Non-contrast cardiac CT is used worldwide to assess coronary artery calcium (CAC), a subclinical marker of coronary atherosclerosis. Manual quantification of regional CAC scores includes identifying candidate regions, followed by thresholding and connected component labeling. We aimed to develop and validate a fully-automated, algorithm for both overall and regional measurement of CAC scores from non-contrast CT using a hybrid multi-atlas registration, active contours and knowledge-based region separation algorithm. A co-registered segmented CT atlas was created from manually segmented non-contrast CT data from 10 patients (5 men, 5 women) and stored offline. For each patient scan, the heart region, left ventricle, right ventricle, ascending aorta and aortic root are located by multi-atlas registration followed by active contours refinement. Regional coronary artery territories (left anterior descending artery, left circumflex artery and right coronary artery) are separated using a knowledge-based region separation algorithm. Calcifications from these coronary artery territories are detected by region growing at each lesion. Global and regional Agatston scores and volume scores were calculated in 50 patients. Agatston scores and volume scores calculated by the algorithm and the expert showed excellent correlation (Agatston score: r = 0.97, p < 0.0001, volume score: r = 0.97, p < 0.0001) with no significant differences by comparison of individual data points (Agatston score: p = 0.30, volume score: p = 0.33). The total time was <60 sec on a standard computer. Our results show that fast accurate and automated quantification of CAC scores from non-contrast CT is feasible.