Novel biomarkers are needed to improve current imperfect risk prediction models for cancer-associated thrombosis (CAT). We recently identified an RNA-sequencing profile that associates with CAT in colorectal cancer (CRC) patients, with REG4, SPINK4, and SERPINA1 as the top-3 upregulated genes at mRNA level. In the current study, we investigated whether protein expression of REG4, SPINK4 and alpha-1 antitrypsin (A1AT, encoded by SERPINA1) in the tumor associated with CAT in an independent cohort of CRC patients. From 418 patients with resected CRC, 18 patients who developed CAT were age, sex, and tumor stage-matched to 18 CRC patients without CAT. Protein expression was detected by immunohistochemical staining and scored blindly by assessing the H-score (percentage positive cells*scoring intensity). The association with CAT was assessed by means of logistic regression, using patients with an H-score below 33 as reference group. The odds ratios (ORs) for developing CAT for patients with A1AT^high, REG4^high, SPINK4^high tumors were 3.5 (95%CI 0.8–14.5), 2.0 (95%CI 0.5–7.6) and 2.0 (95%CI 0.5–7.4) when compared to A1AT^low, REG4^low, SPINK4^low, respectively. The OR was increased to 24.0 (95%CI 1.1–505.1) when two proteins were combined (A1AT^high/REG4^high). This nested case–control study shows that combined protein expression of A1AT and REG4 associate with CAT in patients with colorectal cancer. Therefore, REG4/A1AT are potential biomarkers to improve the identification of patients with CRC who may benefit from thromboprophylaxis.

需要新的生物标志物来改进当前不完善的癌症相关血栓形成（CAT）风险预测模型。我们最近确定了与结直肠癌 (CRC) 患者中 CAT 相关的 RNA 测序图谱，其中REG4、SPINK4和SERPINA1是 mRNA 水平上调最多的 3 个基因。在当前的研究中，我们在独立的 CRC 患者队列中研究了肿瘤中 REG4、SPINK4 和 α-1 抗胰蛋白酶（A1AT，由SERPINA1编码）的蛋白表达是否与 CAT 相关。在 418 名切除的 CRC 患者中，18 名出现 CAT 的患者与 18 名未患 CAT 的 CRC 患者的年龄、性别和肿瘤分期相匹配。通过免疫组织化学染色检测蛋白质表达，并通过评估H评分（阳性细胞百分比*评分强度）进行盲评分。使用 H 分数低于 33 的患者作为参考组，通过逻辑回归评估与 CAT 的关联。^A1AT高、REG4^高、SPINK4^高肿瘤患者发生 CAT 的比值比 (OR)分别为 3.5 (95%CI 0.8–14.5)、2.0 (95%CI 0.5–7.6) 和 2.0 (95%CI 0.5–7.4) ) 分别与 A1AT^低、REG4^低、SPINK4^低相比。当两种蛋白质组合时（A1AT^高/REG4^高），OR 增加至 24.0 (95% CI 1.1–505.1)。这项巢式病例对照研究表明，结直肠癌患者中 A1AT 和 REG4 的联合蛋白表达与 CAT 相关。因此，REG4/A1AT 是潜在的生物标志物，可提高对可能受益于血栓预防的 CRC 患者的识别。