人源性食管癌移植瘤模型的建立、评价及其应用的初步研究

目的用，晦床新鲜肿瘤组织移植于免疫缺陷小鼠以建立人源性食管癌移植瘤模型并初步评估其应用性。方法将36例食管癌手术切除组织分别移植于SCID小鼠皮下，待移植瘤长成后对其进行裸鼠间连续传代。当每例模型的移植瘤体积生长至500-1000mm3时，收集肿瘤组织作4种处理，一部分继续用于鼠间传代，一部分组织冻存于液氮中用于将来肿瘤组织复苏模型重建，一部分组织作速冻处理用于DNA／RNA提取，一部分组织固定于福尔马林溶液中作石蜡包埋和HE染色用于病理学研究。选用其中4例模型ESX00l、Esx002、ESX003和ESX004进行体外药敏实验和体内药效实验以评价4例人源性食管癌移植瘤对常用抗肿瘤药物的敏感性。结果36例食管癌手术切除组织接种和传代，成功23例，成功率为63．89％。异种移植瘤生长稳定并能连续传代，各代移植瘤组织形态学特征与患者肿瘤组织一致。23例液氮保存的人源食管癌移植瘤组织复苏后移植于SCID小鼠皮下，21例成功生长肿瘤，复苏成功率为91．30％。体外药敏实验表明4例人食管癌模型对临床上常用的抗肿瘤药物索拉非尼（Sorafenib）、5-氟尿嘧啶（5．Fu）、阿霉素（Doxorubicin）、特罗凯（Tarceva）、和伊立替康（Irinotecan）敏感；对多西紫杉醇（Docetaxel）敏感性存在病例差异，其中ESX003和ESX004对多西紫杉醇无反应。体内药效实验表明Irinotecan在100mg／kg剂量下对4例人食管癌裸鼠移植瘤模型有较强的抑制作用，相对肿瘤增殖率分别为22．02％、39．63％、21．69％和39．70％。结论成功建立了人源性食管癌小鼠移植瘤模型，移植瘤保留了，临床食管癌的病理特征。冻存的肿瘤组织，经复苏后仍可建立小鼠移植瘤模型，提示本研究建立的人源性食管癌移植瘤模型具有可持续应用性。4例人源性食管癌移植瘤模型的体外药敏实验和体内药效实验均表明人源性食管癌移植瘤模型具有很好的应用价值，可用于抗肿瘤药物的筛选及转化医学研究。

Preliminary Research in Establishment, Characterization, and Application of Human Primary Esophageal Cancer Xenograft Models

Objective To establish and characterize human primary esophageal xenografts by subcu- taneous implantation of fresh resected esophageal cancer （EC） specimens in immunodeficient mice with unchanged tumor histological features and validate their application value. Methods Thirty-six fresh resected human esophageal cancer specimens were subcutaneously implanted into SCID mice. After tumor developed, theywere serially passaged with nude mice using the same methods. For each case, when the xenograft tumors reached size 500-1000ram3, tumors were excised and devided into four parts：the first part was implanted into nude mice for the growth of next generation; the second part of tumor was viably saved in liquid nitrogen for future model recovery; the third part of tumor was snap frozen for DNA/RNA extraction; and the last part of tumor was fixed in formalin and embedded into paraffin （FFPE） for H＆E staining and pathology analysis. Using four cases of human primary EC xenografts passaged in nude mice, chemosensitivity study in vitro and efficacy study in vivo were performed to evaluate the application value of these models. Results Twenty-three in 36 cases by fresh EC speci- mens implantation were successfully developed into tumor in immunodeficient mice, the implantation success rate was 63.89%. These xenografts were grown up stably and could be continuous passaged to next generation. Histopathology examination of transplanted tumors showed that these xenografts were stable with similar histological features compared with the original donors. Twenty-one in 23 models were successfully established in SCID mice when implanted with recovering cryopreserved xenograft tissues, the recovery success rate was 91.30%. In in vitro chemo-sensitivity assay, the selected four cases were sensitive to Sorafenib, 5-FU, Doxorubicin, Tarceva and Irinotecan; however, there were individual differences in sensitivity to Docetaxel, even more, ESX003 and ESX004 were showed no response to Docetaxel. At dosage of 100 mg/kg, Irinotecan could significantly inhibit the growth of these four human primary EC xenografts in nude mice with relative tumor proliferation rate （T/C） of 22.02%, 39.63%, 21.69% and 39.70%, respectively. Conclusions Human primary EC xenograft models could be established with holding original donor＇s histological features. The cryopreserved xenografts tissues could be recovered successfully in SCID mice, indicating that these xenograft models can be used repeatedly. The results in chemosensitivity study in vitro and efficacy study in vivo with four selected EC xenograft models show that the human primary EC xenograft models are valuable and very helpful for anti-cancer drugs screening and translational research of human esophageal cancer.