ASP8273, a novel mutant-selective irreversible EGFR inhibitor, inhibits growth of non-small cell lung cancer (NSCLC) cells with EGFR activating and T790M resistance mutations
ASP8273，一种新颖的突变选择性不可逆EGFR抑制剂，抑制发生EGFR敏感性（原发性活性突变）突变和继发性T790M突变的非小细胞肿瘤细胞的生长。
BACKGROUND: Reversible EGFR TKIs, gefitinib and erlotinib, have shown antitumor efficacy in NSCLC patients with activating mutations in EGFR kinase domain. But the clinical efficacy of these agents is limited by the development of acquired drug resistance, which is most commonly caused by T790M resistance mutation in EGFR. This mutation has been detected in approximately 50% to 60% of patients. The 2nd generation irreversible EGFR inhibitors inhibit EGFR with T790M, but their clinical efficacy to NSCLC patients with T790M appears to be limited due to severe adverse effects caused by concomitant WT EGFR inhibition. Therefore, an EGFR TKI which inhibits T790M mutant EGFR selectively with less activity against WT EGFR may be beneficial. Here we report ASP8273, a novel, small molecule EGFR TKI that inhibits the kinase activity of EGFR containing the activating and T790M resistance mutations with less activity against WT EGFR.
背景：可逆EGFR抑制剂吉非替尼和厄洛替尼虽然对发生EGFR活性突变的患者已经表现出抗肿瘤活性，但是由于发生继发性突变（T790M）而出现第二次耐药，限制了它们的临床应用。在这些突变里面，最常见的突变是T790M耐药突变，这种突变约占总耐药人数的50-60%。第二代不可逆抑制剂对T790M突变耐药虽也有效，由于它同时也对正常的野生型EGFR（WT）产生抑制，因而出现了严重的不良反应，在一定程度也限制了它的临床应用。因此，一种可以抑制T790M突变，而对野生型抑制活性比较低的选择性抑制剂可能会更具有临床价值。。
METHODS: The inhibitory effect and the selectivity of ASP8273 were evaluated against mutant EGFR (L858R, del ex19, L858R/T790M and del ex19/T790M) and WT EGFR using in vitro enzymatic and cell-based assay. Binding mode of ASP8273 to EGFR was assessed by mass spectrometry. Antitumor activity of ASP8273 was evaluated in xenograft models using PC-9 (del ex19), HCC827 (del ex19), NCI-H1975 (L858R/T790M) and PC-9ER (Erlotinib Resistant)(del ex19/T790M) NSCLC cells.
方法：ASP8273的选择性抑制活性是通过体外的激酶和细胞试验测试其对EGFR (L858R, del ex19, L858R/T790M and del ex19/T790M)以及野生型的EGFR的抑制活性，ASP8273的键合模型是通过质谱进行检测，体内活性是通过ASP8273测试对PC-9 (del ex19), HCC827 (del ex19), NCI-H1975 (L858R/T790M) and PC-9ER (Erlotinib Resistant)(del ex19/T790M)的动物模型的抑制。.
RESULTS: ASP8273 inhibited mutant EGFR containing del ex19 or L858R activating mutations as well as the T790M resistance mutation with lower IC50 values than WT EGFR. Mass spectrometry analysis revealed that ASP8273 is covalently bound to a mutant EGFR(L858R/T790M) via C797 in the kinase domain of EGFR. In NCI-H1975 cells, ASP8273 induced long-lasting inhibition of EGFR phosphorylation for 24 h after washout of compound.
结果：ASP8273不仅对19号外显子的缺失，L858R和T790M突变有效，而且对野生型的活性较低。质谱分析表明，ASP8273与H1975细胞的797位的半胱氨酸发生共价键结合而发生作用。清洗掉化合物后，ASP8273依然能持续抑制EGFR的磷酸化达24小时之久。
In assays using endogenously EGFR-dependent cells, ASP8273 inhibited the growth of PC-9(del ex19), HCC827(del ex19), NCI-H1975(del ex19/T790M) and PC-9ER(del ex19/T790M) with IC50 values of 8-33 nM, more potently than that of NCI-H1666(WT) with IC50 value of 230 nM.
In mouse xenograft studies, ASP8273 induced tumor regression in NCI-H1975 (L858R/T790M), HCC827 (del ex19) and PC-9 (del ex19) xenograft models by repeated oral dosing in a dose-dependent manner. Dosing schedules did not affect the efficacy of ASP8273. In an NCI-H1975 xenograft model, complete regression of tumor was achieved after 14-days of ASP8273 treatment. Complete regression was maintained in 50% of mice more than 85 days after cessation of ASP8273 treatment.
在所造的老鼠模型中，ASP8273重复口服给药后以剂量依赖的形式诱导双突变(L858R/T790M)模型，19号外显子缺失突变HCC827和PC-9模型动物肿瘤的缩小。14天连续ASP8273给药后，肿瘤完全缩小。停药后，有50%的老鼠保持完全缩小状态达85天以上。
CONCLUSIONS: ASP8273 inhibits the growth of NSCLC cells with EGFR activating and T790M resistance mutations with evidence of tumor regression. Therefore, ASP8273 may show therapeutic efficacy in NSCLC patients with EGFR mutations. Clinical trials of ASP8273 in NSCLC patients are planned in the US/EU and Asia.
结论：ASP8273抑制同时含有EGFR活性突变和T790M耐药突变的非小细胞肺癌细胞，动物实验也表明肿瘤有明显的缩小。因此，对于含有EGFR突变的非小细胞肺癌病人，ASP8273也许会有疗效。ASP8273的临床试验正在美国/欧盟和亚洲进行。
ASP8273目前开了I期临床：
http://clinicaltrials.gov/show/NCT02113813
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