Objective:To investigate the changes of circulating tumor DNA (ctDNA), circulating B cell-specific Moloney leukemia virus insertion site 1 mRNA (Bmi-1 mRNA) and microRNA-21 (miR-21) before and after treatment with epidermal growth factor receptor (EGFR) monoclonal antibody in advanced colorectal cancer, and analyze their association with treatment response.Methods:The clinical data of 98 patients with advanced colorectal cancer from March 2019 to March 2021 in Yantai Yuhuangding Hospital were retrospectively analyzed. After treatment with cetuximab, complete remission was in 4 cases, partial remission in 26 cases, stable disease in 39 cases, and progressive disease in 29 cases. The patients with complete remission and partial remission were classified as remission group (30 cases), the stable disease and progressive disease were classified as non-remission group (68 cases). Before treatment and after 2 cycles of treatment, the plasma level of ctDNA was detected by high-throughput sequencing; the levels of Bmi-1mRNA and miR-21 were detected by real-time fluorescence quantitative polymerase chain reaction. Spearman correlation was used to analyze the relationship between ctDNA, Bmi-1mRNA, miR-21 and treatment responsiveness after 2 cycles of treatment; multivariate Logistic regression was used to analyze the independent risk factors affecting treatment responsiveness; receiver operating characteristic (ROC) curve was drawn to evaluate the value of ctDNA, Bmi-1mRNA and miR-21 in predicting remission after 2 cycles of treatment.Results:There were no significant differences in ctDNA, Bmi-1mRNA and miR-21 before treatment between 2 groups ( P>0.05); the ctDNA, Bmi-1mRNA and miR-21 after 2 cycles of treatment in remission group were significantly lower than those in non-remission group: (10.03 ± 3.32) μg/L vs. (15.33 ± 5.14) μg/L, 0.13 ± 0.04 vs. 0.19 ± 0.05 and 0.81 ± 0.26 vs. 1.08 ± 0.24, and there were statistical differences ( P<0.01). Spearman correlation analysis result showed that ctDNA, Bmi-1mRNA and miR-21 were negatively correlated with treatment response ( r = -0.500, -0.506 and -0.531; P<0.01). Multivariate Logistic regression analysis result showed that, after controlling for the number of distant metastatic organs and clinical stage, ctDNA, Bmi-1mRNA and miR-21 were still independent risk factors for treatment response in patients with advanced colorectal cancer ( OR = 3.342, 2.725 and 1.838; 95% CI 3.116 to 3.584, 2.647 to 2.805 and 1.768 to 1.911; P<0.01). ROC curve analysis result showed that the area under the curve (AUC) of ctDNA, Bmi-1mRNA combined with miR-21 after 2 cycles of treatment to predict the treatment response was the largest with 0.922. Conclusions:The changes of ctDNA, Bmi-1mRNA and miR-21 in patients with advanced colorectal cancer before and after treatment with EGFR monoclonal antibody are related to the treatment response. Combined detection is helpful for screening patients sensitive to EGFR-targeted therapy, and can provide reference for new targets of molecular intervention.