Wang Chaojie, Delcros Jean-Guy, Biggerstaff John, Phanstiel Otto
Groupe de Recherche en Therapeutique Anticancéreuse, Faculté de Médecine, 2, Avenue du Professeur Léon Bernard, 35043 Rennes, France.
J Med Chem. 2003 Jun 19;46(13):2672-82. doi: 10.1021/jm020598g.
A series of nine N(1)-(9-anthracenylmethyl)tetraamines (e.g., Ant-4,4,4-tetraamine) were synthesized and evaluated for cytotoxicity in L1210, alpha-difluoromethylornithine (DFMO)-treated L1210, Chinese hamster ovary (CHO), and CHO-MG cell lines. Surprisingly, the 3,3,4- and 3,4,3-tetraamine motifs had the same or decreased cytotoxicity in DFMO-treated L1210 cells, whereas the rest of the tetraamine systems were usually more cytotoxic and gave lower IC(50) values in this treated cell line. The most sensitive derivatives to DFMO treatment were the Ant-4,4,3- and Ant-4,4,4-tetraamine analogues, which were 7 and 5 times more cytotoxic in DFMO-treated L1210 cells, respectively. K(i) values for each of the anthracenylmethyl(Ant)-polyamine conjugates were determined in L1210 cells and revealed that these systems are high-affinity ligands for the polyamine transporter (PAT). Mixed results were observed in the CHO and CHO-MG assays. The 4,4,4- and 5,4,4-tetraamine motifs were 3 times more toxic to CHO cells with active polyamine transporters. For example, the Ant-4,4,4-tetraamine conjugate displayed IC(50) values of 11 microM in CHO cells and 33 microM in CHO-MG cells, a PAT-deficient cell line. This suggested that these derivatives used the PAT in part to access cells. However, most of the other tetraamine derivatives had similar potencies in both the CHO and CHO-MG cell lines. In terms of vector design, higher affinity for the PAT (lower K(i) values) did not translate into higher potency for the tetraamine conjugate. In contrast, the related triamine systems, which had micromolar K(i) values in L1210 cells, were more efficacious and selective. In one case, the 4,4-triamine motif imparted 150-fold higher potency in CHO cells than the CHO-MG mutant. A deconvolution microscopy study in A375 melanoma cells revealed a rapid internalization of the Ant-4,4-triamine as fluorescent vesicles, whereas the Ant-4,4,4-tetraamine remained mostly at the cell surface. These findings help define the key characteristics required for selective delivery of polyamine-drug conjugates into cell types with active polyamine transporters.
合成了一系列9种N(1)-(9-蒽甲基)四胺（如Ant-4,4,4-四胺），并在L1210、α-二氟甲基鸟氨酸（DFMO）处理的L1210、中国仓鼠卵巢（CHO）和CHO-MG细胞系中评估了它们的细胞毒性。令人惊讶的是，在DFMO处理的L1210细胞中，3,3,4-和3,4,3-四胺基序具有相同或降低的细胞毒性，而其余的四胺系统通常具有更高的细胞毒性，并且在该处理的细胞系中给出更低的半数抑制浓度（IC(50)）值。对DFMO处理最敏感的衍生物是Ant-4,4,3-和Ant-4,4,4-四胺类似物，它们在DFMO处理的L1210细胞中的细胞毒性分别高7倍和5倍。测定了每种蒽甲基(Ant)-多胺缀合物在L1210细胞中的抑制常数（K(i)）值，结果表明这些系统是多胺转运蛋白（PAT）的高亲和力配体。在CHO和CHO-MG测定中观察到了混合结果。4,4,4-和5,4,4-四胺基序对具有活性多胺转运蛋白的CHO细胞的毒性高3倍。例如，Ant-4,4,4-四胺缀合物在CHO细胞中的IC(50)值为11微摩尔，在PAT缺陷细胞系CHO-MG细胞中的IC(50)值为33微摩尔。这表明这些衍生物部分通过PAT进入细胞。然而，大多数其他四胺衍生物在CHO和CHO-MG细胞系中的效力相似。在载体设计方面，对PAT的更高亲和力（更低的K(i)值）并没有转化为四胺缀合物的更高效力。相比之下，相关的三胺系统在L1210细胞中的K(i)值为微摩尔级别，更有效且更具选择性。在一个案例中，4,4-三胺基序在CHO细胞中的效力比CHO-MG突变体高150倍。在A375黑色素瘤细胞中进行的去卷积显微镜研究表明，Ant-4,4-三胺作为荧光囊泡迅速内化，而Ant-4,4,4-四胺大多保留在细胞表面。这些发现有助于确定将多胺-药物缀合物选择性递送至具有活性多胺转运蛋白的细胞类型所需的关键特征。
