿瘤细胞系显示出高抗增殖活性。Dp44mT显示出最大的抗肿瘤效能，其IC50范围从0.005到0.4 μM。Dp44mT在28种细胞类型上的平均IC50为0.03±0.01 μM，显著低于Triapine的平均IC50（1.41±0.37 μM）[3]。

Triapine 在以体重百分比表达的情况下显著增加脾重（增加1.7倍），达到1.02±0.06%（n=25），相比之下，对照组小鼠的脾重为0.6±0.03%（n=27）。在长期实验组中，经Dp44mT（每天0.4 mg/kg）处理后，心脏重量显著增加，达到0.8±0.06%（n=4），而对照组小鼠为0.5±0.01%（n=6）。Dp44mT 和 Triapine（每天12 mg/kg）处理的动物，肝脏中Ndrg1、TfR1和VEGF1的表达显著减少。这种表达的减少可能与Dp44mT 和 Triapine 处理的小鼠肝脏中铁含量的增加有关[3]。

Ribonucleotide reductase assay: CDP reductase is assayed using Dowex 1-borate ion-exchange chromatography. The assay mixture contains 0.02 μCi of [14C]CDP (52.9 mCi/mmol), 3 mM dithiothreitol, 6 mM MgCl2, 30 mM HEPES, 5 mM ATP, 0.15 mM unlabeled CDP, and 10 μL of cellular extract in a final volume of 0.02 mL. The incubation time for the reaction is 60 min, during which time the reaction is linear.

Triapine is dissolved in DMSO and diluted with appropriate media[2]. An MTT assay is used to determine cell growth inhibition of CHO cells. Human leukemia K562 cells and K/VP.5 cells (a 26-fold etoposide-resistant K562-derived sub-line with decreased levels of topoisomerase IIα mRNA and protein) are maintained as suspension cultures in "MEM (Minimal Essential Medium Alpha, Invitrogen) containing 10% fetal calf serum (FCS). For growth inhibition assays, K562 and K/VP.5 cells are plated at a concentration of 1.5×105 cell/mL, and incubated 5 d with various concentrations of Dp44mT, Triapine or vehicle (DMSO) for 48 h, after which cells are counted on a model ZBF Coulter counter. The IC50 growth inhibitory concentration for each cell line is calculated from a non-linear least-squares fit to a 2-parameter logistic equation[2].