【摘要】 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。更多还原

【Abstract】 Objective:To investigate the effect of non-A5.1 type MHC class Ⅰ-related molecules A(MICA) transmembrane allele on the treatment efficacy of chemotherapy combined with NK cell immunotherapy in advanced esophageal cancer patients. Methods: One hundred and fifty-two patients with advanced esophageal cancer( TNM Ⅲ-Ⅳa) underwent systemic chemotherapy after palliative surgery between April,2013 and October,2015 were included in this study. According to whether the MICA transmembrane allele was A5.1,patients were divided into four groups to observe the clinical efficacy: A5.1 chemotherapy + NK cell therapy( A5.1+NK group); A5.1 chemotherapy alone( A5.1 group); non-A5.1chemotherapy+NK cell therapy( non A5.1+NK group); non-A5.1 chemotherapy alone( non A5.1 group). Eukaryotic expression vector p TE-1A5.1 carrying MICA allele A5.1 was constructed and used to transfect esophageal cancer TE-1 cell line. Western blotting was used to detect the effect of p TE-1A5.1 transfection on the expression of MICA protein in TE-1cells. The sensitivity change of TE-1 cells to NK cells before and after the transfection was detected by LDH. ELISA was used to detect the soluble MICA in serum of esophageal cancer patients and the soluble MICA content in the supernatant of TE-1 cells before and after transfection with p TE-1A5.1. Results: After 3 years of follow-up,the medium overall survival(m OS) of A5.1+NK group,A5.1group,non A5.1+NK group and non A5.1 group were 15.0 months,16.0 months,22.4months and 16.0 months,respectively. The m OS of non A5.1+NK group was significantly longer than that of other groups(P<0.05). There was no significant correlation between age,sex,ECOG score,genotype and prognosis by Cox multivariate regression analysis( P>0.05). The cross analysis of genotype and NK therapy showed that the m OS of non A5.1 +NK group was significantly longer than that of the other three groups( P<0.05). The expression of MICA in TE-1 cells was significantly increased after transfection 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). Compared with pre-transfection,the killing rate of NK cells on TE-1 cells over-expressing MICA was significantly decreased([29.5 ± 7.2]% vs [42.5 ± 7.1]%,P < 0.05).Conclusion: Compared with esophageal patients with MICA-A5. 1 allele,the efficacy of combined treatment of NK cell therapy and chemotherapy for patients with MICA-non A5.1 is better,which is closely related to low level of soluble MICA.更多还原