Psilocybin is the psychoactive substance contained in the psilocybe (hallucinogenic) mushroom, which has received considerable attention among the scientific community in recent years. Human studies have demonstrated that even a single-dose of psilocybin can improve debilitating physical and psychological symptoms with durable long-term effects 1,2. A systematic review of 28 patient studies with psilocybin reported sustained long-term benefits (from 3 weeks to 4.5 years) following a single-dose 1,2. To date,136 clinical studies with psilocybin have been completed or are ongoing for various indications, including psychiatric, neurodegenerative, chronic pain, and more. However, despite overwhelming clinical evidence for the therapeutic effects of psilocybin, the underlying molecular mechanisms responsible for its beneficial actions remain enigmatic. Studies with psilocybin have overwhelmingly focused on neurological impacts and/or behavioral outcomes; few studies have evaluated other mechanisms by which it exerts beneficial effects. Given that the short elimination half-life of psilocin 3, how does a single dose lead to sustained benefits across numerous indications that can last for years?
It has been postulated that therapeutic use of psilocybin promotes beneficial effects on aging; this theory is based on a large corpus of studies that have established strong correlations between mental health and aging 4. Psychological state impacts telomere length. Positive mental psychological states are correlated with longer telomeres 5. In contrast, negative psychological conditions are associated with telomere attrition 5. Accumulating evidence indicate that clinical depression accelerates aging, with quantifiable impacts on telomere length 6-8; chronic stress, anxiety, and depression are associated with shorter telomeres 5,6,9-11. Accordingly, the “psilocybin-telomere hypothesis” asserts that psilocybin interventions exert quantifiable positive impacts on telomere length 4. As psilocybin reduces anxiety and depression 12,13, it is plausible that psilocybin may impact telomere length. Although it has been speculated that psilocybin treatment impacts aging, no prior studies have experimentally investigated the impact of psilocybin on senescence or aging.
To evaluate the impact of psilocybin on cellular senescence, we employed a validated model of replicative senescence using human lung fibroblasts, previously developed in our laboratory 14. Cells were cultured for ~3 months to induce replicative senescence with different concentrations of psilocybin continuously supplemented in the culture media. Psilocybin treatment led to a dose-dependent decrease in cell cycle arrest markers (p21, p16, and p53) and an increase in markers of DNA replication (phosphorylated-Retinoblastoma protein; pRB) and proliferation (PCNA), as compared to vehicle-treated cells (Fig. 1A). Senescent cells also secrete high levels of inflammatory cytokines, known as senescence-associated secretory phenotype (SASP). Psilocybin treatment also led to a decrease in secreted SASP (Fig. 1B); macrophage colony-stimulating factor (M-CSF), monocyte chemoattractant protein 1 (MCP-1), tumor necrosis factor-alpha (TNF-a), and transforming growth factor-beta (TGF-b) are all known SASP markers 15.
Since cellular senescence is associated with oxidative stress, we evaluated the impact of psilocybin treatment on oxidative stress levels. We found that psilocybin treatment led to a dose-dependent reduction in oxidative stress levels (Fig. 1C). Since no published studies have evaluated the effects of psilocybin on oxidative stress levels, we evaluated the possibilities that psilocybin may directly target pathways involved in reactive oxygen species (ROS) production and/or may have antioxidant properties. NADPH-oxidase-4 (Nox4) is ubiquitously expressed and is a major cellular source of ROS production 16. Cells that stably over-express Nox4 (HEK/Nox4 cells), which generate high levels of ROS, were treated with psilocybin; psilocybin showed no efficacy for inhibiting Nox4-dependent ROS (Fig. 1D). To evaluate if psilocybin mediates Nox-independent mechanisms of ROS production, we evaluated the effects of psilocybin on xanthine oxidase, which generates ROS (superoxide and H2O2) independent of Nox activity. However, psilocybin showed no inhibitory activity in this assay (Fig. 1E), indicating that psilocybin does not inhibit Nox-independent ROS production. Given that psilocybin demonstrated no activity in both assays (even at high concentrations, up to 300mM; Fig. 1D), these data also suggest that psilocybin does not exhibit antioxidant properties.
Senolytics are a class of “senescence destroying” agents, which selectively induce death of senescent cells. We next evaluated the possibility that psilocybin may exhibit senolytic properties (selective killing of senescent cells), which could potentially explain our findings of decreased senescence following psilocybin treatment. “Young” (cells at low population doublings) and senescent (cells that have undergone replicative-induced senescence) were treated with psilocybin and cell viability was assessed. However, psilocybin treatment did not alter cell viability in young or senescent endothelial cells (Fig. 1F) or fibroblasts (Fig. 1G), indicating that psilocybin is not a senolytic agent.
In summary, we provide the first experimental evidence suggesting that psilocybin may decelerate cellular senescence. Our studies suggest that the reduced oxidative stress levels observed (Fig. 1C) are not attributed to direct inhibition of Nox4-dependent ROS production, antioxidant properties, orsenolytic properties of psilocybin. Instead, these data are consistent with the concept that “young” cells exhibit lower steady-state levels of ROS, as compared to senescent cells. One unknown aspect of these findings is the mechanisms by which psilocybin leads to decelerated senescence. Psilocybin is a potent serotogenic agonist, which interacts with the serotonin receptor (5-HT2A) and multiple 5-HT receptor subtypes; the 5-HT2A receptor is expressed in multiple organs and cell types, including neurons, fibroblasts, smooth and skeletal muscle cells, cardiomyocytes, endothelial cells, epithelial cells, macrophages, and T-cells 17. A recent study demonstrated that in cortical neurons, 5-HT2A receptor stimulation induced sirtuin1 (SIRT1)-dependent expression of antioxidant enzymes, which led to decreased cellular ROS and neuroprotection against excitotoxic and oxidative stress 18. It is possible that psilocybin-dependent 5-HT2A receptor activation may increase cellular antioxidant responses and/or decrease ROS production (via Nox4-independent mechanisms), which may impact the onset of senescence. Further, studies have demonstrated that SIRT1 is a critical mediator of senescence, and increased expression of SIRT1 is sufficient to extend lifespan in yeast, c. elegans, and mice 19. Thus, psilocybin may mediate SIRT1-dependent pathways which impact cellular senescence. Overall, the finding that psilocybin decelerates cellular senescence could potentially help explain its durable effects for multiple disease indications, and identifying the mechanisms responsible for these effects warrant further investigation.
Although psilocybin was widely used in experimental research in 1960s, during this time, psilocybin-containing mushrooms became a popular recreational drug, culminating in regulatory controls classifying it as a schedule I drug in 1971 20. This led to a sharp decline in psilocybin research funding, drug production, and the discontinuation of all human subjects research. During the last 30 years, there has been a progressive resurgence in psilocybin research, fueled by an appreciation for its therapeutic potential 21. However, regulatory barriers imposed by its schedule I drug designation have made research progress challenging, which in part explains our limited understanding of the mechanisms underlying its therapeutic benefits 21. Accordingly, funding for psilocybin research remains limited 22. It is important to note that the schedule I designation of psilocybin is likely to be reclassified in the near future. There are 2 ongoing Phase III clinical trials and the FDA has already designated psilocybin as a “breakthrough therapy” (an action that is meant to accelerate the FDA-approval process) for the treatment of depression. Its breakthrough therapy designation also highlights the safety profile of psilocybin, as minimal adverse effects have been reported 23-26. FDA-approval of psilocybin for any indication would be an accepted medical use, which would justify re-classification. Further, legislative reform is currently underway to decriminalize and/or legalize psilocybin. In 2020, Oregon became the first state to both decriminalize psilocybin and legalize it for medical use. >100 cities have already decriminalized psilocybin or have introduced legislation in support of this. The Biden administration has announced plans for the legalization of psilocybin therapies within 2 years, which is being driven by a federal task force 27. Overall, given the safety profile, imminent FDA-approval, and legislative reform underway, the re-classification of psilocybin from its schedule I status is eminent. Once this occurs, many more people will have access to psilocybin. Thus, there is an urgent need for studies that shed light on its mechanisms of action. To our knowledge, these data are the first experimental evidence to suggest that psilocybin may decelerate cellular senescence. Given that senescence contributes to the pathogenesis of numerous age-related diseases, these studies could lay the foundation for the use of psilocybin as a therapeutic strategy for new disease indications. These limited and yet striking findings warrant further investigation, including in vivo validation of its impacts on senescence and lifespan. Psilocybin may represent a “disruptive” pharmacotherapy for several age-related disease indications and/or as a novel geroprotective agent.