Prashad N, Lotan D, Lotan R
Cancer Res. 1987 May 1;47(9):2417-24.
Dibutyryl cyclic adenosine 3':5'-monophosphate (Bt2cAMP) and beta-all-trans retinoic acid (RA) have been shown separately, and in some cases in combination, to modulate the growth, differentiation, and cAMP-dependent protein kinase (PK-A) activity of various tumor cells. The effects of Bt2cAMP and RA on a cholinergic clone (S20) of C1300 mouse neuroblastoma cells were explored in the present study. Treatment of these cells with 1 mM Bt2cAMP for 3 or more days resulted in 93% inhibition of cell proliferation in monolayer cultures and in 98% inhibition of colony formation in semisolid medium (0.5% agarose). In contrast, treatment of the cells with 1 or 10 microM RA had no inhibitory effects on cell proliferation in monolayer cultures but enhanced colony formation in agarose by up to 130%. The growth of cells treated with a combination of Bt2cAMP and RA was inhibited, although less so than with Bt2cAMP alone. Cells treated with Bt2cAMP alone or Bt2cAMP and RA extended long, neurite-like, cellular processes indicative of differentiation, whereas only a few untreated or RA-treated cells produced such extensions. The amount of [3H]cAMP-binding protein increased gradually up to 2-fold during a 3-day treatment with Bt2cAMP; in contrast it decreased by nearly 2-fold during RA treatment. These changes occurred in the level of the type I regulatory subunit (RI) of PK-A as determined by photoaffinity labeling with 8-azidoadenosine cyclic 3':5'-[32P]monophosphate. The increase in RI following Bt2cAMP treatment was corroborated by DEAE-cellulose chromatography. This analysis also demonstrated that type I PK-A is the predominant kinase in the untreated S20 cells and that RI exists as a free subunit in Bt2cAMP-treated cells. The activity of PK-A decreased by about 20% following treatment with either Bt2cAMP or RA and by 45% following treatment with a combination of both agents. These results suggest that the distinct effects of Bt2cAMP and RA on the anchorage-independent growth of S20 cells may be related to their opposite effects on the level of RI.
二丁酰环磷腺苷（Bt2cAMP）和全反式维甲酸（RA）已被分别证明，在某些情况下两者联合使用时，可调节各种肿瘤细胞的生长、分化以及环磷腺苷依赖性蛋白激酶（PK-A）的活性。本研究探讨了Bt2cAMP和RA对C1300小鼠神经母细胞瘤细胞的胆碱能克隆（S20）的影响。用1 mM Bt2cAMP处理这些细胞3天或更长时间，导致单层培养中的细胞增殖受到93%的抑制，在半固体培养基（0.5%琼脂糖）中集落形成受到98%的抑制。相比之下，用1或10 microM RA处理细胞对单层培养中的细胞增殖没有抑制作用，但使琼脂糖中的集落形成增加了高达130%。用Bt2cAMP和RA联合处理的细胞生长受到抑制，尽管比单独使用Bt2cAMP时抑制作用小。单独用Bt2cAMP或用Bt2cAMP和RA处理的细胞会伸出长的、类似神经突的细胞突起，这表明细胞发生了分化，而未处理或仅用RA处理的细胞中只有少数会产生这种突起。在用Bt2cAMP处理3天的过程中，[3H]cAMP结合蛋白的量逐渐增加，最高可达2倍；相比之下，在RA处理过程中，其含量下降了近2倍。通过用8-叠氮腺苷环3':5'-[32P]单磷酸进行光亲和标记测定，这些变化发生在PK-A的I型调节亚基（RI）水平上。通过DEAE-纤维素色谱法证实了Bt2cAMP处理后RI的增加。该分析还表明，I型PK-A是未处理的S20细胞中的主要激酶，并且在Bt2cAMP处理的细胞中RI以游离亚基的形式存在。用Bt2cAMP或RA处理后，PK-A的活性下降了约20%，而用两种药物联合处理后，PK-A的活性下降了45%。这些结果表明，Bt2cAMP和RA对S20细胞非锚定依赖性生长的不同影响可能与其对RI水平的相反作用有关。
