Background: Artificial intelligence (AI) systems are proposed as a replacement of the first reader in double reading within mammography screening. We aimed to assess cancer detection accuracy of an AI system in a Danish screening population. Methods: We retrieved a consecutive screening cohort from the Region of Southern Denmark including all participating women between Aug 4, 2014, and August 15, 2018. Screening mammograms were processed by a commercial AI system and detection accuracy was evaluated in two scenarios, Standalone AI and AI-integrated screening replacing first reader, with first reader and double reading with arbitration (combined reading) as comparators, respectively. Two AI-score cut-off points were applied by matching at mean first reader sensitivity (AI_sens) and specificity (AI_spec). Reference standard was histopathology-proven breast cancer or cancer-free follow-up within 24 months. Coprimary endpoints were sensitivity and specificity, and secondary endpoints were positive predictive value (PPV), negative predictive value (NPV), recall rate, and arbitration rate. Accuracy estimates were calculated using McNemar’s test or exact binomial test. Results: Out of 272,008 screening mammograms from 158,732 women, 257,671 (94.7%) with adequate image data were included in the final analyses. Sensitivity and specificity were 63.7% (95% CI 61.6%-65.8%) and 97.8% (97.7-97.8%) for first reader, and 73.9% (72.0-75.8%) and 97.9% (97.9-98.0%) for combined reading, respectively. Standalone AI_sens showed a lower specificity (-1.3%) and PPV (-6.1%), and a higher recall rate (+ 1.3%) compared to first reader (p < 0.0001 for all), while Standalone AI_spec had a lower sensitivity (-5.1%; p < 0.0001), PPV (-1.3%; p = 0.01) and NPV (-0.04%; p = 0.0002). Compared to combined reading, Integrated AI_sens achieved higher sensitivity (+ 2.3%; p = 0.0004), but lower specificity (-0.6%) and PPV (-3.9%) as well as higher recall rate (+ 0.6%) and arbitration rate (+ 2.2%; p < 0.0001 for all). Integrated AI_spec showed no significant difference in any outcome measures apart from a slightly higher arbitration rate (p < 0.0001). Subgroup analyses showed higher detection of interval cancers by Standalone AI and Integrated AI at both thresholds (p < 0.0001 for all) with a varying composition of detected cancers across multiple subgroups of tumour characteristics. Conclusions: Replacing first reader in double reading with an AI could be feasible but choosing an appropriate AI threshold is crucial to maintaining cancer detection accuracy and workload.