Observational studies and clinical trials have evaluated the associations between vitamin D supplementation and cancer incidence/mortality and obtained mixed results. Previous meta-analyses have also yielded inconsistent conclusions. In this paper, we conduct an updated meta-analysis by including current randomized clinical trials (RCTs) to assess the association between vitamin D supplementation and cancer incidence and mortality. The PubMed, Scopus and Embase databases were systematically searched from their inception to 6 February 2022. Fixed-effects meta-analyses were conducted. Trial sequential analyses were performed using a risk ratio reduction threshold of 10% for cancer incidence and mortality. Twenty-six RCTs were eligible, and pooled results indicated that vitamin D supplementation, compared to placebo with/without calcium, was not associated with a reduction in total cancer incidence (risk ratio: 0.98, 95% CI: 0.94, 1.02; I-2 = 0%). In contrast, vitamin D supplementation significantly reduced total cancer mortality (risk ratio: 0.88, 95% CI: 0.8, 0.96; I-2 = 0%). Moreover, trial sequential analysis provided reliable evidence that supplementation with vitamin D lowered the relative risk of total cancer mortality by 10%. Our updated meta-analysis suggested that vitamin D supplementation did not reduce total cancer incidence but significantly lowered total cancer mortality. Supplemental data for this article is available online at https://doi.org/10.1080/10408398.2022.2056574 .