The development of targeted therapies using FKHRL1 for the treatment of leukemia

FKHRL 1（也称为FOXO 3a）是Forkhead转录因子FOXO亚家族的成员，位于细胞因子信号通路的下游。该分子被激活的Akt/Gsk磷酸化，并作为“非活性形式”保留在细胞质中。在这项研究中，我们发现FKHRL 1在慢性粒细胞白血病（CML）中作为磷酸化的“非活性”形式在Bcr-Abl酪氨酸激酶下游发挥作用。Bcr-Abl酪氨酸激酶抑制剂伊马替尼通过将CML衍生细胞系中的FKHRL 1从磷酸化的“非活性”形式转化为去磷酸化的“活性”形式，诱导细胞周期停滞和随后的凋亡。为了检查“活性”FKHRL 1是否可以克服对伊马替尼的抗性，我们在CML衍生的伊马替尼抗性细胞系中产生了4-羟基他莫昔芬（4-OHT）诱导的“活性”FKHRL 1（FKHRL 1-TM ; FKHRL 1的三重突变体，其中所有三个Akt磷酸化位点都已突变）-雌激素受体融合蛋白表达系统。4-OHT抑制细胞生长和细胞周期进程，诱导细胞凋亡，并上调肿瘤坏死因子相关凋亡诱导配体（TRAIL）。此外，我们还发现全反式维甲酸（all-transretinoicacid，ATRA）通过FKHRL 1介导的TRAIL产生诱导对ATRA敏感的APL细胞株NB 4的凋亡。与CML细胞一样，过表达“活性”FKHRL 1不仅在ATRA敏感细胞系NB 4中诱导凋亡，而且在ATRA耐药细胞系NB 4 R中也诱导凋亡。因此，我们的研究结果表明，“活性”FKHRL 1可以克服伊马替尼耐药和ATRA耐药的CML和APL细胞系，分别通过部分TRAIL的生产。

FKHRL1 (also called FOXO3a) is a member of the FOXO subfamily of Forkhead transcription factors and lies downstream of cytokine signaling pathways. This molecule is phosphorylated by activated Akt/Gsk and remains in the cytoplasm as an "inactive form". In this study, we found that FKHRL1 functions downstream of Bcr-Abl tyrosine kinase as a phosphorylated "inactive" form in chronic myelogenous leukemia (CML). The Bcr-Abl tyrosine kinase inhibitor imatinib induced cell cycle arrest and subsequent apoptosis via the conversion of FKHRL1 from the phosphorylated "inactive" form to the dephosphorylated "active" form in CML-derived cell lines. To examine whether "active" FKHRL1 can overcome the resistance to imatinib, we generated a 4-hydroxytamoxifen (4-OHT)-inducible "active" FKHRL1 (FKHRL1-TM ; a triple mutant of FKHRL1 in which all three Akt phosphorylation sites have been mutated)-estrogen receptor fusion protein expression system in CML-derived imatinib-resistant cell lines. 4-OHT inhibited the cell growth and cell cycle progression, and subsequently induced apoptosis, accompanied by up-regulation of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). Thus, "active" FKHRL1 antagonized deregulated proliferation and induced apoptosis in these cell lines.Moreover, we also found that all-trans retinoic acid (ATRA) induced apoptosis via FKHRL1-mediated TRAIL production in APL-derived cell line NB4 which was sensitive to ATRA. Like CML cells, over-expression of "active" FKHRL1 induced apoptosis not only in ATRA-sensitive cell line NB4 but also in ATRA-resistant NB4R cell line. Therefore, our results suggest that "active" FKHRL1 can overcome imatinib resistance and ATRA resistance in CML and APL cell lines, respectively, in part via TRAIL production.