Beckwith M, Urba W J, Longo D L
Biological Carcinogenesis and Development Program, Program Resources Inc./DynCorp Inc., Frederick, Md.
J Natl Cancer Inst. 1993 Mar 17;85(6):483-8. doi: 10.1093/jnci/85.6.483.
Dolastatins 10 and 15 are small peptides isolated from the marine sea hare Dolabella auricularia. In vitro studies of these peptides have demonstrated antimitotic and antiproliferative activity and growth inhibition in hematopoietic progenitor cells.
The purpose of our in vitro study was to determine the biological effects of these marine peptides on growth of human lymphoma cell lines and to investigate mechanisms by which the dolastatins may act.
Cell lines DB, HT, RL, and SR were grown from the ascites or pleural effusion of four patients with lymphoma. The DB, HT, and RL cell lines are of B-cell origin, and the SR cell line appears to be a less differentiated lymphoid cell type. Cells from these lines were cultured in the presence of vincristine or dolastatin 10 or 15. [3H]Thymidine-uptake assays were used to measure effects on DNA synthesis. Cell cycle analysis using propidium iodide was performed to measure drug-induced cell-cycle arrest. DNA fragmentation was used as an assay for drug-induced apoptosis and was measured by agarose gel electrophoresis.
In the three B cell lines, dolastatin 10 was more effective than dolastatin 15. Values for concentrations required for inhibition of proliferation by 50% (IC50) were .00013-.0013 nM for dolastatin 10 in each cell line; values for dolastatin 15 were approximately .13 nM in DB and HT cells and .0013-.013 nM in RL cells. SR cells were more sensitive to dolastatin 15 than to dolastatin 10 (IC50 = .00013-.0013 nM versus .0013-.013 nM). Both dolastatins arrested more than 70% of cells in mitosis in all cell lines. This effect was reversed if the drug was removed by 4 hours, but by 8 hours of exposure, reversal was not possible. Both dolastatins 10 and 15 produced apoptosis in DB and HT cells but not in the other two cell lines.
We have demonstrated that dolastatins 10 and 15 have a profound antiproliferative effect on four different human lymphoma cell lines and that the dolastatins are approximately 3-4 logarithms more effective as antiproliferative compounds, on a molar basis, than vincristine--a clinically useful, antiproliferative agent. These data support the hypothesis that apoptosis, as measured by DNA fragmentation, appears to be a cell-specific response and may not be directly related to the antimitotic effect of the dolostatins.
Our results suggest that these compounds may be good candidates for development as antineoplastic agents.
多拉司他汀10和15是从海洋海兔耳状多肋海蛞蝓中分离出的小肽。对这些肽的体外研究已证明其具有抗有丝分裂和抗增殖活性以及对造血祖细胞的生长抑制作用。
我们体外研究的目的是确定这些海洋肽对人淋巴瘤细胞系生长的生物学效应，并研究多拉司他汀可能起作用的机制。
从4例淋巴瘤患者的腹水或胸腔积液中培养出细胞系DB、HT、RL和SR。DB、HT和RL细胞系起源于B细胞，而SR细胞系似乎是一种分化程度较低的淋巴细胞类型。将这些细胞系的细胞在长春新碱、多拉司他汀10或15存在的情况下进行培养。采用[3H]胸腺嘧啶核苷摄取试验来测量对DNA合成的影响。使用碘化丙啶进行细胞周期分析以测量药物诱导的细胞周期停滞。DNA片段化用作药物诱导凋亡的检测方法，并通过琼脂糖凝胶电泳进行测量。
在三个B细胞系中，多拉司他汀10比多拉司他汀15更有效。在每个细胞系中，多拉司他汀10抑制增殖50%（IC50）所需的浓度值为0.00013 - 0.0013 nM；多拉司他汀15在DB和HT细胞中的值约为0.13 nM，在RL细胞中的值为0.0013 - 0.013 nM。SR细胞对多拉司他汀15比对多拉司他汀10更敏感（IC50 = 0.00013 - 0.0013 nM对0.0013 - 0.013 nM）。两种多拉司他汀在所有细胞系中均使超过7�%的细胞停滞在有丝分裂期。如果在4小时内去除药物，这种效应可以逆转，但在暴露8小时后，逆转则不可能。多拉司他汀10和15在DB和HT细胞中均诱导凋亡，但在其他两个细胞系中未诱导凋亡。
我们已经证明，多拉司他汀10和15对四种不同的人淋巴瘤细胞系具有显著的抗增殖作用，并且在摩尔基础上，多拉司他汀作为抗增殖化合物比长春新碱（一种临床上有用的抗增殖剂）有效约3 - 4个对数。这些数据支持这样的假设，即通过DNA片段化测量的凋亡似乎是一种细胞特异性反应，可能与多拉司他汀的抗有丝分裂作用没有直接关系。
我们的结果表明，这些化合物可能是开发为抗肿瘤药物的良好候选物。
