Stromal Expression of Heat-Shock Protein 27 Is Associated with Worse Clinical Outcome in Patients with Colorectal Cancer Lung Metastases

Background
Pulmonary metastases are common in patients with primary colorectal cancer (CRC). Heat-shock protein 27 (Hsp27) is upregulated in activated fibroblasts during wound healing and systemically elevated in various diseases. Cancer-associated fibroblasts (CAFs) are also thought to play a role as prognostic and predictive markers in various malignancies including CRC. Surprisingly, the expression of Hsp27 has never been assessed in CAFs. Therefore we aimed to investigate the expression level of Hsp27 in CAFs and its clinical implications in patients with CRC lung metastases.
Methods
FFPE tissue samples from 51 pulmonary metastases (PMs) and 33 paired primary tumors were evaluated for alpha-SMA, CD31, Hsp27 and vimentin expression by immunohistochemistry and correlated with clinicopathological variables. 25 liver metastases served as control group. Moreover, serum samples (n=10) before and after pulmonary metastasectomy were assessed for circulating phospho-Hsp27 and total Hsp27 by ELISA.
Results
Stromal expression of Hsp27 was observed in all PM and showed strong correlation with alpha-SMA (P<0.001) and vimentin (P<0.001). Strong stromal Hsp27 was associated with higher microvessel density in primary CRC and PM. Moreover, high stromal Hsp27 and αSMA expression were associated with decreased recurrence-free survival after pulmonary metastasectomy (P=0.018 and P=0.008, respectively) and overall survival (P=0.031 and P=0.017, respectively). Serum levels of phospho- and total Hsp27 dropped after metastasectomy to levels comparable to healthy controls.
Conclusions
Herein we describe for the first time that Hsp27 is highly expressed in tumor stroma of CRC. Stromal α-SMA and Hsp27 expressions correlate with the clinical outcome after pulmonary metastasectomy. Moreover, serum Hsp27 might pose a future marker for metastatic disease in CRC.

研究背景
原发性结直肠癌（CRC）患者中，肺转移较为常见。热休克蛋白27（Hsp27）在伤口愈合过程中的活化成纤维细胞中表达上调，并在多种疾病中呈全身性升高。癌相关成纤维细胞（CAFs）也被认为在包括结直肠癌在内的多种恶性肿瘤中可作为预后及预测标志物。令人意外的是，目前尚未有针对癌相关成纤维细胞中热休克蛋白27表达的相关研究。因此本研究旨在探讨结直肠癌肺转移患者癌相关成纤维细胞中热休克蛋白27的表达水平及其临床意义。

研究方法
本研究采用免疫组化法检测51例肺转移灶（PMs）及33例配对原发肿瘤的福尔马林固定石蜡包埋（FFPE）组织样本中α-平滑肌肌动蛋白（alpha-SMA）、CD31、热休克蛋白27及波形蛋白的表达，并分析其与临床病理参数的相关性；以25例肝转移灶作为对照组。此外，采用酶联免疫吸附试验（ELISA）检测10例患者肺转移切除术前后的血清样本中循环磷酸化Hsp27及总Hsp27水平。

研究结果
所有肺转移灶均检测到热休克蛋白27的间质表达，且其表达与α-平滑肌肌动蛋白（P<0.001）及波形蛋白（P<0.001）的表达呈显著正相关。间质高表达热休克蛋白27与结直肠癌原发灶及肺转移灶的微血管密度升高相关。此外，间质α-平滑肌肌动蛋白及热休克蛋白27高表达与肺转移切除术后更短的无复发生存期（分别为P=0.018及P=0.008）及总生存期（分别为P=0.031及P=0.017）相关。肺转移切除术后，患者血清中磷酸化Hsp27及总Hsp27水平均降至与健康对照组相当的水平。

研究结论
本研究首次证实热休克蛋白27在结直肠癌肿瘤间质中呈高表达；间质α-平滑肌肌动蛋白及热休克蛋白27的表达与肺转移切除术后的临床结局相关。此外，血清热休克蛋白27有望成为结直肠癌转移性疾病的潜在新型标志物。