The slow waves of non-rapid eye movement (NREM) sleep reflect experience-dependent plasticity and play a direct role in the restorative functions of sleep. Importantly, slow waves behave as traveling waves, and their propagation is assumed to occur through cortico-cortical white matter connections. In this light, the corpus callosum (CC) may represent the main responsible for cross-hemispheric slow-wave propagation. To verify this hypothesis, we performed overnight high-density (hd)-EEG recordings in five patients who underwent total callosotomy due to drug-resistant epilepsy (CPs; two females), in three noncallosotomized neurologic patients (NPs; two females), and in a sample of 24 healthy adult subjects (HSs; 13 females). In all CPs slow waves displayed a significantly reduced probability of cross-hemispheric propagation and a stronger inter-hemispheric asymmetry. In both CPs and HSs, the incidence of large slow waves within individual NREM epochs tended to differ across hemispheres, with a relative overall predominance of the right over the left hemisphere. The absolute magnitude of this asymmetry was greater in CPs relative to HSs. However, the CC resection had no significant effects on the distribution of slow-wave origin probability across hemispheres. The present results indicate that CC integrity is essential for the cross-hemispheric traveling of slow waves in human sleep, which is in line with the assumption of a direct relationship between white matter integrity and slow-wave propagation. Our findings also revealed a residual cross-hemispheric slow-wave propagation that may rely on alternative pathways, including cortico-subcortico-cortical loops. Finally, these data indicate that the lack of the CC does not lead to differences in slow-wave generation across brain hemispheres.SIGNIFICANCE STATEMENT The slow waves of NREM sleep behave as traveling waves, and their propagation has been suggested to reflect the integrity of white matter cortico-cortical connections. To directly assess this hypothesis, here we investigated the role of the corpus callosum in the cortical spreading of NREM slow waves through the study of a rare population of totally callosotomized patients. Our results demonstrate a causal role of the corpus callosum in the cross-hemispheric traveling of sleep slow waves. Additionally, we found that callosotomy does not affect the relative tendency of each hemisphere at generating slow waves. Incidentally, we also found that slow waves tend to originate more often in the right than in the left hemisphere in both callosotomized and healthy adult individuals.