Hisatake J, Kubota T, Hisatake Y, Uskokovic M, Tomoyasu S, Koeffler H P
Division of Hematology/Oncology, Cedars-Sinai Research Institute, UCLA School of Medicine, Los Angeles, California 90048, USA.
Cancer Res. 1999 Aug 15;59(16):4023-9.
The 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] is the physiologically active form of vitamin D3 that inhibits proliferation and induces differentiation of a variety of malignant cells. We evaluated a newly synthesized vitamin D3 analogue [1,25(OH)2-16-ene-5,6-trans-D3 (Ro 25-4020)] that has a novel 5,6-trans motif. Dose-response studies showed that 1,25(OH)2-16-ene-5,6-trans-D3 had 10-100-fold greater antiproliferative activities than 1,25(OH)2D3 when measuring clonal growth of breast (MCF-7) and prostate (LNCaP) cancer cell lines as well as a myeloid leukemia cell line (HL-60). Because the chief toxicity of vitamin D3 is hypercalcemia, we examined the calcemic activity of 1,25(OH)2-16-ene-5,6-trans-D3 in mice. Remarkably, 1,25(OH)2-16-ene-5,6-trans-D3 was at least 40-fold less calcemic as compared with 1,25(OH)2D3 and 1,25(OH)2-16-ene-D3 (Ro 24-2637). To explore the mechanism by which the 1,25(OH)2-16-ene-5,6-trans-D3 analogue mediated its antiproliferative activity, several studies were performed. Pulse-exposure studies showed that a 4-day pulse exposure to 1,25(OH)2-16-ene-5,6-trans-D3 (10(-7) M) in liquid culture was adequate to achieve a 40% inhibition of MCF-7 clonal growth in the absence of the analogue, suggesting that the growth inhibition mediated by 1,25(OH)2-16-ene-5,6-trans-D3 was at least in part irreversible. Cell cycle studies showed that 1,25(OH)2-16-ene-5,6-trans-D3 increased the proportion of MCF-7 cells in the G0-G1 phase and decreased those in the S phase. Furthermore, 1,25(OH)2-16-ene-5,6-trans-D3 induced an elevated expression of the cyclin-dependent kinase inhibitors, p21waf1 and p27kip1. In addition, 1,25(OH)2-16-ene-5,6-trans-D3 almost completely inhibited telomerase activity, as measured by telomeric repeat amplification protocol assay and human telomerase reverse transcriptase mRNA. For each of the growth-related parameters that were examined, the vitamin D3 analogue was more active than 1,25(OH)2D3. In contrast, 1,25(OH)2D3 was more calcemic than 1,25(OH)2-16-ene-5,6-trans-D3. In summary, 1,25(OH)2-16-ene-5,6-trans-D3, having a novel 5,6-trans motif, strongly inhibited clonal proliferation and reduced telomerase activity with low calcemic activity, suggesting further testing in in vivo cancer models. This analogue may gain a therapeutic niche for selected malignancies.
1,25 - 二羟基维生素D3 [1,25(OH)2D3] 是维生素D3的生理活性形式，可抑制多种恶性细胞的增殖并诱导其分化。我们评估了一种新合成的具有新型5,6 - 反式基序的维生素D3类似物 [1,25(OH)2 - 16 - 烯 - 5,6 - 反式 - D3 (Ro 25 - 4020)]。剂量反应研究表明，在检测乳腺癌（MCF - 7）、前列腺癌（LNCaP）细胞系以及髓系白血病细胞系（HL - 60）的克隆生长时，1,25(OH)2 - 16 - 烯 - 5,6 - 反式 - D3的抗增殖活性比1,25(OH)2D3高10 - 100倍。由于维生素D3的主要毒性是高钙血症，我们检测了1,25(OH)2 - 16 - 烯 - 5,6 - 反式 - D3在小鼠体内的血钙活性。值得注意的是，与1,25(OH)2D3和1,25(OH)2 - 16 - 烯 - D3（Ro 24 - 2637）相比，1,25(OH)2 - 16 - 烯 - 5,6 - 反式 - D3的血钙活性至少低40倍。为了探究1,25(OH)2 - 16 - 烯 - 5,6 - 反式 - D3类似物介导其抗增殖活性的机制，我们进行了多项研究。脉冲暴露研究表明，在液体培养中对1,25(OH)2 - 16 - 烯 - 5,6 - 反式 - D3（10(-7) M）进行4天的脉冲暴露足以在无该类似物的情况下实现对MCF - 7克隆生长40%的抑制，这表明1,25(OH)2 - 16 - 烯 - 5,6 - 反式 - D3介导的生长抑制至少部分是不可逆的。细胞周期研究表明，1,25(OH)2 - 16 - 烯 - 5,6 - 反式 - D3增加了MCF - 7细胞在G0 - G1期的比例，降低了S期的比例。此外，1,25(OH)2 - 16 - 烯 - 5,6 - 反式 - D3诱导细胞周期蛋白依赖性激酶抑制剂p21waf1和p27kip1的表达升高。另外，通过端粒重复序列扩增法和人端粒酶逆转录酶mRNA检测发现，1,25(OH)2 - 16 - 烯 - 5,6 - 反式 - D3几乎完全抑制了端粒酶活性。对于所检测的每个与生长相关的参数，该维生素D3类似物都比1,25(OH)2D3更具活性。相比之下，1,25(OH)2D3的血钙活性比1,25(OH)2 - 16 - 烯 - 5,6 - 反式 - D3更高。总之，具有新型5,6 - 反式基序的1,25(OH)2 - 16 - 烯 - 5,6 - 反式 - D3强烈抑制克隆增殖并降低端粒酶活性，且血钙活性较低，这表明其在体内癌症模型中有待进一步测试。这种类似物可能在某些特定恶性肿瘤的治疗中占据一席之地。
