||The role of ribosomal protein S6 kinase 1(S6K1) on cisplatin resistance in chronic myeloid leukemia (CML)
||Institute of Basic Medical Sciences
95%慢性骨髓性白血病 (chronic myeloid leukemia, CML)病人皆帶有BCR-ABL融合基因。BCR-ABL融合基因酪氨酸激酶抑制劑 (tyrosine kinase inhibitors, TKIs)成為治療慢性骨髓性白血病標靶藥物，但復發的或在急性轉化期 (blast crisis phase, BC phase)的病人對於此抑制劑並無反應或產生抗藥性。因而尋找不同的治療方法對於治療慢性骨髓性白血病還是重要的。環磷酸腺苷(cyclic AMP, cAMP)可藉由調控核糖體蛋白S6激酶1 ( ribosomal protein S6 Kinase 1, S6K1)影響細胞增生。另外，抑癌基因p53的表現除了與慢性骨髓性白血病的病程相關和抗藥性也有間接關係。所以此篇研究的目的在於探討環磷酸腺苷、核糖體蛋白S6激酶1和p53三者的關係是否影響化療藥物對於慢性骨髓性白血病細胞的毒性。我們利用環磷酸腺苷促效劑(cAMP agonists)增加環磷酸腺苷濃度後，發現慢性骨髓性白血病細胞產生對化療藥物cisplatin的抗藥性。接著，我們分別利用蛋白激酶A(protein kinase A, PKA)和環磷酸腺苷激活蛋白( exchange protein directly activated by cAMP, EPAC)活化劑得知環磷酸腺苷是藉由蛋白激酶A調控cisplatin抗藥性。我們也看到增加環磷酸腺苷後造成磷酸化核糖體蛋白S6激酶1和核醣體蛋白S6(ribosomal protein S6, RPS6)程度下降。同時，藉由磷酸化核糖體蛋白S6激酶1抑制劑以及基因減量(knockdown)證實當磷酸化核糖體蛋白S6激酶1受到抑制時會造成cisplatin抗藥性。而S6K1則是藉由調控DNA-dependent protein kinase, catalytic subunit (DNA-PKcs), H2A histone family member X (H2AX)和Poly ADP-ribose polymerase (PARP) 而造成cisplatin 抗藥性。除此之外，p53在慢性骨髓性白血病細胞的表現量則會影響核糖體蛋白S6激酶1所調控的cisplatin抗藥性現象。綜合以上，當慢性骨髓性白血病人在使用化療藥物進行治療時必須考慮環磷酸腺苷的濃度。更重要的是我們發現p53的表現對於是否欲選擇核糖體蛋白S6激酶1抑制劑作為化療藥物的佐劑(adjuvant agent)是很重要的。
More than 95% chronic myeloid leukemia (CML) patients carry the constitutively activated tyrosine kinase, BCR-ABL fusion protein. Therefore, tyrosine kinase inhibitors (TKIs) are designed for CML treatment. However, some patients have no response to TKIs especially for those patients with relapsed or diagnosed at blast crisis (BC) phase. Thus, it is needed for developing alternative approaches for CML treatment. Cyclic AMP (cAMP) regulates cell proliferation via ribosomal protein S6 kinase 1 (S6K1). The expression of tumor suppressor gene, p53, is involved in the disease progression and affects drug resistance in CML. The purpose of our study is to investigate whether the relationship between cAMP, S6K1 and p53 in drug resistance in CML cells. We observed that increased level of cAMP conferred cisplatin resistance in CML cells. From the experiments of the activators of protein kinase A (PKA) and exchange protein directly by cAMP (EPAC), we observed that PKA is the effector on cAMP-induced cisplatin resistance. We also found that increased level in cAMP reduced the formation of pS6K1 and pRPS6. Furthermore, inhibition and knockdown of S6K1 lead to cisplatin resistance in CML cells. Next, we observed that S6K1 acts via DNA-PKcs, H2AX and PARP regulating cisplatin resistance. Last but not the least, p53 expression attenuates the effect of S6K1 on cisplatin resistance in CML cells. In conclusion, the level of cAMP should be considered when choosing chemotherapeutic drug such as cisplatin for CML treatment. More importantly, we suggest that p53 expression is crucial for choosing the inhibitor of S6K1 signaling pathway as the adjuvant agent with chemotherapeutic drug in CML treatment.
List of Figures v
Experimental Designs 11
Specific Aims 12
Materials and Method 16
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