Mariano Thomas M, Vetrano Anna M, Gentile Shannon L, Heck Diane E, Whittemore Marilyn S, Guillon Christophe D, Jabin Ivan, Rapp Robert D, Heindel Ned D, Laskin Jeffrey D
Department of Environmental and Community Medicine, UMDNJ-Robert Wood Johnson Medical School, 675 Hoes Lane, Piscataway, NJ 08854, USA.
Biochem Pharmacol. 2002 Jan 1;63(1):31-9. doi: 10.1016/s0006-2952(01)00855-3.
Psoralens such as 8-methoxypsoralen and 4,5',8-trimethylpsoralen (TMP) are used in photochemotherapy for the treatment of a variety of epidermal proliferative diseases. Sequential treatments of the skin with psoralens plus ultraviolet light in the range of 320-400 nm (UVA light), referred to as PUVA therapy, results in the suppression of abnormal keratinocyte growth. With the recognition that the psoralens are phototoxic and carcinogenic, presumably due to their ability to intercalate into DNA and photo cross-link pyrimidine bases following UVA light activation, it is clear that the development of biologically active analogs lacking this activity would be of significant therapeutic benefit. Towards this goal we have characterized active 4'- and 5'-pyridinium derivatives of 4',5'-dihydro-TMP (H2TMP), a psoralen analog that does not form DNA cross-links. These analogs, which are charged at physiological pH and cannot penetrate cells, are unique in that they retain biological activity as inhibitors of keratinocyte cell growth when activated by UVA light. However, they do not appear to cross-link or damage DNA as determined by plasmid DNA unwinding and nicking experiments, in intact cells using fluorescent analysis of DNA unwinding assays, and by thymidine uptake studies. Reverse transcription-polymerase chain reaction and western blotting demonstrated that, unlike TMP and H2TMP, when activated by UVA light, the pyridinium derivatives were not inhibitors of transcription since interferon-gamma-inducible nitric oxide synthase mRNA and protein in the keratinocytes were unaffected. Taken together, our data suggest that uptake of the compounds by the cells and DNA cross-link formation are not required for growth inhibition. These findings further support the model that the cell membrane is an important target for the psoralens.
补骨脂素如8-甲氧基补骨脂素和4,5',8-三甲基补骨脂素（TMP）用于光化学疗法治疗多种表皮增生性疾病。用补骨脂素加320 - 400纳米范围内的紫外线（UVA光）对皮肤进行序贯治疗，即PUVA疗法，可抑制异常角质形成细胞的生长。鉴于补骨脂素具有光毒性和致癌性，推测这是由于它们能够插入DNA并在UVA光激活后使嘧啶碱基发生光交联，显然开发缺乏这种活性的生物活性类似物将具有显著的治疗益处。为了实现这一目标，我们对4',5'-二氢-TMP（H2TMP）的活性4'-和5'-吡啶鎓衍生物进行了表征，H2TMP是一种不形成DNA交联的补骨脂素类似物。这些类似物在生理pH下带电荷且不能穿透细胞，其独特之处在于当被UVA光激活时，它们作为角质形成细胞生长抑制剂保留生物活性。然而，通过质粒DNA解旋和切口实验、在完整细胞中使用DNA解旋测定的荧光分析以及胸苷摄取研究确定，它们似乎不会交联或损伤DNA。逆转录-聚合酶链反应和蛋白质印迹表明，与TMP和H2TMP不同，当被UVA光激活时，吡啶鎓衍生物不是转录抑制剂，因为角质形成细胞中干扰素-γ诱导型一氧化氮合酶的mRNA和蛋白质不受影响。综上所述，我们的数据表明细胞对化合物的摄取和DNA交联形成不是生长抑制所必需的。这些发现进一步支持了细胞膜是补骨脂素重要靶点的模型。
