新型IDO1和IDO2双重抑制剂的设计、合成与抗肿瘤作用机制研究

IDO1 inhibitors can reverse tumor immune tolerance, and inhibit tumor growth. However, available IDO1 inhibitors have been limited to broadly application due to their unsatisfactory effect, obvious side effects and drug resistance which is easily generated after long term use. When IDO2 and IDO1 are involved in tumorigenesis and immune tolerance together, simultaneously inhibition of IDO1 and IDO2 result in synergistic effect of anti-tumor and it is possible to effectively overcome drug resistance of IDO1 inhibitor. In our previous work, INCB024360, a selective IDO1 inhibitor which is discovered by Incyte Corporation through HTS, was chosen as the lead compound to discover novel IDO1 inhibitor. The novel molecule skeleton FFA9 was identified as potent IDO1 and IDO2 dual inhibitors for the first time. On the basis of this result, FFA9 was optimized as lead compound in this project firstly. Meanwhile, the oxime group of the structure of FFA9 is replaced by propionic acid group lead to another series of novel compounds. 80 compounds will be designed and synthesized. The inhibitory activity against IDO1 and IDO2 in vitro will be tested for all compounds. Some potent compounds will be selected to evaluate their anti-tumor activity and mechanism in vitro and in vivo. It is our goal to identify one or two novel and potent antitumor candidates which is not easy generate drug resistance. And the structure-activity relationship will be analyzed for further research on IDO1 and IDO2 dual inhibitor.

IDO1抑制剂能够逆转IDO1介导的免疫抑制，抑制肿瘤生长。但现有IDO1抑制剂单药疗效有限，副作用比较明显，长期使用容易对免疫疗法产生抗性。IDO2与IDO1共同作用参与肿瘤发生和免疫耐受，同时抑制IDO1和IDO2对肿瘤免疫治疗具有协同增效作用，有可能有效克服IDO1抑制剂介导的免疫疗法产生的抗性。课题组前期以选择性IDO1抑制剂INCB024360为先导物，进行结构优化，首次发现了具有IDO1和IDO2双重抑制活性的分子骨架FFA9。本项目以FFA9为先导物，首先对其进行结构优化，同时运用生物电子等排原理进行药效团替换，将羟肟替换成丙酸，设计合成两个系列共计80个化合物，通过对IDO1和IDO2酶活性测试、体内外抗肿瘤活性筛选以及抗肿瘤作用机制研究，期望获得1~2个抗肿瘤活性强、不易产生抗性、结构新颖的候选化合物，分析构效关系，为IDO1和IDO2双靶点抑制剂的深入研究提供理论依据。

IDO1抑制剂能够逆转IDO1介导的免疫逃逸，发挥抑制肿瘤生成的作用。但现有IDO1抑制剂的体内活性普遍不够强、毒副作用明显。长期使用有可能导致癌症免疫疗法所存在的抗性问题。IDO1/2在肿瘤的免疫治疗方面可能有着协同增效的作用，同时抑制IDO1和IDO2有可能在提高肿瘤免疫治疗效果的同时克服（或部分克服）癌症免疫疗法抗性。课题组前期以IDO1抑制剂epa为先导化合物，进行结构优化，首次发现了具有IDO1和IDO2双重抑制获得化合物FFA9。为了获得活性更好，对两个靶点抑制活性均衡的、新的IDO1/2双靶点抑制剂，本项目以FFA9为先导化合物，对其进行结构优化，同时运用生物电子等排等原理进行药效团替换，将羟肟结构用丙酸替换，共设计合成了三个系列化合物82个，所有化合物均经1H-NMR、13C-NMR和HRMS进行了结构确证。通过对IDO1/2酶活性的测试发现了两个候选化合物IIIa-14和IIIa-22。候选化合物表现出对IDO1/2均具有较好的体外酶抑制活性，激酶选择性强。成药性质优良，SD大鼠的药代动力学试验结果显示，灌胃给予化合物IIIa-14和IIIa-22时，Cmax分别为2733 ng/mL和1043 ng/mL，Tmax分别为0.42 h和1.3 h，t1/2分别为1.3 h和1.9 h，口服生物利用度F分别为20.3%和12.9%。小鼠体内抗肿瘤活性的测试结果显示，以50 mg/kg剂量每天灌胃给药两次，连续28天，与对照组相比，化合物IIIa-14和IIIa-22和阳性药Epacadostat在CT26小鼠移植瘤模型上均能够抑制肿瘤的生长，且化合物IIIa-14和IIIa-22的抑瘤率达60%以上，明显优于同剂量下阳性药Epacadostat的抑瘤率（40%）；在HCT116裸鼠异种移植瘤模型上，分别以100mg/kg和300mg/kg剂量每天灌胃给药两次，连续给药21天，化合物IIIa-22并没有显示出抗肿瘤活性，初步说明化合物是通过激活免疫达到抗肿瘤作用的。进一步通过初步的机制研究显示，化合物IIIa-22是选择性地作用于IDO1和IDO2，阻止色氨酸向犬尿氨酸的代谢途径，从而促进肿瘤组织、脾脏和引流淋巴细胞中Treg细胞和CD4+T细胞的凋亡，达到肿瘤免疫治疗的目的。本研究为IDO1/2双靶点抑制剂的研究提供了理论基础和研究经验。