Improved pharmacokinetic properties of tryptamine prodrugs

Abstract

The present invention describes an injectable composition comprising a compound of formula (I): # imgabs0 # (I) or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier wherein the compound of formula (I) is present in a dose of from about 30 mg to about 50 mg calculated on a free base. Also described is a method of treating a psychiatric disorder comprising the step of administering to a patient in need thereof a pharmaceutical composition comprising a compound of formula (I) or a pharmaceutically acceptable salt thereof.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims the benefit under 35 U.S.C. §119(e) of provisional patent application serial number 63 / 478,593 filed on January 5, 2023, provisional patent application serial number 63 / 505,237 filed on May 31, 2023, and provisional patent application serial number 63 / 507,600 filed on June 12, 2023, the contents of each of which are hereby incorporated by reference into this document. Background Art Technical Field

[0003] In one aspect, the present invention relates to a method for treating a psychiatric disorder in a patient. In another aspect, the present invention relates to an injectable composition, preferably for use in a method for treating a psychiatric disorder in a patient.

[0004] Description of the existing technology

[0005] Tryptamines are 3-aminoethyl indoles that bind to and activate serotonin receptors (also known as 5HT receptors). Activation of the 5HT2A subtype of serotonin receptors by 5HT2A receptor agonist compounds can lead to hallucinations. The endogenous receptor is 5-hydroxytryptamine (serotonin). Tryptamine (3-(2-aminoethyl)-indole) is also an endogenous neurotransmitter.

[0006] The serotonin receptor system is associated with depression and depressive states, which are often treated with 5HT1A receptor antagonists (Affective Disorders: Depression in Neuropsychopharmacology and Therapeutics, 1st edition, Chapter 6, Ivor S. Ebenezer, 2015). Recently, 5HT2A receptor agonists have shown potential as depression medications (Carhart-Harris 2018 Psychopharmacology).

[0007] Tryptamine molecules, which produce hallucinogenic states and have been used in traditional medicine, may have therapeutic potential in treating mood disorders, psychological distress, depression, and other conditions. For example, ayahuasca, a naturally occurring form of dimethyltryptamine (DMT), can be ingested when combined with monoamine oxidase inhibitors and produce a hallucinogenic state of varying intensity and duration, with effects lasting from 6 to 15 hours. DMT is also found naturally in the brain (in smaller amounts) and may function as a neurotransmitter.

[0008] Lysergic acid diethylamide (LSD), a diethylamide derivative of a naturally occurring substance from a fungus in rye grain, also produces a hallucinogenic state lasting up to 8 to 12 hours.

[0009] Psilocybin is a naturally occurring, plant-derived tryptamine found in Psilocybe mushrooms that produces a hallucinogenic state lasting approximately six to eight hours. Psilocybin was first synthesized in 1958 and is currently being studied as a treatment for depression. Psilocybin is a prodrug whose active substance in the body is psilocybin dephosphate. Psilocybin contains a phosphate group bound to the 4-hydroxyl group of psilocybin dephosphate. When Psilocybe mushrooms or the drug substance are taken orally, this phosphate group is cleaved in the intestines:

[0010]

[0011] Simple monofunctional organic esters of psilocybin dephosphodiester have been reported. Lower alkoxy-modified psilocybin dephosphodiester have also been described. Sulfate-conjugated psilocybin dephosphodiester have been prepared, and other monoprotic and diprotic inorganic acid-modified psilocybin dephosphodiester have also been described. Psilocybin dephosphodiester acetate is known and has been used in underground psychedelic subcultures.

[0012] Psychedelic substances have been shown to be effective in treating depression, and when used in combination with psychotherapy, their effectiveness in treating depression is even better (Watts 2020 Journal of Contextual Behavioral Science).

[0013] Since perhaps the earliest recorded work by Albert Hoffman, only a limited number of synthetic tryptamines have been prepared. Structure-activity relationships for a number of tryptamines have been described (Claire 1988).

[0014] Succinate and other diacid functional groups have been explored as components of prodrug delivery systems for the preparation of water-soluble, injectable forms of hydrophobic or poorly water-soluble drug substances such as testosterone, haloperidol, chloramphenicol, or estradiol (Silverman and Holladay, Chapter 9.2: Prodrugs and Drug Delivery Systems in The Organic Chemistry of Drug Design and Drug Action (3rd ed., 2014). Tetrahydrocannabinol succinate has been patented for the treatment of glaucoma. However, ester cleavage is not always rapid, its rate is unpredictable, and may depend on the structure of the group attached to the drug, so relevant research is necessary (Anderson 1984 J Pharma Sci). Esterase enzymes are responsible for the active cleavage of the prodrug ester group in vivo, and differences in esterase content and specificity in tissues of different species complicate the research and optimization process (Bahar 2012 J Pharm Sci).

[0015] 4-Valeryl-3-(2-diisopropylaminoethyl)-1H-indole hydrochloride (also referred to herein as RE104HCl, formerly codenamed FT-104) is a prodrug of the synthetic hallucinogenic drug 3-(2-diisopropylaminoethyl)-1H-indole-4-ol (also known as 4-OH-DiPT or hydroxyisopropyltryptamine). This molecule is structurally related to dephosphorylated psilocybin:

[0016]

[0017] RE104 HCl is converted to 4-OH-DiPT in the body by esterases and is being developed as a subcutaneous injection to ensure rapid and complete absorption and that the hallucinogenic experience is brief and reproducible.

[0018] The active molecule, 4-OH-DiPT, acts as a serotonin (5-HT) agonist, specifically at the 5-HT2A and 5-HT2B subtypes. These receptors are thought to contribute to 4-OH-DiPT-mediated activity. 4-OH-DiPT belongs to the tryptamine class of compounds, more specifically the 4-substituted tryptamines—a class that also includes the well-known psychedelic compound psilocybin. Psilocybin is a natural product produced by various species of Psilocybin mushrooms. The phosphate group at the 4-position of tryptamine in psilocybin is enzymatically cleaved in vivo to produce dephosphopsilin, an agonist at multiple serotonin receptors, most notably the 5-HT2A receptor, which is thought to mediate the compound's hallucinogenic activity (Nichols, 2004). In RE104 HCl, the glutaric acid group at the 4-position of tryptamine is hydrolyzed to form 4-OH-DiPT, which mediates its 5-HT2A receptor agonist activity.

[0019] Classical serotonergic psychedelics (a class of compounds that bind to serotonin (5-HT) receptors) are characterized by their ability to induce changes in perception, mood, thinking, and self-perception, ultimately leading to adaptive remodeling of psychological function (Vollenweider, 2001; Kometer et al., 2012; Vollenweider et al., 1998), and are known as mystical experiences that occur during psilocybin therapy. These changes have been repeatedly observed to predict subsequent effects on behavior and mood, including reductions in depressive and anxious behaviors (Griffiths et al., 2011; Griffiths et al., 2016; Ross et al., 2016).

[0020] Multiple lines of evidence suggest that serotonergic hallucinogens, such as psilocybin dephosphate and hydroxyisopropyltryptamine (4-OH-DiPT), have the clinical potential to induce therapeutically beneficial behavioral changes in a variety of psychiatric disorders. Lasting changes in attitudes, depression, anxiety, well-being, substance abuse, and mindfulness have been documented following administration of hallucinogens. Mystical experiences, feelings of connection, emotional breakthroughs, and increased neural entropy have been associated with long-term changes in these psychological functions (Aday et al., 2020). Furthermore, growing evidence suggests that hallucinogen-assisted psychotherapy may be an effective treatment for depression and other psychological disorders (Carhart-Harris et al., 2016).

[0021] It has also been reported that cancer patients often experience chronic, clinically significant depression and anxiety symptoms, which are associated with adverse psychiatric and medical outcomes (Swift et al., 2017; Griffiths et al., 2016; Ross et al., 2016).

[0022] Furthermore, encouraging results were obtained from a study (Moreno et al., 2006) of patients with obsessive-compulsive disorder (OCD). In this study, subjects received up to four psilocybin administrations, separated by at least one week, in an ascending dose sequence (100 μg / kg, 200 μg / kg, and 300 μg / kg). Reductions in OCD symptom scores were observed in all subjects during one or more testing sessions, with reductions ranging from 23% to 100%.

[0023] The efficacy of psilocybin has also been investigated in alcohol dependence (Bogenschutz et al., 2015). A recent (2021) review evaluated the clinical efficacy of serotonergic hallucinogens and their potential for treating mental health conditions (Andersen et al., 2021). The review included 16 papers representing 10 independent trials of hallucinogen-assisted therapy (7 using psilocybin, 2 using ayahuasca, and 1 using LSD) involving 188 patients with cancer or disease-related anxiety and depression, including major depressive disorder, OCD, or substance use disorder. Although most of the reviewed studies were small and open-label, which requires caution when evaluating the efficacy of hallucinogen-assisted therapy, two larger controlled studies reported comparable efficacy—these studies evaluated depression (and anxiety) symptoms in cancer patients using either an inactive placebo or a low-dose psilocybin control (Andersen et al., 2021).

[0024] Finally, Shulgin provides anecdotal evidence regarding the effects of 4-OH-DiPT (hydroxyisopropyltryptamine) (10 to 20 mg taken orally), noting that its effects when used recreationally are roughly comparable to those of other serotonergic hallucinogens, such as psilocybin dephosphate, but that it is unique in that its duration of action is relatively short (Shulgin and Shulgin, 1997). Regarding oral administration of hydroxyisopropyltryptamine, Shulgin (Shulgin and Shulgin, 1997) noted that he "doubts whether any other psychedelic drug, whatever it is, can rival this drug in terms of speed, intensity, and shortness of action." Studies have shown that the onset and duration of hydroxyisopropyltryptamine's effects are dose-dependent: after oral administration of 10 mg, only mild somatic sensations occur within 1 hour, while the onset of action is much faster (within 15 minutes) with a 20 mg dose. After oral administration of 20 mg, the recovery time is suggested to be 3 hours. However, controlled studies on the safety, tolerability, and pharmacokinetic properties of 4-OH-DiPT in humans are lacking, partly due to the compound's poor stability (susceptible to air oxidation) and extremely low solubility (<2 mg / ml in PBS).

[0025] Conversion to the prodrug RE104 provides improved stability and solubility, while also generating the active compound 4-OH-DIPT in situ (in vivo) via an esterase-mediated cleavage pathway [see US Pat. No. 11292765]. However, there remains a need to more clearly elucidate the pharmacokinetic properties of RE104 or a pharmaceutically acceptable salt thereof (such as the hydrochloride) in humans, and to conduct comparative studies with 4-OH-DiPT and psilocybin. It is also desirable to more clearly elucidate the dosage characteristics of RE104 or a pharmaceutically acceptable salt thereof (such as the hydrochloride) in humans. Summary of the Invention

[0026] It is an object of the present invention to obviate or mitigate at least one of the above-mentioned disadvantages of the prior art.

[0027] One object of the present invention is to provide improved drug delivery and pharmacokinetic properties following administration of RE104 or a pharmaceutically acceptable salt thereof, such as the hydrochloride salt.

[0028] Another object of the present invention is to provide a shortened duration of hallucinogenic experience following administration of a RE104 prodrug or a pharmaceutically acceptable salt thereof (such as the hydrochloride salt) at a dose level that produces a complete mystical experience in most subjects (defined as a score of ≥ 60% on all dimensions of the Mystical Experience Questionnaire (MEQ-30)). [See Ko, K et al., Front. Psychiatry 2022, 13, 917-199].

[0029] It is another object of the present invention to provide a shortened duration of a hallucinogenic experience following administration of a prodrug of RE104 or a pharmaceutically acceptable salt thereof, such as the hydrochloride salt, at a dosage level expected to produce a therapeutic effect, compared to administration of an effective dosage level of psilocybin.

[0030] Yet another object of the present invention is to reduce side effects, particularly blood pressure fluctuations and headaches, following administration of a prodrug of RE104 or a pharmaceutically acceptable salt thereof (such as the hydrochloride salt) at a dosage level expected to produce a therapeutic effect, compared to administration of an effective dosage level of psilocybin.

[0031] It is also an object of the present invention to provide improved safety and tolerability following administration of a prodrug of RE104 or a pharmaceutically acceptable salt thereof, such as the hydrochloride salt, at a dosage level expected to produce a therapeutic effect, compared to administration of an effective dosage level of psilocybin.

[0032] In one aspect, the present invention relates to a composition comprising a compound as described herein and a pharmaceutically acceptable excipient. In some embodiments, the composition comprises an oral formulation or an injectable formulation.

[0033] In another aspect, the present invention includes a method for treating a psychiatric disorder comprising the step of administering an effective amount of a compound described herein. In some embodiments, the psychiatric disorder is a depressive state, including unipolar and bipolar depressive states, such as, but not limited to, depression, generalized anxiety disorder, major depressive disorder, treatment-resistant depression, and postpartum depression.

[0034] In another aspect, the present invention relates to the use of a compound as described herein for treating a psychotic disorder, or in the manufacture of a medicament for treating a psychotic disorder, such as depression.

[0035] In another aspect, the present invention relates to a method for treating a mental disorder, comprising the step of administering to a patient in need of treatment a pharmaceutical composition comprising a compound of formula (I):

[0036]

[0037] or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, wherein the compound of formula (I) is present in a dosage of about 30 mg to about 50 mg calculated as the free base.

[0038] In another aspect, the present invention relates to a method for treating a mental disorder, comprising the step of administering to a patient in need of treatment a pharmaceutical composition comprising a compound of formula (I):

[0039]

[0040] or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, wherein the compound of formula (I) is present in a dosage of 30 mg or about 40 mg calculated as the free base.

[0041] In another aspect, the present invention relates to an injectable composition comprising a compound of formula (I):

[0042]

[0043] or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, wherein the compound of formula (I) is present in a dose of 1 mg / kg calculated as a free base.

[0044] In another aspect, the present invention relates to an injectable composition comprising a compound of formula (I):

[0045]

[0046] or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, wherein the compound of formula (I) is present in a dosage of about 30 mg to about 50 mg calculated as the free base.

[0047] In another aspect, the present invention relates to an injectable composition comprising a compound of formula (I):

[0048]

[0049] or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, wherein the compound of formula (I) is present in a dosage of about 30 mg or about 40 mg calculated as the free base.

[0050] Administration of the RE104 prodrugs described herein or pharmaceutically acceptable salts thereof (such as the hydrochloride salt) can be helpful in treating psychiatric disorders, such as depressive states, including unipolar and bipolar depressive states, such as, but not limited to, depression, generalized anxiety-induced depression, major depressive disorder, refractory depression, and postpartum depression. BRIEF DESCRIPTION OF THE DRAWINGS

[0051] Embodiments of the present invention will now be described with reference to the accompanying drawings, in which:

[0052] Figure 1 A schematic diagram showing the design of the human clinical study described in Example 2;

[0053] Figure 2 The modified Drug Effect Questionnaire (DEQ) used in the human clinical study described in Example 2 is presented;

[0054] Figure 3 The revised Mystical Experience Questionnaire (MEQ) used in the human clinical study described in Example 2 is presented;

[0055] Figure 4 The following table shows the changes in the mean DEQ excitement scores over time for different dose groups in the human clinical study described in Example 2;

[0056] Figure 5 The mean MEQ scores for each dimension and the total scores for each RE104 dose level in the human clinical study described in Example 2 are shown;

[0057] Figure 6 The present invention shows the changes over time in mean plasma 4-OH-DiPT concentration and DEQ hyperactivity score of the RE104 33 mg dose group in the human clinical study described in Example 2;

[0058] Figure 7 The relationship between the mean plasma 4-OH-DiPT concentration and the DEQ hyperactivity score for all cohorts in the human clinical study described in Example 2 is shown;

[0059] Figure 8 The RE104 5-HT2A receptor binding profile studied in Example 3 is shown;

[0060] Figure 9 The 5-HT2A receptor binding of 4-OH-DiPT studied in Example 3 is shown;

[0061] Figure 10 The RE104 5-HT2B receptor binding profile studied in Example 3 is shown;

[0062] Figure 11The RE104 5-HT2A receptor activation studied in Example 3 is shown;

[0063] Figure 12 Shows the changes in plasma RE104 concentration over time in mice studied in Example 3;

[0064] Figure 13 Shows the changes in plasma RE104 concentration over time in rats studied in Example 3;

[0065] Figure 14 Shows the changes in plasma RE104 concentration over time in dogs studied as described in Example 3;

[0066] Figure 15 Shows the changes in human plasma RE104 concentration over time in the study described in Example 3;

[0067] Figure 16 The present invention shows the time course of mean 4-OH-DiPT plasma concentrations and mean number of head impulse responses (HTRs) counted at 10-minute intervals following intravenous (IV) and subcutaneous (SC) administration of RE104 or 4-OH-DiPT to groups of three rats in the study described in Example 3.

[0068] Figure 17 Shows the correlation between HTR and mean 4-OH-DiPT plasma concentrations following intravenous administration of 2 mg / kg RE104 in the study described in Example 3 (the line represents the best-fit curve);

[0069] Figure 18 Shows the correlation between HTR and mean 4-OH-DiPT plasma concentrations following subcutaneous administration of 2 mg / kg RE104 in the study described in Example 3 (the line represents the best-fit curve);

[0070] Figure 19 Shows the correlation between HTR and mean 4-OH-DiPT plasma concentrations following intravenous administration of 1.39 mg / kg 4-OH-DiPT in the study described in Example 3 (the line represents the best-fit curve);

[0071] Figure 20 shows the correlation between HTR and mean 4-OH-DiPT plasma concentrations following subcutaneous administration of 1.39 mg / kg 4-OH-DiPT in the study described in Example 3 (the line represents the best fit curve); and

[0072] Figures 21 to 23 The effects of RE104 and RE109 (psilocybin dephosphate glutarate) on specific behaviors in the forced swim test were demonstrated separately. Figure 21 For the immobile time, Figure 22 It's time to swim. Figure 23 =Climbing time) (n=12 male Wistar rats per treatment group) [Error bars represent standard error of the mean. *P<0.05, **P<0.01, ***P<0.001, relative to the vehicle group, one-way ANOVA followed by Dunnett's multiple comparison test.] The independent sample t test was used for comparison with the control group. relative to RE109]. DETAILED DESCRIPTION

[0073] In one aspect, the present invention relates to a method for treating a mental disorder, comprising the step of administering to a patient in need of treatment a pharmaceutical composition comprising a compound of formula (I):

[0074]

[0075] or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier at a dose of about 30 mg to about 50 mg, and a pharmaceutically acceptable carrier, wherein the compound of formula (I) is present in a dose of about 30 mg to about 50 mg calculated as the free base (e.g., about 30 mg or about 40 mg calculated as the free base).

[0076] Preferred embodiments of the method may include any one of the following features, or a combination of any two or more of the following features:

[0077] The compound of formula (I) is used in the form of its hydrochloride;

[0078] When used as the hydrochloride salt, the compound of formula (I) is used in a dose of about 33 mg or about 44 mg;

[0079] The duration of action of the compound of formula (I) is from about 2.5 hours to about 4.5 hours after administration;

[0080] The duration of action of the compound of formula (I) is about 3.5 hours or 3.7 hours after administration;

[0081] The compound of formula (I) in the patient's body max in the range of about 1000 ng / mL to about 2000 ng / mL;

[0082] The compound of formula (I) in the patient's body max within the range of about 1500 ng / mL to about 1800 ng / mL;

[0083] The T of the compound of formula (I) in the patient max In the range of about 20 minutes to 30 minutes;

[0084] The T of the compound of formula (I) in the patient max In about 15 minutes;

[0085] The compound of formula (I) is converted into the compound of formula (II) after administration

[0086]

[0087] And the compound of formula (II) is C in the patient's body max in the range of about 100 ng / mL to about 300 ng / mL;

[0088] The compound of formula (I) is converted into the compound of formula (II) after administration

[0089]

[0090] And the compound of formula (II) is C in the patient's body max in the range of about 120 ng / mL to about 300 ng / mL;

[0091] The compound of formula (I) is converted into the compound of formula (II) after administration

[0092]

[0093] And the compound of formula (II) has a T max In the range of about 60 minutes to about 150 minutes.

[0094] The compound of formula (I) is converted into the compound of formula (II) after administration

[0095]

[0096] And the compound of formula (II) has a T max in the range of about 60 minutes to about 80 minutes;

[0097] The method demonstrates a mean MEQ30 score in at least 50% of patients; and / or

[0098] The method showed mean MEQ30 scores in at least 60% of patients;

[0099] The pharmaceutical composition is administered to the patient by injection; and / or

[0100] The pharmaceutical composition is administered to the patient by subcutaneous injection.

[0101] As used herein, the term "mental disorder" includes conditions that may be diagnosed as psychological or psychiatric disorders by a mental health professional, including those that can be diagnosed by reference to the Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition (DSM-5).

[0102] As used herein, the term "treatment" includes both prophylactic (ie, preventive) treatment and palliative treatment (ie, to alleviate, lessen or slow the progression of a patient's disease, disorder or condition).

[0103] As used herein, a "hallucinogenic state" is an altered state of consciousness experienced by an individual that can include heightened perceptions, perceptual distortions or hallucinations, and / or feelings of euphoria or despair. Hallucinogenic states have been described as being caused by hallucinogenic drugs, such as DMT (dimethyltryptamine), LSD, mescaline, or psilocybin. Other known hallucinogenic drugs include N-methyl-N-isopropyltryptamine (MiPT) and the 4-hydroxy analogue of N,N-diisopropyltryptamine (DiPT).

[0104] The present invention includes the use of prodrugs of hydroxyindole 5HT2A receptor agonists (such as RE104) that induce a hallucinogenic state or provide beneficial therapeutic effects without the associated hallucinogenic state. These prodrugs can be used in combination with other treatments known to be effective for treating mental disorders, such as psychotherapy, electroconvulsive therapy, and / or other pharmaceutical compounds, for example, tricyclic antidepressants (TCAs), selective serotonin reuptake inhibitors (SSRIs), selective norepinephrine reuptake inhibitors (SNRIs), monoamine oxidase inhibitors (MOAIs), or other antidepressants. In preferred embodiments, the treatment can produce a long-lasting effect, for example, an effect that persists for more than 1 month after a single treatment, preferably for more than 3 months, and more preferably for more than 6 months. In some embodiments, additional therapy may not be required.

[0105] Compound

[0106] When used herein, "compound" includes any pharmaceutically acceptable derivative or variant, encompassing conformational isomers (e.g., cis-trans isomers), all optical isomers (e.g., enantiomers and diastereomers), racemates, diastereomeric mixtures and other mixtures of such isomers, as well as solvates, hydrates, isomers, polymorphs, tautomers, esters, salt forms and prodrugs. The expression "prodrug" refers to a compound that is a drug precursor, which releases the drug (or "active ingredient") in vivo after administration via a chemical or physiological process (e.g., hydrolysis, enzymatic cleavage or hydrolysis, or metabolic conversion to the desired drug form). The present invention includes within its scope pharmaceutically acceptable salts of the compounds of the present invention. Therefore, unless explicitly indicated to the contrary, the phrase "or a pharmaceutically acceptable salt thereof" is implicit in the description of all compounds described herein.

[0107] In some embodiments, the compounds employed in the therapeutic compositions and methods of treatment described herein include prodrug compounds that are easily purified, easily formulated, and stable, preferably providing highly soluble drug substances with rapid onset and clearance for ease of use in clinical settings. In some embodiments, these compounds can be produced in the form of zwitterions, which can be converted into pharmaceutically acceptable salts.

[0108] In some embodiments, the compounds of the present invention preferably enable rapid cleavage of the prodrug group in vivo to release the active pharmacophore, for example, 90% conversion can be completed within 4 hours, preferably less than 2 hours, and more preferably less than 1 hour. The prodrug itself may have low, little, or no pharmacological activity, but after administration to a patient, it can be converted into an active compound through processes such as hydrolytic cleavage.

[0109] Formulations and compositions

[0110] The present invention also provides pharmaceutically acceptable compositions comprising a therapeutically effective amount of one or more compounds described herein, formulated together with one or more pharmaceutically acceptable carriers (additives) and / or diluents, and optionally comprising one or more additional therapeutic agents. Although the compounds described herein can be administered alone, it is preferred to administer the compounds in the form of a pharmaceutical composition.

[0111] The term "pharmaceutical composition" means a composition comprising a combination of a compound of the present invention and at least one additional pharmaceutically acceptable carrier. A "pharmaceutically acceptable carrier" refers to a medium recognized in the art for delivering biologically active agents to animals (particularly mammals), specifically including (that is) adjuvants, excipients or vehicles (such as diluents, osmotic supplements, preservatives, fillers, flow regulators, disintegrants, wetting agents, emulsifiers, suspending agents, sweeteners, flavorings, fragrances, antibacterial agents, antifungal agents, lubricants, polymers, solubilizers, stabilizers, antioxidants and dispensing agents, etc.), the specific type of which depends on the nature of the administration method and the dosage form. Each carrier must be "acceptable", that is, compatible with the other ingredients of the formulation and harmless to the patient.

[0112] As used herein, "oral" administration includes administration by swallowing into the stomach or intestine, and also includes lingual, sublingual, buccal and oropharyngeal administration. The compounds of the present invention can be administered by any suitable route to achieve any use or method described herein, for example: oral administration, such as tablets, capsules (each of which can contain sustained-release or timed-release formulations), pills, powders, granules, elixirs, suspensions (including nanosuspensions, microsuspensions, spray-dried dispersions), syrups and emulsions; sublingual administration (for example, as a film, effervescent tablet or sublingual self-dissolving tablet); parenteral administration, such as by subcutaneous, intravenous, intramuscular injection or infusion technology (for example, as a sterile injectable aqueous solution or non-aqueous solution / suspension); nasal administration, including nasal mucosal administration, such as by inhalation spray; or rectal administration, such as in the form of a suppository.

[0113] The preferred formulation allows for subcutaneous injection. Likewise, the preferred route of injection is subcutaneous injection.

[0114] Of course, the dosage regimen of the compounds described herein will vary depending on known factors, such as the pharmacokinetic and pharmacodynamic properties of the specific drug and its mode and route of administration; the species, age, sex, health condition, disease state, and weight of the recipient; the nature and severity of symptoms; the type of concomitant therapy; the frequency of treatment; the route of administration, the patient's renal and hepatic function; and the desired effect. The selected dosage level may also depend on additional factors, including the activity of the specific compounds and pharmaceutical compositions described herein; whether the compound used contains an ester, salt, or amide substituent; the time of administration; the rate of excretion or metabolism of the specific compound used; the rate and extent of absorption; the duration of treatment; other drugs that the patient may be taking; other compounds and / or materials used in combination with the specific compound used; and similar factors well known in the medical arts.

[0115] Generally speaking, when used to indicate therapeutic efficacy, the dosage of the prodrug in a single treatment will be in the range of about 0.001 mg / dose to about 500 mg / dose, preferably about 0.01 mg / dose to about 200 mg / dose, preferably about 0.1 mg / dose to about 50 mg / dose, such as 10 mg, 20 mg, 30 mg, 40 mg, 50 mg, 100 mg or 200 mg. In a more preferred embodiment, when used to indicate therapeutic efficacy, the dosage of the prodrug in a single treatment is about 30 mg / dose to about 50 mg / dose, such as 33 mg or 44 mg.

[0116] Intravenously, the most preferred doses will range from about 0.01 mg / kg / min to about 10 mg / kg / min during a constant rate infusion.

[0117] The compounds of the present invention may be administered as a single daily dose, or the total daily dose may be divided into multiple doses, such as twice, three or four times daily. Alternatively, the dosage may be provided weekly, biweekly or monthly. In a preferred embodiment, only one or two doses are required to achieve an antidepressant effect, which may last for 1 month, 2 months, 3 months, 6 months or longer.

[0118] For tablet dosage forms, the drug may comprise 1% to 80% of the dosage form weight, more typically 5% to 60%, depending on the dosage. In addition to the drug, tablets typically contain a disintegrant. Examples of disintegrants include sodium starch glycolate, sodium carboxymethylcellulose, calcium carboxymethylcellulose, croscarmellose sodium, crospovidone, polyvinylpyrrolidone, methylcellulose, microcrystalline cellulose, lower alkyl-substituted hydroxypropyl cellulose, starch, pregelatinized starch, and sodium alginate. Generally, the disintegrant will comprise 1% to 25%, preferably 5% to 20%, of the dosage form weight.

[0119] Binders are generally used to impart cohesive properties to tablet formulations. Suitable binders include microcrystalline cellulose, gelatin, sugars, polyethylene glycol, natural and synthetic gums, polyvinyl pyrrolidone, pregelatinized starch, hydroxypropyl cellulose and hydroxypropyl methylcellulose. Tablets can also include diluents such as lactose (monohydrate, spray-dried monohydrate, anhydrous matter, etc.), mannitol, xylitol, glucose, sucrose, sorbitol, microcrystalline cellulose, starch and dihydrated calcium phosphate.

[0120] Tablets may also optionally contain surfactants (such as sodium lauryl sulfate and polysorbate 80) and glidants (such as silicon dioxide and talc). If present, surfactants are typically used in amounts of 0.2% to 5% by weight of the tablet and glidants are typically used in amounts of 0.2% to 1% by weight of the tablet.

[0121] Tablets also typically contain lubricants such as magnesium stearate, calcium stearate, zinc stearate, sodium stearyl fumarate, and a mixture of magnesium stearate and sodium lauryl sulfate. Lubricants are typically present in an amount of 0.25% to 10%, preferably 0.5% to 3%, by weight of the tablet.

[0122] Other conventional ingredients include antioxidants, colorants, flavorings, preservatives, and taste-masking agents.

[0123] Exemplary tablets contain up to about 80% by weight drug, about 10% to about 90% by weight binder, about 0% to about 85% by weight diluent, about 2% to about 10% by weight disintegrant, and about 0.25% to about 10% by weight lubricant.

[0124] Tablet blends can be compressed directly or by roller compaction into tablets. Tablet blends, or portions of such blends, can alternatively be processed by wet granulation, dry granulation, melt granulation, melt coacervation, or extrusion prior to tableting. The final formulation can comprise one or more layers and can be coated or uncoated, or encapsulated.

[0125] Tablet formulations are discussed in detail in "Pharmaceutical Dosage Forms: Tablets," Vol. 1, H. Lieberman and L. Lachman, Marcel Dekker, NY, NY, 1980 (ISBN 0 8247 6918X).

[0126] A typical capsule for oral administration contains at least one compound of the invention (eg, 25 mg), lactose (eg, 75 mg), and magnesium stearate (eg, 15 mg).The mixture is passed through a 60-mesh screen and filled into size 1 gelatin capsules.

[0127] Liquid preparations include suspensions, solutions, syrups, and elixirs. Such preparations can be used as fillers in soft or hard capsules and typically contain a carrier (e.g., water, ethanol, polyethylene glycol, propylene glycol, methylcellulose, or a suitable oil), and one or more emulsifiers and / or suspending agents. Liquid preparations can also be prepared by reconstitution of a solid, such as from a powder bag.

[0128] The compounds of the present invention can also be administered directly into the bloodstream, muscle, or internal organs. Suitable routes for parenteral administration include intravenous, intraarterial, intraperitoneal, intrathecal, intraventricular, intraurethral, ​​intrasternal, intracranial, intramuscular, and subcutaneous administration. Suitable devices for parenteral administration include needle (including microneedle) syringes, needle-free syringes, and infusion technology equipment.

[0129] Parenteral preparations are typically aqueous solutions that may contain excipients such as salts, carbohydrates, pH adjusters, or buffers (pH preferably 3.0 to 7.0, preferably 4.0 to 6.0, more preferably 4.5 to 5.5). However, for some applications, they may be more suitably formulated as sterile non-aqueous solutions or dry preparations (needing to be used with a suitable vehicle such as sterile pyrogen-free water or pre-prepared, ready-to-mix aqueous buffers). Osmotic agents may be added to control osmotic pressure.

[0130] Parenteral kits for reconstitution at the patient site under sterile conditions may be readily prepared using standard pharmaceutical techniques well known to those skilled in the art (eg, by lyophilization).

[0131] A typical injectable formulation is prepared by the following steps: at least one compound of the present invention (e.g., 25 mg, 33 mg, or 44 mg) is aseptically added to a vial in the form of a sterile-filtered solution, followed by aseptic lyophilization and sealing. Upon use, the vial contents are mixed with, for example, 2 mL of normal saline for injection, and an appropriate amount of an osmotic supplement and a pH adjuster are optionally added to achieve a weakly acidic to neutral pH (e.g., pH 4-7), thereby preparing a low-irritation injectable formulation while maintaining the solubility and / or stability of the prodrug.

[0132] The compounds used in the present invention may be used in combination with soluble macromolecular entities (such as cyclodextrins and suitable derivatives thereof or polyethylene glycol-containing polymers) to improve their solubility, dissolution rate, taste masking, bioavailability and / or stability, thereby being suitable for any of the aforementioned modes of administration.

[0133] For example, it has been found that drug cyclodextrin complexes are generally useful in most dosage forms and routes of administration. Both inclusion and non-inclusion complexes can be employed. As an alternative to directly forming complexes with drugs, cyclodextrins can also be used as auxiliary additives, i.e., serving as carriers, diluents, or solubilizers. The most commonly used for these purposes are α-, β-, and γ-cyclodextrins, examples of which can be found in International Publication Nos. WO 91 / 11172, WO 94 / 02518, and WO 98 / 55148.

[0134] Regardless of the route of administration selected, the compounds of the present invention (which may be in a suitable hydrated form) and / or the pharmaceutical compositions of the present invention are formulated into pharmaceutically acceptable dosage forms by conventional methods known to those skilled in the art. The actual dosage level of the active ingredient in the pharmaceutical compositions of the present invention can be adjusted to obtain an amount of active ingredient that is effective to achieve the desired therapeutic response for a particular patient, a particular composition, and a particular mode of administration.

[0135] A person having ordinary skill in the art (e.g., a nurse, nurse practitioner, physician, or veterinarian) can readily determine and prescribe the effective amount of the desired pharmaceutical composition. For example, such a person can initially administer a dose of the compound of the invention employed in the pharmaceutical composition that is lower than the dose required to achieve the desired therapeutic effect, and gradually increase the dose until the desired effect is achieved.

[0136] In general, a suitable daily dose of a compound of the invention will be the lowest dose of the compound effective to produce a therapeutic effect. Such an effective dose will generally depend on the factors described above.

[0137] As used herein, a "therapeutically effective amount" refers to an amount of a compound administered that will alleviate to some extent one or more of the symptoms of the condition being treated. With respect to the treatment of depression, a therapeutically effective amount refers to an amount that has the effect of reducing the severity of depression. The severity of depression can be assessed using well-known structured assessment tools, such as the Structured Clinical Interview for DSM-5 (SCID-5) and the GRID-Hamilton Depression Rating Scale (GRID-HAMD). A therapeutically effective amount may be less than the amount required to induce a hallucinogenic state.

[0138] An effective dose can be administered via a single or multiple administration regimen. For the purposes of the present invention, an effective dose of a drug, compound, or pharmaceutical composition is an amount sufficient to directly or indirectly achieve a prophylactic or therapeutic treatment. As understood in the clinical context, an effective dose of a drug, compound, or pharmaceutical composition can be achieved with or without the use of another therapy, drug, compound, or pharmaceutical composition.

[0139] Treatment methods and uses

[0140] Treatment with the compounds of the present invention (such as RE104 described herein) can substantially alleviate clinical or subclinical depression and prevent relapse, particularly when used in conjunction with psychotherapy for depression treatment. It is known that administration of effective doses of psilocybin produces a rapid and substantial reduction in depressive symptoms, with many subjects achieving remission within a four-week follow-up period (Davis et al.). Without being bound by theory, it is believed that hallucinogenic states are associated with beneficial effects, but certain compounds that are 5HT2A agonists can provide the desired therapeutic effects without the hallucinogenic state. One aspect of the present invention includes prodrugs of these 5HT2A agonists that provide beneficial therapeutic states.

[0141] Generally speaking, the present invention includes: the use of the compounds of the invention herein for treating any disease or condition that can be alleviated by a 5HT2A agonist; or the use of the compounds of the invention herein for the manufacture of a medicament for treating any disease or condition that can be alleviated by a 5HT2A agonist; or a method of treating any disease or condition that can be alleviated by a 5HT2A agonist.

[0142] In some embodiments, the present invention may include the use of the compounds described herein to treat psychiatric disorders. In some embodiments, the present invention may include the use of the compounds of the present invention to treat depression, particularly major depressive disorder, postpartum depression, and refractory depression. Other conditions that may be treated include: anxiety disorders, including anxiety in terminal illness (e.g., cancer) and generalized anxiety disorder; cluster headaches; obsessive-compulsive disorder; personality disorders (including conduct disorder); substance disorders, including alcohol dependence, nicotine dependence, opioid dependence, cocaine dependence; and other addictions, including gambling disorder, eating disorders, and body dysmorphic disorder; chronic pain; or chronic fatigue.

[0143] In some embodiments, the present invention may include a method of treating a mental disorder comprising administering to a subject in need thereof a therapeutically effective amount of a compound of the present invention. In one embodiment, a method of treating depression is provided comprising administering to a subject in need thereof a therapeutically effective amount of a compound of the present invention. The depression may be treatment-resistant depression or major depressive disorder.

[0144] For example, patients diagnosed with depression can be screened prior to treatment and then prepared for the dosing process by a trained psychotherapist. During the dosing process, the compounds of the invention can be administered by injecting the patient with a sterile solution at a rate of 0.01 mg / kg to 0.3 mg / kg. The patient preferably remains seated and wears an eye mask for the duration of the dosing process. For safety reasons, a trained healthcare professional can monitor the patient throughout the dosing process, which may last up to 12 hours. In some cases, music can be played for the patient. Once the healthcare professional is able to determine that the drug substance has cleared, the trained healthcare professional can assist the patient in resolving any issues related to the hallucinogenic experience, after which the patient can be allowed to leave.

[0145] To further alleviate any anxiety that may occur during treatment, physicians may prefer to split the therapeutic dose, thereby lessening the initial onset of psychoactivity before administering the full dose to achieve the full effect.

[0146] In some embodiments, treatment with the compounds of the present invention can be combined with concomitant treatment using another antidepressant, which is implemented simultaneously or sequentially. In a preferred embodiment, treatment with the compounds of the present invention is combined with psychotherapy, which can be implemented before or after treatment. If implemented before treatment, the treatment process can focus the patient on the purpose of treatment. If implemented after treatment, psychotherapy is preferably performed within 48 hours after the administration process to help the patient integrate any feelings, emotions, images or thoughts that may occur during the treatment process, and to enable the psychotherapist to provide advice on how to best change thinking or behavior patterns to improve the antidepressant effect. Psychotherapy can be continued after the administration process as needed (e.g., for up to an additional 3 months) to assist the patient in integrating any experience or cognition gained during the administration process.

[0147] The present invention can be described with reference to the following examples. These examples are provided for illustrative purposes only. All terms, names, abbreviations, or acronyms are in accordance with the conventional understanding of those skilled in the art. Compounds shown as zwitterionic forms can be readily assumed by those skilled in the art to be in neutral forms, and vice versa. These examples should not be used to interpret or limit the scope of the present invention.

[0148] Example 1

[0149] Preliminary studies and nonclinical pharmacology

[0150] RE104 hydrochloride (RE104 HCl) is a prodrug of the synthetic hallucinogenic drug 3-(2-diisopropylaminoethyl)-1H-indol-4-ol (4-OH-DiPT, also known as hydroxytryptamine), a molecule structurally related to psilocybin dephosphate. 4-OH-DiPT has been reported as a substance of abuse, producing subjective experiences similar to psilocybin in recreational settings, but with shorter-lasting psychoactive properties (Shulgin and Shulgin, 1997). No formal human studies of 4-OH-DiPT have been documented, but the literature does contain limited pharmacological and nonclinical studies. 4-OH-DiPT acts as a serotonin (5-hydroxytryptamine [5-HT]) agonist, particularly at the 2A and 2B subtypes. Multiple lines of evidence suggest that serotonergic hallucinogens, such as those produced after administration of psilocybin (the active ingredient is psilocybin dephosphate) and hydroxytryptamine, have clinical potential to induce therapeutically beneficial behavioral changes in a variety of psychiatric disorders.

[0151] For all studies, placebo was a commercially available 0.9% saline solution. All drug administrations in the studies described below were single subcutaneous (SC) injections, e.g., in the patient's upper arm. Regardless of route of administration, subcutaneous injection may generally be preferred because it offers certain advantages over oral administration, such as: (1) ensuring dose absorption, whereas oral absorption is often associated with nausea and dose loss due to vomiting; (2) ensuring complete and reproducible absorption, whereas gastrointestinal absorption may vary with several parameters (including individual metabolic characteristics and recent food intake); (3) ensuring the fastest and most complete systemic availability by avoiding delays associated with gastrointestinal transit; and (4) these combined factors result in greater reproducibility of absorption rates and overall pharmacokinetic profiles, which also ensures that the duration and intensity of drug effect are as consistent as possible from treatment to treatment.

[0152] Nonclinical Pharmacology

[0153] Similar to psilocybin and dephosphopsilin, RE104 and 4-OH-DiPT are 5-HT2A agonists. Receptor binding studies (5-HT2A and 5-HT2B agonist radioligand binding assays) have demonstrated that the active moiety, 4-OH-DiPT, is more potent (IC50 values ​​of 150 nM and 42 nM at 5-HT2A and 5-HT2B, respectively) than the prodrug, RE104 (IC50 values ​​of 1600 nM and 740 nM at 5-HT2A and 5-HT2B, respectively). Furthermore, in in vitro radioligand binding assays, RE104 can interact with other receptors (e.g., the 5-HT2A transporter, kappa and μ opioid receptors), as can 4-OH-DiPT (5HT2A transporter, kappa opioid receptor, 5-HT1A, 5-HT1B, μ opioid receptor, and muscarinic M2 receptor). However, for both RE104 and 4-OH-DiPT, binding at both 5-HT2A and 5-HT2B receptors was dominant compared to all other receptors included in the screening assay.

[0154] When administered to rats, 5-HT2A agonists (such as psilocybin, dephosphopsilin, RE104, and 4-OH-DiPT) induce behaviors such as head shaking, wet dog shakes, tremors, hindlimb abduction, and backward walking due to excessive pharmacological effects of serotonin receptors (Halberstadt et al., 2011; Haberzettl et al., 2013; Rickli 2016 Eur Neuropharmacol; Klien 2020 ACS Pharmacol Transl Sci). Head shaking behavior in rodents has been associated with hallucinogenic reactions similar to human hallucinogenic effects (Willins and Meltzer, 1997). Excessive pharmacological effects associated with acute or chronic overstimulation of serotonin receptors have been reported to produce a condition known as serotonin syndrome (SS). Serotonin syndrome is an adverse drug reaction caused by excessive serotonergic activity that is observed at very high dose levels (such as those evaluated in nonclinical toxicology studies described in the Investigator's Manual) and may occur in humans when they experience excessive acute or chronic exposure (Boyer and Shannon, 2005). Acute SS associated with serotonin agonists (such as psilocybin) evolves roughly with the plasma levels of the active drug and resolves within a short period of time (<24 hours). Chronic SS may produce more serious health effects and should be avoided. RE104 was developed as a single-use drug, in which repeated, intermittent use will be limited, so only mild to moderate manifestations of excessive pharmacological effects can be expected. See, for example, Franciscangeli 2019 Int J Mol Sci.

[0155] Safety Pharmacology

[0156] In behavioral studies in rats, a single SC administration of 10 mg / kg, 25 mg / kg, or 50 mg / kg of RE104HCl induced significant acute and reversible neuropharmacological effects; however, all observations were attributed to excessive pharmacological effects of the test article at serotonin receptors (i.e., SS). The incidence and duration of the observations were dose-related. At the 10 mg / kg dose, the effects were observed within 30 minutes (min) to 1 hour (h); at the 25 mg / kg dose, the effects lasted up to 6 hours; at the 50 mg / kg dose, the effects lasted up to 6 hours after dosing but resolved before 24 hours.

[0157] In a respiratory function study in male rats, single SC doses of 10 mg / kg, 25 mg / kg, and 50 mg / kg of RE104 HCl caused increases in respiratory rate (8% to 28%), decreases in tidal volume (28% to 30%), and decreases in minute ventilation (24% to 31%). All of these changes were transient and returned to pre-dose values ​​by the time the reading was taken 6 hours after dosing. Based on the small magnitude and transient nature of these changes, they are not considered adverse effects.

[0158] In cardiovascular studies, a single subcutaneous administration of 1 mg / kg of RE104 HCl resulted in a small, transient increase in heart rate, but no effect on blood pressure, in conscious, telemetrically monitored beagle dogs. Transient increases in heart rate and blood pressure (systolic, diastolic, and mean) were observed at doses of 3 mg / kg and 8 mg / kg. No significant, biologically relevant effects on heart rhythm, electrocardiographic (ECG) parameters, or body temperature were observed at subcutaneous doses of 1 mg / kg, 3 mg / kg, or 8 mg / kg of RE104 HCl.

[0159] At the highest concentration tested (30 μM, calculated as zwitterion), RE104 HCl exhibited minimal effect in the human ether-a-go-go related gene (hERG) tail current assay, resulting in a 5.9% ± 1.6% decrease in hERG current amplitude.

[0160] toxicology

[0161] A non-GLP single-dose dose-ranging study of RE104 HCl was conducted in rats and dogs to explore toxicity and determine the maximum tolerated dose (MTD) by SC injection. The rat study explored the administration of two formulations: water for injection (WFI) at a pH of 3-4 and phosphate-buffered saline (PBS) at a final formulation pH of 4.5-5. Single doses of 20 mg / kg to 300 mg / kg formulated in WFI produced dose-dependent, pharmacologically relevant systemic serotonergic clinical signs. Similarly, 25 mg / kg to 225 mg / kg RE104 formulated in PBS induced SS clinical signs approximately 1 hour after SC administration, which recovered within 24 hours. Reduced food intake, weight loss, and decreased body temperature (females) were observed at a dose of 225 mg / kg.

[0162] Injection site reactions were observed at all WFI doses (20 mg / kg to 150 mg / kg) and at doses above 75 mg / kg in PBS. Clinical signs were reversible by recovery day 13 (study day 14). Pathological changes at the microinjection site were observed after a single dose of 225 mg / kg. The MTD of RE104 in rats is 150 mg / kg in WFI and 75 mg / kg in PBS.

[0163] In a 14-day extended single SC dose toxicity study of GLP in rats, doses of 0 (PBS), 10 mg / kg, 25 mg / kg, and 75 mg / kg were evaluated. Animals were necropsied on study day 2 and day 14 post-dose (also known as recovery day 13).

[0164] Systemic exposure of RE104 HCl and 4-OH-DiPT (C max and AUC 0-t ) No significant sex differences (>2-fold) were observed. Across the 10 mg / kg to 75 mg / kg dose range, systemic exposures of RE104 and 4-OH-DiPT generally increased in a greater-than-dose-proportional manner with increasing doses, with the exception of the Cmax of 4-OH-DiPT in males, which showed a less-than-dose-proportional increase.

[0165] Acute, mild clinical signs of SS (“excessive pharmacological effects at the serotonin 2A receptor”) were evident in all RE104 HCl dose groups, accompanied by reversible injection site changes including edema / inflammatory infiltrates in all treatment groups at both 25 mg / kg and 75 mg / kg RE104.

[0166] In male rats, the changes in all groups resolved by day 2, and the animals returned to normal.

[0167] Preclinical Pharmacokinetics and Metabolism

[0168] The pharmacokinetic (PK) properties of RE104 and 4-OH-DiPT were characterized in rats following a single intravenous (IV) or subcutaneous (SC) dose of 2 mg / kg RE104 HCl. Following IV administration, RE104 was only measurable in plasma from 5 minutes to 0.25 hours post-dose, and following SC administration, only at 5 minutes post-dose; therefore, PK parameters for RE104 could not be determined in this study. The active substance, 4-OH-DiPT, exhibited a short plasma elimination half-life (t½) following all routes of administration: t½ of 0.426 hours following IV administration and t½ of 0.678 hours following SC administration. The absolute bioavailability of RE104 following SC administration was nearly 100% in rats. In monkeys, RE104 was also rapidly converted to 4-OH-DiPT, with half-lives of 0.631 hours and 0.950 hours following IV and SC administration, respectively. In dogs, RE104 undergoes a slow conversion to 4-OH-DiPT. Following subcutaneous administration, the half-life of the parent drug, RE104, is 1.3 hours, while the half-life of 4-OH-DiPT is 0.888 hours. Following IV administration, the half-life of 4-OH-DiPT is 0.863 hours. Similarly, the absolute bioavailability of RE104 in dogs is close to 100% following subcutaneous administration.

[0169] The active substance 4-OH-DiPT metabolized by RE104 HCl has a low binding rate to animal plasma proteins (binding rate is approximately 30% to 60%), but has a higher binding rate to human plasma proteins in a concentration-dependent manner (binding rate is approximately 80% to 93% in the concentration range of 1μM to 20μM).

[0170] In hepatocytes, the in vitro intrinsic clearance (CLint) of RE104 was the fastest in rats and the slowest in dogs. The CLint values ​​of RE104 in rats, rabbits, humans, monkeys and dogs were 6.66, 4.88, 3.45, 1.14 and 0.87 mL / h / 10, respectively. 6 cells. In human, rat, rabbit, dog, and monkey hepatocytes, the metabolite profiles of RE104 were qualitatively similar across species, with only minor differences. The metabolic profile of 4-OH-DiPT in human hepatocytes was also qualitatively similar to that of RE104 in rat, rabbit, dog, and monkey hepatocytes, with only minor differences. In in vitro assays using recombinant human enzymes (Supersomes), which contain multiple cytochrome P450 (CYP) enzymes, flavin-containing monooxygenases (FMOs), and monoamine oxidases (MAOs), it was shown that RE104 was not a substrate for any cytochrome P450, FMO, or MAO isoforms. The enzymatic conversion of RE104 to 4-OH-DiPT is likely achieved by plasma esterases such as cholinesterase.

[0171] Drug administration rationale and exposure limits

[0172] The no-observed-adverse-effect level (NOAEL) for local injection-site changes was established at 10 mg / kg for male and female rats. The NOAEL for systemic effects was 10 mg / kg. The 10 mg / kg dose in males was considered a low adverse-effect level (LOAEL). The 10 mg / kg dose in rats corresponds to a human equivalent dose (HED) of approximately 97 mg (as zwitterion), based on a 60 kg human body weight and a standard (2005 FDA guidance) body surface area (BSA) conversion factor. In this study, this HED dose is approximately 19-fold greater than the proposed starting dose of 5 mg (as zwitterion) and approximately 2-fold greater than the maximum proposed dose of 48 mg (as zwitterion). At the 10 mg / kg dose, the maximum plasma concentration (Cmax) of RE104 was 0.751 ng / mL. The area under the concentration-time curve (AUC0-t) from time 0 to the final time point could not be determined due to its rapid conversion to 4-OH-DiPT; the mean Cmax of 4-OH-DiPT was 0.751 ng / mL. max 542 ng / mL, AUC 0-tThe MTD was 1230 h·ng / mL. To determine the MTD, a non-GLP dose-ranging study was conducted in dogs. Single subcutaneous doses of 7.5 mg / kg, 15 mg / kg, and 20 mg / kg were evaluated. Overt clinical signs of SS were observed across the entire dose range, with hypersalivation and behavioral and postural changes occurring at all doses.

[0173] A GLP 14-day extended single SC dose toxicity study was conducted in dogs at doses of 0 mg / kg, 3 mg / kg, 8 mg / kg, and 20 mg / kg RT-104HCl. Systemic exposure (C max and AUC 0-t ) showed no sex differences. Within the dose range of 3 mg / kg to 20 mg / kg, exposure to RE104 HCl increased approximately dose-proportionally, while exposure to 4-OH-DiPT increased greater than dose-proportionally. Clinical signs were limited to serotonin syndrome.

[0174] The NOAEL for dogs was determined to be 8 mg / kg. The HED for dogs at a dose of 8 mg / kg is approximately 267 mg (as zwitterion), based on a 60 kg human body weight and a standard BSA conversion factor. In this study, this HED provides an approximately 53-fold safety margin relative to the proposed starting dose of 5 mg (as zwitterion) and an approximately 5-fold safety margin relative to the proposed maximum dose of 48 mg (as zwitterion). At a dose of 8 mg / kg in dogs, the C max and AUC 0-t They are 3300 ng / mL and 4540 h·ng / mL respectively; and 4-OH-DiPT is 360 ng / mL and 1010 h·ng / mL respectively.

[0175] Based on the NOAEL of 10 mg / kg in rats and 8 mg / kg in dogs, and applying a 10-fold safety margin, the maximum recommended safe starting dose (MRSD) of RE104 HCl for the RT-104-101 study was 9.7 mg (Table 1). The actual proposed starting dose for the first-in-human (FIH) study was 5 mg (RE104 free base), thus providing a higher safety margin relative to the starting dose.

[0176] Based on a comparison of the effective dose ranges of psilocybin / psilocybin dephosphate in various psychiatric disorders (21 mg to 30 mg orally), and considering the approximately 53% oral bioavailability of psilocybin dephosphate in humans and its affinity for the 5-HT2A receptor, the effective dose of 4-OH-DiPT is estimated to be in the range of 10 mg to 30 mg. RE104 HCl has 100% bioavailability after subcutaneous administration in animals, and thus the parent drug is expected to fall within the same dose range. Therefore, 5.5 mg (RE104 HCl free base) is considered a conservative dose and is expected to serve as a safe starting dose.

[0177] Based on the NOAEL and MRSD in Table 2, proposed dose levels are proposed for human cohorts 1 to 4 of the RE104-101 study program described in Example 2.

[0178] Example 2

[0179] Research rationale

[0180] This study is the first clinical study conducted using RE104 HCl. It aims to investigate the safety, tolerability, pharmacokinetic (PK), and pharmacodynamic (PD) properties of a single subcutaneous (SC) dose of RE104 HCl in healthy female subjects. Data from this study will provide a foundation for further clinical development of RE104 HCl and will be used to support dose selection for future Phase 2 studies in the target patient population.

[0181] Purpose

[0182] Characterize the safety, tolerability, PD, and PK profile of RE104 in a Phase 1, first-in-human (FIH) study.

[0183] design

[0184] The research design diagram is as follows Figure 1 shown.

[0185] This study is a Phase 1, double-blind, randomized, placebo-controlled, single-dose escalation study in healthy women with up to six cohorts. Subject eligibility was assessed during a screening period of up to 28 days prior to enrollment. During the treatment period, subjects were admitted to the Clinical Research Unit (CRU) one day prior to dosing (Day -1). On Day 1, all subjects received a SC injection of their assigned RE104 HCl or placebo in a fed state (subjects stayed overnight in the CRU on Day -1 and consumed their planned dinner; breakfast was consumed before dosing on Day 1). Four hours after dosing, subjects were allowed to choose a snack from a planned menu. Subjects were discharged on Day 2 (approximately 24 hours after dosing) after completing all required study procedures and having their health status confirmed by medical evaluation. Subjects returned to the clinic for follow-up on Day 10 (±2 days).

[0186] A total of six (6) cohorts were included in this study. Figure 1 As shown in Table 3, six (6) single-dose doses were tested in 6 cohorts (Cohort 1 to Cohort 6), each containing 8 healthy female subjects (6 active drug and 2 placebo).

[0187] The starting dose was 5.5 mg of RE104 HCl. All cohorts used a sentinel subject design. In each cohort, two subjects were initially treated: one subject received RE104 HCl and one subject received placebo. If the principal investigator (PI) determined that no clinically significant safety issues occurred within at least 24 hours after dosing the sentinel subject, the remaining six subjects in each cohort were dosed (five active drug and one placebo).

[0188] This study is overseen by a Safety Review Committee (SRC). The SRC is designed to ensure that treatment does not pose an excessive risk to subjects. Before escalating to the next dose level (Table 3), the SRC will evaluate safety, tolerability, and available PK data across cohorts. After each cohort completes dosing, the SRC will hold a meeting to discuss dose escalation. Dose escalation to the next higher dose level will only occur after the evaluation of safety and tolerability data for all evaluable subjects in the previous dose cohort, up to and including at least 24 hours, has been completed. Available PK results will be required to determine dose escalation, as needed.

[0189] Inclusion criteria

[0190] The subject population included healthy adult subjects who met all of the following inclusion criteria:

[0191] --Before any research-related process, the subject must sign an informed consent form in a language that the subject can understand.

[0192] --Healthy female subjects whose age during the screening period was between 18 and 65 years (inclusive).

[0193] - Subjects whose body weight at the screening period was at least 60 kg and whose BMI was between 18.0 kg / m2 and 30.0 kg / m2 (inclusive).

[0194] --The subject self-reported at least one significant past recreational use of hallucinogens or hallucinogen-like compounds (excluding cannabis products), with the most recent use occurring more than 90 days before study drug administration.

[0195] --Women of childbearing potential (WOCBP) must be non-lactating, have negative pregnancy tests at screening and admission, and be willing to undergo additional pregnancy testing as required during the study.

[0196] Women of reproductive potential (WOCBP) who have sexual intercourse with intact male partners must agree to consistently and correctly use a double-barrier, highly effective contraceptive method (e.g., condoms and oral contraceptive pills [OCPs], long-acting hormonal implants, hormonal injections, vaginal rings, or intrauterine devices [IUDs]) evaluated by the PI during the study and for at least 90 days after study drug administration. If using hormonal contraception, it must have been started for at least 30 days before study drug administration. Women who abstain from heterosexual intercourse as a daily routine and do not plan to become pregnant are also eligible.

[0197] Non-WOCBP women must meet any of the following criteria: surgical sterilization (e.g., tubal occlusion, hysterectomy, bilateral salpingectomy, bilateral oophorectomy), or be postmenopausal. For amenorrheic women, menopausal status will be confirmed by measuring follicle-stimulating hormone (FSH) levels (≥30 IU / mL) during the screening period.

[0198] --Female subjects must abstain from egg donation from the screening period until at least 90 days after study drug administration.

[0199] Exclusion criteria

[0200] Subjects were excluded if they met any of the following criteria:

[0201] - The subject currently has or has a history of any clinically significant disease, such as cardiovascular, neurological, pulmonary, hepatic, renal, metabolic, gastrointestinal, urinary, immune, endocrine or psychiatric disease / disorder, or other abnormalities that may affect safety or potentially interfere with study results.

[0202] --The subject currently has or has a history of a clinically significant psychiatric disorder, as assessed by the Mini-International Neuropsychiatric Interview (MINI) questionnaire and an interview with a qualified medical professional (clinical psychologist or psychiatrist) during the screening period.

[0203] - The subject is at risk for suicide (ideation score of 4 or 5 or any suicidal behavior) according to the Columbia-Suicide Severity Rating Scale (C-SSRS) during the screening and baseline periods; or is clinically judged to be at risk by a qualified medical professional (clinical psychologist or psychiatrist); or has a history of suicidal ideation or suicidal behavior.

[0204] --A direct (first-degree) blood relative or individual currently or previously diagnosed with a psychotic disorder or bipolar disorder.

[0205] - Any clinically significant illness, medical / surgical procedure, or trauma within 4 weeks prior to screening.

[0206] - Subjects with a history of cancer (except basal cell carcinoma that has been in remission for at least 5 years before screening). - Subjects with a history of major adverse reactions to hallucinogens or hallucinogen-like drugs (excluding cannabis products) as determined by qualified / trained investigators.

[0207] - Use of any medication (including over-the-counter (OTC) drugs) within 28 days or 5 half-lives (whichever is longer) prior to study drug administration, except for the following:

[0208] - Hormonal contraceptives used by WOCBP.

[0209] - ≤3000mg paracetamol or ≤1200mg ibuprofen daily for a maximum of three days.

[0210] -Topical steroids for acute use.

[0211] - Use of ayahuasca, cambo, yubo, ibogaine, psilocybin, dimethyltryptamine (DMT), 5-methoxy-N,N-dimethyltryptamine (5-MeO-DMT), lysergic acid diethylamide (LSD), Syrian rue, or other synthetic or naturally occurring hallucinogens or mixtures, as well as amphetamines, opioids, or 3,4-methylenedioxymethamphetamine (MDMA) within 90 days prior to screening until EOS / ET (end of study / early termination).

[0212] - Use of synthetic or naturally occurring cannabinoids from 28 days prior to study drug administration until EOS / ET, and agreement not to use during this period.

[0213] --During the screening period or on Day -1, the subject tested positive for cotinine.

[0214] - Positive breath alcohol test or urine drug of abuse screen during the screening period or on Day -1.

[0215] - The subject has a history of substance abuse according to the most recent edition of the Diagnostic and Statistical Manual of Mental Disorders (DSM) available at the start of the study, or as determined by a medically qualified investigator, or has engaged in substance abuse within 1 year prior to screening.

[0216] Excessive alcohol consumption within 180 days prior to screening (regular alcohol intake ≥14 units / week for female subjects). 1 unit (8 gr) is equivalent to 1 / 2 pint (280 mL) of beer, 1 shot (25 mL) of liquor, or 1 small glass (125 mL) of wine. Alcohol consumption during the 48 hours prior to study drug administration until the last PK sample collection.

[0217] Consumption of products containing caffeine, xanthine, or poppy seeds during the period 48 hours before study drug administration until the last PK sample collection.

[0218] - Participation in another clinical study involving investigational therapy within 30 days or 5 half-lives (whichever is longer) prior to screening.

[0219] - Any vaccination during the period from 28 days before study drug administration until EOS / ET.

[0220] Use of any psychoactive medications (e.g., selective serotonin reuptake inhibitors (such as paroxetine or citalopram), haloperidol, any medication with MAO activity (such as isocarboxazid, phenelzine, selegiline or tranylcypromine, linezolid, and methylene blue), or any medication labeled in the label as a potential inducer of serotonin syndrome) during the 28 days prior to study drug administration until EOS / ET. See Tables 4 and 5 in Section 5.3.1.1 for a list of contraindicated medications.

[0221] --Any positive result of serum hepatitis B surface antigen (HbsAg), hepatitis C antibody and human immunodeficiency virus (HIV) during the screening period.

[0222] --Female subjects are pregnant, breastfeeding, or test positive for pregnancy during the screening period or on Day -1.

[0223] - During the screening period or at any time before the administration of study drug, resting (at least 5 minutes) systolic blood pressure >140 mmHg or <90 mmHg, or resting diastolic blood pressure >90 mmHg or <40 mmHg; resting (at least 5 minutes) pulse rate is outside the range of <40 beats / minute or >100 beats / minute.

[0224] - Any clinically significant abnormality in resting electrocardiogram (ECG) rhythm, conduction, or morphology during the screening period or at any time point before study drug administration, and any clinically important 12-lead ECG abnormality that the investigator believes may interfere with the interpretation of corrected QT interval (QTc) changes; QTc ≥ 480ms for female subjects during the screening period or on Day -1.

[0225] --Estimated glomerular filtration rate (eGFR) ≤ 60 ml / min / 1.73 m during the screening period.

[0226] - Subject has any skin condition, application site abnormality, or tattoo that would preclude SC administration (e.g., local or systemic infection) and / or assessment of local site reactions.

[0227] --The subject's peripheral venous access condition is poor.

[0228] --The subject is an immediate family member (e.g., spouse, parent, child, sibling) of a research center employee, or has a subordinate relationship with a research center employee involved in the implementation of this study, or may be coerced into agreeing to participate.

[0229] -A positive test for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2; COVID-19) within 7 days prior to admission to the CRU.

[0230] --The subject has not completed the full course of COVID-19 vaccination, as defined by CRU guidelines and requirements at the national, provincial, and local levels.

[0231] --Current or past COVID-19 or influenza-like illness, defined as fever (>37.5°C) within 7 days prior to administration and accompanied by ≥2 of the following symptoms: cough, sore throat, runny nose, sneezing, limb / joint pain, headache, vomiting / diarrhea without clear cause, and not caused by influenza or COVID-19 infection.

[0232] Study Limitations – Drugs and Dietary Products

[0233] Table 4 provides a list of contraindicated medications and dietary products for Study RE104-101. Table 5 reports a list of specific medications that may induce serotonin syndrome and were contraindicated in this study.

[0234] Study drug and matching placebo

[0235] The active investigational product is RE104 HCl, a prodrug of the synthetic psychedelic drug 3-(2-diisopropylaminoethyl)-1H-indol-4-ol (4-OH-DiPT, also known as hydroxyisopropyltryptamine), a molecule structurally related to dephosphorylated psilocybin. 4-OH-DiPT acts as a 5-HT agonist, specifically targeting the 5-HT2A and 5-HT2B subtypes. RE104 HCl is administered as a solution for injection. RE104 HCl consists of two components: LYO-RE104 HCl, which contains lyophilized RE104 HCl, and RE-PBS diluent (phosphate-buffered saline solution), which is used to prepare the final injection solution. Placebo is a volume-matched 0.9% sterile saline solution.

[0236] Formulation, storage, preparation and handling

[0237] RE104 HCl for SC Injection is supplied as a sterile lyophilized solid (LYO-RE104 HCl) in vials for reconstitution. Mix with an aqueous diluent according to the pharmacy manual. Each vial of LYO-RE104 HCl contains an amount of active ingredient equivalent to the amount of RE104 zwitterion.

[0238] For example, the vial used in cohort 4 contained 36.3 mg of lyophilized drug substance, equivalent to 33 mg of RE104 zwitterion. A diluent consisting of a near-stoichiometric amount of sodium phosphate dibasic was used to dissolve and neutralize LYO-RE104 HCl, resulting in an injectable solution with a pH of approximately 5. Prior to injection, 1.1 mL of RE-PBS was added to the vial containing 36.3 mg of LYO-RE104 HCl to create a 33 mg / mL RT-104 HCl solution. This is equivalent to 30 mg / mL RE104 zwitterion (free base). The remaining cohorts were treated similarly.

[0239] Store LYO-RE104 HCl vials at -20°C. The diluent, RE-PBS, is prepared sterile by mixing water for injection, sodium phosphate dibasic (60 mM), and sodium chloride (40 mM) into a solution. This formulation is selected so that the sodium phosphate dibasic neutralizes the carboxylic acid in RE104 HCl, resulting in the formation of the RE104 zwitterion in a pH 5 solution. Store RE-PBS vials at room temperature (15°C to 25°C). Once RE104 and RE-PBS are mixed, the solution must be administered within 60 minutes.

[0240] Application and application

[0241] Dosing schedules were implemented after approval by the study pharmacy and investigators. RE104 HCl or placebo was administered via subcutaneous injection in the upper arm. Subjects received RE104 HCl or placebo in the fed state. The time of RE104 HCl or placebo administration was recorded in the eCRF (electronic case report form). The regimen used is summarized in Table 3.

[0242] In each cohort, two sentinel subjects were initially treated: one subject received RE104 HCl and one subject received placebo. If no clinically significant safety issues were observed for at least 24 hours after dosing the sentinel subject, as judged by the PI, the remaining six subjects in that cohort were dosed (five active drug and one placebo). All administrations were performed under direct medical supervision. During the observation period for potentially psychoactive study drug, study subjects were supervised by a professional trained in monitoring hallucinogenic experiences.

[0243] Safety and tolerability

[0244] No serious adverse events (AEs) occurred in any of the treated participants, and all participants completed dosing (Table 6).

[0245] The most common treatment-related adverse events with RE104 were nausea, sinus tachycardia (asymptomatic, with a maximum recorded value of 115 beats / minute), restlessness, and headache.

[0246] Two participants (one on 38 mg and one on 44 mg) experienced a serious adverse event, "painful reaction," and received midazolam. Both adverse events resolved after administration of concomitant medication.

[0247] No injection site adverse reactions were reported, but one adverse event of mild bruising related to the administration procedure was recorded.

[0248] No significant blood pressure effects were observed during the study period, and no clinically significant vital sign, clinical laboratory, or electrocardiographic findings were observed.

[0249] Pharmacodynamics

[0250] The “Feel / High” scale of the Drug Effect Questionnaire (DEQ) is widely used in studies of acute subjective reactions to various substances to assess two key aspects of subjective experience: (1) the intensity of the substance’s effects, and (2) the degree to which the substance’s effects are desired (Morean et al., 2013). The revised DEQ assesses the extent to which subjects (1) perceive any substance effect and (2) experience a high. This version of the DEQ contains the following items: “Do you feel the effects of the drug at this moment?” (FEEL); “Do you feel a high at this moment?” (HIGH). The revised DEQ used is detailed in Figure 2 .

[0251] The revised DEQ uses an 11-point numerical rating scale (0-10) and is administered by qualified personnel. After administration of the study drug, participants are asked to verbally describe their perceived intensity of the drug effect and degree of euphoria on a scale of 0 to 10 (Kollltveit et al., 2020).

[0252] The Mystical Experience Questionnaire (MEQ30) is a validated questionnaire used to assess the intensity of individual mystical experiences induced by classic hallucinogens (Barrett et al., 2015). The validated MEQ30 assesses four experiential dimensions with 30 questions: (i) mystical (including items on inner unity, outer unity, intuitive authenticity, and sacredness); (ii) positive emotions; (iii) sense of transcendence in time and space; and (iv) ineffability. Participants will rate each MEQ30 item on a 6-point scale. The revised MEQ30 used is detailed in the Figure 3 .

[0253] The mean changes in DEQ excitement scores over time showed a dose-dependent effect (see Figure 4 ).

[0254] For doses ≥33 mg, DEQ peak euphoria scores ranged from 7 to 10, with a mean time to peak score of 1.1 hours; the mean duration of the experience for the 33 mg dose was 3.7 hours, with all participants scoring ≤1 four hours after dosing.

[0255] A dose-related increase in the frequency of MEQ responders was observed: based on the MEQ total score (Table 7), 66.7%, 83.3% and 100% of participants in the RE104 33 mg, 38 mg and 44 mg treatment groups, respectively, achieved a "complete" mystical experience (defined as a MEQ total score ≥ 60%), which is indicative of clinical efficacy.

[0256] The proportion of MEQ responders in each dimension was divided into different groups according to the medication group. Figure 5 Shown in.

[0257] Pharmacokinetics

[0258] Plasma concentrations of RE104 and hydroxytryptamine were determined using a validated liquid chromatography-tandem mass spectrometry (LC-MS / MS) assay.

[0259] The half-life of the RE104 prodrug was approximately 0.5 hours at all dose levels, and RE104 was not detected in plasma within 5 hours after administration at any dose.

[0260] After RE104 administration, the maximum plasma concentration of 4-OH-DiPT is proportional to the RE104 dose.

[0261] The results of the PK testing are reported in Table 8.

[0262] The PK characteristics of 4-OH-DiPT were consistent with its PD characteristics, and its peak plasma concentration coincided with the time of peak PD effect; however, the subjective drug effect disappeared faster than the decrease in 4-OH-DiPT plasma concentration (for example, in the 33 mg dose group). Figure 6 shown in ).

[0263] When examining PK-PD correlations across all cohorts and dose levels, mean 4-OH-DiPT concentrations within the 40 ng / mL to 100 ng / mL range were correlated with mean DEQ euphoria scores ( Figure 7 ).

[0264] When the concentration was lower than 40 ng / mL, the mean DEQ excitement score did not exceed 2 points.

[0265] RE104 Phase 1 FIH Results Summary

[0266] Overall, RE104 HCl was generally well tolerated, and potent pharmacodynamic (PD) effects were observed at doses ≥33 mg that were well consistent with the pharmacokinetic (PK) profile of 4-OH-DiPT.

[0267] The observed Drug Effect Questionnaire (DEQ) euphoria scores and Mystical Experience Questionnaire (MEQ) response rates at doses ≥33 mg of RE104 suggest its potential for therapeutic efficacy in treatment trials, given that the intensity and quality of subjective drug experiences can predict response to psilocybin therapy in patients with depression (Griffiths et al.; Roseman et al.; and Ross et al.).

[0268] The average duration of the subjective experience with 33 mg of RE104 HCl was 3.7 hours, indicating that its psychoactive experience was 50% shorter than historical evidence for psilocybin, and the duration was conducive to clinical monitoring.

[0269] The adverse event (AE) profile of RE104 HCl was similar to that of psilocybin (Goodwin et al.; and Carbonaro et al.), with no serious AEs occurring in the ≤44 mg dose range and no clinically significant changes in vital signs, clinical laboratory, or electrocardiographic findings.

[0270] Two adverse events were observed at doses ≥38 mg.

[0271] These data served as the basis for selecting a 33mg dose of RE104 HCl for a planned Phase 2 randomized, active-dose controlled trial in women with moderate to severe postpartum depression (PPD).

[0272] At therapeutically relevant dose levels (33 mg), subcutaneous (SC) administration of RE104 HCl exhibited an improved safety and tolerability profile compared to psilocybin administration; SC administration of this prodrug resulted in faster absorption and earlier peak effect (approximately 1 hour) with minimal (optimal) duration of psychoactive activity, which has clinical practicality and operational convenience for patients, physicians, and payers.

[0273] RE104 HCl prodrug administration exhibited improved pharmacokinetic characteristics compared to psilocybin administration, including low variability and dose proportionality.

[0274] Administration of RE104 HCl at the 33 mg dose level demonstrated potent and broadly distributed pharmacodynamic effects, with a shorter duration of experience relative to 25 mg of psilocybin.

[0275] After administration of 33 mg of RE104 HCl prodrug, the MEQ total score and each dimension score reached ≥60% (indicating clinical efficacy), which was better than the psilocybin administration group.

[0276] Example 3

[0277] This example describes preclinical characterization data for RE104, including pharmacology, pharmacokinetic (PK), pharmacodynamic, and metabolic studies. To further its clinical development as a potential treatment for depression, this example reports the results of a 28-day, two-group, single-dose forced swim test (FST) study in rats, comparing the efficacy of RE104 with that of desphosphate psilocybin-4-glutarate (RE109), a desphosphate psilocybin prodrug that also contains a glutarate moiety.

[0278] Materials and methods

[0279] Molecular synthesis

[0280] The synthesis of prodrugs RE104 and RE109 was performed according to the teachings of US Patent 11,292,765 B2 [Bryson].

[0281] Receptor pharmacology

[0282] Receptor binding and agonist activity assays (performed by Eurofins Cerep, Celle-Lévescault, France) were performed using methods previously described (Bryant et al.; Choi et al.; and Porter et al.). Increasing concentrations of 4-OH-DiPT or RE104 were combined with 0.1 nM [ 125I] 1-(4-iodo-2,5-dimethoxyphenyl)isopropylamine (DOI) was co-incubated in HEK-293 cell culture system expressing human recombinant 5-HT2A receptor. Increasing concentrations of RE104 were combined with 0.2 nM [ 125 I] DOI was co-incubated in a CHO cell culture system expressing human recombinant 5-HT2B receptor.

[0283] The radioligand concentration was measured by scintillation counting; the compound binding activity was expressed as [ 125 Calculation of percent inhibition of DOI binding. For the agonist activity assay, test concentrations of RE104 were incubated with HEK-293 cells expressing recombinant human 5-HT2A receptors. IP1 production was measured using a homogeneous time-resolved fluorescence assay. The degree of activation was calculated as a percentage of the response to the positive control (10 μM 5-HT). Further details are provided in the "Supplementary Methods" section below.

[0284] Prodrug cleavage

[0285] The stability of RE104 in 2 mL of male CD-1 mouse plasma, male Sprague Dawley rat plasma, mixed-sex beagle dog plasma, and mixed-sex human plasma was assessed using liquid chromatography-tandem mass spectrometry (LC-MS / MS) (InterVivo Solutions, Inc, Mississauga, Ontario, Canada) by measuring the concentrations of RE104 and 4-OH-DiPT after incubation at 37°C for 0, 0.5, 1, 2, and 4 hours. Data are reported as the mean of three replicates. Further details are provided in the Supplementary Methods section.

[0286] Pharmacokinetics and pharmacodynamics

[0287] Plasma samples from adult male Sprague Dawley rats were analyzed to determine RE104 and 4-OH-DiPT concentrations following intravenous (IV) administration of 2 mg / kg RE104 (InterVivo Solutions, Inc.). Test compound concentrations in plasma were determined by LC-MS / MS. The RE104 dose remained at 2 mg / kg for both subcutaneous (SC) and oral (PO) administration, while the 4-OH-DiPT dose was adjusted to 1.39 mg / kg to match the molar dose of RE104 (5.34 μmol / kg). Pharmacokinetic parameters for each analyte and route of administration were estimated based on the plasma concentration-time profiles of each animal. The head-twitch response (HTR), also known as wet-dog shakes, was assessed by observing the rats' rapid alternating head rotation automatism (González-Maeso et al. and Halberstadt et al.). The HTR was monitored by manual counting at 10-minute intervals for 2 hours after dosing and again at 3, 3.5, and 4 hours after dosing.

[0288] The plasma PK profiles of RE104 and 4-OH-DiPT (QPS Taiwan China, New Taipei City, Taiwan) were evaluated in male beagle dogs following three consecutive doses (IV injection of 1 mg / kg, SC injection of 1 mg / kg, and oral gavage of 5 mg / kg RE104). The interval between doses was 7 days. Plasma samples were analyzed by LC-MS / MS (limit of quantification for both analytes was 1.00 ng / mL).

[0289] Two male cynomolgus monkeys (Calvert Laboratories, Inc., Scott Township, PA; QPS, LLC, Newark, DE, USA) were treated three times using a crossover design. Treatment 1: 1 mg / kg RE104 administered as an IV bolus; Treatment 2: 0.5 mg / kg RE104 administered as a subcutaneous injection; and Treatment 3: 5 mg / kg RE104 administered orally. All formulations were prepared in phosphate-buffered saline. Following each treatment, plasma PK profiles of RE104 and 4-OH-DiPT were analyzed using LC-MS / MS from blood samples.

[0290] For details on the blood sampling time points, see the Supplementary Methods section. Pharmacokinetic parameters were calculated using Phoenix WinNonlin version 6.3 or 8.2 software (Certara, Princeton, NJ, USA).

[0291] Metabolite profiling and identification

[0292] Metabolite profiling and identification were performed on plasma samples from rats, dogs, and nonhuman primates after a single subcutaneous injection of RE104 (QPS, LLC, Newark, DE, USA). Twelve Sprague Dawley rats (6 males and 6 females, 8 to 10 weeks of age at the time of dosing) were grouped and received subcutaneous injections of 10 mg / kg, 25 mg / kg, or 75 mg / kg of RE104. Six beagle dogs (3 males and 3 females, 9 to 10 months of age at the time of dosing) were grouped and received subcutaneous injections of 3 mg / kg, 8 mg / kg, or 20 mg / kg of RE104. Two male cynomolgus monkeys received a single subcutaneous injection of 0.5 mg / kg of RE104.

[0293] Blood collection time points are detailed in the Supplementary Methods section. Blood samples were centrifuged to obtain plasma. Plasma samples from rat and dog studies were pooled by time point, with one pooled sample prepared for each dose and sex; monkey plasma samples were pooled to create one pooled sample.

[0294] In vitro metabolite profiling was performed using cryopreservable primary human hepatocytes (QPS, LLC, Newark, DE, USA) from a pooled sample (10 donors) in a humidified incubator at 37°C with 5% CO₂. The cells were incubated in triplicate with 1 μM 4-OH-DiPT and in duplicate with 1 μM lorazepam (positive control) for up to 24 hours. Metabolite production was monitored at 0, 4, 8, and 24 hours. A negative control group was incubated in duplicate with 1 μM 4-OH-DiPT in the absence of hepatocytes. Positive and negative control samples were assayed at 0, 8, and 24 hours. RE104 and its metabolites were identified and analyzed by ultra-performance liquid chromatography ("Supplementary Methods").

[0295] Forced swim test (FST)

[0296] The FST is a commonly used rat model of antidepressant-like activity that measures immobility, swimming, and climbing behaviors following water exposure; this model was adapted from a previously described dephosphopsilin protocol (which employed a standard FST for up to 35 days after a single administration of a hallucinogenic drug) and revalidated using RE109 as a source of active dephosphopsilin (Hibicke et al. and Slattery et al.).

[0297] Adult male Wistar rats (8 to 10 weeks old) were randomly divided into groups and given a single SC intraperitoneal injection of vehicle, 1 mg / kg RE104, or an equivalent dose of RE-109 (1 mg / kg) (n = 12 / treatment group). The rats were acclimated to the laboratory for 2 days and FST measurements were performed on days 7, 14, and 28. 24 hours before the test, each rat was placed in a cylinder (20 cm diameter, 45 cm high) filled with water for 15 minutes for initial exposure. On the test day, the animals were individually placed in a glass cylinder containing 35 cm of water and maintained at 25°C ± 1°C. The test process was recorded with a video camera for 5 minutes.

[0298] Immobility time, swimming time, and climbing time (in seconds) were measured during the 5-minute test by a blinded observer using a stopwatch based on the recorded video. Statistical analysis was performed using GraphPad Prism-5 software, using one-way analysis of variance followed by a Dunnett's test / Tukey post hoc test or an independent-samples t-test. For trend analysis, a linear repeated-measures model was used for each endpoint. The intercept (model-estimated dosing day), slope (change in swimming, climbing, and immobility time across study days), and P values ​​for the slope parameters were calculated.

[0299] Animal Welfare

[0300] All experiments involving animals were performed according to protocols approved by the Institutional Animal Care and Use Committee of the research institution or equivalent organization.

[0301] result

[0302] Receptor pharmacology

[0303] RE104 prodrug and active compound 4-OH-DiPT have the following chemical structures:

[0304]

[0305] The binding affinity of RE104 and 4-OH-DiPT to the 5-HT2A receptor was determined in HEK-293 cells by radioligand inhibition binding assay. The Ki values ​​of RE104 and 4-OH-DiPT binding at the 5-HT2A receptor were 4300 nM ( Figure 8 ) and 120nM( Figure 9 The binding of RE104 to the 5-HT2B receptor was also evaluated in CHO cells, and the results showed a similar Ki value to that of the 5-HT2A receptor (1800 nM; Figure 10 ).

[0306] To determine whether RE104 prodrug can functionally activate 5-HT2A receptors, a cell agonist effect assay was performed in HEK-293 cells ( Figure 11 The EC50 value of RE104 at the 5-HT2A receptor was >30,000 nM, indicating that RE104 has very low potency at the 5-HT2A receptor.

[0307] Prodrug cleavage

[0308] The kinetics of conversion of the RE104 prodrug to active 4-OH-DiPT were measured in solution and plasma. While RE104 exhibited slow conversion in aqueous solution with changes in pH, conversion was rapid (<30 min) and complete (>95%) in mouse, rat, and human plasma when incubated at 37°C ( Figures 12 to 15 cleavage of the RE104 prodrug was significantly slower in canine plasma (39% remained after 4 hours of incubation). The rate and amount of 4-OH-DiPT formed in all plasma matrices were consistent with hydrolysis of the RE104 glutarate leaving group; 4-OH-DiPT is oxidatively unstable in the medium, as previously reported for the structurally related compound desphosphopsilocybin (Brown et al. and Hasler et al.). The desphosphopsilocybin prodrug, RE109, exhibited similar results to RE104 when incubated with human plasma, with rapid cleavage to desphosphopsilocybin (data not shown).

[0309] Pharmacokinetic studies were conducted in rats, dogs, and nonhuman primates. Estimated mean PK parameters for 4-OH-DiPT following IV, PO, and SC administration of 2 mg / kg RE104 (prodrug) in Sprague Dawley rats are shown in Supplementary Table 1. Following IV administration, RE104 was almost completely metabolized (96.7%) to 4-OH-DiPT within 5 minutes, and RE104 was not measurable in plasma within 5 minutes of administration by any route of administration. The half-life of 4-OH-DiPT was similar following IV and SC administration of RE104 (0.60 and 0.67 hours, respectively). The mean total plasma exposure (area under the curve [AUC]) to 4-OH-DiPT formed from RE104 was slightly higher after SC administration than after IV administration (AUC0-last [standard deviation]: 203 [42.8] h·ng / mL vs. 146 [4.62] h·ng / mL). This may be due to the higher concentration of the SC dosing solution (2.09 mg / mL) compared to the IV dosing solution (1.68 mg / mL) or to variability associated with the small number of animals per treatment group (n=3). Following PO administration, the total plasma exposure to 4-OH-DiPT was extremely low, indicating that RE104 is poorly absorbed in the gastrointestinal tract of rats.

[0310] The PK of 4-OH-DiPT in rats was determined by direct administration of 4-OH-DiPT (1.39 mg / kg IV, PO, and SC). The estimated mean PK parameters of 4-OH-DiPT following administration to rats are summarized in Supplementary Table 2. The lower administered dose was equimolar to 2 mg / kg RE104, which accounts for the smaller molecular weight of the active drug and allows for direct comparison. When administered IV to rats, 4-OH-DiPT exhibited a short half-life (0.74 hours), a steady-state volume of distribution of 5.9 L / kg, and a systemic clearance of 9.4 L / h / kg. Following SC administration, plasma concentrations peaked at 0.61 hours, with a mean bioavailability of 88.6%, whereas PO administration produced peak plasma concentrations at 0.25 hours and a lower mean bioavailability (0.825%).

[0311] The PK of RE104 and 4-OH-DiPT was determined in dogs following IV, SC, and PO administration. Consistent with in vitro cleavage studies, conversion of the prodrug to the active drug was slower in dogs than in rats. RE104 was rapidly absorbed following SC (1 mg / kg) and PO (5 mg / kg) administration, with plasma concentrations exhibiting a rapid decline following an IV bolus (Supplementary Table 3). The mean absolute bioavailability of RE104 was 121% after a single SC dose of 1 mg / kg RE104 and 0.854% after a single PO dose of 5 mg / kg RE104. The mean metabolic ratios of 4-OH-DiPT to RE104 were 0.339, 0.332, and 7.04 after single doses of 1 mg / kg IV RE104, 1 mg / kg SC RE104, and 5 mg / kg PO RE104, respectively.

[0312] Similarly, RE104 was rapidly absorbed after IV (1 mg / kg) and SC (0.5 mg / kg) administration in nonhuman primates, with plasma concentrations of 4-OH-DiPT decreasing from t max The mean terminal half-life of 4-OH-DiPT after IV and SC administration of RE104 was 0.63 hours and 0.95 hours, respectively. Taken together, these data demonstrate that the half-life of the active compound, 4-OH-DiPT, is less than one hour after IV and SC administration of RE104 in vivo and support SC administration of RE104.

[0313] Pharmacodynamics

[0314] The HTR in rodents is a pharmacodynamic measure of 5-HT2A receptor agonism, which is associated with hallucinogenic or psychedelic effects (González-Maeso et al. and Halberstadt et al. (2013)). To determine potential PK-pharmacodynamic relationships, we measured HTR in rats after administration of RE104 and 4-OH-DiPT. The mean number of HTR counts and mean 4-OH-DiPT plasma concentrations in rats at 10-minute intervals are plotted as a function of time. Figure 16 Following IV administration of 2 mg / kg RE104 and 1.39 mg / kg 4-OH-DiPT, HTR peaked immediately and declined over the 4-hour observation period. Following SC administration of 2 mg / kg RE104 or 1.39 mg / kg 4-OH-DiPT, the mean HTR peaked within 1 hour and declined similarly over the observation period. The duration of HTR reflects the PK characteristics of 4-OH-DiPT following IV and SC administration of 2 mg / kg RE104 and 1.39 mg / kg 4-OH-DiPT. In addition, following IV and SC administration of 2 mg / kg RE104 and 1.39 mg / kg 4-OH-DiPT, the mean HTR correlated with 4-OH-DiPT plasma concentrations (r 2 =0.7019; Figures 17 to 20 ).

[0315] metabolism

[0316] The metabolic profile of RE104 was determined in pooled plasma samples from rats, dogs, and nonhuman primates following a single subcutaneous dose of RE104. Hydrolysis to 4-OH-DiPT represented the major metabolic pathway identified in pooled plasma samples from rats administered 10 mg / kg, 25 mg / kg, or 75 mg / kg of RE104 between 0 and 10 hours (accounting for >67% of total metabolites in plasma at all doses). The second most abundant metabolite in rat plasma was 4-hydroxyindole-2-acetic acid (hydrolysis and N-dealkylation), which accounted for up to approximately 18% of total metabolites in pooled plasma samples at all doses of RE104. Other minor metabolic pathways in rats (defined as <10% of total peak area) included hydrolysis followed by (1) oxidation, (2) dual oxidation, (3) glucuronidation, (4) oxidation and glucuronidation, or (5) dual oxidation, dehydrogenation, and glucuronidation.

[0317] In pooled plasma from 0 to 12 hours after subcutaneous administration of 3 mg / kg, 8 mg / kg, and 20 mg / kg of RE104 in dogs, the major metabolite was 4-OH-DiPT, accounting for 3.34% to 16.5% of the total fraction at all doses; the majority of the drug remained in the RE104 form (78.5% to 96.0% at all doses). Other metabolic pathways were minor and produced multiple unidentified products with molecular weights consistent with single and multiple oxidation, glucuronidation, and hydrogenation steps. No other metabolites in this class reached >3% of the total plasma fraction at all doses.

[0318] In pooled plasma samples collected from monkeys 0 to 12 hours after dosing, unchanged RE104 parent molecule accounted for 71.1% of the total peak area, while the active compound 4-OH-DiPT accounted for 4.42% of the total peak area. The most abundant metabolite identified represented the glucuronidation product of 4-OH-DiPT (accounting for 24.5% of the total peak area). As expected for rapid conversion to the active drug, incubation of 1 μM 4-OH-DiPT with mixed plate-cultured human hepatocytes produced primarily the O-glucuronide and negligible amounts of other metabolites (including 4-hydroxyindole-2-acetic acid), indicating that RE104 metabolism is qualitatively similar across the species tested.

[0319] Antidepressant-like activity in the FST

[0320] Antidepressant-like effects were evaluated using a modified FST specifically developed in rats to evaluate the lasting effects of a single dose of hallucinogenic drugs (Hibicke et al.). When RE104 and RE109 were administered, unique serotonergic behaviors (e.g., HTR, piloerection, hindlimb rearing, and body flat posture) were observed on the day of administration for approximately 3 hours; these responses were not observed in vehicle-treated animals. The first FST was performed 7 days after the single dose. Significant reductions in the mean immobility time were observed on days 7, 14, and 28 for RE104, and on days 14 and 28 for RE109 (P < 0.05; Table 2). Figures 21 to 23 ), demonstrating that both compounds have sustained effects.

[0321] The reduction in immobility time was attributed to increased swimming and climbing escape behaviors (no diving was observed). Compared with the vehicle group, the mean swimming time was significantly increased for both RE104 (P < 0.01) and RE109 (P < 0.05). The trend from day 7 to day 28 showed a steady increase in swimming (P = 0.08) and a gradual decrease in climbing (P < 0.004) for RE109, while for RE104, climbing gradually increased (P < 0.02) and swimming decreased (P < 0.001) over 28 days; for both compounds, immobility time remained constant and different from the vehicle group (P < 0.05). By day 28, differences were observed between RE104 and RE109 in the amount of swimming (P < 0.05) and climbing (P < 0.001), with swimming and climbing accounting for 55% and 45% of the escape behavior, respectively, in RE104 and 74% and 26%, respectively, in RE109.

[0322] discuss

[0323] The results presented in the Examples demonstrate that the active compound 4-OH-DiPT exhibits 5-HT2A receptor binding affinity comparable to that of a previously described psilocybin active compound (psilocybin dephosphate), supporting a similar pharmacological profile (Rickli et al.). The prodrug RE104 is rapidly metabolized to 4-OH-DiPT in preclinical models, and the half-life and PK profile of 4-OH-DiPT after RE104 administration correlate with the duration of the pharmacodynamic effect (HTR). Furthermore, a single administration of RE104 in rats elicited sustained antidepressant-like activity over 28 days (assessed by the FST), comparable to that of depsilocybin formed in situ after administration of the psilocybin prodrug RE109. Taken together, these results demonstrate that RE104 administration, likely through the active compound 4-OH-DiPT, induces behavioral effects and antidepressant-like activity comparable to those of psilocybin (González-Maeso et al. and Hibicke et al.).

[0324] The in vitro pharmacology data presented here are consistent with previously observed binding of 4-OH-DiPT to the 5-HT2A receptor, with a Ki value in the nanomolar range (Rickli et al.). In contrast, RE104 exhibited significantly reduced binding to both 5-HT2A and 5-HT2B compared to 4-OH-DiPT, with Ki values ​​in the micromolar range. Functional activation of the 5-HT2A receptor by 4-OH-DiPT has been previously reported, with EC50 values ​​of 6.82 nM and 93 nM (Rickli et al. and Gatch et al.). The EC50 value of RE104 at the 5-HT2A receptor was >30,000 nM, indicating minimal potency that may not be relevant for in vivo pharmacology. These results are also consistent with previous observations that 4-O-acetyltryptamine compounds exhibit reduced 5-HT2A activation compared to their 4-hydroxy counterparts (Klein et al.). Taken together, these results confirm that 4-OH-DiPT may be the sole active agent following administration of RE104.

[0325] 4-OH-DiPT plasma concentrations after RE104 administration correlated with pharmacodynamic HTR responses in rats, similar to the PK-pharmacodynamic and pharmacodynamic-receptor occupancy correlations observed for psilocybin in human volunteers (Madsen et al.). The 4-OH-DiPT half-life observed after IV or SC administration of RE104 in rats was approximately one-third the half-life of dephosphopsilin reported after oral administration of the dephosphopsilin-containing mushroom species Gymnopilus spectabilis (the terminal half-life of 4-OH-DiPT is approximately 40 minutes, while the distribution and elimination half-lives of dephosphopsilin are 117 and 148 minutes, respectively) (Chen et al.). The observed PK and pharmacodynamic profiles are also consistent with anecdotal reports of a 2- to 3-hour duration of hallucinogenic effects of 4-OH-DiPT in humans (Shulgin), suggesting that administration of RE104 in a clinical setting could reduce the need for clinical monitoring compared to administration of psilocybin (6- to 8-hour duration) (von Rotz et al.; Goodwin et al.; and Carbonaro et al.).

[0326] In summary, the proposed distribution-metabolism model for RE104 is that the prodrug is enzymatically converted to 4-OH-DiPT. It is believed that the circulating prodrug is confined to the periphery as a charged zwitterion at neutral pH and cannot readily diffuse into the brain. Active monoamines can penetrate the central nervous system, causing near-immediate but transient behavioral changes (e.g., HTR) and longer-term antidepressant-like behaviors. Circulating 4-OH-DiPT is metabolized to either a glucuronide (major metabolite) or indoleacetic acid (minor metabolite, in rats), both of which are likely inactive and eliminated. Generally speaking, the metabolic profile of RE104 in animal models is similar to that of psilocybin and serotonin (Dinis-Oliveira), although indoleacetic acid formation may be reduced in higher species, particularly humans, due to the difficulty of monoamine oxidase in metabolizing the isopropyl derivative. Combined with in vitro metabolic profiling in human hepatocytes, these data support that RE104 is eliminated primarily as glucuronide metabolites, most likely through urine and feces, again mirroring the situation with psilocybin (Dinis-Oliveira). The low absolute oral bioavailability of RE104 relative to IV or SC administration supports the use of an injectable formulation of RE104.

[0327] RE104 exhibited antidepressant-like activity similar to that of dephosphopsilin in the FST model of prolonged efficacy (Hibicke et al.). Both RE104 and RE109 reduced immobility time compared to the vehicle group, and the effect was maintained for 28 days after a single dose. The findings with RE109 are consistent with previously reported sustained antidepressant-like effects observed in the FST after a single dose of psilocybin, and it is noteworthy that these results have been successfully translated into the clinic (Goodwin et al. and Hibicke et al.). Furthermore, these data are consistent with previous reports that the antidepressant-like effects of serotonergic agonists and reuptake inhibitors are specifically reflected in a reduction in immobility behavior in the FST, further suggesting that the antidepressant effects of serotonergic hallucinogens are mediated by serotonergic modulation (Slattery et al. and Cryan et al.). Although increased swimming and climbing behaviors dominated the remainder of the test period after RE104 and RE109 administration, a trend toward increased climbing with RE104 and decreased climbing with RE109 was observed over the 28-day period (reaching significance on day 28), with complementary trends observed, with increased swimming with RE109 and decreased swimming with RE104. Previous studies have shown that swimming and climbing behaviors respond differently to selective serotonin reuptake inhibitors and selective norepinephrine reuptake inhibitors, respectively, and are therefore attributed to differential modulation of the serotonergic and adrenergic systems (Detke et al. and Lucki et al.). The preference for swimming over climbing in the FST has been observed in longitudinal FST studies of rats administered 5-HT1A agonists and initially psilocybin (Hibicke et al. and Lucki et al.); notably, dephosphopsilin is a more potent 5-HT1A agonist than 4-OH-DiPT (Rickli et al.). Furthermore, in rodents, climbing (but not swimming) is the opposite of immobility (a proxy for depression and / or despair) under a prolonged FST dosing regimen (Masuda et al.). The differences observed in climbing and swimming behaviors in the FST may indicate subtle but persistent differences in the specific receptor system interactions and the location and extent of downstream neuroplastic modifications induced by psychedelic molecules (Rickli et al. and Detke et al.).

[0328] The results of this example demonstrate that the novel prodrug RE104 rapidly cleaves to the 5-HT2A-activating compound 4-OH-DiPT, resulting in short-lived pharmacodynamic effects indicative of hallucinogenic activity in preclinical models. Furthermore, antidepressant-like activity of RE104 was confirmed in a translational proof-of-concept assay. Taken together, these studies demonstrate that RE104 has the potential to provide therapeutic efficacy in human depression, similar to that observed in recent randomized clinical trials of psilocybin, but with a shortened duration of the psychoactive state.

[0329] Supplementary methods

[0330] Receptor pharmacology

[0331] For receptor binding assays, test compound concentrations were 3 nM, 30 nM, 100 nM, 300 nM, 1 μM, 3 μM, 10 μM, 30 μM (4-OH-DiPT) and 10 nM, 30 nM, 100 nM, 300 nM, 1 μM, 3 μM, 10 μM, and 30 μM (RE104). The half-maximal inhibitory concentration (IC50) and Hill coefficient were determined by nonlinear regression analysis of competition curves generated using the Hill equation curve fit of averaged replicate (n=2) values. The inhibition constant was calculated using the Cheng-Prusoff equation: Ki=IC50 / (1+L / KD), where L=radioligand concentration and KD=affinity of the radioligand for the receptor (determined by Scatchard plot). In the functional activity assay, RE104 was tested at concentrations of 10 nM, 30 nM, 100 nM, 300 nM, 1 μM, 3 μM, 10 μM, and 30 μM. EC50 and IC50 values ​​were determined by nonlinear regression analysis of concentration-response curves generated using Hill equation curve fitting of averaged replicate values. For all receptor pharmacology assays, data were analyzed using software developed by Cerep (Celle-Lévescault, France) (Hill software) and analyzed by integration with commercial software. 4.0 (for )( 1997SPSS Inc.) to compare and verify the results.

[0332] Prodrug cleavage

[0333] Stock solutions of 0.1 mg / mL 4-OH-DiPT and RE104 were prepared in dimethyl sulfoxide; the stock solutions were diluted 100-fold to 1 μg / mL with 2 mL of mixed-sex human, mixed-sex beagle dog, male CD-1 mouse, or male Sprague Dawley rat plasma (containing K2EDTA as an anticoagulant). Three aliquots (50 μL) of each spiked plasma solution were immediately added to 200 μL of ice-cold methanol / acetonitrile (50:50 v / v) containing the internal standard to generate time 0 samples. The spiked plasma solutions were incubated at 37°C on an orbital shaker set to 75 rpm, and three 50 μL samples were removed in triplicate after 30 minutes and 1, 2, and 4 hours of incubation. Each sample was immediately added to 200 μL of ice-cold methanol / acetonitrile containing the internal standard to terminate the reaction. Samples were mixed by vortexing and stored at −80°C until analysis.

[0334] Because RE104 was observed to be unstable in mouse, rat, and human plasma, plasma containing calibration standards could not be used for absolute quantification; therefore, the analyte / internal standard peak area ratio is reported as a function of incubation time. For 4-OH-DiPT stability samples, mouse, dog, and human plasma samples were analyzed using rat plasma calibration standards. Calibration standards were prepared in plasma for each species and analyzed in batches by LC-MS / MS. The sample batch consisted of three replicates of the system suitability standard (containing the analyte and internal standard), the appropriate calibration standards in ascending order (including a blank sample without internal standard, a zero sample (containing internal standard), and at least six non-zero standards), the assay samples, and finally three replicates of the system suitability sample.

[0335] PK sampling methods and time points

[0336] For rat PK evaluation, approximately 0.25 mL of blood was collected continuously at 8 time points (5 minutes, 15 minutes, 30 minutes, 45 minutes, and 1 hour, 2 hours, 4 hours, and 6 hours after administration) over 6 hours via a surgically placed carotid artery catheter and immediately transferred to a tube containing K2EDTA placed on ice. For dog PK evaluation, approximately 0.5 mL of blood was collected via a superficial vein before administration and 5 minutes (only for IV and SC injections), 15 minutes, 30 minutes, and 1 hour, 2 hours, 4 hours, 8 hours, 16 hours, 24 hours, 48 ​​hours, and 72 hours after administration, and collected into a tube containing K2EDTA, wherein 10 mM dichlorvos was added as an anticoagulant and stabilizer, respectively. For nonhuman primate PK assessments, blood samples (approximately 1 mL) were collected via femoral vein puncture into tubes containing K2EDTA and 10 mM dichlorvos before dosing and at 5, 15, 30 minutes and 1, 1.5, 2, 4, 8, and 12 hours after dosing.

[0337] Metabolite Profiling and Identification For experiments conducted in rats, blood samples were collected before dosing and at 15 minutes, 30 minutes, and 1 hour, 2 hours, 4 hours, 6 hours, and 10 hours after dosing. Blood samples were collected from dogs before dosing and at 5 minutes, 30 minutes, and 1 hour, 2 hours, 4 hours, 8 hours, 12 hours, and 24 hours after dosing. Blood samples were collected from monkeys before dosing and at 5 minutes, 15 minutes, 30 minutes, and 1 hour, 1.5 hours, 2 hours, 4 hours, 8 hours, and 12 hours after dosing. To the pooled plasma samples, a volume of acetonitrile:methanol:formic acid (50:50:0.1, v / v / v) equal to three times the plasma volume was added, vortexed for 3 minutes, and then centrifuged at 3000 rpm for 10 minutes at 4°C. The supernatant was evaporated in a 25°C water bath under a stream of nitrogen. The dried residue was reconstituted with acetonitrile:methanol:water:formic acid (15:15:70:0.1, v / v / v / v), centrifuged at 3000 rpm for 10 min, and then transferred to a vial for ultra-high performance liquid chromatography (UHPLC) analysis.

[0338] For plate-cultured human hepatocytes, all samples were quenched with an equal volume of acetonitrile:methanol:formic acid (50:50:0.1, v / v / v) after incubation with 4-OH-DiPT. Quenched samples were mixed by pipetting, collected, and stored at -70°C or below until analysis by liquid chromatography-tandem mass spectrometry. Positive control lorazepam samples were processed identically to 4-OH-DiPT. Samples incubated with plate-cultured human hepatocytes were immediately frozen at ≤ -70°C or below and thawed to room temperature before sample processing. A 500 μL aliquot of each sample was transferred to a 1.4 mL matrix tube, followed by the addition of 500 μL of a 50:50:0.1, v / v / v, mixture of acetonitrile:methanol:formic acid. The resulting mixture was vortexed for 2 minutes and then centrifuged at 3000 rpm for 10 minutes. The supernatants from the three replicate incubations were combined to generate a pooled sample (1.5 mL each) for LC-MS analysis.

[0339] For all metabolite identification and profiling experiments, Shimadzu Nexera TM UHPLC system (coupled with high-resolution mass spectrometer SCIEX 6600Triple ) RE104 and its metabolites were identified and profiled by UHPLC. Mass spectrometry analysis was performed using an electrospray ionization source operating in positive ion mode, employing full-scan TOF MS and MS / MS modes. Proposed metabolite structures were based on the observed accurate masses of the molecular ions and their associated mass fragmentation patterns. The fragmentation patterns of the proposed metabolites were compared with common fragment ions obtained from RE104 and 4-OH-DiPT reference standards. Unknown metabolites were searched using Metabolite Pilot software from Sciex. Molecular ions of potential metabolites were searched based on metabolic pathways.

[0340] The percentages of RE104 and its metabolites relative to the total area of ​​compound-related substances were obtained by integrating the extracted ion chromatogram peaks on the full-scan mass chromatogram. The percentage of each component's peak area relative to the total peak area was determined using the following formula: Component % Peak Area = (Component Peak Area × 100%) / Total Integrated Peak Area of ​​Compound-Related Substances.

[0341] *********************

[0342] Although the present invention has been described with reference to illustrative embodiments and examples, this description is not intended to be construed in a limiting sense. Therefore, various modifications of these illustrative embodiments, as well as other embodiments of the present invention, will become apparent to those skilled in the art upon reference to this description. It is therefore intended that the appended claims will cover any such modifications or embodiments.

[0343] All publications, patents, and patent applications cited herein are incorporated by reference in their entirety to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference in its entirety.

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[0405] von Rotz R,Schindowski EM,Jungwirth J,Schuldt A,Rieser NM,ZahoranszkyK,et al.Single-dose psilocybin-assisted therapy in major depressive disorder:A placebo-controlled,double-blind,randomised clinicaltrial.EClinicalMedicine.2023;56:101809.

[0406] Willins DL,Meltzer HY.Direct injection of 5-HT2A receptor agonistsinto the medial prefrontal cortex produces a head-twitch response in rats.JPharmacol Exp Ther.1997Aug;282(2):699-706.PMID:9262333.

[0407] Definitions and Interpretations

[0408] The description of the present invention is presented for the purpose of illustration and description, but is not intended to be exhaustive or to limit the invention to the disclosed form. Many modifications and variations will be apparent to those of ordinary skill in the art without departing from the scope and spirit of the invention. The selection and description of the embodiments are in order to best explain the principles and practical applications of the invention and to enable others of ordinary skill in the art to understand that the invention has various modifications suitable for the specific purposes envisioned in various embodiments. To the extent that the following description relates to specific embodiments or specific purposes of the invention, it is intended to be illustrative only and not to limit the invention as claimed.

[0409] The structures, materials, acts, and equivalents thereof corresponding to all means or step plus function elements in the claims appended hereto are intended to include any structure, material, or act for performing the function, in combination with other claimed elements, as specifically described in the claims.

[0410] References in the specification to "one embodiment" or the like indicate that the embodiment being described may include a particular aspect, feature, structure, or characteristic, but not every embodiment necessarily includes that aspect, feature, structure, or characteristic. Furthermore, such phrases may, but do not necessarily, refer to the same embodiment referenced elsewhere in the specification. In addition, when a particular aspect, feature, structure, or characteristic is described in conjunction with one embodiment, those skilled in the art will appreciate that that aspect, feature, structure, or characteristic may be combined with, connected to, or used to influence other embodiments, regardless of whether such connection or combination is explicitly described. In other words, in different embodiments, any element or feature may be combined with any other element or feature unless there is an obvious or inherent incompatibility between the two, or they are specifically excluded.

[0411] It is further noted that the claims may be drafted to exclude any optional element. Therefore, this statement is intended to serve as a precondition for the use of exclusive terminology (such as "only," "only," etc.) in the recitation of claim elements or the use of "negative" limitations. The terms "preferably," "preferred," "preferably," "optionally," "may," and similar terms are used to indicate that the referenced element, item, condition, or step is an optional (non-essential) feature of the present invention.

[0412] The singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. The term "and / or" means any one, any combination of items, or all of the items associated with it.

[0413] As will be understood by those skilled in the art, for any and all purposes, particularly in terms of providing a written description, all ranges cited herein also encompass any and all possible subranges and combinations of subranges thereof, as well as the individual values, particularly integer values, constituting the ranges. The ranges cited (e.g., weight percent or carbon group) include each particular value, integer, decimal, or equivalent within the range. Any listed range can be readily understood to fully describe and enable the same range to be decomposed into at least equal half, one-third, one-quarter, one-fifth, or one-tenth. As a non-limiting example, each range discussed herein can be readily decomposed into a lower third, a middle third, and an upper third, etc.

[0414] As will also be understood by those skilled in the art, all ranges described herein and all language such as "between," "up to," "at least," "greater than," "less than," "more than," "or more," etc., include the recited numbers and that such terms refer to ranges that can subsequently be broken down into sub-ranges as discussed above.

[0415] Implementation Plan List

[0416] The following is a list of non-limiting embodiments of the present invention:

[0417] 1. An injectable composition comprising a compound of formula (I):

[0418]

[0419] or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, wherein the compound of formula (I) is present in a dosage of about 30 mg to about 50 mg calculated as the free base.

[0420] 2. An injectable composition comprising a compound of formula (I):

[0421]

[0422] or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, wherein the compound of formula (I) is present in a dosage of 30 mg or 40 mg calculated as a free base.

[0423] 3. The injectable composition according to embodiment 1 or embodiment 2, wherein the compound of formula (I) is its hydrochloride salt.

[0424] 4. The injectable composition according to embodiment 2, wherein the compound of formula (I) is its hydrochloride salt present in a dose of 33 mg or 44 mg.

[0425] 5. The injectable composition according to any one of embodiments 1 to 4, wherein the injectable composition is a subcutaneously injectable composition.

[0426] 6. A method for treating a mental disorder, comprising administering to a patient in need of treatment a pharmaceutical composition comprising a compound of formula (I):

[0427]

[0428] and a pharmaceutically acceptable carrier, the dosage of the compound of formula (I) is about 30 mg to about 50 mg calculated as the free base.

[0429] 7. A method for treating a mental disorder, comprising administering to a patient in need of treatment a pharmaceutical composition comprising a compound of formula (I):

[0430]

[0431] and a pharmaceutically acceptable carrier, the dosage of the compound of formula (I) is about 30 mg or about 40 mg calculated as the free base.

[0432] 8. The method according to embodiment 6 or embodiment 7, wherein the compound of formula (I) is its hydrochloride salt.

[0433] 9. The method according to embodiment 7, wherein the compound of formula (I) is its hydrochloride salt present in a dose of 33 mg or 44 mg.

[0434] 10. A method for treating a mental disorder, comprising administering to a patient in need of treatment a pharmaceutical composition comprising a compound of formula (I):

[0435]

[0436] and a pharmaceutically acceptable carrier, the dosage of the compound of formula (I) is 1 mg / kg calculated as the free base.

[0437] 11. The method of any one of embodiments 6 to 10, wherein the duration of action of the compound of formula (I) is from about 2.5 hours to about 4.5 hours after administration.

[0438] 12. The method according to any one of embodiments 6 to 10, wherein the duration of action of the compound of formula (I) is about 3.5 hours or 3.7 hours after administration.

[0439] 13. The method according to any one of embodiments 6 to 12, wherein the compound of formula (I) is in the patient's body C maxIn the range of about 1000 ng / mL to about 2000 ng / mL.

[0440] 14. The method according to any one of embodiments 6 to 12, wherein the compound of formula (I) is in the patient's body C max In the range of about 1500 ng / mL to about 1800 ng / mL.

[0441] 15. The method according to any one of embodiments 6 to 14, wherein the compound of formula (I) is in the patient's body at T max In the range of about 20 minutes to 30 minutes.

[0442] 16. The method according to any one of embodiments 6 to 14, wherein the compound of formula (I) is in the patient's body at T max In about 15 minutes.

[0443] 17. The method according to any one of embodiments 6 to 16, wherein the compound of formula (I) is converted to the compound of formula (II) after administration

[0444]

[0445] And the compound of formula (II) is C in the patient's body max In the range of about 100 ng / mL to about 300 ng / mL.

[0446] 18. The method according to any one of embodiments 6 to 16, wherein the compound of formula (I) is converted to the compound of formula (II) after administration

[0447]

[0448] And the compound of formula (II) is C in the patient's body max In the range of about 120 ng / mL to about 300 ng / mL.

[0449] 19. The method according to any one of embodiments 6 to 18, wherein the compound of formula (I) is converted to the compound of formula (II) after administration

[0450]

[0451] And the compound of formula (II) has a T max In the range of about 60 minutes to about 150 minutes.

[0452] 20. The method according to any one of embodiments 6 to 18, wherein the compound of formula (I) is converted to the compound of formula (II) after administration

[0453]

[0454] And the compound of formula (II) has a T max In the range of about 60 minutes to about 80 minutes.

[0455] 21. The method of any one of embodiments 6 to 20, which shows a mean MEQ30 score in at least 50% of patients.

[0456] 22. The method of any one of embodiments 6 to 20, which shows a mean MEQ30 score in at least 60% of patients.

[0457] 23. The method according to any one of embodiments 6 to 22, wherein the pharmaceutical composition is an injectable pharmaceutical composition.

[0458] 24. The method according to any one of embodiments 6 to 22, wherein the pharmaceutical composition is a subcutaneously injectable pharmaceutical composition.

[0459] Table 1 Maximum Recommended Safe Starting Dose (MRSD) of RE104 (zwitterion)

[0460]

[0461] Abbreviations: BSA = body surface area; HED = human equivalent dose; NOAEL = no observed adverse effect level; MRSD = maximum recommended safe starting dose.

[0462] a. HED is calculated based on the animal-to-human BSA conversion factors detailed in the 2005 FDA guidance.

[0463] b. Based on 60kg body weight.

[0464] c. Based on 1.62m 2 BSA.

[0465] d. The NOAEL for female rats is 10 mg / kg

[0466] e. The NOAEL for both male and female dogs is 8 mg / kg.

[0467] Table 2 – Proposed dose levels for cohorts 1 to 4

[0468] queue <![CDATA[Proposed RE104 HCl dose (mg) a > Proportional NOAEL dose reduction by X times b 1 5.5 19 2 11 9.7 3 22 4.9 4 33 3.2

[0469] a. The doses shown in the table are all calculated in terms of RE104 HCl. These doses are equivalent to 5 mg, 10 mg, 20 mg, and 30 mg of RE104 zwitterion form, respectively.

[0470] b. Example calculation for Cohort 1: 97.2 mg / 5 mg = 19-fold reduction in the proportional NOAEL dose.

[0471] Table 3 - Dosing regimen for study RE104-101

[0472]

[0473] Table 4 Prohibited drugs

[0474]

[0475]

[0476] Abbreviations: 5-MeO-DMT = 5-methoxy-N,N-dimethyltryptamine; AE = adverse event; DMT = dimethyltryptamine; EOS = end of study; ET = premature termination; LSD = lysergic acid diethylamide (LSD); OTC = over-the-counter; PK = pharmacokinetics.

[0477] Table 5: Agents that can induce serotonin syndrome, alone or in combination with other serotonergic agents

[0478]

[0479]

[0480]

[0481] source: https: / / www.uspharmacist.com / article / drug-induced-serotonin- syndrome and https: / / www.uptodate.com / contents / serotonin-syndrome-serotonin- toxicity#H3 .

[0482] Table 6 – Summary of AEs

[0483]

[0484] AEs classified by preferred term (≥2 participants across all dose levels)

[0485]

[0486] SAE, serious adverse event

[0487] Table 7 – Summary of MEQ responders by dose group

[0488]

[0489] MEQ, Mystical Experience Questionnaire.

[0490] Table 8 – Mean Pharmacokinetics (Cohorts 1 to 6)

[0491]

[0492] C max , maximum plasma concentration; CV, coefficient of variation; t 1 / 2 , half-life; T max , the time to reach maximum plasma concentration.

[0493] Supplementary Appendix

[0494] Supplementary Table 1. Summary of estimated mean pharmacokinetic parameters of 4-OH-DiPT in rats after administration of 2 mg / kg RE104

[0495]

[0496]

[0497] 4-OH-DiPT, 4-hydroxy-N,N-diisopropyltryptamine; AUC0-, area under the curve extrapolated from the time of administration to infinity; AUC0-last, area under the curve from the time of administration to tlast; Cmax, maximum concentration observed after non-parenteral administration; fm, fraction metabolized (AUC iv [Parent] Prodrug / AUC iv [Parent] parent drug, at equimolar doses); IV, intravenous; MRTO, mean residence time extrapolated to infinity from the time of administration; PO, oral; NA, not applicable; SC, subcutaneous; SD, standard deviation; t1 / 2, half-life; tmax, time to maximum concentration observed after non-parenteral administration.

[0498] aCalculated based on the AUC0--(IV) of 4-OH-DiPT administered at equimolar doses (152 h·ng / mL).

[0499] Supplementary Table 2. Estimated pharmacokinetic parameters of 4-OH-DiPT in rats after administration of 1.39 mg / kg 4-OH-DiPT Value Summary

[0500]

[0501] 4-OH-DiPT, 4-hydroxy-N,N-diisopropyltryptamine; AUC0-, area under the curve extrapolated from the time of administration to infinity; AUC0-last, area under the curve from the time of administration to tlast; C0, concentration extrapolated to time 0 after IV administration; CL, clearance; Cmax, maximum concentration observed after non-parenteral administration; F, oral bioavailability ([dose IV ×AUC PO ] / [dose PO ×AUC IV]×100); IV, intravenous; MRTO, mean residence time extrapolated to infinity from the time of administration; PO, oral; NA, not applicable; SC, subcutaneous; SD, standard deviation; t1 / 2, half-life; tmax, time to maximum concentration observed after non-parenteral administration; Vss, volume of distribution at steady state.

[0502]

Claims

1. An injectable composition comprising a compound of formula (I): (I) or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, wherein the compound of formula (I) is present in a dosage of about 30 mg to about 50 mg calculated as the free base.

2. An injectable composition comprising a compound of formula (I): (I) or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, wherein the compound of formula (I) is present in a dosage of 30 mg or 40 mg calculated as a free base.

3. An injectable composition according to claim 1 or claim 2, wherein the compound of formula (I) is its hydrochloride salt.

4. The injectable composition according to claim 2, wherein the compound of formula (I) is its hydrochloride salt present in a dose of 33 mg or 44 mg.

5. The injectable composition according to any one of claims 1 to 4, wherein the injectable composition is a subcutaneously injectable composition.

6. A method for treating a mental disorder, comprising administering to a patient in need of treatment a pharmaceutical composition comprising a compound of formula (I): (I) and a pharmaceutically acceptable carrier, the dosage of the compound of formula (I) is about 30 mg to about 50 mg calculated as the free base.

7. A method for treating a mental disorder, comprising administering to a patient in need of treatment a pharmaceutical composition comprising a compound of formula (I): (I) and a pharmaceutically acceptable carrier, the dosage of the compound of formula (I) is about 30 mg or about 40 mg calculated as the free base.

8. The method according to claim 6 or claim 7, wherein the compound of formula (I) is its hydrochloride salt.

9. The method according to claim 7, wherein the compound of formula (I) is its hydrochloride salt present in a dosage of 33 mg or 44 mg.

10. A method for treating a mental disorder, comprising administering to a patient in need of treatment a pharmaceutical composition comprising a compound of formula (I): (I) and a pharmaceutically acceptable carrier, the dosage of the compound of formula (I) is 1 mg / kg calculated as the free base.

11. The method of any one of claims 6 to 10, wherein the duration of action of the compound of formula (I) is from about 2.5 hours to about 4.5 hours after administration.

12. The method of any one of claims 6 to 10, wherein the duration of action of the compound of formula (I) is about 3.5 hours or 3.7 hours after administration.

13. The method according to any one of claims 6 to 12, wherein the compound of formula (I) is in the patient's body C max In the range of about 1000 ng / mL to about 2000 ng / mL.

14. The method according to any one of claims 6 to 12, wherein the compound of formula (I) is in the patient's body C max In the range of about 1500 ng / mL to about 1800 ng / mL.

15. The method according to any one of claims 6 to 14, wherein the compound of formula (I) is in the patient's body at T max In the range of about 20 minutes to 30 minutes.

16. The method according to any one of claims 6 to 14, wherein the compound of formula (I) is in the patient's body at T max In about 15 minutes.

17. The method according to any one of claims 6 to 16, wherein the compound of formula (I) is converted to the compound of formula (II) after administration (II) And the compound of formula (II) is C in the patient's body max In the range of about 100 ng / mL to about 300 ng / mL.

18. The method according to any one of claims 6 to 16, wherein the compound of formula (I) is converted to the compound of formula (II) after administration (II) And the compound of formula (II) is C in the patient's body max In the range of about 120 ng / mL to about 300 ng / mL.

19. The method according to any one of claims 6 to 18, wherein the compound of formula (I) is converted to the compound of formula (II) after administration (II) And the compound of formula (II) has a T max In the range of about 60 minutes to about 150 minutes.

20. The method according to any one of claims 6 to 18, wherein the compound of formula (I) is converted to the compound of formula (II) after administration (II) And the compound of formula (II) has a T max In the range of about 60 minutes to about 80 minutes.

21. The method according to any one of claims 6 to 20, which shows a mean MEQ30 score in at least 50% of patients.

22. The method according to any one of claims 6 to 20, which shows a mean MEQ30 score in at least 60% of patients.

23. The method according to any one of claims 6 to 22, wherein the pharmaceutical composition is an injectable pharmaceutical composition.

24. The method according to any one of claims 6 to 22, wherein the pharmaceutical composition is a subcutaneously injectable pharmaceutical composition.

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