Background: Natural orifice specimen extraction (NOSE) surgery has been widely implemented in colorectal cancer surgery due to its good short-term efficacy. However, anastomotic leakage (AL) is a serious postoperative complication in colorectal cancer, and the risk factors for this complication after NOSE surgery have rarely been investigated. The aim of this study was to explore the predictive factors for AL after laparoscopic resection with transrectal NOSE for rectal cancer and sigmoid colon cancer. Methods: A total of 208 patients who underwent total laparoscopic resection with transrectal NOSE for rectal cancer and sigmoid colon cancer from January 2014 to June 2019 were systematically reviewed. Univariate and multivariate analyses were performed to identify the relevant risk factors. Results: The rate of AL was 10.1% (21 of 208 patients). The univariate analyses showed that male sex (85.7% vs 57.8%, P = .013), the distance from the anal verge (10.5 vs 14.5 cm, P = .011), and a duration of operation ≥140 min (71.4% vs 29.4%, P<.001) were associated with an increased incidence of AL. The multivariate analysis showed that a duration of operation ≥140 min (OR = 5427, 95% CI = 1.355-21.727, P = .017) was an independent risk factor for AL. Conclusion: A duration of operation ≥140 min is a possible risk factor for AL after total laparoscopic resection with transrectal NOSE for colorectal cancer.