外周血循环肿瘤细胞及人表皮生长因子受体2胞外区表达在进展期胃癌治疗疗效评估中的价值

Value of serum human epithelial growth factor receptor 2 extracellular domain and circulating tumor cells in evaluating therapeutic response in advanced gastric cancer

目的研究人血清中表皮生长因子受体2胞外区（HER2 ECD）与外周血循环肿瘤细胞（CTC）的相关性，并进一步研究组织学HER2阴性或阳性胃癌患者中，HER2 ECD动态变化及与治疗疗效的关系。方法将北京大学肿瘤医院消化肿瘤内科2012年1月至2014年1月期间收治、并曾入组中国进展期胃癌CTC临床研究（ClinicalTrial gov. ID：NCT01625702）的53例患者入组本回顾性研究。入组条件：除病理组织学确诊为进展期胃癌外，至少接受2个周期以氟尿嘧啶为基础的化疗或联合靶向治疗，并同时进行了治疗前后外周血CTC的计数；按实体瘤的疗效评价标准（RECIST），治疗前至少具有一个可测量的靶病灶；患者需经医院伦理委员会同意及患者知情同意。留取患者治疗前及治疗2周期后的血清，利用化学发光免疫分析法进行血清样本中HER2 ECD水平的检测。HER2 ECD阳性阈值为≥15 ng/ml；CTC计数阳性阈值为≥3个/7.5 ml。采用Log-rank检验不同组间无进展生存期（PFS）及总体生存期（OS）的差异。结果53例进展期胃癌患者HER2阴性39例，HER2阳性的9例（17.0%），另5例检测状态未知。均接受氟尿嘧啶为基础的化疗，9例HER2阳性患者在化疗基础上联合抗HER2靶向治疗。治疗前全组血清HER2 ECD中位浓度为10.45（8.0-83.2）ng/ml；HER2 ECD阳性7例（13.2%），其中4例为HER2阳性患者外，3例HER2阴性患者同样存在血清HER2 ECD阳性；CTC中位数为2（0-668）个/7.5 ml，CTC阳性25例（47.2%）。HER2阴性患者中有10例治疗后血清HER2 ECD阳性，其中有2例患者在治疗前其HER2 ECD分别为83.3 ng/ml和46.9 ng/ml，治疗后则为22.4 ng/ml和20.4 ng/ml；另8例治疗前HER2 ECD均〈15 ng/ml（10.3-14.5 ng/ml），但在治疗2个周期后发生获得性HER2 ECD水平升高（15.1-19.5 ng/ml）。治疗后血清HER2 ECD水平升高患者外周血CTC水平通常较低，但相关性未达统计学差异。HER2阴性病例，治疗前血清HER2 ECD阳性患者与HER2 ECD阴性患者的PFS（7.6月比4.4月，P = 0.328）及OS（13.6比10.9，P = 0.679）比较，差异无统计学意义。HER2阳性病例，治疗前HER2 ECD阳性者其PFS（10.7月）和OS（16.5月）均显著长于HER2 ECD阴性者（4.2月和8.9月），差异有统计学意义（P = 0.025，P = 0.015）。HER2阴性病例，治疗前CTC计数阴性者PFS和OS（5.3月和14.3月）显著长于CTC计数阳性患者（3.3月和7.6月），差异有统计学意义（P = 0.049，P = 0.001）。而HER2阳性者，CTC计数多少均与PFS及OS无关（均P 〉 0.05）。HER2阴性的8例治疗2周期后发生获得性HER2 ECD水平升高者，其HER2 ECD水平升高与PFS及OS无关（P 〉 0.05）。9例HER2阳性患者，4例治疗后HER2 ECD水平下降、CTC计数下降或不变者，其PFS（7.5-15.3月）和OS（11.0-26.3月），长于治疗后HER2 ECD水平上升的2例患者，其PFS分别为3.0和4.8月，OS则分别为7.3和8.6月。结论HER2阳性胃癌患者治疗前HER2 ECD水平升高预示更好的PFS及OS，且治疗过程中HER2 ECD获得性升高预示疗效不佳。HER2 ECD水平可在HER2阴性患者中获得性升高，且与外周血CTC计数水平变化存在相关性。

Objective To examine the correlation between serum human epithelial growth factor receptor 2 extracellular domain （HER2 ECD） and circulating tumor cells （CTC）, as well as the dynamic variation of HER2 ECD and its correlation to the therapeutic efficacy. Methods Fifty-three advanced gastric cancer （AGC） patients who treated in Peking University Cancer Hospital and ever enrolled into CTC study （ClinicalTrial gov. ID： NCT01625702） were retrospectively included in this study. Inclusion criteria： the patients were histologically confirmed as locally advanced or recurrent and/ or metastatic adencarcinoma; they received two or more cycles of fluorouracil-based chemotherapy or combination targeted therapy; serum CTC was counted before and after therapy; the clinical response was evaluated every 2 cycles of treatment by the presence of at least one measurable lesion according to RECIST version 1.1 criteria. This study was approved by Ethics Committee of Peking University Cancer Hospital, and informed consents were signed by patients. The sera before and after two cycles of treatment were collected for CTC enumeration and HER2 ECD detection, in which the levels of HER2 ECD were measured by chemiluminescence immnnoassays method. The positive threshold value of HER2 ECD and CTC number were ≥ 15 μg/L and ≥3 CTCs/7.5 ml respectively. The progression-free survival （PFS） and overall survival （OS） were compared among different groups using Log-rank tests. Results In 53 enrolled patients, 39 were histologically identified as negative HER2, 9 as positive HER2 and another 5 cases were unknown. All the patients received fluorouracil-based chemotherapy, and 9 positive HER2 patients received combined anti-HER2 targeted therapy. Before therapy, the median HER2 ECD concentration of 53 cases was 10.45 （8.0 to 83.2） μg/L. Seven patients exhibited positive HER2 ECD levels, in whom 4 were histologically HER2 positive, but 3 were histologically HER2 negative. The median CTC number of 53 cases was 2 （0 to 668） CTCs/7.5 ml, and the positive rate of CTC was 47.2% （25/53）. Following 2 cycles of therapy, a total of l0 histologically HER2 negative patients exhibited positive HER2 ECD levels, in whom 2 also possessed positive HER2 ECD levels, 83.3 μg/L and 46.9 μg/L before therapy, and 22.4 μg/L and 20.4μg/L after therapy respectively, whereas another 8 patients （10.3 to 14.5 μg/L before therapy） acquired the elevated expression of HER2 ECD following therapy （15.1 to 19.5 μg/L）. It seems that the increased level of HER2 ECD after therapy was, though not statistically significant, correlated to low number of CTCs. In histologically HER2 negative patients, pretherapeutic HER2 ECD level （positive vs. negative） was not significantly correlated to PFS （7.6 months vs. 4.4 months, P=0.328） and OS （13.6 months vs. 10.9 months, P = 0.679）. However, in histologically HER2 positive patients, patients with positive HER2 ECD level before therapy exhibited longer PFS （10.7 months vs. 4.2 months, P = 0.025） and OS （16.5 months vs. 8.9 months, P= 0.015） compared to those with negative HER2 ECD level. Additionally, CTC number was significantly correlated to prognosis in histologically HER2 negative patients. Patients with positive pretherapeutic CTC number showed longer PFS （5.3 months vs. 3.3 months, P= 0.049） and OS （14.3 months vs. 7.6 months, P = 0.001） as well. While in histologically HER2 positive patients, CTC number was not obviously correlated to the PFS and OS. In above 8 negative HER2 patients acquiring elevated expression of HER2 ECD following therapy, the increased HER2 ECD level was not correlated to PFS and OS （all P 〉 0.05）. In 9 histologically HER2 positive patients, 4 patients who exhibited decreased HER2 ECD level and reduced or constant CTC number had longer PFS （7.5 to 15.3 months） and OS （11.0 to 26.3 months） compared with those 2 patients who suffered from acquired HER2 ECD level following therapy （PFS 3.0 to 4.8 months and OS 7.3 to 8.6 months）. Conclusions In histologically HER2 positive patients, increased pretherapeutic HER2 ECD level predicts better prognosis. The acquired elevated HER2 ECD level following therapy is correlated to inefficient therapeutic response. The acquirement of elevated HER2 ECD level can also be found in histologically HER2 negative patients, which may be correlated to the corresponding variation of CTC number.