Abstract: Pimecrolimus and tacrolimus are topical calcineurin inhibitors developed specifically for the treatment of atopic eczema. Experience with long-term use of topical calcineurin inhibitors is limited and the risk of rare but serious adverse events remains a concern. We have previously demonstrated the absence of carcinogenic effect of tacrolimus alone and in combination with simulated solar radiation (SSR) on hairless mice. The aim of this study is to determine whether pimecrolimus accelerates photocarcinogenesis in combination with SSR or pimecrolimus and tacrolimus accelerate photocarcinogenesis in combination with UVA. We used 11 groups of 25 hairless female C3.Cg/TifBomTac immunocompetent mice (n = 275). Pimecrolimus cream or tacrolimus ointment was applied on their dorsal skin three times weekly followed by SSR (2, 4, or 6 standard erythema doses, SED) or UVA (25 J/cm2) 3–4 h later. This was done up to 365 days in the SSR-treated groups and up to 500 days in the UVA-treated groups. Pimecrolimus did not accelerate the time for development of the first, second or third tumor in any of the groups. Median time to the first tumor was 240 days for the control-2SED group compared with pimecrolimus-2SED group (233 days), control-4SED group (156 days) compared with pimecrolimus-4SED group (163 days) and control-6SED group (162 days) compared with pimecrolimus-6SED group (170 days). Only one mouse in each of the three UVA groups developed a tumor. We conclude that pimecrolimus in combination with SSR and both pimecrolimus and tacrolimus in combination with UVA do not accelerate photocarcinogenesis in hairless mice.