Purpose: To investigate whether pretreatment serum carbohydrate antigen 19-9 (CA 19-9) levels are associated with pathological responses to preoperative chemoradiotherapy (CRT) in patients with rectal cancer. Materials and Methods: In total, 260 patients with locally advanced rectal cancer (cT3-4NanyM0) who underwent preoperative CRT and radical surgery were analyzed retrospectively. CRT consisted of 50.4 Gy pelvic radiotherapy and concurrent chemotherapy. Radical surgery was performed at a median of 7 weeks after CRT completion. Pathological CRT response criteria included downstaging (ypStage 0-I) and ypT0-1. A discrimination threshold of CA 19-9 level was determined using a receiver operating characteristics analysis. Results: The median CA 19-9 level was 8.0 (1.0-648.0) U/mL. Downstaging occurred in 94 (36.2%) patients and ypT0-1 in 50 (19.2%). The calculated optimal threshold CA 19-9 level was 10.2 U/mL for downstaging and 9.0 U/mL for ypT0-1. On multivariate analysis, CA 19-9 (${leq}9.0U/mL$) was significantly associated with downstaging (odds ratio, 2.089; 95% confidence interval, 1.189-3.669; P=0.010) or ypT0-1 (OR, 2.207; 95%CI, 1.079-4.512; P=0.030), independent of clinical stage or carcinoembryonic antigen. Conclusions: This study firstly showed a significant association of pretreatment serum CA 19-9 levels with pathological CRT responses of rectal cancer. The CA 19-9 level is suggested to be valuable in predicting CRT responses of rectal cancer cases before treatment.