Topical calcineurin inhibitors (TCI) are commonly used for atopic dermatitis and other inflammatory dermatoses. The U.S. Food and Drug Administration issued a black box warning in 2006 indicating a potential risk of malignancy with TCI use based primarily on case reports, animal studies, and systemic tacrolimus use in organ transplant recipients. Since then, large epidemiologic studies have examined the association between TCIs and cancer; we conducted this systematic review and meta-analysis of observational studies to synthesize the evidence. We searched Medline, Embase, and Web of Science from inception to August 2020. We included observational studies investigating the association between treatment with TCIs (tacrolimus, pimecrolimus) and development of cancer, with non-active or active comparators. A total of 8 cohort studies (408,366 treated participants, 1,764,313 non-active comparator controls, 1,067,280 controls using topical corticosteroids) and 3 case-control studies (3,898 cases and 14,026 cancer-free controls) were included. There was no association between TCI use and cancer overall compared to non-active comparators (relative risk (RR) 1.03, 95% confidence interval 0.92 to 1.16). Lymphoma risk was elevated with TCI use in studies with non-active (RR 1.86, 1.39 to 2.49) more than topical corticosteroid comparators (RR 1.35, 1.13 to 1.61), suggesting the relationship may be partly confounded by indication. No significant association was found between TCI use and skin cancer. In summary, we found TCI use to be associated with a modestly elevated risk of lymphoma but not with other cancers. Given the low absolute risk of lymphoma, patients and clinicians should be reassured by these findings.