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目的:探讨非A5.1型MHC-Ⅰ类相关分子A(MHC classⅠ-related molecules A,MICA)穿膜区等位基因对中晚期食管癌患者术后化疗联合NK细胞免疫治疗疗效的影响。方法:选取福建省肿瘤医院2013年4月至2015年10月收治的中晚期(TNMⅢ-Ⅳa期)食管癌患者152例,均在姑息性手术后行全身化疗,根据患者MICA穿膜区等位基因是否为A5.1分为4组:(1)A5.1化疗+NK细胞治疗(A5.1+NK)组;(2)A5.1单纯化疗(A5.1)组;(3)非A5.1化疗+NK细胞治疗(no A5.1+NK)组;(4)非A5.1单纯化疗(no A5.1)组,观察各组患者临床疗效。构建食管癌组织标本MICA等位基因A5.1的真核表达载体p TE-1A5.1,转染食管癌TE-1细胞株;Western blotting检测p TE-1A5.1转染对TE-1细胞MICA蛋白表达的影响,LDH法检测TE-1细胞转染p TE-1A5.1前后对NK细胞杀伤敏感性的变化,ELISA法检测食管癌患者血清可溶性MICA及转染p TE-1A5.1前后TE-1细胞上清中可溶性MICA含量。结果:经过3年的随防,A5.1+NK组患者中位总生存期(medium overall survival,m OS)为15.0个月,A5.1组为16.0个月,no A5.1+NK组为22.4个月,no A5.1组为16.0个月,no A5.1+NK组m OS明显长于其他3组(P<0.05)。经Cox多因素回归分析发现,患者年龄、性别、ECOG评分及基因型与预后均无明显相关(P>0.05),将基因类型与是否NK细胞治疗进行交叉分析,no A5.1+NK组m OS明显长于其他3组(P<0.01)。转染p TE-1A5.1后TE-1细胞内MICA表达显著升高,培养液上清可溶性MICA分泌明显增加[(256.2±45.3)vs(45.3±11.5)pg/ml,P<0.01];与转染前相比,NK细胞对过表达MICA的TE-1细胞的杀伤率明显降低[(29.5±7.2)%vs(42.5±7.1)%,P<0.05]。结论:相对中晚期食管癌MICA穿膜区A5.1等位基因患者,非A5.1基因患者手术后化疗联合NK细胞的疗效较好,其机制与其不容易产生可溶性MICA密切相关。
Aims: To investigate the effect of non-A5.1 MHC class I-related molecules A (MICA) on the efficacy of postoperative chemotherapy and NK cell immunotherapy in advanced esophageal cancer patients. Methods: A total of 152 patients with advanced stage (TNMⅢ-Ⅳa) esophageal cancer who were treated in Fujian Tumor Hospital from April 2013 to October 2015 were enrolled in this study. All patients underwent palliative surgery with systemic chemotherapy. According to the MICA (1) A5.1 chemotherapy + NK cell therapy (A5.1 + NK) group; (2) A5.1 chemotherapy alone group (A5.1); (3) A5.1 chemotherapy + NK cell therapy (no A5.1 + NK) group; (4) Non-A5.1 chemotherapy alone (no A5.1) group, to observe the clinical efficacy of each group. The eukaryotic expression vector p TE-1A5.1 of esophageal cancer tissue MICA allele A5.1 was constructed and transfected into esophageal cancer TE-1 cell line. The expression of p TE-1A5.1 was detected by Western blotting in TE-1 cells MICA protein expression before and after TE-1 cells transfected with p TE-1A5.1 were detected by LDH method. Serum soluble MICA and the expression of p TE-1A5.1 in esophageal cancer patients were detected by ELISA. The content of soluble MICA in TE-1 cell supernatant. RESULTS: After 3 years of follow-up, the median overall survival (m OS) was 15.0 months in the A5.1 + NK group, 16.0 months in the A5.1 group, no A5.1 + NK group Was 22.4 months, no A5.1 group was 16.0 months, no A5.1 + NK group m OS was significantly longer than the other three groups (P <0.05). Cox regression analysis showed that age, gender, ECOG score, genotype and prognosis had no significant correlation (P> 0.05), and the genotypes were analyzed with NK cell therapy. No A5.1 + NK group m OS was significantly longer than the other three groups (P <0.01). MICA expression was significantly increased in TE-1 cells transfected with p TE-1A5.1, and the soluble MICA secretion in culture supernatant was significantly increased [(256.2 ± 45.3) vs (45.3 ± 11.5) pg / ml, P <0.01]; NK cell killing rate of TE-1 cells over-expressing MICA was significantly lower than that before transfection [(29.5 ± 7.2)% vs (42.5 ± 7.1)%, P <0.05]. Conclusions: Compared with the A5.1 allele in the MICA transmembrane region of advanced stage esophageal cancer patients, the non-A5.1 gene chemotherapy combined with NK cells has a better curative effect. The mechanism is closely related to its non-susceptibility to soluble MICA.