Study Description and Overview Thirty participants (males and females) ages 20 to 65 inclusive, who, at Screening, meet ICD-10 criteria for major depressive disorder (MDD), have a current depressive episode of at least 30-day duration, have a Screening Montgomery-Asberg Depression Rating Scale (MADRS) total score \>= 22 and meet all other inclusion/exclusion criteria will be randomized with a 1-to-1 allocation under double-blind conditions to receive a single 25 mg oral dose of psilocybin or a single 100 mg oral dose of niacin. Niacin will serve as an active placebo control that provides an acute physiological response (flushing) that is intended to aid in blinding of intervention allocation. All randomized participants will be included in the Full Analysis Set (FAS) population used in analyzing primary and secondary study endpoints. Only participants who meet depressive symptom severity criteria at web screening (MADRS self-rating (MADRS-S) score \> =19) and who do not show an unacceptably large degree of symptom improvement between the web screening and in-person screening (indexed by change in MADRS-S (improvement) 30% (MADRS representing web screening will be approximated to MADRS-S + 3) will be eligible for randomization. This is to minimize the risk for spontaneous remission before dosing. Participants deemed eligible following successful completion of all screening assessments including a structural Magnetic Resonance Imaging (MRI) examination will be determined as eligible. Eligible participants at Baseline will submit cerebrospinal fluid (CSF), submit blood samples, be examined with positron emission tomography (PET) and the radioligand \[11C\]UCB-J and receive one preparation session (see further below) to be eligible for randomization on Dosing (Day 0) to receive either psilocybin or niacin active-placebo. They will complete follow-up visits, including outcome measures assessments, on study Day 1, 8, 15, 42 and 365 (within corresponding visit windows). At day 15 the sampling of CSF, blood and \[11C\]UCB-J PET will be repeated. PSIPET Protocol 5 200821 Page 15 After day 42, all participants will be given follow-up visits at Norra Stockholms Psykiatri for up to one year after dosing, to study dedicated physicians or nurses at a frequency determined by the health care professional. If needed to reach/stay in remission, the participants will be provided antidepressant treatment in accordance with the regional guidelines for antidepressant treatment (https://psykiatristod.se/regionala-vardprogram/ depression). At least monthly the participants will be asked to provide on-line symptom rating data (via 1177.se). At the 365-day visit, symptoms will be evaluated using MADRS, Clinical Global Impression Improvement (CGI-I) and Severity (CGI-S) scales. After completing the study (one year or withdrawal), participants will be subject to standard care, including referral in accordance with regional guidelines. The study outcome measures will be used to assess depressive symptoms, clinical global functioning, functional disability, anxiety symptoms and health-related quality of life. Safety outcome measures will be collected at all assessment time points from the time of consent through the end of study. To enhance participant safety, the current study proposes to test psilocybin within a "set and setting" (SaS) protocol similar to the protocol that has been used in all modern studies of psilocybin in both diseased and normal healthy populations. The SaS protocol for this study includes: 1) a preparation with session Facilitators (licensed psychologists) prior to dosing; 2) administration of study medications in an aesthetically neutral room under the supervision of two Facilitators who are present throughout the session (with the exception of short, temporary allowances for facilitator breaks; e.g. bathroom breaks); and 3) three post-dose integration sessions during which participants are encouraged to discuss their intervention experience with the Facilitators. To evaluate the Facilitators' adherence to the study manual, and the role of Facilitators' and participants' in-session behaviors for treatment outcome, all five sessions in the trial with Facilitators present will be recorded. The SaS will be identical for those randomized to psilocybin or niacin active placebo. Study Duration The planned maximum study duration for each participant will be approximately one year, with variation primarily dependent on the length of the screening period, the number of days between baseline and dosing, and the visit windows provided for each post-dose assessment. For each participant, the study will be divided into two phases: Phase A or treatment phase (day 0 to and including day 42), and phase B or follow-up phase (day 43 -365). The primary objective of this study is to evaluate the efficacy of a single 25 mg oral dose of psilocybin for major depressive disorder (MDD) compared to an active placebo (niacin) in otherwise medically-healthy participants between the ages of 20 and 65, assessed as the difference between groups in changes in depressive symptoms. Primary Outcome Measure Change in blinded rater MADRS total score from Baseline to Day 8.