Establishment of Vitamin K2 Therapy in Myelodysplastic Syndromes

我们最初报道了维生素K2（VK 2）类似物，包括甲基萘醌4（MK 4），而不是维生素K1，有效地诱导各种类型的原代培养的白血病细胞和白血病细胞系在体外凋亡。一些病例报告也证明了使用VK 2治疗白血病和骨髓增生异常综合征（MDS）患者的临床获益。有趣的是，已经报道了在有/没有维生素D3（D3）的情况下VK 2治疗对于改善MDS中患有难治性贫血（RA）的患者的血细胞减少是有效的，尽管潜在的机制仍不清楚。我们在这里集中在VK 2和D3对白血病细胞的联合作用。用VK 2加1α，25-二羟基维生素D3（D3）加VK 2处理HL-60和U937细胞显著增强单核细胞分化--流式细胞术显示，与单独用VK 2或D3处理的细胞相比，CD 14的平均强度增加了20倍以上--并且还实现了单核细胞的完全成熟。 ...更多信息 n通过形态学评估。此外，在暴露于VK 2加D3 48至72小时后检测到G1期阻滞的累积增强。这些综合效应远远超过了每种维生素单独最佳浓度下的最大分化诱导能力。令人感兴趣的是，沿着细胞分化，细胞变得抵抗各种凋亡刺激，包括VK 2-和H2 O2-处理和血清剥夺。此外，在用VK 2加D3处理96小时后，检测到细胞质p21 <CIP1>^沿着细胞核p21^的消失<CIP1>而显著积累。这种抗凋亡作用部分通过阻断JNK活性来介导，JNK活性由p21 β<CIP1>和ASK-1（JNK途径中的上游丝氨酸/苏氨酸激酶）的相互作用来调节。U937-ΔNLS-p21^的稳定转染子<CIP1>缺乏p21^的核定位信号<CIP1>，导致细胞质p21^过表达<CIP1>，而没有单核细胞分化，显示出对细胞凋亡诱导的抵抗力。这些数据表明，随着分化，p21 CIP的细胞内分布从细胞核到细胞质的变化沿着，似乎起着抗凋亡行为的作用。我们的数据表明，VK 2加D3可能是AML和MDS基于分化的治疗的有效组合。此外，在强制分化/成熟诱导过程中的抗凋亡作用可能解释了RA患者血细胞减少的改善，其血细胞减少是通过细胞凋亡介导的。少

We originally reported that vitamin K2 (VK2) analogs, including menaquinone 4 (MK4) but not vitamin K1, effectively induce apoptosis in various types of primary cultured leukemia cells and leukemia cell lines in vitro. Several case reports also demonstrated the clinical benefits of using VK2 for the treatment of patients with leukemia and myelodysplastic syndrome (MDS). It is of interest that VK2 treatment with/ without vitamin D3 (D3) has been reported to be effective for improvement of cytopenia in patients with refractory anemia (RA) in MDS, although the underlying mechanism is still unclear. We here focus on the combined effects of VK2 and D3 on leukemia cells. Treatment of HL-60 and U937 cells with VK2 plus 1α, 25-dihydroxy-vitamin D3 (D3) plus VK2 dramatically enhanced monocytic differentiation -a more than 20-fold increase of the mean intensities of CD14 as compared to the cells treated with either VK2 or D3 alone by flow cytometry -and also achieved complete monocytic maturatio … More n as assessed by morphology. In addition, enhanced accumulation of G1 arrest was detected after 48 to 72 hr-exposure to VK2 plus D3. These combined effects far exceed the maximum differentiation-inducing ability at the optimal concentrations of each vitamin alone. It was of interest that, along with cell differentiation, the cells became resistant against various apoptosis stimuli including VK2-and H2O2-treatment and serum deprivation. Furthermore, dramatic accumulation of cytoplasm p21^<CIP1> along with disappearance of nuclear p21^<CIP1> was detected in response to 96-hr treatment with VK2 plus D3. This anti-apoptotic effect was mediated in part by blocking JNK activity, which is regulated by interaction of p21^<CIP1> and ASK-1, an upstream serine/threonine kinese in the JNK pathway. A stable transfectant of U937-ΔNLS-p21^<CIP1>, which lacks a nuclear localizing signal of p21^<CIP1> and results in over-expression of cytoplasm p21^<CIP1> without monocytic differentiation, showed resistance against apoptosis inductions. These data suggest that a change of intracellular distribution of p21CIP from nucleus to cytoplasm along with differentiation appears to be functioning as anti-apoptotic behavior. Our data suggest that VK2 plus D3 might be a potent combination for the differentiation-based therapy for AML and MDS. Furthermore, an anti-apoptotic effect during enforced differentiation/maturation induction might explain the improvement of cytopenia in RA patients, whose cytopenias are mediated through apoptosis. Less