HCT 116和NCI - H630人结肠癌细胞对1-β-D-阿拉伯呋喃糖基胞嘧啶敏感性的决定因素

Determinants of sensitivity to 1-beta-D-arabinofuranosylcytosine in HCT 116 and NCI-H630 human colon carcinoma cells.

作者信息

Grem J L, Geoffroy F, Politi P M, Cuddy D P, Ross D D, Nguyen D, Steinberg S M, Allegra C J

机构信息

NCI-Navy Medical Oncology Branch, Division of Cancer Treatment, National Cancer Institute, National Naval Medical Center, Bethesda, Maryland 20889, USA.

出版信息

Mol Pharmacol. 1995 Aug;48(2):305-15.

PMID:7651364

Abstract

The cytotoxicity and metabolism of 1-beta-D-arabinofuranosylcytosine (AraC) and its effects on DNA synthesis and integrity were studied in HCT 116 and NCI-H630 human colon cancer cells. In 116 cells, 0.1 microM AraC decreased colony formation by approximately 50%, whereas 1 microM was required in H630 cells. AraCTP levels after a 24-hr AraC exposure were 2.3- to 3.5-fold lower in H630 cells due to increased ability to deaminate AraCMP. AraC DNA levels increased in proportion to AraCTP pools (r = 0.99) and were 2-fold higher in 116 cells after a 24-hr exposure to 0.1 and 1 microM AraC. Although the half-life of AraCTP was < 1 hr in both lines, > 80% of AraC DNA was retained at 24 hr after drug removal. Clonogenic capacity was inversely related to the extent of AraC DNA incorporation. Interference with nascent DNA chain elongation increased with increasing AraC concentration x time. A 24-hr AraC exposure produced a dramatic shift in the elution profile of nascent DNA during a 15-hr elution at pH 12.1; these effects were greater in 116 cells (DNA retained on filter [percentage of control]): 78%, 23%, and 9% with 0.1, 1, and 10 microM AraC versus 84%, 42%, and 18% in H630 cells, respectively. The extent of nascent DNA damage was proportional to AraC DNA content. Net DNA synthesis was potently inhibited during AraC exposure in both lines. H630 cells had partial recovery of DNA synthesis at 24 hr after drug removal, whereas persistent inhibition was noted in 116 cells. A slight excess of double-strand breaks in parental DNA was detected after a 24-hr exposure to 10 microM AraC in 116 cells. The extent of DNA fragmentation was more pronounced 16 hr after drug removal and was greater in 116 cells: 8.5%, 19%, and 21% with 0.1, 1, and 10 microM AraC DNA content, magnitude of nascent DNA damage, duration of DNA synthetic inhibition, and induction of double-stranded DNA fragmentation appeared to be the crucial determinants of lethality.

摘要

在HCT 116和NCI-H630人结肠癌细胞中研究了1-β-D-阿拉伯呋喃糖基胞嘧啶（AraC）的细胞毒性、代谢及其对DNA合成和完整性的影响。在116细胞中，0.1μM AraC使集落形成减少约50%，而在H630细胞中则需要1μM。由于脱氨生成AraCMP的能力增强，H630细胞在AraC暴露24小时后的AraCTP水平低2.3至3.5倍。AraC DNA水平与AraCTP池成比例增加（r = 0.99），在116细胞中，在暴露于0.1和1μM AraC 24小时后，AraC DNA水平高2倍。尽管在这两种细胞系中AraCTP的半衰期均小于1小时，但在去除药物后24小时仍保留了超过80%的AraC DNA。克隆形成能力与AraC DNA掺入程度呈负相关。随着AraC浓度×时间增加，对新生DNA链延伸的干扰增强。在pH 12.1条件下进行15小时洗脱期间，24小时的AraC暴露使新生DNA的洗脱曲线发生显著变化；这些影响在116细胞中更大（滤膜上保留的DNA[相对于对照的百分比]）：0.1、1和10μM AraC分别为78%、23%和9%，而在H630细胞中分别为84%、42%和18%。新生DNA损伤程度与AraC DNA含量成比例。在AraC暴露期间，两种细胞系中的净DNA合成均受到强烈抑制。在去除药物后24小时，H630细胞的DNA合成有部分恢复，而在116细胞中则观察到持续抑制。在116细胞中，暴露于10μM AraC 24小时后，检测到亲本DNA中双链断裂略有增加。在去除药物后16小时，DNA片段化程度更明显，且在116细胞中更大：0.1、1和10μM AraC时分别为8.5%、19%和21%。DNA含量、新生DNA损伤程度、DNA合成抑制持续时间以及双链DNA片段化的诱导似乎是致死性的关键决定因素。