Background and Objective: Long-term opioid treatment (L-TOT) of chronic non-cancer pain (CNCP) patients has been suspected to alter the endocrine system. This systematic review and meta-analysis aimed at investigating the published evidence of L-TOT effects on the endocrine system in adult CNCP patients. Databases and Data Treatment: A systematic search of the literature in MEDLINE, EMBASE, Cochrane Central Register of Controlled Trials and the CINAHL was performed. Studies examining measures of endocrine function of the hypothalamic-pituitary-gonadal, -adrenal, -thyroid, -somatotropic and -prolactin axis in adult CNCP patients in L-TOT (≥4 weeks of use) were included. Outcomes and the level of evidence were analyzed (The Cochrane Collaboration Tool, modified version of the Newcastle-Ottawa Scale and Rating of Recommendations Assessment, Development and Evaluation working group). Results: A total of 2,660 studies were identified; 1981 excluded and finally thirteen studies (one randomized controlled trial (RCT), three longitudinal- and nine cross-sectional studies) were analyzed. L-TOT was associated with low insulin, suppression of the hypothalamic-pituitary-gonadal axis and alterations of the hypothalamic-pituitary-adrenal axis in both men and women with CNCP compared to different control groups (CNCP or healthy pain-free). No other significant differences were reported. The studies had a high risk of bias and the overall quality of evidence was low. Conclusion: There seems to be an impact of L-TOT in CNCP patients on several components of the endocrine system, but the level of evidence is weak. Given the high prevalence of L-TOT use systematic studies of larger patient populations are urgently needed. Significance: This systematic review and meta-analysis suggested that long-term opioid treatment may suppress the hypothalamic-pituitary-gonadal axis, and result in lower insulin levels and alter the glucocorticoid adrenal axis in adult chronic non-cancer pain patients. This adds to the need of more research of both clinical and paraclinical outcomes and their association when initiating and maintaining long-term opioid treatment.