We have previously reported the generation of Rassf6 knockout mice using CRISPR/Cas technology. Furthermore, we have reported that RASSF6 is implicated in the regulation of the canonical nuclear factor (NF)-κB signaling pathway and that 7,12-dimethylbenz(a)anthracene induced skin inflammation in Rassf6 knockout mice more remarkably than in wild-type mice. In this study, we investigated the role of RASSF6 in skin tumorigenesis using a two-stage carcinogenesis model in Rassf6 knockout mice. Rassf6 knockout mice initially developed significantly more papillomas than did the control wild-type mice. Additionally, macrophages were detected and the canonical NF-κB signal was elevated in papillomas of Rassf6 knockout mice. These results indicated that canonical NF-κB signaling may be involved in papilloma formation. The suppression of NF-κB signaling may have implications for preventing cancer initiation and progression. The findings of this study indicate a tumor suppressive role of RASSF6.