基于人源肿瘤组织异种移植模型研究萘普替尼的抗肿瘤药效

Pharmacodynamics of neptinib based on the human-derived tumor xenograft (PDX) model of lung adenocarcinoma

目的:以表皮生长因子受体(epithelial growth factor receptor, EGFR)野生型肺腺癌人源肿瘤组织异种移植(patient-derived tumor xenograft,PDX)模型为研究对象,探索萘普替尼的抗肿瘤活性。方法:本研究将临床手术切除的新鲜EGFR野生型肺腺癌组织经无菌条件处理后移植到高免疫缺陷小鼠的皮下。并将第一代PDX模型(F1)肿瘤组织在裸鼠体内传代~第4代(F4),分别将F1~F4代裸鼠的肿瘤组织进行HE染色和免疫组织化学检测,验证人源肿瘤组织在裸鼠体内传代过程中组织结构和生物活性表达的稳定性。将F4代EGFR野生型肺腺PDX模型小鼠随机分为模型组、阿法替尼、萘普替尼-LD、萘普替尼-HD组,分别连续灌胃(ig)给予生理盐水、阿法替尼10 mg·kg^-1、萘普替尼0.7和2.1 mg·kg^-1 28 d,qd。每周2次测量体重、肿瘤体积,计算相对肿瘤体积、绘制肿瘤生长曲线。给药结束后脱颈椎处死各组小鼠,摘取肿瘤组织,称量肿瘤湿重,计算肿瘤湿重抑制率。结果:F1~F4代的肺腺癌PDX模型的肿瘤组织分别进行HE染色及IHC检测,肿瘤组织中细胞异形性明显,并且出现病理性核分裂现象和局部组织的坏死,免疫组化中TTF-1,CK7,NAPsina IHC检测均呈阳性反应,证明了人源肿瘤组织在免疫缺陷小鼠体内稳定遗传,肺腺癌PDX模型建立成功。药效实验结果显示,阿法替尼、萘普替尼-LD、萘普替尼-HD组肿瘤体积生长均显著低于模型组,给药28 d后,阿法替尼、萘普替尼-HD组,萘普替尼-LD组的相对肿瘤增殖率均低于40%,同时连续给药28 d中实验组体重与模型组比较无显著差异。连续给药28 d后摘取肿瘤称重,并计算肿瘤湿重抑制率,阿法替尼、萘普替尼-LD、萘普替尼-HD组分别为(69.26±13.82)%,(38.98±42.64)%,(65.85±20.43)%,与模型组比较有显著性差异(P<0.05)。结论:萘普替尼对EGFR野生型肺腺癌PDX模型在2.1 mg·kg^-1给药剂量即可产生与进口药物阿法替尼10 mg·kg^-1剂量相同的抗肿瘤药效,且用药期间未产生对小鼠的毒性作用,具有较好的临床应用前景。

Objective: To explore the antitumor activity of neptinib with the epithelial growth factor receptor(EGFR) wild type lung adenocarcinoma model as the research object. Methods: EGFR wild-type lung adenocarcinoma tumors derived from primary surgical resection were sliced into fragments, and then implanted into the subcutaneous tissues of severe immunodeficiency mice. With the tumor tissues of the first-generation PDX model(F1) serial passaging in nude mice until the fourth-generation(F4), the tumor tissues of the F1~F4 generation mice were respectively subjected to hematoxylin-eosin staining and immunohistochemical detection to verify the stability of the tissue structure and biological activity expression of tumor tissues during the passage in nude mice. The F4 PDX model mice of EGFR wild-type lung adenocarcinoma were randomly divided into the model group, afatinib, neptinib-LD and neptinib-HD group, and were given normal saline, afatinib 10 mg·kg^-1, neptinib 0.7 mg·kg^-1 and 2.1 mg·kg^-1 respectively for 28 days, once a day. The tumor volume and weight were measured twice a week, the relative tumor volume was calculated, and the tumor growth curve was plotted. After the end of administration, the mice were sacrificed, the tumors were taken and weighted, and the inhibition rate of tumor wet weight was calculated. Results: The cells in the tumor tissues of F1~F4 generations mice showed obvious atypia, and pathologic mitosis and local tissue necrosis were observed. TTF-1, CK7 and NAPsina IHC were all positive in the PDX model of lung adenocarcinoma of F1~F4 generations, which proved that the human tumor tissue was stably hereditary in immunodeficient mice, and the PDX model of lung adenocarcinoma was successfully established. The tumor volume growth in afatinib, neptinib-LD and neptinib-HD groups was significantly lower than that in the model group. After 28 days of administration, the relative tumor proliferation rate of afatinib, neptinib-HD and neptinib-LD groups were lower than 40%. Moreover, there was no significant difference in body weight between each group and the model group during 28 days of administration. After 28 days of continuous administration, the inhibition rates of tumor wet weight of afatinib, neptinib-LD and neptinib-HD groups were(69.26±13.82)%,(38.98±42.64)% and(65.85±20.43)%, respectively, which were significantly different from the model group(P<0.05). Conclusion: Neptinib at 2.1 mg·kg^-1 produces the same anti-tumor effect as afatinib at 10 mg·kg^-1 in mice, without toxic effect during the treatment period, indicating that it has good clinical application prospects.