靶向酪氨酸激酶ALK耐药突变G1202R的新型抑制剂的设计、合成及生物活性研究

Tyrosine kinase ALK is an important molecular target for non-small cell lung cancer. The clinical use of current ALK inhibitors is limited by the occurrence of resistant mutations, especially the point mutation G1202R, which both the first and second generation ALK inhibitors failed to combat. Since G1202R mutation results in a positively charged, bulky arginine residue at this position that can cause severe atomic collisions with the ligand, our group has recently focused on the development of novel ALK inhibitors by targeting ALK G1202R mutant as well as a variety of other frequently observed mutants. Compound SOMCL-15-665 bearing a flexible amide chain with negative electricity was identified to potently inhibit ALK G1202R mutant with IC50 value of 65.0 nM. Based on this result, the current project is proposed to conduct a diverse medicinal chemistry modification by focusing on the ligand structure nearby the positively charged, bulky arginine residue. New compounds with high potency against ALK G1202R as well as a variety of other frequently observed mutations inhibitors are expected. These compounds will be further evaluated both in vitro and in vivo, which will serve as the basis for the development of new generation ALK inhibitors for clinical use.

间变性淋巴瘤（ALK）激酶是非小细胞肺癌药物研发重要的分子靶标，然而现有的ALK抑制剂由于耐药严重制约其临床应用。点突变G1202R是现有ALK抑制剂难以克服的最顽固的耐药突变。因此，研究能克服G1202R突变的新型ALK抑制剂具有重要临床意义。针对突变后R1202的大位阻及电性作用迫使抑制剂不能与ALK激酶结合导致活性丧失，本课题组近期基于R1202的位阻及电性，对配体的临近结构区域进行多样性药化修饰，包括负电性基团引入、柔性链替换、取代基移位等，发现化合物SOMCL-15-665对ALK G1202R突变有效，IC50达到65.0 nM。本项目将基于现有抑制剂与ALKG1202R的对接信息，对先导物SOMCL-15-665进行多角度多参数优化，获得靶向ALKG1202R高活性抑制剂，并对活性化合物进行系统的体内外活性及成药研究，为进一步研发新一代ALK抑制剂奠定基础。

ALK-G1202R突变是ALK代第一和第二代抑制剂难以克服且发生率最高的突变。研究能克服G1202R突变的新型ALK抑制剂具有重要临床意义。突变后精氨酸具有较大大位阻，加上电性作用迫使抑制剂ALK-G1202R亲和力大大下降甚至丧失，我们从课题组以前期开发的氨基嘧啶类二代ALK抑制剂（化合物2）为基础，结合化合物2与ALKG1202R突变蛋白的结合模式，发现化合物2中的甘氨酸侧链柔性侧链及所处的位置优势，Arg1202与侧链甘氨酸的位阻作用较小，加之结合口袋的溶剂区域存在许多极性氨基酸残基。基于此，我们在甘氨酸末端通过酰胺键引入间二苯酚片段，令人惊喜的是，该化合物对ALK激酶的活性明显高于化合物2（0.8 nM vs 2.7 nM），对ALK依赖的H3122细胞活性有稍微提高（84 nM vs 97 nM）。我们以该化合物作为新的苗头化合物进行构效分析和优化，最终得到化合物6-5d，其对野生型ALK激酶IC50值为1.7 nM，对ALKL1196M和ALKG1202R突变的酶活分别为3.5 nM和1.8 nM；对BaF3/ALK-L1196M和BaF3/ALK-G1202R突变细胞的活性都小于1.5 nM。化合物6-5d不仅提高了对L1196M和G1202R突变的活性，而且改善了原化合物6-5d的hERG毒性（hERG IC50 > 40 μM）。令人遗憾的是，由于化合物的PK性质不好导致该类化合物的体内抗肿瘤活性欠佳，然而该类化合物体外显示对ALKG1202R突变的高抑制活性使其可作为ALK耐药研究的工具药。

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