MK-2206联合顺铂对乳腺癌4T1/DDP移植瘤耐药性的影响

Effect of MK-2206 plus cisplatin on the drug resistant 4T1/DDP breast tumor in mice

目的探讨蛋白激酶B(protein kinase B,Akt)抑制剂MK-2206联合顺铂对三阴性乳腺癌4T1顺铂耐药(4T1/cisplatin,4T1/DDP)移植瘤耐药性的影响。方法采用顺铂(cisplatin,DDP)低剂量诱导及体内外交叉致瘤相结合的方法建立三阴性乳腺癌4T1/DDP耐药小鼠移植瘤模型,给予DDP腹腔注射(0.3 g/L,1次/d)单独或联合MK-2206灌胃(12 g/L,1次/周)治疗。给药28 d后处死小鼠,小动物成像检测观察肿瘤生长情况;Western blot法检测肿瘤组织中磷酸化Akt(phosphorate-Akt,p-Akt)和总Akt(total-Akt,t-Akt)蛋白水平变化;免疫组织化学法分析耐药相关蛋白P-糖蛋白(P-glycoprotein,P-gp)、乳腺癌耐药蛋白(breast cancer resistance protein,BCRP)、基质金属蛋白酶7(matrix metalloproteinase 7,MMP7)表达差异。结果建立的三阴性乳腺癌耐药小鼠模型达中等耐药程度。建立非耐药小鼠与耐药小鼠肿瘤组织生长速度未见明显区别(P>0.05)。分别给予这两种模型小鼠相同剂量(0.3 g/L)的DDP治疗后,耐药小鼠对DDP的敏感性低于非耐药小鼠(P<0.01)。将MK-2206(12 g/L)与DDP(0.3 g/L)联合应用后小鼠肿瘤较单独应用DDP缩小,且可降低P-gp、BCRP、MMP7蛋白的表达(均P<0.05),并降低Akt蛋白磷酸化水平(P<0.01)。结论 MK-2206联合顺铂能够逆转4T1/DDP移植瘤耐药性,且与抑制Akt蛋白磷酸化相关。

Objective To investigate the effect of protein kinase B （Akt） inhibitor MK-2206 combined with cisplatin on the drug resistance of 4T1/cisplatin （DDP） transplantation breast tumor. Methods Drug resistant 4T1/DDP transplantation tumor model was established with low-dose DDP induction and in vivo/in vitro tumorigenic approach in mice. The mice were then treated with intraperitoneal injection of DDP alone （0.3 g/L, once per day）, or in combination with MK- 2206 gavage （ 12 g/L, once per week）. The mice were sacrificed 28 days afterwards. Small animal live imaging technology was applied to detect tumor growth. Western blots were used to examine the protein levels of phosphorylated-Akt and total- Akt. Immunohistochemistry was used to examine the expression of P-glycoprotein （P-gp）, breast cancer resistance protein （BCRP） , and matrix metalloproteinase 7 （MMP7）. Results 4T1/DDP transplantation tumor mice model developed me- dium level of drug resistance. The tumor growth rates between non-resistant and resistant mice showed no significant differ- ence （ P 〉 0.05 ）. After similar dose of DDP （0.3 g/L） treatment, drug resistant mice showed lower sensitivity than non- resistant mice （P 〈0.01 ）. However, in the drug resistant mice treated with MK-2206（ 12 g/L） and DDP （0.3 g/L）, the tumor size were significantly reduced, compared to DDP alone group （P 〈 0.01 ）. Moreover, the expression levels of P-gp, BCRP and MMP7 were all significantly reduced （ P 〈 0.05 ） in MK-2206 and DDP treated mice. The phosphorylation level of Akt was also decreased （P 〈 0.01 ）. Conclusion MK-2206 in combination with cisplatin can reverse the drug resist- ance of 4T1/DDP transplantation tumor, which may be mediated through the inhibition of Akt phosphorylation.