According to the World Health Organization, there are more than 300 million depressed patients worldwide, of which about 2/3 of patients have had suicidal ideation, and about 1/4 of patients have had suicidal behavior. Suicide is also one of the leading causes of death among young people today, and nearly half of them occur in people with depression. Traditional antidepressants currently on the market often take up to weeks or even months to take effect and have no improving effect on one-third of people with refractory depression.
In recent years, multiple hallucinogens have shown some potential in the treatment of depression1. Psilocybin, a natural hallucinogen extracted from the "magic mushroom", was awarded a breakthrough therapy for the treatment of major depressive disorder and drug-resistant depression1-3 by the US FDA in 2019. The results of the phase II clinical study show that psilocybin can greatly improve the symptoms of depressed patients within one day, and the effect can last for more than three months4. However, the hallucinogenic side effects of hallucinogens greatly limit their clinical research and widespread application. Therefore, scientists around the world have been working to find and develop new antidepressants that are non-hallucinogenic and can work quickly.
On January 28, 2022, Wang Sheng's research group from the Center for Excellence in Molecular and Cell Science (Institute of Biochemistry and Cell Biology) of the Chinese Academy of Sciences and the Cheng Jianjun Research Group of the iHuman Institute of ShanghaiTech University published a research paper entitled Structure-based discovery of non-hallucinogenic psychedelic analogs in the journal Science. Proceeding from the elucidation of the complex structure of the hallucinogen and its target serotonin 2A receptor (5-HT2AR), the work found that in addition to the previously predicted classical binding mode with 5-HT2AR, there is another lipid-regulated binding mode located above the classical binding mode. The researchers found that in the presence of lipid molecules, the second binding mode of hallucinogens is crucial for the preferential activation of the β-arrestin signal downstream of 5-HT2AR; further experimental results show that the hallucinogenic effect of hallucinogens is closely related to the simultaneous and efficient activation of two downstream signals, G protein and β-arrestin. Based on the second binding mode, the researchers designed and synthesized a series of 5-HT2AR β-arrestin signal preference agonists represented by IHCH-7086 and IHCH-7079; and demonstrated in animal experiments that the compounds had no hallucinogenic effect, but had a rapid antidepressant effect similar to that of hallucinogens.
1
Activation of lipids on serotonin 2A receptors
Psilocybin (psilocybin) is metabolized in the human body to psiloccin (dephosphate psilocybin), which works in combination with 5-HT2AR. Wang Sheng's research group first analyzed the crystal structure of 5-HT2AR and four complexes of endogenous ligand serotonin (serotonin), hallucinogen psiloccin, hallucinogen LSD (diethylamide of lysergic acid), and non-hallucinogen lisuride (ergoacetine, Parkinson's disease drug). The binding position of the ergoline skeleton of LSD and lisuride is similar to that of other ergolin drugs analyzed in the laboratory, both at the orthosteric binding pocket (OBP)5,6; unlike the previously predicted orthosteric binding site, serotonin and psilocin are lipid-regulated and bind to the extended binding pocket (EBP). Further experimental results show that the lipid is monoolein (glycerol monooleate), which can occupy the side-extended pocket (SEP) unique to 5-HT2AR, and has a certain polar interaction with the residues of TM5. Experiments show that monoolein enters OBP through the G2385.42 residue position, activating the G protein signal downstream of 5-HT2AR and the activation of this signal can be inhibited by the selective inhibitor MDL100907 of 5-HT2AR.
Fig. 1 Activation of lipids on 5-HT2AR
2
The second binding pattern of serotonin and psilocin in serotonin 2A receptors
The researchers believe that serotonin and psilocin in two different forms of binding in 5-HT2AR (OBP and EBP) may have different effects on receptor function. Experiments have found that monoolein alone cannot activate the β-arrestin signal downstream of 5-HT2AR, but can activate the β-arrestin signaling dose-dependent in the presence of serotonin. In contrast, in the presence of LSD, monoolein that fails to penetrate obopus cannot activate β-arrestin signal. The results showed that the second binding mode of serotonin and psilocin was critical for the activation of their β-arrestin signals.
Figure 2 Serotonin and psilocin binding in EBP under lipid presence
Comparisons of the 5-HT2AR-LSD and 5-HT2AR-lisuride structures showed that both ethyl groups in the LSD were exposed to residue Y3707.43 in EBP, while lisuride had only one ethyl interacting with it. Experiments have found that the 5-HT2AR-Y3707.43W mutant can significantly improve the β-arrestin signal of lisuride, but has little effect on G protein signaling. However, since Y3707.43 interacts directly with the two ethyl groups of LSD, the Y3707.43W mutant simultaneously reduces LSD-mediated G protein signaling and β-arrestin signaling. The results show that although LSD and lisuride have similar binding patterns in OBP, the difference in binding modes in EBP leads to different interactions with TM7, which affects the downstream β-arrestin signal.
Figure 3 Effect of Y3707.43 on lisuride and LSD activation of 5-HT2AR downstream signals
Various early studies have shown that the affinity of hallucinogens for 5-HT2AR is closely related to their psychoactivity7,8. Animal behavioral models do not accurately capture the perturbations of perception, cognition, and emotion produced by hallucinogens in humans. However, studies have shown that the head twitch response (HTR) in mice is closely related to hallucinations induced by human hallucinogens9. The researchers further verified that 5-HT2AR-Y3707.43W spot mutation mice can successfully erase the HTR effect of LSD while retaining the antidepressant effect of LSD by constructing a model based on magnetic signal detection mouse HTR. The results showed that the psychoactive effect of hallucinogens required 5-HT2AR-mediated high-efficiency signal transduction, and the hallucinogenic effects and antidepressant effects of hallucinogens could be separated by altering the pharmacological activity of 5-HT2AR agonists.
Figure 4 Effect of 5-HT2AR-Y3707.43W mice on the hallucinogenic and antidepressant effects of LSD
3
Structure-oriented synthesis of β-arrestin signaling preference compounds
Using the above data, the researchers speculate that targeting EBP can enhance the 5-HT2AR-mediated β-arrestin signal, which is conducive to designing β-arrestin signal preference ligands. Preliminary laboratory findings suggest that the head groups of antipsychotics are all bound to EBPs of their GPCR-targeted receptors10,11 and that 5-HT2AR is one of the important targets of atypical antipsychotics. By analyzing dozens of commercially available antipsychotic drugs, the researchers targeted the 2019 FDA-approved lumateperone (an inhibitor of 5-HT2AR)12; and resolved its crystal structure with a complex of 5-HT2AR, finding that the tetracyclic skeleton of the lumateperone head group binds to EBP at 5-HT2AR. The researchers designed and synthesized the head group based on the structure: IHCH-7113. Further data show that IHCH-7113 has a weak β-arrestin signal preference and a high activation efficiency in both downstream signaling pathways of 5-HT2AR, which can induce HTR in mice.
Fig. 5 Structural design ideas, activation signals, and hallucinogenic effects of IHCH-7113
Further through the rational design of structure-oriented compounds, the researchers synthesized IHCH-7086 and its series of derivatives by introducing methoxyphenyl on the basis of IHCH-7113; making its fluorophenyl relative to lumateperone avoid the PIF domain of 5-HT2AR (which determines the inhibitor effect of lumateperone); at the same time, according to the design, the series of compounds will be mainly bound EBP (β-arrestin preference), While avoiding lipid-binding pockets that bind to OBP (G protein preference). By analyzing the crystal structure of the complexes of IHCH-7086 and 5-HT2AR, the binding pattern of IHCH-7086 in 5-HT2AR is indeed as the researchers envisioned, and IHCH-7086 exhibits a significant preference for β-arrestin signals.
Figure 6 Activation signal of IHCH-7086 and complex structure bound to 5-HT2AR
4
Effects of 5-HT2AR-mediated downstream signaling in hallucinogenic and antidepressant behaviors
The researchers further detected the hallucinogenic effects of IHCH-7086, IHCH-7079 (IHCH-7086 analogues) in a model based on magnetic signal detection of mouse HTR, and the results showed that IHCH-7086 and IHCH-7079, which have a clear preference for β-arrestin signals, do not induce HTR in mice when injected with high doses (10 mg/kg).
Subsequently, the researchers verified the antidepressant effect of IHCH-7086 and IHCH-7079 in mouse depression-like models constructed by different methods, and the experimental results showed that IHCH-7079 and IHCH-7086 had similar antidepressant effects to LSD previously reported, which could significantly improve depressive behavior in mice, and the selective inhibitor MDL100907 of 5-HT2AR could inhibit their improvement.
Fig. 7 Hallucinogenic and antidepressant effects of IHCH-7086 and IHCH-7079
The work not only clarifies the molecular mechanism of hallucinogenicity, but also helps to understand the pharmacological mechanism of hallucinogens to alleviate depression, and provides theoretical guidance for the development of new fast-acting and long-acting antidepressants. It should be pointed out that compounds are not equivalent to drugs, and the researchers of the research team are conducting systematic compound druggability verification and continuous optimization of lead compounds, striving to make them drugs that benefit patients as soon as possible. The road to "reforming evil and returning to righteousness" of hallucinogens is still "a long road and a long way to repair", and researchers will continue to "seek up and down".
Cao Dongmei, phD candidate of Wang Sheng Research Group of Molecular Cell Excellence Center, Yu Jing, assistant researcher, Wang Huan, assistant researcher of Cheng Jianjun Group of ShanghaiTech University, Professor Luo Zhipu of Soochow University, and Liu Xinyu, phD candidate of Li Jinsong Research Group of Center of Molecular Cell Excellence, are the co-first authors of the paper, and Wang Sheng, researcher of Molecular Cell Excellence Center of Excellence, and Cheng Jianjun, researcher of iHuman Institute of ShanghaiTech University, are the co-corresponding authors of the paper. At the same time, this work was also assisted by Academician Li Jinsong of the Center of Excellence for Molecular Cells and Dr. Lin He of the Third Research Institute of the Ministry of Public Security. The research work has been funded by the Key R&D Project of the Ministry of Science and Technology, the Strategic Pilot Project of the Chinese Academy of Sciences, the National Natural Science Foundation of China, the Youth Thousand Talents Fund of the Ministry of Science and Technology, and the Sunshine Program of the Shanghai Municipal Science and Technology Commission. The research effort was supported by both the Molecular cell center of excellence for chemical biology technology platform and the animal experiment technology platform.
Group photo of Wang Sheng's research team
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Source: Center for Excellence in Molecular and Cell Science, Chinese Academy of Sciences