Hochreiter R, Weiger T M, Colombatto S, Langer T, Thomas T J, Cabella C, Heidegger W, Grillo M A, Hermann A
Department of Molecular Neurobiology and Cellular Physiology, Institute of Zoology, University of Salzburg, Austria.
Naunyn Schmiedebergs Arch Pharmacol. 2000 Mar;361(3):235-46. doi: 10.1007/s002109900188.
A series of diamines with the general structure NH2(CH2)xNH2, x=2-12, was tested for their potential effects on cell proliferation of cultured rat C6 glioma cells in comparison to natural polyamines. Long chain diamines reduced cell number after 48 h in culture with a sequence of 1,12-diaminododecane (1,12-DD) >1,10-diaminodecane >1,9-diaminononane. Polyamines (putrescine, spermidine and spermine) as well as diamines up to a CH2-chain length of x=8 were found to be ineffective. The spermine analogue 1,12-DD was the most effective molecule in reducing cell number in an irreversible, dose-dependent manner (EC50=3 microM under serum-free conditions). In further experiments we investigated the mechanisms of action of 1,12-DD. The compound had only a minor effect on cell cycle and did not affect free internal calcium concentration. Under physiological conditions 1,12-DD interacts with triplex DNA but not with duplex DNA. Ornithine decarboxylase activity as well as the concentration of internal polyamines were found to be reduced by 1,12-DD. Polyamine application, however, was not able to reverse the effect of 1,12-DD, indicating a polyamine-independent or non-competitive mechanism of action. 1,12-DD reduced cell number by induction of apoptosis as well as necrosis. In molecular modeling studies it was found that a minimal hydrophobic intersegment of at least 4 A was required to make a diamine an effective drug in respect to cellular growth. A hydrophobic gap of this size fits the minimum requirement expected from molecular modeling to provide space for hydrophobic interactions with parts of proteins like a CH3-group. Our results show that 1,12-DD acts as a potent drug, reducing the number of C6 glioma cells, and suggest that its spatial and hydrophobic properties are responsible for its mechanism of action.
为了研究一系列通式为NH2(CH2)xNH2（x = 2 - 12）的二胺与天然多胺相比，对培养的大鼠C6胶质瘤细胞增殖的潜在影响，我们进行了实验。长链二胺在培养48小时后可减少细胞数量，其作用顺序为1,12 - 二氨基十二烷（1,12 - DD）> 1,10 - 二氨基癸烷> 1,9 - 二氨基壬烷。多胺（腐胺、亚精胺和精胺）以及碳链长度x = 8以下的二胺均无效。精胺类似物1,12 - DD是最有效的分子，它以不可逆的剂量依赖性方式减少细胞数量（无血清条件下的EC50 = 3 microM）。在进一步的实验中，我们研究了1,12 - DD的作用机制。该化合物对细胞周期影响较小，且不影响细胞内游离钙浓度。在生理条件下，1,12 - DD与三链DNA相互作用，但不与双链DNA相互作用。发现1,12 - DD可降低鸟氨酸脱羧酶活性以及细胞内多胺浓度。然而，多胺的应用并不能逆转1,12 - DD的作用，这表明其作用机制与多胺无关或为非竞争性机制。1,12 - DD通过诱导凋亡和坏死来减少细胞数量。在分子建模研究中发现，二胺要成为影响细胞生长的有效药物，至少需要4 Å的最小疏水片段。这种大小的疏水间隙符合分子建模预期的最小要求，可为与蛋白质部分（如甲基基团）的疏水相互作用提供空间。我们的结果表明，1,12 - DD作为一种强效药物，可减少C6胶质瘤细胞数量，并表明其空间和疏水特性决定了其作用机制。
