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WO2025231265A1 - Combinaison comprenant un agoniste du récepteur adrénergique alpha-2 - Google Patents

Combinaison comprenant un agoniste du récepteur adrénergique alpha-2

Info

Publication number
WO2025231265A1
WO2025231265A1 PCT/US2025/027331 US2025027331W WO2025231265A1 WO 2025231265 A1 WO2025231265 A1 WO 2025231265A1 US 2025027331 W US2025027331 W US 2025027331W WO 2025231265 A1 WO2025231265 A1 WO 2025231265A1
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WO
WIPO (PCT)
Prior art keywords
alkylene
compound
pain
yield
chosen
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
PCT/US2025/027331
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English (en)
Inventor
Lingyu Zhu
Zhiqiang Cheng
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Alpherabio LLC
Original Assignee
Alpherabio LLC
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Filing date
Publication date
Application filed by Alpherabio LLC filed Critical Alpherabio LLC
Publication of WO2025231265A1 publication Critical patent/WO2025231265A1/fr
Pending legal-status Critical Current
Anticipated expiration legal-status Critical

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Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/41—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with two or more ring hetero atoms, at least one of which being nitrogen, e.g. tetrazole
    • A61K31/4164—1,3-Diazoles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/02—Drugs for disorders of the nervous system for peripheral neuropathies
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D403/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00
    • C07D403/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings
    • C07D403/10—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings linked by a carbon chain containing aromatic rings

Definitions

  • ⁇ 2AR agonists for the treatment or prevention of diseases thereof, either usded alone or in combination with a second therapeutical agent.
  • CNS central nervous system
  • Clonidine was first developed to manage hypertension. Administered orally, clonidine diffuses into the CNS and activates the ⁇ 2AR in the nucleus tractus solitarii (NTS), which in turn triggers a pathway inhibiting excitatory cardiovascular neurons. This cascade effectively reduces sympathetic outflow from the CNS, leading to a clinical decrease in arterial blood pressure.
  • NTS nucleus tractus solitarii
  • clonidine was found to induce sedation by acting through the activation of central pre- and postsynaptic ⁇ 2AR in the locus coeruleus (LC), a nucleus in the medial dorsal pons, thereby inducing sedative effects.
  • LC locus coeruleus
  • clonidine has been approved for epidural use under the trade name Duraclon, marking a significant advancement in the treatment of cancer pain.
  • Duraclon has been documented to induce centrally mediated sedation, hypotension, bradycardia, and depression of its applications, which persist throughout the analgesic treatment process. Such sedation effect significantly limits the dosages that can be administered safely.
  • the present disclosure relates to methods of treating or preventing a disease in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a peripherally selective alpha-2 adrenergic receptor ( ⁇ 2AR) agonist.
  • ⁇ 2AR alpha-2 adrenergic receptor
  • the peripherally selective ⁇ 2AR agonist activates at least one sub type of ⁇ 2AR, particularly ⁇ 2A AR, ⁇ 2B AR, or ⁇ 2C AR.
  • the peripherally selective ⁇ 2AR agonist has a Kp,uu,brain is lower than 0.05, 0.02, or 0.01.
  • the disease is chosen from pain, rosacea, spasticity, and aging.
  • the peripherally selective ⁇ 2AR agonist causes reduced biological effects mediated by CNS, such as sedation, hypotension, and bradycardia, than treating with a non-peripherally selective ⁇ 2AR agonist. 2 Attorney Docket No.: 071741.11025/5WO1 [0013]
  • the present disclosure provides a peripherally selective ⁇ 2AR agonist that comprises an ⁇ 2AR activation moiety covalently linked to a peripheral distribution moiety, and its uses in the treatment of a disease.
  • the present disclosure relates to methods of treating or preventing a disease in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of a peripherally selective ⁇ 2AR agonist, wherein the peripherally selective ⁇ 2AR agonist comprises an ⁇ 2AR activation moiety covalently linked to a peripheral distribution moiety.
  • the peripherally selective ⁇ 2AR agonist causes less sedation than treating with a non-peripherally selective ⁇ 2AR agonist.
  • the present disclosure relates to a compound of formula (I-D): N NH T , or a ste er, pharmaceutically acceptable salt or solvate thereof, wherein, Y 1 is CH, or N; X 1 is chosen from H, D, and halogen; RT is defined as above in formula (I-A).
  • the present disclosure relates to a compound of formula (II): R T B , or a ste r, tautomer, pharmaceutically acceptable salt or solvate thereof, wherein, A is one chosen from: S 1 ; each R 1 is independently chosen from hydrogen, halogen, haloalkyl, hydroxyl, hydroxyalkyl, alkoxy, alkyl, and -COOH; B is one chosen from: 6 Attorney Docket No.: 071741.11025/5WO1 H N N H X S N N H; oaryl, wherein the C3-12 cycloalkyl or C2-12 heterocyclyl is optionally fused with an aryl; r is 1 or 2; n2 is 0, 1, or 2; each R 2 is independently chosen from hydrogen, halogen, hydroxyl, and alkoxy; R 3 is chosen from CN, hydroxy, alkoxy, -C(O)-C 0-12 alkylene-CN, -C 0-12 alkylene-
  • the present disclosure relates to a pharmaceutical composition
  • a pharmaceutical composition comprising a compound as described herein or a stereoisomer, tautomer, pharmaceutically acceptable salt or solvate thereof, and a pharmaceutically acceptable carrier.
  • the present disclosure relates to the use of a compound as described herein or a stereoisomer, tautomer, pharmaceutically acceptable salt or solvate thereof, for treating or preventing a disease, including pain, glaucoma, spasticity, nasal congestion, rosacea, rhinitis, anesthesia, presbyopia, acute kidney injury, insomnia, inflammatory disease, cancer, etc. in a subject in need thereof.
  • the present disclosure relates to a pharmaceutical combination comprising a first therapeutical agent and a second therapeutical agent, wherein the first therapeutical agent is the peripherally selective ⁇ 2AR agonist described herein.
  • the pharmaceutical combination further comprises a third therapeutic agent.
  • the second therapeutic agent is chosen from opioids, antidepressant medications, antiepileptic medications, local analgesics, nonsteroidal anti- inflammatory drugs (NSAIDs), acetaminophen (paracetamol), corticosteroids, NMDA antagonists, and selective NaV1.8 inhibitors.
  • the present disclosure relates to a method of treating or preventing a disease in a subject in need thereof, the method comprising administering to the subject (i) an effective amount of a first therapeutic agent, and (ii) a second therapeutic agent, wherein the first therapeutic agent is a peripherally selective alpha-2 adrenergic receptor ( ⁇ 2AR) agonist.
  • the method further comprises administering to the subject an effective amount of a third therapeutic agent.
  • the disease is chosen from glaucoma, analgesia, spasticity, nasal congestion, rosacea, rhinitis, anesthesia, menopausal hot flashes, dysmenorrhea, presbyopia, acute kidney injury, insomnia, inflammatory disease, and cancer.
  • the second therapeutic agent is chosen from opioids, antidepressant medications, antiepileptic medications, local analgesics, nonsteroidal anti- inflammatory drugs (NSAIDs), acetaminophen (paracetamol), corticosteroids, NMDA receptor antagonists, and selective NaV1.8 inhibitors.
  • the administration of the first therapeutic agent and the second therapeutic agent provides a synergistic effect in treating the disease.
  • the administration of the first therapeutic agent and the second therapeutic agent provides an additive effect in treating the disease.
  • the present disclosure relates to a method for treating or preventing pain in a subject in need thereof, the method comprising administering to the subject an effective amount of a first analgesic agent, and an effective amount of a second analgesic agent, wherein first analgesic agent is a peripherally selective alpha-2 adrenergic receptor ( ⁇ 2AR) agonist.
  • ⁇ 2AR peripherally selective alpha-2 adrenergic receptor
  • the method for treating or preventing pain further comprising administering to the subject an effective amount of a third analgesic agent.
  • the pain is neuropathic pain, nociceptive pain, nociplastic pain, or mixed pain (a mixture of nociceptive, neuropathic, and/or nociplastic pain).
  • the second analgesic agent is any analgesic agent other than a peripherally selective alpha-2 adrenergic receptor ( ⁇ 2AR) agonist.
  • the third analgesic agent is any analgesic agent other than a peripherally selective alpha-2 adrenergic receptor ( ⁇ 2AR) agonist, and the third analgesic agent is different from the second analgesic agent.
  • ⁇ 2AR peripherally selective alpha-2 adrenergic receptor
  • the second analgesic agent is chosen from opioids, acetaminophen (paracetamol), local analgesics, nonsteroidal anti-inflammatory drugs (NSAIDs), antiepileptic medication, antidepressant medication, topical analgesic agents, NMDA receptor antagonists, neurotoxins, cannabinoids, voltage-gated sodium channel inhibitors, adenosine agonists, transient receptor potential (TRP) channel modulators, NGF inhibitors, purinergic receptor antagonists, adenosinergic pathway modulators, sigma-1 receptor antagonists, KCC2 enhancers, glutamate transport enhancers, TrkA receptor antagonists, somatostatin receptor ligands, Human Adenylyl Cyclase Associated Protein 1 (CAP1) modulators, and angiotensin II receptor antagonists, corticosteroids, and any combinations thereof.
  • opioids opioids
  • acetaminophen paracetamol
  • NSAIDs nonsteroidal anti-inflammatory drugs
  • the second analgesic agent is chosen from GGG tri-agonist (e.g., retatrutide), metabotropic glutamate receptors (mGluR) modulator (e.g., basimglurant), TGF beta-1 Inhibitor (e.g., Vicatertide (SB-01)), gonadotropin-releasing hormone receptor(GnRH) antagonist (e.g., linzagolix), 11 ⁇ -HSD1 inhibitor (e.g., clofutriben), 17-HSD-1 inhibitor (e.g., OG-6219), Dual ENKephalinase Inhibitor (DENKI) (e.g., PL37), CCR2 receptor antagonist (e.g., CNTX-6970), IL- 10 receptor agonist (e.g., XT-150), nAChR dual alpha 4/beta 2 subunit stimulator (e.g., ATA-104), allosteric NEK7
  • GGG tri-agonist
  • the second analgesic agent is chosen from IRX-101, MR-107A-02, 3-VM-1001, lysergide assisted therapy, SIL-1002, TRN-261, HR-1405-01, HRF-2105, TTAX-03, KP-910, LYT-503, LL-50, ZeP-3, YR-1702, YZJ-1495, FB-1003, MK-4318, PZH-2108, HEC- 137076MsOH, VVZ-2471, SYNP-101, Pudafensine, BIOS-0618, and HSK-36357.
  • the present disclosure relates to a method for treating or preventing a disease in a subject in need thereof, the method comprising administering to the subject an effective amount of a codrug that comprises a first therapeutic agent and a second therapeutic agent, wherein first therapeutic agent is a peripherally selective alpha-2 adrenergic receptor ( ⁇ 2AR) agonist, such as the peripherally selective ⁇ 2AR agonist described herein.
  • first therapeutic agent is a peripherally selective alpha-2 adrenergic receptor ( ⁇ 2AR) agonist, such as the peripherally selective ⁇ 2AR agonist described herein.
  • ⁇ 2AR peripherally selective alpha-2 adrenergic receptor
  • the method for treating or preventing the disease has a synergistic effect or additive effect.
  • the method for treating or preventing the disease has a sparing effect on the second therapeutic agent or the second analgesic agent.
  • FIGs.1A-1H PWT value of sham and spared nerve injury SNI mouse model, vehicle and drug treatment groups at 1 hour after dosing.
  • FIG.1A 3 mg/mL pregabalin p.o.
  • FIG.1B 1 mg/mL morphine s.c.
  • FIG.1C 1 mg/mL compound 1-B p.o. and 10mg/mL compound 1-B p.o.
  • FIG.1D 1 mg/mL compound 10-B p.o.
  • FIG.1E 1 mg/mL compound 44-B p.o. and 1 mg/mL compound 45-B p.o.
  • FIG.1F 1 mg/mL compound 46-B p.o.
  • FIGs.2A-2D PWT value of sham and bone cancer pain mouse (BCP) model, vehicle and drug treatment groups at 1 hour after dosing.
  • FIG.2A 3 mg/mL pregabalin p.o.
  • FIG.2B 1 mg/mL morphine s.c.
  • FIG.2C 1 mg/mL compound 44-B p.o.
  • FIG.2D 20 mg/mL compound 1-B p.o. and : 20 mg/mL compound 44-B p.o.
  • FIGs.3A-3C PWT value of sham and post-surgery pain mouse model, vehicle and drug treatment groups at 1 hour after dosing.
  • FIG.3A 10 mg/mL compound 1-B p.o.
  • FIG.3B 10 mg/mL compound 44-B p.o.
  • FIG.3C 3 mg/mL morphine s.c.
  • FIGs.4A-4O Analgesic ratio (%) following single-drug and combination administrations in SNI and BCP mouse models.
  • FIG.4A 1 mg/mL 1-B p.o., 1 mg/mL morphine s.c. and their combination in SNI
  • FIG.4B 1 mg/mL 1-B p.o., 3 mg/mL suzetrigine p.o.
  • FIG.4C 1 mg/mL 1-B p.o., 3 mg/mL pregabalin p.o. and their combination in SNI FIG.4D: 1 mg/mL 44-B p.o., 1 mg/mL morphine s.c. and their combination in SNI 12 Attorney Docket No.: 071741.11025/5WO1
  • FIG.4E 1 mg/mL 44-B p.o., 3 mg/mL pregabalin p.o. and their combination in SNI FIG.4F: 1 mg/mL 44-B p.o., 3 mg/mL suzetrigine p.o.
  • FIG.4G 1 mg/mL 44-B p.o., 7 mg/mL duloxetine p.o. and their combination in SNI
  • FIG.4H 1 mg/mL 44-B p.o., 7 mg/mL amitriptyline p.o. and their combination in SNI
  • FIG.4I 1 mg/mL 44-B p.o., 10 mg/mL ketorolac i.p. and their combination in SNI FIG.4J: 1 mg/mL 44-B p.o., 10 mg/mL ketorolac p.o.
  • FIG.4k 1 mg/mL 44-B p.o., 1 mg/mL morphine s.c. and their combination in BCP
  • FIG.4L 1 mg/mL 44-B p.o., 3 mg/mL pregabalin p.o. and their combination in BCP
  • FIG.4M 1 mg/mL 44-B p.o., 7 mg/mL amitriptyline p.o. and their combination in BCP
  • FIG.4N 1 mg/mL 44-B p.o., 7 mg/mL duloxetine p.o.
  • FIG.4O 1 mg/mL 44-B p.o., 10 mg/mL ketorolac i.p. and their combination in BCP
  • FIGs.5A-5C Body weight curve (FIG.5A), tumor volume growth curve (FIG.5B) ), and tumor volume in day 17 (FIG.5C) of mice in each group in subcutaneous colorectal cancer syngeneic model MC38, including group 1 (control group, 0mg/kg, p.o., QD*Day0-17), group 2 (clonidine, 5mg/kg, p.o., QD*Day0-3; 2mg/kg, p.o., QD* Day4-17), group 3 (compound 1-B HCl, 5mg/kg, p.o., BID*Day0-17), and group 4 (compound 1-B HCl, 10mg/kg, p.o., BID*Day0-3; 5mg/kg,
  • FIGs.6A-6C Body weight curve (FIG.6A), tumor volume growth curve (FIG.6B), and tumor volume in day 17 (FIG.6C) of mice in each group in subcutaneous colorectal cancer syngeneic model MC38, including group 1 (control group, 0mg/kg, p.o., QD*Day0-15), group 2 (Anti-mPD-1, 10mg/kg, i.p., BIW ⁇ 5 doses), group 3 (clonidine, 2mg/kg, p.o., QD*Day0-15), group 4 (44-B HCl, 2mg/kg, p.o., QD*Day0-15), group 5 (44-B HCl, 5mg/kg, p.o., QD*Day0-15), group 6 (clonidine, 2mg/kg, p.o., QD*Day0-15, and Anti-mPD-1, 10mg
  • the phrase “at least one of A, B, and C” means that only A is present, only B is present, only C is present, both A and B are present, both A and C are present, both B and C are present, or each of A, B, and C is present.
  • “at least one of” preceding a series of elements can also encompass situations in which any one or more of the elements is present in greater than one instance, e.g., “at least one of A, B, and C” can also encompass situations in which A is present in duplicate alone or further in combination with any one or more of elements B and C.
  • the conjunctive term “and/or” between multiple recited elements is understood as encompassing both individual and conjuntive options.
  • a first option refers to the applicability of the first element without the second.
  • a second option refers to the applicability of the second element without the 14 Attorney Docket No.: 071741.11025/5WO1 first.
  • a third option refers to, conjunctively,the applicability of the first and second elements together. Any one of these options is understood to fall within the meaning, and therefore satisfy the requirement of the term “and/or” as used herein.
  • any numerical value such as a concentration or a concentration range described herein, are to be understood as being modified in all instances by the term “about.”
  • a numerical value typically includes ⁇ 10% of the recited value.
  • the recitation of “10-fold” includes 9-fold and 11-fold.
  • the use of a numerical range expressly includes all possible permutations and combinations of subranges, all individual numerical values within that range, including integers within such ranges and fractions of the values unless the context clearly indicates otherwise.
  • subject means any animal, such as a mammal, particularly a human, to whom will be or has been treated by a method described herein.
  • mammal as used herein, encompasses any mammal. Examples of mammals include, but are not limited to, cows, horses, sheep, pigs, cats, dogs, mice, rats, rabbits, guinea pigs, and non-human primates (NHPs), such as monkeys or apes, humans, etc.
  • pharmaceutically acceptable salt(s) means those salts of a compound of interest that are safe and effective for topical use in mammals and that possess the desired biological activity.
  • Pharmaceutically acceptable salts include salts of acidic or basic groups present in the specified compounds.
  • Pharmaceutically acceptable acid addition salts include, but are not limited to, hydrochloride, hydrobromide, hydroiodide, nitrate, sulfate, bisulfate, phosphate, acid phosphate, isonicotinate, carbonate, bicarbonate, acetate, lactate, salicylate, citrate, tartrate, propionate, butyrate, pyruvate, oxalate, malonate, pantothenate, bitartrate, ascorbate, succinate, maleate, gentisinate, fumarate, gluconate, glucaronate, saccharate, formate, benzoate, glutamate, methanesulfonate, ethanesulfonate, benzensulfonate, p-toluenesulfonate and pamoate (i.e., 1,1′- methylene-bis-(2-hydroxy-3
  • Suitable base salts include, but are not limited to, aluminum, calcium, lithium, magnesium, potassium, sodium, zinc, bismuth, and diethanolamine salts.
  • Suitable base salts include, but are not limited to, aluminum, calcium, lithium, magnesium, potassium, sodium, zinc, bismuth, and diethanolamine salts.
  • alkyl means a saturated, monovalent, unbranched or branched hydrocarbon chain. An alkyl group can be unsubstituted or substituted with one or more suitable substituents.
  • alkyl groups include, but are not limited to, methyl (Me), ethyl (Et), propyl (e.g., n-propyl, isopropyl), butyl (e.g., n-butyl, isobutyl, tert-butyl), and pentyl (e.g., n- pentyl, isopentyl, neopentyl), etc.
  • An alkyl group can have a specified number of carbon atoms. When numbers appear in a subscript after the symbol “C”, the subscript defines with more specificity the number of carbon atoms which that particular alkyl can contain.
  • C 1 to C10 alkyl or “C1-10 alkyl” is intended to include alkyl groups having 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10 carbon atoms.
  • C 1 to C 8 alkyl or “C 1-8 alkyl” denotes an alkyl having 1, 2, 3, 4, 5, 6, 7, or 8 carbon atoms.
  • alkenyl refers to an unbranched or branched hydrocarbon chain containing at least one carbon-carbon double bond. An alkenyl group can be unsubstituted or substituted with one or more suitable substituents.
  • alkenyl groups include ethenyl, propenyl, butadienyl (including 1,2-butadienyl and 1,3-butadienyl).
  • C alkenyl
  • C 2-10 alkenyl is intended to include alkenyl groups having 2, 3, 4, 5, 6, 7, 8, 9, and 10 carbon atoms.
  • C2 to C8 alkenyl or “C2-8 alkenyl” denotes an alkenyl having 2, 3, 4, 5, 6, 7, or 8 carbon atoms.
  • alkynyl refers to an unbranched or branched hydrocarbon chain containing at least one carbon-carbon triple bond.
  • An alkynyl group can be unsubstituted or substituted with one or more suitable substituents.
  • alkynyl also includes those groups having one triple bond and one double bond. When numbers appear in a subscript after the symbol “C”, the subscript defines with more specificity the number of carbon atoms which that particular alkynyl can contain. For example, “C2 to C10 alkynyl” or “C2-10 alkynyl” is intended to include alkynyl groups having 2, 3, 4, 5, 6, 7, 8, 9, and 10 carbon atoms.
  • C 2 to C8 alkynyl or “C2-8 alkynyl” denotes an alkynyl having 2, 3, 4, 5, 6, 7, or 8 carbon atoms.
  • cycloalkyl refers to any stable monocyclic or polycyclic saturated hydrocarbon ring system.
  • a cycloalkyl group can be unsubstituted or substituted with one or more suitable substituents.
  • a cycloalkyl group can have a specified number of carbon atoms.
  • C3 to C6 cycloalkyl or “C3-6 cycloalkyl” includes cycloalkyl groups having 3, 4, 5, 16 Attorney Docket No.: 071741.11025/5WO1 or 6 ring carbon atoms, i.e., cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl.
  • Polycyclic cycloalkyls include bridged, fused, and spiro ring structures in which all ring atoms are carbon atoms.
  • a “spiro ring” is a polycyclic ring system in which two rings share one carbon atom, referred to as the “spiro atom,” which is typically a quaternary carbon atom.
  • a “fused ring” is a polycyclic ring system in which two rings share two adjacent atoms, referred to as “bridgehead atoms,” i.e., the two rings share one covalent bond such that the bridgehead atoms are directly connected.
  • a “bridged ring” is a polycyclic ring system in which two rings share three or more atoms separating the bridgehead atoms by a bridge containing at least one atom. Examples of cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, etc.
  • aryl as used herein is a group that contains any carbon-based aromatic group including, but not limited to, phenyl, naphthyl, anthracenyl, phenanthranyl, and the like.
  • Aryl moieties are well known and described, for example, in Lewis, R. J., ed., Hawley’s Condensed Chemical Dictionary, 13th Edition, John Wiley & Sons, Inc., New York (1997).
  • An aryl group can be substituted or unsubstituted with one or more suitable substituents.
  • An aryl group can comprise a single ring structure (i.e., monocyclic) or multiple ring structures (i.e., polycyclic, e.g., bicyclic or tricyclic).
  • an aryl group can be a monocyclic aryl group, e.g., phenyl.
  • heterocyclyl includes stable monocyclic and polycyclic hydrocarbons that contain at least one heteroatom ring member, such as sulfur, oxygen, or nitrogen, wherein the ring structure is saturated or partially unsaturated, provided the ring system is not fully aromatic.
  • a heterocyclyl group can be unsubstituted, or substituted with one or more suitable substituents at any one or more of the carbon atom(s) and/or nitrogen heteroatom(s) of the heterocyclyl.
  • a heterocyclyl can comprise a single ring structure (i.e., monocyclic) or multiple ring structures (i.e., polycyclic, e.g., bicyclic).
  • Polycyclic heterocyclyls include bridged, fused, and spiro ring structures in which at least one ring atom of at least one of the rings of the polycyclic ring system is a heteroatom, for instance oxygen, nitrogen, or sulfur, wherein bridged, fused, and spiro rings are as defined above.
  • a heterocyclyl ring can be attached to the parent molecule at any suitable heteroatom (typically nitrogen) or carbon atom of the ring.
  • the term “4- to 9-membered monocyclic or bicyclic heterocyclyl” includes any four, five, six, seven, eight, or nine membered monocyclic or bicyclic ring structure containing at least one heteroatom ring member selected from oxygen, nitrogen, and sulfur, or independently selected from oxygen and nitrogen, optionally 17 Attorney Docket No.: 071741.11025/5WO1 containing one to three additional heteroatoms independently selected from oxygen, nitrogen, and sulfur, or independently selected from oxygen and nitrogen, wherein the ring structure is saturated or partially unsaturated, provided the ring structure is not fully aromatic.
  • heterocyclyl refers to 4-, 5-, 6-, or 7-membered monocyclic groups and 6-, 7-, 8-, or 9- membered bicyclic groups which have at least one heteroatom (O, S, or N) in at least one of the rings, wherein the heteroatom-containing ring(s) typically has 1, 2, or 3 heteroatoms, such as 1 or 2 heteroatoms, independently selected from O, S, and/or N, or independently selected from O and N.
  • the subscript defines with more specificity the number of carbon atoms which that particular heterocycly can contain, in addition to the heteroatoms which that particular heterocycly can contain.
  • C1 to C10 heterocycl or “C1-10 heterocycl” is intended to include heterocycl groups having 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10 carbon atoms.
  • C 1 to C 8 heterocycly or “C 1-8 heterocycly” denotes a heterocycl having 1, 2, 3, 4, 5, 6, 7, or 8 carbon atoms.
  • Examples of monocyclic heterocyclyl groups include, but are not limited to azetidinyl, oxetanyl, tetrahydrofuranyl, pyrrolidinyl, imidazolidinyl, pyrazolidinyl, oxazolidinyl, isoxazolidinyl, thiazolidinyl, isothiazolidinyl, dioxolanyl, dithiolanyl, piperidinyl, piperazinyl, dioxanyl, morpholinyl, azepanyl, oxepanyl, oxazepanyl (e.g., 1,4-oxazepanyl, 1,2-oxazepanyl) and the like.
  • bicyclic heterocyclyl groups include, but are not limited to, 2-aza- bicyclo[2.2.1]heptanyl, 8-aza-bicyclo[3.2.1]octanyl, 2-aza-spiro[3.3]heptanyl, 3- azabicyclo[2.2.2]octanyl, 3-oxa-9-azabicyclo[3.3.1]nonanyl, 2-oxa-5-azabicyclo[2.2.1]heptanyl, 7- oxa-2-azaspiro[3.5]nonanyl, and 5-azaspiro[2.3]hexanyl and the like.
  • heteroaryl includes stable monocyclic and polycyclic aromatic hydrocarbons that contain at least one heteroatom ring member such as sulfur, oxygen, or nitrogen.
  • a heteroaryl group can be unsubstituted or substituted with one or more suitable substituents.
  • a heteroaryl can comprise a single ring structure (i.e., monocyclic) or multiple ring structures (i.e., polycyclic, e.g., bicyclic or tricyclic).
  • Each ring of a heteroaryl group containing a heteroatom can contain one or two oxygen or sulfur atoms and/or from one to four nitrogen atoms provided that the total number of heteroatoms in each ring is four or less and each ring has at least one carbon atom.
  • Heteroaryl groups which are polycyclic, e.g., bicyclic or tricyclic must include at least one fully aromatic ring, but the other fused ring or rings can be aromatic or non-aromatic.
  • the fused rings completing the bicyclic group can contain only carbon atoms and can be saturated, partially saturated, or unsaturated.
  • heteroaryl can be attached to the parent molecule at any available nitrogen or carbon atom of any ring of the heteroaryl group.
  • heteroaryl refers to 5- or 6-membered monocyclic groups and 9- or 10- membered bicyclic groups which have at least one heteroatom (O, S, or N) in at least one of the rings, wherein the heteroatom-containing ring typically has 1, 2, or 3 heteroatoms, such as 1 or 2 heteroatoms, selected from O, S, and/or N.
  • a heteroaryl group can be unsubstituted, or substituted with one or more suitable substituents at any one or more of the carbon atom(s) and/or nitrogen heteroatom(s) of the heteroaryl.
  • the nitrogen and sulfur heteroatom(s) of a heteroaryl can optionally be oxidized (i.e., N ⁇ O and S(O)r, wherein r is 0, 1 or 2).
  • N ⁇ O and S(O)r wherein r is 0, 1 or 2.
  • the subscript defines with more specificity the number of carbon atoms which that particular heteroaryl can contain, in addition to the heteroatoms which that particular heteraryl can contain.
  • “C 1 to C 10 heteroaryl” or “C1-10 heteroaryl” is intended to include heteroaryl groups having 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10 carbon atoms.
  • C1 to C8 heteroaryl or “C1-8 heteroaryl” denotes a heteroaryl having 1, 2, 3, 4, 5, 6, 7, or 8 carbon atoms.
  • Exemplary monocyclic heteroaryl groups include, but are not limited to, pyrrolyl, pyrazolyl, pyrazolinyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, thiadiazolyl, isothiazolyl, furanyl, thiophenyl, oxadiazolyl, pyridinyl, pyrazinyl, pyrimidinyl, pyridazinyl, and triazinyl.
  • Exemplary bicyclic heteroaryl groups include, but are not limited to, indolyl, benzothiazolyl, benzodioxolyl, benzoxazolyl, benzothienyl, quinolinyl, tetrahydroisoquinolinyl, isoquinolinyl, benzimidazolyl, benzopyranyl, indolizinyl, benzofuranyl, chromonyl, coumarinyl, benzopyranyl, cinnolinyl, quinoxalinyl, indazolyl, pyrrolopyridinyl, furopyridinyl, dihydroisoindolyl, and tetrahydroquinolinyl.
  • alkoxy refers to an –O-alkyl group, wherein alkyl is as defined above.
  • An alkoxy group is attached to the parent molecule through a bond to an oxygen atom.
  • An alkoxy group can have a specified number of carbon atoms.
  • C1 to C10 alkoxy or “C1-10 alkoxy” is intended to include alkoxy groups having 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10 carbon atoms.
  • C 1 to C 4 alkoxy or “C 1-4 alkoxy” denotes an alkoxy having 1, 2, 3, or 4 carbon atoms.
  • alkoxy examples include, but are not limited to, methoxy, ethoxy, propoxy (e.g., n-propoxy, isopropoxy), butoxy (e.g., n-butoxy, isobutoxy, tert-butoxy), pentyloxy 19 Attorney Docket No.: 071741.11025/5WO1 (e.g., n-pentyloxy, isopentyloxy, neopentyloxy), etc.
  • An alkoxy group can be unsubstituted or substituted with one or more suitable substituents.
  • alkylthio or “thioalkoxy” represents an alkyl group as defined above attached to the parent molecule through a bond to a sulfur atom, for example, -S-methyl, -S-ethyl, etc.
  • Representative examples of alkylthio include, but are not limited to, -SCH3, -SCH2CH3, etc.
  • halogen means fluorine, chlorine, bromine, or iodine.
  • halo means fluoro, chloro, bromo, and iodo.
  • Haloalkyl is intended to include both branched and straight-chain saturated aliphatic hydrocarbon radicals substituted with one or more halogen atoms.
  • haloalkyl examples include, but are not limited to, fluoromethyl, difluoromethyl, trifluoromethyl, chloromethyl, dichloromethyl, trichloromethyl, pentafluoroethyl, pentachloroethyl, 2,2,2- trifluoroethyl, heptafluoropropyl, and heptachloropropyl.
  • fluoroalkyl in particular include, but are not limited to, -CF3, -CHF2, -CH2CF3, -CF2CF3, and the like.
  • alkylamino groups have one or both hydrogen atoms of an amino group replaced with an alkyl group and is attached to the parent molecule through a bond to the nitrogen atom of the alkylamino group.
  • alkylamino includes methylamino (-NHCH3), dimethylamino (-N(CH3)2), -NHCH2CH3 and the like.
  • aminoalkyl as used herein is intended to include both branched and straight- chain saturated aliphatic hydrocarbon groups substituted with one or more amino groups.
  • C1-4 aminoalkyl is intended to include alkyl groups having 1, 2, 3, or 4 carbon atoms 20 Attorney Docket No.: 071741.11025/5WO1 substituted with one or more amino groups.
  • Aminoalkyl groups are attached to the parent molecule through a bond to a carbon atom of the alkyl moiety of the aminoalkyl group.
  • Representative examples of aminoalkyl groups include, but are not limited to, -CH 2 NH 2 , -CH 2 CH 2 NH 2 , and – CH2CH(NH2)CH3.
  • amido refers to –C(O)N(R)2, wherein each R is independently an alkyl group (including both branched and straight-chain alkyl groups) or a hydrogen atom.
  • amido groups include, but are not limited to, -C(O)NH 2 , -C(O)NHCH 3 , and –C(O)N(CH 3 ) 2 .
  • hydroxyl-substituted alkyl “hydroxylalkyl” and “hydroxyalkyl” are used interchangeably, and refer to a branched or straight-chain aliphatic hydrocarbon group substituted with one or more hydroxyl groups.
  • Hydroxyalkyl groups are attached to the parent molecule through a bond to a carbon atom of the alkyl moiety of the hydroxyalkyl group.
  • a hydroxyalkyl group can have a specified number of carbon atoms.
  • “C1 to C10 hydroxyalkyl” or “C1- 10 hydroxyalkyl” is intended to include hydroxyalkyl groups having 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10 carbon atoms.
  • C1 to C4 hydroxylalkyl” or “C1-4 hydroxyalkyl” denotes a hydroxyalkyl group having 1, 2, 3, or 4 carbon atoms.
  • hydroxyalkyl examples include, but are not limited to, hydroxylmethyl (-CH 2 OH), hydroxylethyl (-CH 2 CH 2 OH), etc.
  • amide refers to –N(R’)C(O)R, wherein each R and R’ is independently chosen from hydrogen, alkyl, cycloalkyl, aryl, and heteroaryl.
  • amide groups include, but are not limited to, -NHC(O)CH 3 , -NHC(O)CH 2 CH 3 , and – N(CH 3 )C(O)CH 3 .
  • carbamide refers to –N(R’)C(O)N(R) 2 , wherein each R and R’ is independently chosen from hydrogen, alkyl, cycloalkyl, aryl, and heteroaryl.
  • carbamide groups include, but are not limited to, -NHC(O)NH 2 , -NHC(O)NHCH 3 (methyl carbamide), and –NHC(O)NH(Ph).
  • sulfonamide refers to –N(R’)SO2-R, wherein each R and R’ is independently chosen from hydrogen, alkyl, cycloalkyl, aryl, and heteroaryl.
  • sulfonamide groups include, but are not limited to, -NHSO 2 CH 3 (methyl sulfonamide), and –NH SO2Ph.
  • sulfonamide groups include, but are not limited to, -NHSO 2 CH 3 (methyl sulfonamide), and –NH SO2Ph.
  • substituted as used herein with respect to any organic radical (e.g., alkyl, cycloalkyl, heteroaryl, aryl, heterocyclyl, etc.) means that at least one hydrogen atom is replaced with a non-hydrogen group, provided that all normal valencies are maintained and that the substitution results in a stable compound.
  • that group can have one or more substituents, such as from one to five substituents, one to three substituents, or one to two substituents, independently selected from the list of substituents.
  • substituents independently when used in reference to substituents, means that when more than one of such substituents is possible, such substituents can be the same or different from each other.
  • suitable substituents include, but are not limited to, alkyl, halo, haloalkyl, alkoxy, amido, hydroxy, hydroxyalkyl, amino, carboxyl, ester, oxo, cyano and the like.
  • a group is shown to be substituted with 0-3 R groups, then said group can be optionally substituted with up to three R groups, and at each occurrence, R is selected independently from the definition of R.
  • R is selected independently from the definition of R.
  • the terms “optional” or “optionally” mean that the event or circumstance described can, but need not, occur, and such a description includes the situation in which the event or circumstance does or does not occur.
  • “optionally substituted heterocyclyl” means that a substituent group can be, but need not be, present, and such a description includes the situation of the heterocyclyl group being substituted by a suitable substituent and the heterocyclyl group not being substituted by any substituent.
  • Stereoisomers includes enantiomers and diastereomers. Enantiomers are stereoisomers that are non-super-imposable mirror images of each other. A 1:1 mixture of a pair of enantiomers is a racemate or racemic mixture.
  • Diastereomers are stereoisomers that are not 22 Attorney Docket No.: 071741.11025/5WO1 enantiomers, i.e., they are not related as mirror images, and occur when two or more stereoisomers of a compound have different configurations at one or more of the equivalent stereocenters and are not mirror images of each other.
  • Substituent groups e.g., alkyl, heterocyclyl, etc.
  • Certain examples contain chemical structures that comprise (R) or (S) terminology.
  • Stereochemically pure isomeric forms can be obtained by techniques known in the art in view of the present disclosure.
  • diastereoisomers can be separated by physical separation methods such as fractional crystallization and chromatographic techniques, and enantiomers can be separated from each other by the selective crystallization of the diastereomeric salts with optically active acids or bases or by chiral chromatography.
  • Pure stereoisomers can also be prepared synthetically from appropriate stereochemically pure starting materials, or by using stereoselective reactions.
  • tautomer refers to compounds that are interchangeable forms of a particular compound structure and that vary in the displacement of hydrogen atoms and electrons. Tautomers are constitutional isomers of chemical compounds that readily interconvert, usually resulting in relocation of a proton (hydrogen). Thus, two structures can be in equilibrium through the movement of pi electrons and an atom (usually hydrogen). All tautomeric forms and mixtures of tautomers of the compounds described herein are included with the scope of the present disclosure. [0095] Compounds described herein can exist in solvated and unsolvated forms.
  • solvate means a physical association, e.g., by hydrogen bonding, of a compound described herein with one or more solvent molecules.
  • the solvent molecules in the solvate can be present in a regular arrangement and/or a non-ordered arrangement.
  • the solvate can comprise either a stoichiometric or nonstoichiometric amount of the solvent molecules.
  • “Solvate” encompasses both solution-phase and isolable solvates.
  • Compounds described herein can form solvates with water (i.e., hydrates) or common organic solvents.
  • Exemplary solvates include, but are not limited to, hydrates, ethanolates, methanolates, and isopropanolates.
  • isotopes of atoms occurring in the compounds described herein, including intermediates and final products.
  • isotopes include those atoms having the same atomic number but different mass numbers.
  • isotopes of hydrogen include deuterium and tritium.
  • isotopes of carbon include 13 C and 14 C.
  • the present disclosure further includes isotopically-labeled compounds.
  • isotopically- labeled or “radio-labeled” compound is a compound of the present disclosure where one or more atoms are replaced or substituted by an atom having an atomic mass or mass number different from the atomic mass or mass number typically found in nature (i.e., naturally occurring).
  • isotopically- labeled compounds can generally be prepared by conventional techniques known to those skilled in the art or by processes analogous to those described herein, using an appropriate isotopically- labeled reagent in place of the non-labeled reagent otherwise employed.
  • the name of a compound is intended to encompass all possible existing isomeric forms, including stereoisomers (e.g., enantiomers, diastereomers, racemate or racemic mixture, and any mixture thereof) of the compound.
  • the present disclosure relates to methods of treating or preventing a disease in a subject in need thereof, the method comprising administering to the subject an effective amount of a peripherally selective ⁇ 2AR agonist.
  • the peripherally selective ⁇ 2AR agonist activates at least one sub type of ⁇ 2AR, particularly ⁇ 2A AR, ⁇ 2B AR, or ⁇ 2C AR.
  • the disease is glaucoma, pain, spasticity, nasal congestion, rosacea, rhinitis, anesthesia, presbyopia, acute kidney injury, insomnia, inflammatory disease, cancer, etc.
  • the disease is chosen from pain, rosacea, spasticity, and aging.
  • treating with the peripherally selective ⁇ 2AR agonist causes less sedation than treating with a non-peripherally selective ⁇ 2AR agonist, such as at similar or comparable dosage.
  • the peripherally selective ⁇ 2AR agonist comprises an ⁇ 2AR activation moiety covalently linked to a peripheral distribution moiety.
  • the present disclosure provides a peripherally selective ⁇ 2AR agonist that comprises an ⁇ 2AR activation moiety covalently linked to a peripheral distribution moiety, and its uses in the treatment or prevention of a disease.
  • a disease is glaucoma, pain, spasticity, nasal congestion, rosacea, rhinitis, anesthesia, presbyopia, acute kidney injury, insomnia, inflammatory disease, cancer, etc.
  • the disease is pain.
  • Pain Pain, as a complex and multidimensional sensory and emotional experience, poses a significant challenge to human health.
  • neuropathic pain is caused by a lesion or disease of the somatosensory nervous system. Neuropathic pain can be divided into central neuropathic pain and peripheral neuropathic pain. Central neuropathic pain includes spinal cord injury, post-stroke pain, and MS pain, while peripheral neuropathic pain includes diabetic neuropathy, postherpetic neuralgia, HIV-associated pain, chemotherapy-induced peripheral neuropathy, and post-surgical neuropathic pain.
  • first-line treatment drugs include Gabapentinoids, tricyclic antidepressants, and noradrenaline/serotonin uptake inhibitors. Although these drugs can relieve pain to some extent, the side effects of long-term use still cause a decrease in the quality of life of patients.
  • Second-line treatment drugs such as opiate receptor agonist, not only have side effects but also have a high addiction rate, which has caused many social impacts and cannot well address the demand for neuropathic pain drugs.
  • ⁇ 2AR agonists such as clonidine and dexmedetomidine, are considered an important method for treating pain in academic research and clinical applications. Scientists have found that the intraspinal administration of ⁇ 2AR agonists can effectively relieve pain.
  • ⁇ 2AR agonists are not without limitations.
  • Existing ⁇ 2AR agonists are often associated with a range of biological reactions, including sedation, hypotension, bradycardia, drowsiness, dizziness, depression, bradycardia, orthostatic hypotension, constipation, nausea, gastric upset, dry mouth (xerostomia), dry nasal mucosa, impotence, fluid retention, edema, and pupil size.
  • These other biological effects, especially sedation set limits on the dosages that can be safely administered, thereby constraining the wide-scale utility of these drugs in long-term pain management. This not only affects the quality of life for patients but also restricts the applicability of these drugs for various types and levels of pain symptoms.
  • an effective amount means an amount of a composition or compound that elicits a biological or medicinal response in a tissue system or subject that is being sought by a researcher, veterinarian, medical doctor or other professional, which can include alleviation of the symptoms of the disease, disorder, or condition being treated.
  • An effective amount can vary depending upon a variety of factors, such as the physical condition of the subject, age, weight, health, etc.; and the particular disease, disorder, or condition to be treated. An effective amount can readily be determined by one of ordinary skill in the art in view of the present disclosure.
  • an effective amount refers to the amount of a composition or compound described herein which is sufficient to activate ⁇ 2AR.
  • an effective amount refers to the amount of a composition or compound described herein which is sufficient to treat or prevent the disease or alleviate the symptoms associated with the disease.
  • the pain is neuropathic pain, nociceptive pain, nociplastic pain, or mixed pain.
  • the term “neuropathic pain” refers to pain caused by a lesion or disease affecting the somatosensory nervous system. It results from abnormal processing of pain signals due to nerve damage or dysfunction, either in the peripheral nervous system (e.g., peripheral neuropathy) or the central nervous system (e.g., spinal cord injury, multiple sclerosis).
  • nociceptive pain refers to pain that arises from actual or threatened damage to non-neural tissue and is due to the activation of nociceptors (pain-sensing nerve fibers). It is the body’s natural response to harmful stimuli such as injury, inflammation, or mechanical stress.
  • nociplastic pain refers to pain that arises from altered nociception.
  • mixed pain refers to a mixture of nociceptive, neuropathic, and/or nociplastic pain.
  • the neuropathic pain is chosen from diabetic peripheral neuropathy, postherpetic neuralgia, chemotherapy-induced peripheral neuropathy, Fabry disease– associated pain, trigeminal neuralgia, glossopharyngeal neuralgia, occipital neuralgia, HIV- 26 Attorney Docket No.: 071741.11025/5WO1 associated neuropathy, hereditary sensory and autonomic neuropathy, Charcot-Marie-Tooth disease–related pain, small fiber neuropathy, brachial plexus avulsion, phantom limb pain, post- surgical neuropathic pain, carpal tunnel syndrome, other compression or entrapment neuropathies, amyloid neuropathy, vasculitic neuropathy, leprosy-associated pain, radiation-induced neuropathy, drug-induced neuropathy, metabolic neuropathies, endocrine-related neuropathies, toxic neuropathies, immune-mediated neuropathies, traumatic peripheral nerve injury, and heritable neuropathies, as well as post-stroke pain syndrome
  • the nociceptive pain is somatic (i.e., from skin, muscles, or joints (e.g., cuts, fractures)) or visceral (i.e., from internal organs (e.g., appendicitis)).
  • the nociceptive pain is osteoarthritis-related rheumatoid pain, arthritis-associated post-traumatic pain, musculoskeletal pain, postoperative pain, tendonitis, myofascial pain, visceral inflammatory pain (e.g., appendicitis, cystitis, pancreatitis), dysmenorrhea, or inflammatory bowel disease-related pain.
  • the mixed pain is cancer-related pain (e.g., bone metastasis pain, tumor infiltration pain, perineural invasion), complex regional pain syndrome type I, failed back surgery syndrome, low back pain with radiculopathy endometriosis-associated pelvic pain, chronic post-surgical pain, chronic pancreatitis-associated pain, interstitial cystitis/bladder pain syndrome, fibromyalgia, temporomandibular disorder, vulvodynia, irritable bowel syndrome with visceral pain, and combinations thereof.
  • the pain is acute pain or chronic pain.
  • the term “chronic pain” refers to pain that is persistent or recurrent pain lasting longer than 3 months.
  • the chronic pain is chronic primary pain, chronic cancer pain, chronic postsurgical and posttraumatic pain, chronic neuropathic pain, chronic headache and orofacial pain, chronic visceral pain, chronic musculoskeletal pain.
  • the pain is cancer pain.
  • the pain is post-surgery pain. 27 Attorney Docket No.: 071741.11025/5WO1 [0126]
  • the peripherally selective ⁇ 2AR agonist comprises an ⁇ 2AR activation moiety covalently linked to a peripheral distribution moiety.
  • the present disclosure relates to methods of treating or preventing a disease in a subject in need thereof, the method comprising administering to the subject an effective amount of a peripherally selective ⁇ 2AR agonist, wherein the peripherally selective ⁇ 2AR agonist comprises an ⁇ 2AR activation moiety covalently linked to a peripheral distribution moiety.
  • the disease is glaucoma, pain, spasticity, nasal congestion, rosacea, rhinitis, anesthesia, presbyopia, acute kidney injury, insomnia, inflammatory disease, cancer, etc.
  • the disease is glaucoma or cancer.
  • the disease is pain.
  • the pain is neuropathic pain, nociceptive pain, nociplastic pain, or mixed pain.
  • the neuropathic pain is chosen from diabetic peripheral neuropathy, postherpetic neuralgia, chemotherapy-induced peripheral neuropathy, Fabry disease– associated pain, trigeminal neuralgia, glossopharyngeal neuralgia, occipital neuralgia, HIV- associated neuropathy, hereditary sensory and autonomic neuropathy, Charcot-Marie-Tooth disease–related pain, small fiber neuropathy, brachial plexus avulsion, phantom limb pain, post- surgical neuropathic pain, carpal tunnel syndrome, other compression or entrapment neuropathies, amyloid neuropathy, vasculitic neuropathy, leprosy-associated pain, radiation-induced neuropathy, drug-induced neuropathy, metabolic neuropathies, endocrine-related neuropathies, toxic neuropathies, immune-mediated neuropathies, traumatic peripheral nerve
  • the nociceptive pain is somatic (i.e., from skin, muscles, or joints (e.g., cuts, fractures)) or visceral (i.e., from internal organs (e.g., appendicitis)).
  • the nociceptive pain is osteoarthritis-related rheumatoid pain, arthritis-associated post-traumatic pain, musculoskeletal pain, postoperative pain, tendonitis, 28 Attorney Docket No.: 071741.11025/5WO1 myofascial pain, visceral inflammatory pain (e.g., appendicitis, cystitis, pancreatitis), dysmenorrhea, or inflammatory bowel disease-related pain.
  • the mixed pain is cancer-related pain (e.g., bone metastasis pain, tumor infiltration pain, perineural invasion), complex regional pain syndrome type I, failed back surgery syndrome, low back pain with radiculopathy endometriosis-associated pelvic pain, chronic post-surgical pain, chronic pancreatitis-associated pain, interstitial cystitis/bladder pain syndrome, fibromyalgia, temporomandibular disorder, vulvodynia, irritable bowel syndrome with visceral pain, and combinations thereof.
  • the pain is acute pain or chronic pain.
  • the term “chronic pain” refers to pain that is persistent or recurrent pain lasting longer than 3 months.
  • the chronic pain is chronic primary pain, chronic cancer pain, chronic postsurgical and posttraumatic pain, chronic neuropathic pain, chronic headache and orofacial pain, chronic visceral pain, chronic musculoskeletal pain.
  • the pain is cancer pain.
  • the pain is post-surgery pain.
  • treating with the peripherally selective ⁇ 2AR agonist causes less side effects than treating with a non-peripherally selective ⁇ 2AR agonist, such as at similar or comparable dosage.
  • treating with the peripherally selective ⁇ 2AR agonist causes no side effects.
  • the side effect is sedation, decreasing heart rate, and decreasing blood pressure, particularly the side effect is sedation.
  • a non-peripherally selective ⁇ 2AR agonist refers to a compound can be readily distributed into the CNS after being administered into a subject, binds to and activates ⁇ 2AR receptor in both the central nervous system (brain and spinal cord) and the peripheral nervous system. Examples of non-peripherally selective ⁇ 2AR agonists include, but not limited to, dexmedetomidine, and clonidine.
  • an ⁇ 2AR agonist binds to and activates ⁇ 2AR in the central nervous system, it can produce the above mentioned side effects in patients, such as sedation, decreased heart rate, blood pressure, depression, bradycardia, orthostatic hypotension, 29 Attorney Docket No.: 071741.11025/5WO1 constipation, nausea, gastric upset, dry mouth (xerostomia), dry nasal mucosa, impotence, fluid retention, edema, and pupil size.
  • a peripherally selective ⁇ 2AR agonist refers to a compound that primarily exerts its effects outside of the central nervous system (CNS), typically because it is impeded by the blood-CNS barrier.
  • Blood-CNS barrier the physical barrier between blood and the CNS, safeguards the CNS from both toxic and pathogenic agents in the blood.
  • the blood-CNS barrier comprises the blood-brain barrier, the blood-spinal cord barrier, and the blood-CSF (cerebrospinal fluid) barrier.
  • a compound may act on the rest of the body with less or no side-effects related to their effects on the brain or spinal cord.
  • peripherally selective ⁇ 2AR agonists include, but not limited to, the compounds described herein, such as compounds of formula (I-A), (I-B), (I-C), (I-D), or (II), described herein.
  • the peripherally selective ⁇ 2AR agonist primarily binds to or activates ⁇ 2AR outside CNS, thus herby producing less or no foregoing side effects, compared to the non-peripherally selective ⁇ 2AR agonists.
  • the present invention satisfies an unmet need, and has developed a series of peripherally selective ⁇ 2AR agonists.
  • the peripherally selective ⁇ 2AR agonist binds to ⁇ 2AR with a Ki ranging from 250nM to1000nM, 50nM to 250nM, 10nM to 50nM, or less than 10nM.
  • the peripherally selective ⁇ 2AR agonist activates ⁇ 2AR with an EC50 ranging from 250nM to1000nM, 50nM to 250nM, 10nM to 50nM, or less than 10nM.
  • the non-peripherally selective ⁇ 2AR agonist and the peripherally selective ⁇ 2AR agonist can be differentiated in terms of blood-brain barrier (BBB) permeability. Drugs that specifically target the central nervous system (CNS) must first traverse the BBB. In contrast, peripherally selective drugs primarily exert their effects outside of CNS, largely because they are impeded by the blood-brain barrier (BBB).
  • BBB blood-brain barrier
  • BBB blood-brain barrier
  • CNS central nervous system
  • Kp the concentration of drug in the brain divided by concentration in the blood.
  • Kp is often calculated as “logBB”, which refers to the logarithmic ratio of the concentration of a compound in the brain and in the blood.
  • Kp is a common 30 Attorney Docket No.: 071741.11025/5WO1 numeric value for describing permeability across the blood-brain barrier.
  • a compound is considered “peripherally selective” if, upon administration to a subject, its Kp is lower than 0.4, 0.2, 0.1, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02, or 0.01.
  • Kp,uu,brain is another common numeric value for describing permeability across the blood-brain barrier.
  • Kp,uu,brain or “Kp,uu”, refers to the unbound brain-to- plasma partition coefficient.
  • Kp,uu provides a more accurate measure of distribution equilibrium between unbound fractions in brain and plasma.
  • Any methods known in the field can be used to measure Kp,uu,brain.
  • AUC Area Under the Curve
  • Another example is Steady-State Concentrations, which uses the steady-state unbound concentrations of the drug in brain interstitial fluid (C_u,brain,ss) and in plasma (C_u,plasma,ss).
  • a compound is considered “peripherally selective” if, upon administration to a subject, its Kp,uu,brain is lower than 0.4, 0.2, 0.1, 0.09, 0.08, 0.07, 0.06, 0.05, 0.04, 0.03, 0.02, or 0.01. In some further embodiments, a compound is considered “peripherally selective” if, upon administration to a subject, its Kp,uu,brain is lower than 0.05, 0.04, 0.03, 0.02, or 0.01. [0154] In some embodiments, the peripherally selective ⁇ 2AR agonist comprises an ⁇ 2AR activation moiety that is covalently linked to a peripheral distribution moiety.
  • the ⁇ 2AR activation moiety is a non-peripherally selective ⁇ 2AR agonist or another peripherally selective ⁇ 2AR agonist.
  • the ⁇ 2AR activation moiety is an ⁇ 2AR agonist that is chosen from (R)-3-nitrobiphenyline, A-193080, ADX-415, AGN 192836, AGN-191103, AGN-197075, AGN-201781, AGN-241622, amitraz, Apraclonidine, AR-08, Bethanidine, Brimonidine, BRL- 48962, Bromocriptine, Cirazoline, Clonidine, Detomidine, Detomidine carboxylic acid, Dexmedetomidine, Dipivefrin, DL-Methylephedrine, Droxidopa, Epinephrine, ergotamine, etilefrine, Etomidate, Fadolmidine, Guanabenz, Guanethidine, Gu
  • the functional derivative of an ⁇ 2AR agonist refers to any compound that is derived from the ⁇ 2AR agonist by a chemical reaction.
  • the derivatives include, but not limited to, acid or base salts, prodrugs, compounds containing protected functional groups such as hydroxyl, amino, carboxyl and carbonyl groups.
  • the ⁇ 2AR activation moiety is a non-peripherally selective ⁇ 2AR agonist, such as dexmedetomidine, brimonidine, and clonidine.
  • the ⁇ 2AR activation moiety is dexmedetomidine.
  • the term “a peripheral distribution moiety” refers to a moiety that can increase or improve the peripheral selectivity of an ⁇ 2AR agonist.
  • the peripheral selectivity is increased or improved so that the ⁇ 2AR agonist is a peripherally selective ⁇ 2AR agonist.
  • the peripheral distribution moiety can be the following chemical fragments: • type A fragments; those that can increase the overall molecular polarity of the compound or reduce the overall lipophilicity of the compound; • type B fragments: those that can increase the overall molecular weight or the molecular size of the compound; and • type C fragments: those that comprises a substrate element of an efflux transporter.
  • the peripheral distribution moiety is a type A fragment.
  • the type A fragment increases the total number of intermolecular hydrogen bond (H-bond) within the compound, such as H-bond donors and H-bond acceptors.
  • the type A fragment is a H-bond donor.
  • the type A fragment increases the overall molecular polarity of the compound.
  • such type A fragments can comprise a polar functional group or a charged group.
  • Examples of the polar functional group include, but not limited to, hydroxyl, amine, amide, sulfonamide, carboxyl, ether, imine, hydroxylamine, ester, aldehyde, ketone, nitro, 32 Attorney Docket No.: 071741.11025/5WO1 phosphate, thioether, and sulfone groups.
  • Examples of the charged group include, but not limited to, quaternary ammonium and organic acids such as carboxylic acids and sulfonic acids.
  • the type A fragment reduces the overall lipophilicity of the compound.
  • type A fragments include, but not limited to, alkyl or acyl that is added to a function group such as hydroxyl and amino.
  • the type A fragment is not tertiary amine or one that can help form an intramolecular H-bond.
  • the peripheral distribution moiety is a type B fragment.
  • the type B fragment is a bulky group, which can increase the overall molecular weight and the molecular size of the compound. Examples of such type B fragments include, but not limited to, long alkyl chains, polyethylene glycol (PEG), large aromatic groups, and extra cyclic or heterocyclic groups.
  • the peripheral distribution moiety is a type C fragment.
  • the type C fragment comprises a substrate element of an efflux transporter, wherein the efflux transporter is P-glycoprotein (P-gp) transporter, breast cancer resistance protein (BCRP) transporter, or multidrug resistance protein 2 (MRP2) transporter.
  • P-gp P-glycoprotein
  • BCRP breast cancer resistance protein
  • MRP2 multidrug resistance protein 2
  • a substrate element of an efflux transporter refers to a fragment that makes the compound to become a substrate of the efflux transporter.
  • the term of “a substrate element of an efflux transporter” refers to a fragment of a substrate of the efflux transporter.
  • the type C fragment comprises a substrate element of P-gp.
  • P-gp efflux is a significant limitation to BBB permeation. Any methods known in the filed can be used to determine whether a compound is a P-gp substrate. For example, the efflux ratio obtained from in vitro P-gp assay, MDCK-MDR1, can be used to identify the substrate of P-gp. A compound is considered as a P-gp substrate if its efflux ratio is greater than 2, 5, 8, 10, 50, or 100.
  • the substrate element for P-gp contains one or more of the structural modifications described above.
  • the substrate element for P-gp is chosen from: O H N O H N O O H O O NH N P S F 3 .
  • the type C fragment comprises a substrate element of MPR2 transporter.
  • the type C fragment does not comprise a substrate element of uptake transporter, such as LAT1, GLUT1, MCT1, CAT1, CNT2, OATP, PEPT1, PEPT2, and OCT.
  • a substrate element of uptake transporter such as LAT1, GLUT1, MCT1, CAT1, CNT2, OATP, PEPT1, PEPT2, and OCT.
  • the peripheral distribution moiety reduces and/or minimizes brain exposure to a peripherally selective ⁇ 2AR agonist.
  • the peripheral distribution moiety decreases passive transcellular BBB permeability by increasing topological polar surface area (TPSA), increasing molecule weight, increasing polarity, or adding hydrogen binding, especially hydrogen bond donor.
  • TPSA topological polar surface area
  • the peripheral distribution moiety introduces an acidic group to the peripherally selective ⁇ 2AR agonist.
  • the peripheral distribution moiety comprises a substrate element for P-gp, wherein the substrate element for P-gp increases P-gp efflux by increasing lipophilicity, increasing hydrogen bond acceptors, removing steric hindrance around hydrogen bind acceptors, or removing electron-withdrawing group adjacent to hydrogen bond acceptor.
  • the peripheral distribution moiety makes a compound to become a dual substrate for both P-gp and BCRP.
  • R L is a moiety that covalently connects two functional groups or moieties within a single molecule.
  • One end of RL is connected to RP and the other end of RL is connceted to Y.
  • RL can be any moiety that serves the linking function, such as the linkers used in proteolysis targeting chimeras (PROTACs) and non-cleavable linkers used in antibody-drug conjugates (ADCs).
  • R L include, but are not limited to, polyethylene glycol (PEG) and alkyl chains of varying lengths, glycols, alkynes, triazoles, saturated heterocycles such as piperazine and piperidine, thioethers, maleimidocaproyl linker.
  • R L is chosen from alkyl, polyethylene glycol, other glycol, cycloalkyl, heterocycle, aryl, and heteroaryl; wherein the cycloalky, heterocycle, aryl, or heteroaryl is optionaly substituted with at least one substituent chosen from halogen, hydroxyl, alkyl, haloalkyl, alkoxy and hydroxyalkyl.
  • RL is one selected from the followings: Z Y N m2 Y Y m2 m2 H , m , ag e s, e ag e s ca e co ec e a y o e .
  • Rp can be the following chemical moieties: • those that can increase the overall molecular weight of the compound, such as bulky functional groups and additional molecular structures, including long alkyl chains, large aromatic groups, and extra cyclic structures like cyclohexane or cyclopentane rings; • those that can increase the overall molecular polarity of the compound, such as hydroxyl, amine, amide, sulfonamide, ether, imine, hydroxylamine, ester, aldehyde, ketone, nitro, phosphate, thioether; and • those that can bring charge to the compound, such as the functional groups that ionize at physiological pH, including carboxylic acid, quaternary ammonium and quaternary phosphonium.
  • RP when RP is not substituted with RC, RP is: O H N O H N O O O H N N . 39 Attorney Docket No.: 071741.11025/5WO1 [0190] In some embodiments, when RP is substituted with RC, RP is: O H O N O H N O O H O O NH N P S S O F 3 . [0191] As u end of RP.
  • RC is chosen from -C0-12 alkylene-C3-12 cycloalkyl, -C0-12 alkylene- C 2-12 heterocyclyl, -C 0-12 alkylene-C 1-12 heteroaryl, -NH-C 0-12 alkylene-C 3-12 cycloalkyl, -NH-C 0-12 alkylene-C2-12 heterocyclyl, -NH-C0-12 alkylene-C1-12 heteroaryl, -O-C0-12 alkylene-C3-12 cycloalkyl, -O-C0-12 alkylene-C2-12 heterocyclyl, -O-C0-12 alkylene-C1-12 heteroaryl, and alkyl substituted with trialkylammonium, wherein each of the C 3-12 cycloalkyl, C 2-12 heterocyclyl, and C 1-12 heteroaryl is optionally substituted with one or more substituents chosen from hydroxy, alkyl, oxo,
  • RC is: Z Z s s Z s Z s N H .
  • Attorney Docket No.: 071741.11025/5WO1 [0194]
  • RC is: H NH N NH NH O O O O
  • Attorney Docket No.: 071741.11025/5WO1 O NH H O O N N O O S
  • the present disclosure relates to a compound of formula (I-D): N NH T , or a stereoisomer, tautomer, pharmaceutically acceptable salt or solvate thereof, wherein, Y 1 is CH, or N; X 1 is chosen from H, D, and halogen; and R T is defined as above in formula (I-A).
  • the compound of formula (I-A) has the formula (I-A-1): R T m , or a stereoisomer, tautomer, pharmaceutically acceptable salt or solvate thereof, wherein, R 1 is chosen from H, D, and halogen; 43 Attorney Docket No.: 071741.11025/5WO1 A is a ring chosen from phenyl, pyridinyl, thienyl, furyl, pyrrolyl, 4H-pyran, or 4H- thiopyran; R 2 , n, R 3 , m, and R T are defined as above in formula (I-A).
  • the compound of formula (I-A) has the formula (I-A-2): R T , or a stereoisomer, tautomer, pharmaceutically acceptable salt or solvate thereof, wherein, A is a ring chosen from phenyl, 1,2,3,4-tetrahydro-1-naphthyl, quinoxalinyl, pyrimidinyl, and 2,1,3-benzothiadiazol; Y is CH, N, -O-CH-, or -C-NH-; when Y is -O-C-, the oxygen atom is connected to A, and the carbon atom is connected X to both ; when arbon atom is connected to both R T and A, and the nitrogen X atom is connecte ; X is NH, O, or S; and R 2 , n, and RT are defined as above in formula (I-A).
  • the present disclosure relates to a compound of formula (II): R T B , or a stereoisomer, tautomer, pharmaceutically acceptable salt or solvate thereof, wherein, A is one chosen from: S 1 ; each R 1 is independently chosen from hydrogen, halogen, haloalkyl, hydroxyl, hydroxyalkyl, alkoxy, alkyl, and -COOH; B is one chosen from: 46 Attorney Docket No.: 071741.11025/5WO1 H N N H X S N N H; oaryl, wherein the C3-12 cycloalkyl or C2-12 heterocyclyl is optionally fused with an aryl; r is 1 or 2; n2 is 0, 1, or 2; each R 2 is independently chosen from hydrogen, halogen, hydroxyl, and alkoxy; R 3 is chosen from CN, hydroxy, alkoxy, -C(O)-C 0-12 alkylene-CN, -C 0-12
  • R 1 is halogen, such as fluorine or chlorine.
  • R 1 is alkoxy, such as -OMe.
  • R 1 is hydroxyl or -COOH or -CH 2 OH.
  • R 1 is haloalkyl, such as trifluoromethyl or -CH2CH2F.
  • B i wherein X is S, O, or NH.
  • n2 [0210] In some embodiments .
  • R 8 n4 [0211] In some embodiments, RT is .
  • ring 12 heteroaryl [0213] In some embodiments, ring M is C 3-12 cycloalkyl or C 2-12 heterocyclyl, wherein the C 3-12 cycloalkyl or C2-12 heterocyclyl is optionally fused with an aryl. [0214] In some embodiments, ring M is phenyl, pyridinyl, pyrimidinyl, thiophenyl, cyclopentyl, or cyclohexyl.
  • R 2 is hydroxyl.
  • R 2 is halogen, such as fluorine or chlorine.
  • the pharmaceutically acceptable salt of the compound of formula (I) is trifluoroacetate or hydrochloride.
  • the compound of formula (II) is a compound of formula (II-A): 49 Attorney Docket No.: 071741.11025/5WO1 R 2 R 3 H , wherein R 1 , R 2 , R 3 , and n1 are defined as above in formula (II).
  • R 1 is halogen, haloalkyl, hydroxyl, alkyl, or -COOH.
  • R 1 is methyl, ethyl, hydroxyl, fluorine, chlorine, trifluoromethyl, -CH 2 CH 2 F, or -COOH.
  • n1 is 2.
  • R 2 is hydrogen, hydroxyl, or halogen.
  • R 2 is fluorine or chlorine.
  • R 3 is -C(O)-NR 4 R 4’ or -SO 2 -NR 4 R 4’ , wherein each of R 4 and R 4’ is independently hydrogen, alkyl, alkoxy, -C0-12 alkylene-N(R 6a )t, -C0-12 alkylene-C3-12 cycloalkyl, - C0-12 alkylene-C2-12 heterocyclyl, -C0-12 alkylene-C1-12 heteroaryl, -C0-12 alkylene-OR 6a , or hydroxyalkyl, and the hydroxyalkyl is optionally substituted with alkoxy; wherein each of the alkyl, C3-12 cycloalkyl, C2-12 heterocyclyl, and C1-12 heteroaryl is optionally substituted with one or more R 4a , wherein R 4a , R 6a , and t are defined as above.
  • each of R 4 and R 4’ is independently hydrogen, alkyl, alkoxy, or hydroxyalkyl. O , ed CH3 O with alkoxy, such as , wherein p is 0, 1, 2, or 3, particularly p is 2.
  • R 3 is -C(O)-NR 4 R 4 or -SO2-NR 4 R 4’ , wherein R 4 and R 4’ , together with the nitrogen atom that they are attached to, form a heterocycle comprising one or more 50 Attorney Docket No.: 071741.11025/5WO1 heteroatoms chosen from O, N, and S, particularly, R 4 and R4’, together with the nitrogen atom that they are attached to, form a six-membered heterocycle.
  • O N [0231]
  • R 3 is h drox l -COOH -CH(CH 3 )-COOH -CN , N .
  • R 4 is hydrogen or alkyl
  • R 5 is amino, alkylamino, C1-12 haloalkyl, -C0-12 alkylene-OR 6a , -C0-12 alkylene-N(R 6a )t, -C0-12 alkylene- SR 6a , -C0-12 alkylene-CN, -C0-12 alkylene-C3-12 cycloalkyl, -C0-12 alkylene-C2-12 heterocyclyl, -C0-12 alkylene-C 1-12 heteroaryl, -C 2-12 alkenyl, or alkyl optionally substituted with cyano or amido; wherein each of the C 3-12 cycloalkyl, C 2-12 heterocyclyl, and C 1-12 heteroaryl is optionally substituted with one or more R 4a ,
  • R 5 is amino, alkylamino, alkoxy, alkyl, or -C2-12 alkenyl.
  • R 5 is alkyl substituted with cyano, such as -CH 2 CN.
  • R 5 is alkyl substituted with amido, such as -CH2CH3CONH2.
  • R 5 is alkyl substituted with alkoxy, trialkylammonium, or thiolate.
  • R 5 when R 5 is -C0-12 alkylene-C3-12 cycloalkyl, -C0-12 alkylene-C2-12 heterocyclyl, or -C0-12 alkylene-C1-12 heteroaryl, the C3-12 cycloalkyl, C2-12 heterocyclyl, and C1-12 , . , , 2 heteroaryl is substituted with one or more R 4a , R 4a is hydroxyl, methyl, oxo, or -C(O)-Me.
  • R 3 is m , wherein m is 0, 1, 2, 3, 4, or 5, and R 6 is sulfonamide, carbamide, or alkyl stituted with cyano.
  • R is sulfonamide of formula –N(R’)SO2-R, wherein each R and R’ is independently chosen from hydrogen and alkyl, particularly R 6 is -NHSO 2 CH 3.
  • R 6 is carbamide of formula –N(R’)C(O)N(R) 2 , wherein each R N N N H and R’ is independently chosen from hydrogen, alkyl, and heteroaryl, particular .
  • R 6 is alkyl optionally substituted with cyano, alkyl optionally substituted with cyano, particularly C1-4 alkyl substituted with cyano.
  • m is 1, 2, or 3, particularly 2.
  • R 7 is hydrogen. N O O S O [0246] In certain embodiments .
  • the compound of formula (II) is a compound of formula (II-B): 52 Attorney Docket No.: 071741.11025/5WO1 R 1 R 8 ( ) n1 (CH2) n3 O n4 , wherein R 1 , R 8 , n1, n3, and n4 are defined as above in formula (II).
  • R 1 is hydrogen or alkyl, such as alkyl, particularly methyl.
  • n1 is 2. [0252] In some embodiments, n3 is 0, 1 or 3. [0253] In some embodiments, n4 is 2, 3, or 5. [0254] In some embodiments, R 8 is alkoxy, such as C1-4 alkoxy, particularly methoxy or ethoxy. [0255] In some embodiments, R 8 is amino. [0256] In some embodiments, R 8 is alkylamino, such as C 1-4 alkylamino, particularly methylamino.
  • R 8 is amide of formula –N(R’)C(O)R, wherein each R and R’ is independently chosen from hydrogen, alkyl, cycloalkyl, aryl, and heteroaryl.
  • R 8 is amide of formula –N(R’)C(O)R, particularly R 8 is - NHCOCH3.
  • R 8 is sulfonamide of formula –N(R’)SO 2 -R, wherein each R and R’ is independently chosen from hydrogen, alkyl, cycloalkyl, aryl, and heteroaryl.
  • R 8 is sulfonamide of formula –N(R’)SO2-R, wherein each R and R’ is independently chosen from hydrogen, -C 0-12 alkylene-C 2-12 heterocyclyl, and alkyl, particularly R 3 is -NHSO 2 CH 3.
  • R 8 is carbamide of formula –N(R’)C(O)N(R)2, wherein each R and R’ is independently chosen from hydrogen, alkyl, cycloalkyl, aryl, and heteroaryl.
  • R 8 is carbamide of formula –N(R’)C(O)N(R) 2 , wherein each R and R’ is independently chosen from hydrogen, alkyl, and heteroaryl, particularly R 3 is - N N 53 Attorney Docket No.: 071741.11025/5WO1 [0263]
  • R 8 is -OCH3, - O O N N H O H O O N N N N N S S .
  • the compound of formula (II) is a compound of formula (II-C): R 2 R 3 H , wherein, n2 is 1 or 2; and R 1 , R 2 , R 3 , and n1 are defined as above in formula (II).
  • R 1 is alkyl, such as methyl.
  • n1 is 2.
  • n2 is 1.
  • R 2 is hydrogen or halogen.
  • R 2 is fluorine.
  • R 3 is -C(O)-NR 4 R 4’ , wherein each of R 4 and R 4’ is independently hydrogen, hydroxy, alkyl, alkoxy, -SO2-NHCH3, -SO2-NH-Ph, -CH2-COOH, -CH2-CH2-COOH, O .
  • R 3 is -SO2-NR 4 R 4’ , wherein each of R 4 and R 4’ is independently hydrogen, hydroxy, or -C0-12 alkylene-C2-12 heterocyclyl.
  • R 3 is -NH-C(O)-R 5 , -N(CH 3 )-C(O)-R 5 or -NH-SO 2 -R 5 , wherein R 5 is alkyl, -C0-12 alkylene-alkoxy, -C0-12 alkylene-C3-12 cycloalkyl, -C0-12 alkylene-NH- C1-12 alkyl, -C0- 12 alkylene-NH- C2-12 heterocyclyl, or -C0-12 alkylene-C2-12 heterocyclyl. [0273] In certain embodiments, R 5 is alkyl, such as methyl.
  • R 5 is -C 0-12 alkylene-alkoxy, such as -CH 2 -OCH 3 . 54 Attorney Docket No.: 071741.11025/5WO1 [0275] In certain embodiments, R 5 is -C 0-12 alkylene-C 3-12 cycloalkyl, such . [0276] In certain embodiments, R 5 is -C0-12 alkylene-NH- C1-12 alkyl, such 5 [0277] In certain embodiments, R is-C0-12 alkylene-NH- C2-12 heterocyclyl, such as O H H N O HN N O .
  • R 3 is -SO2-alkyl, such as -SO2-CH3.
  • R 3 is -C 0-12 alkylene-COOH, such as -COOH, -CH 2 -COOH, - C(Me) 2 -COOH, -CH 2 - CH 2 -COOH.
  • R 3 is -C0-12 alkylene-C1-12 heteroaryl, su , O N HN N N N N N N N N N -R 5
  • the compound of formula (II) is a compound of formula (II-D): 55 Attorney Docket No.: 071741.11025/5WO1 R 2 R 3 H , wherein, n2 is 0 or 1; R 3 is chosen from -C(O)-NHR 4 , -SO2-NHR 4 , -NH-C(O)-R 5 , and -NH-SO2-R 5 , and -NH-R 7 ; R 4 is -C 0-12 alkylene-NHR 6a , -C 0-12 alkylene-C 3-12 cycloalkyl, -C 0-12 alkylene-C 2-12 heterocyclyl, -C0-12 alkylene-C1-12 heteroaryl, -C0-12 alkylene-
  • R 1 is alkyl, such as methyl. [0286] In some embodiments, R 1 is alkoxy, such as -OMe. [0287] In some embodiments, n1 is 1. [0288] In some embodiments, n1 is 2. [0289] In some embodiments, n2 is 0. [0290] In some embodiments, n2 is 1. [0291] In some embodiments, R 2 is hydrogen or halogen.
  • R 3 is -C(O)-NHR 4 or -SO2-NHR 4 , wherein R 4 is -C0-12 alkylene- NHR 6a , -C0-12 alkylene-C3-12 cycloalkyl, -C0-12 alkylene-C2-12 heterocyclyl, -C0-12 alkylene-C1-12 heteroaryl, -C 0-12 alkylene-OR 6a , or alkyl substituted with trialkylammonium; wherein and each of the C3-12 cycloalkyl, C2-12 heterocyclyl, and C1-12 heteroaryl is optionally substituted with one or more R 4a , wherein R 4a and R 6a are defined as above.
  • R 5 is alkyl substituted with trialkylammonium.
  • R 5 is -C0-12 alkylene-C3-12 cycloalkyl, -C0-12 alkylene-C2-12 heterocyclyl, or -C 0-12 alkylene-C 1-12 heteroaryl
  • the C 3-12 cycloalkyl, C 2-12 heterocyclyl, and C 1-12 H heteroaryl is substituted with one or more R 4a , R 4a is hydroxyl, methyl, oxo, or -C(O)-Me.
  • 57 Attorney Docket No.: 071741.11025/5WO1 OMe P [0299] In certain embodiments O .
  • t he compound of formula (II) is a compound of formula (II-E): R 2 R 3 N (II-E) , wherein x is 0 or 1; y is 0 or 1; X is S, O, or NH; and R 1 , R 2 , R 3 , n1, and n2 are defined as above in formula (II).
  • x is 0, y is 1, and X is S, O, or NH.
  • x is 0 or 1
  • y is 0, and X is NH.
  • each R 1 is independently chosen from hydrogen, halogen, alkoxy, and alkyl.
  • R 1 is methyl, chlorine, or methoxy.
  • n2 is 1.
  • n2 is 2.
  • R 2 is hydrogen.
  • R 3 is -C(O)-NR 4 R 4’ , wherein each of R 4 and R 4’ is independently hydrogen or alkoxy.
  • R 3 is -SO2-NR 4 R 4’ , wherein each of R 4 and R 4’ is independently hydrogen or alkyl.
  • R 3 is -NH-C(O)-R 5 or -NH-SO 2 -R 5 , wherein R 5 is alkyl or -C 0-12 alkylene-C2-12 heterocyclyl. 58 Attorney Docket No.: 071741.11025/5WO1 [0311] In certain embodiments, R 5 is alkyl, such as methyl.
  • R 5 is -C0-12 alkylene-C2-12 heterocyclyl, such a or O F): , , R 2 is adjacent to R 3 , and R 2 and R 3 , together with the carbon atoms that they are attached to, form a heterocycle optionally substituted with one or more R 4a ; and R 1 , R 4a , and n1 are defined as above in formula (II). [0314] In some embodiments, R 2 and R 3 , together with the carbon atoms that they are attached to, form a 5- or 6-membered heterocycle optionally substituted with one or more R 4a .
  • the compound of formula (II-F) is a compound of formula (II-F- 1): M1 H ( - - ) , wherein M1 is a heterocycle optionally substituted with one or more R 4a .
  • the compound of formula (II) is a compound of formula (II-G): 59 Attorney Docket No.: 071741.11025/5WO1 R 2 r R 3 H , wherein, each R 2 is independently chosen from hydroxyl and alkoxy; R 3 is chosen from hydroxy and alkoxy; and R 1 , n1 and r are defined as above in formula (II).
  • one R 2 is adjacent to R 3 .
  • each R 2 is independently chosen from hydroxyl and methoxy.
  • R 3 is chosen from hydroxyl and methoxy.
  • the compound of formula (II-G) is a compound of formula (II-G- 1) of (II-G-2): R 2 R 2 R 3 R 3 NH , .
  • the compound of formula (II) is a compound of formula (II-H): R 2 , l; e-COOH, -O-C0-12 alkylene-COOH, -C0-12 alkylene- P(O)(OH) 2 , -C(O)-NH-SO 2 -R 5 , -C(O)-NH-C 0-12 alkylene-COOH, -NH-C 0-12 alkylene-COOH, - 60 Attorney Docket No.: 071741.11025/5WO1 N N N alkylene N ; wherein the -C0-12 alkylene-COOH is optionally osen from amino and alkylamino; and R , R , and n1 are defined as above in formula (II).
  • M is phenyl.
  • M is pyridinyl.
  • Exemplary compounds of formula (I-A), (I-B), (I-C), (I-D), or (II) include, but are not limited to, the compounds described herein, and any tautomer, stereoisomer, pharmaceutically acceptable salt or solvate thereof.
  • Exemplary RT No. structure No. structure No. structure 74 Attorney Docket No.: 071741.11025/5WO1 H N H H 4 N N O 108 O O O 203 NH O O O O 75
  • O N 124 H 216 76
  • stereoisomers e.g., enantiomers, diastereomers
  • racemic mixtures of enantiomers which can be separated from one another using art-known resolution procedures, for instance including liquid chromatography using a chiral stationary phase.
  • stereochemically pure isomeric forms of the compounds described herein can be derived from the corresponding stereochemically pure isomeric forms of the appropriate starting materials, intermediates, or reagents.
  • the compound can be synthesized by stereospecific methods of preparation, which typically employ stereochemically pure starting materials or intermediate compounds.
  • salts of compounds described herein can be synthesized from the parent compound containing an acidic or basic moiety by conventional chemical methods. Generally, such salts can be prepared by reacting the free acid or base forms of these compounds with a stoichiometric amount of the appropriate acid or base in water or in an organic solvent, or in a mixture of the two.
  • suitable organic solvents include, but are not limited to, ether, ethyl acetate (EtOAc), ethanol, isopropanol, or acetonitrile.
  • compositions comprising a compound of formula (I-A), (I-B), (I-C), (I-D), or (II) or a stereoisomer, tautomer, pharmaceutically acceptable salt or solvate thereof, as described herein.
  • Compositions can also comprise a pharmaceutically acceptable carrier.
  • a pharmaceutically acceptable carrier is non-toxic and should not interfere with the efficacy of the active ingredient.
  • Pharmaceutically acceptable carriers can include one or more excipients such as 83 Attorney Docket No.: 071741.11025/5WO1 binders, disintegrants, swelling agents, suspending agents, emulsifying agents, wetting agents, lubricants, flavorants, sweeteners, preservatives, dyes, solubilizers and coatings.
  • excipients such as 83 Attorney Docket No.: 071741.11025/5WO1 binders, disintegrants, swelling agents, suspending agents, emulsifying agents, wetting agents, lubricants, flavorants, sweeteners, preservatives, dyes, solubilizers and coatings.
  • the precise nature of the carrier or other material can depend on the route of administration, e.g., intramuscular, intradermal, subcutaneous, oral, intravenous, cutaneous, intramucosal (e.g., gut), intranasal or intraperitoneal routes.
  • suitable carriers and additives include water, glycols, oils, alcohols, preservatives, coloring agents and the like.
  • suitable carriers and additives include starches, sugars, diluents, granulating agents, lubricants, binders, disintegrating agents and the like.
  • the aqueous solution/suspension can comprise water, glycols, oils, emollients, stabilizers, wetting agents, preservatives, aromatics, flavors, and the like as suitable carriers and additives.
  • compositions can be formulated in any matter suitable for administration to a subject to facilitate administration and improve efficacy, including, but not limited to, oral (enteral) administration and parenteral injections.
  • the parenteral injections include intravenous injection or infusion, subcutaneous injection, intradermal injection, and intramuscular injection.
  • Compositions can also be formulated for other routes of administration including transmucosal, ocular, rectal, long acting implantation, sublingual administration, under the tongue, from oral mucosa bypassing the portal circulation, inhalation, or intranasal.
  • compositions The form of the pharmaceutical compositions, the route of administration, the dosage and the regimen depend upon the condition to be treated, such as the severity of the illness, the age, weight, and sex of the patient.
  • Pharmaceutical compositions can be formulated for different modes of administration such as for topical, oral, intranasal, parenteral, intraocular, intravenous, intramuscular, or subcutaneous administration.
  • a method of preparing a pharmaceutical composition comprising combining a compound of formula (I-A), (I-B), (I-C), (I-D), or (II), or a stereoisomer, tautomer, pharmaceutically acceptable salt or solvate thereof, with at least one pharmaceutically acceptable carrier.
  • compositions can be prepared by any method known in the art in view of the present disclosure, and one of ordinary skill in the art will be familiar with such techniques used to prepare pharmaceutical compositions.
  • a pharmaceutical composition according to the present disclosure can be prepared by mixing a compound of formula (I-A), (I-B), (I-C), (I-D), or (II), with one or more pharmaceutically acceptable carriers according 84 Attorney Docket No.: 071741.11025/5WO1 to conventional pharmaceutical compounding techniques, including but not limited to, conventional admixing, dissolving, granulating, emulsifying, encapsulating, entrapping or lyophilizing processes.
  • the present disclosure relates to a pharmaceutical combination comprising a first therapeutical agent and a second therapeutical agent, wherein the first therapeutical agent is a peripherally selective ⁇ 2AR agonist, such as the peripherally selective ⁇ 2AR agonist described herein.
  • the pharmaceutical combination further comprises a third therapeutic agent.
  • the peripherally selective ⁇ 2AR agonist comprises an ⁇ 2AR activation moiety covalently linked to a peripheral distribution moiety.
  • the ⁇ 2AR activation moiety has formula of m , wherein Y, A, B, R2, R3, m, and n are defined as in formula [0345] In certain embodiments, the ⁇ 2AR activation moiety has formula of Y m , wherein Y, A, B, R2, R3, m, and n are defined as in formula [0346] In certain embodiments, the ⁇ 2AR activation moiety has formula of Y , wherein Y, A, B, R2, R3, m, and n are defined as in formula N [0347] In certain embodiments, the ⁇ 2AR activation moiety has formu , wherein X 1 and Y 1 are defined as in formula (I-D).
  • the ⁇ 2AR activation moiety has formula A B , wherein A and B are defined as in formula (II).
  • R T [0349]
  • the peripheral distribution moiety has formula of , wherein RT is defined as in formula (I-A).
  • R T [0350]
  • the peripheral distribution moiety has formula of , wherein R T is defined as in formula (II).
  • the peripherally selective ⁇ 2AR agonist is a compound of formula (I-A), (I-B), (I-C), (I-D) or (II).
  • the peripherally selective ⁇ 2AR agonist is a compound of formula (II-A), (II-B), (II-C), (II-D), (II-E), (II-F), (II-G), or (II-H).
  • the second therapeutic agent is any therapeutic agent other than a peripherally selective alpha-2 adrenergic receptor ( ⁇ 2AR) agonist.
  • the third therapeutic agent is any therapeutic agent other than a peripherally selective alpha-2 adrenergic receptor ( ⁇ 2AR) agonist, and the third therapeutic agent is different from the second therapeutic agent.
  • the second therapeutic agent is chosen from opioids, acetaminophen (paracetamol), local analgesics, nonsteroidal anti-inflammatory drugs (NSAIDs), antiepileptic medication, antidepressant medication, topical analgesic agents, NMDA receptor antagonists, neurotoxins, cannabinoids, voltage-gated sodium channel inhibitors, adenosine agonists, transient receptor potential (TRP) channel modulators, NGF inhibitors, purinergic receptor antagonists, adenosinergic pathway modulators, sigma-1 receptor antagonists, KCC2 enhancers, glutamate transport enhancers, TrkA receptor antagonists, somatostatin receptor ligands, Human Adenylyl Cyclase Associated Protein 1 (CAP1) modulators, and angiotensin II receptor antagonists, corticosteroids, and any combinations thereof.
  • opioids acetaminophen (paracetamol)
  • NSAIDs nonsteroidal anti-inflammatory drugs
  • antiepileptic medication
  • the opioid is chosen from meperidine, tramadol, codeine, pentazocine, dihydrocodeine, tapentadol, hydrocodone, morphine, oxycodone, methadone, 86 Attorney Docket No.: 071741.11025/5WO1 oxymorphone, hydromorphone, butorphanol, buprenorphine, fentanyl, sufentanil, remifentanil, alfentanil, thebaine, diacetyl morphine, vicodin, dinorbuprenorphine, nalbuphine, and heroin.
  • the opioid is chosen from anrikefon, cebranopadol, STC-007, MR-309, desmetramadol, LPM-3480392, YZJ-4729, RFUS-144, HY-1608, NES-100, PN-6047, Cyt-1010, TRV-250, DMX-101, ADV-502, MEB-1170, TRV-734, KUR-101, 18F-FTC-146, MUM0-1, RFUS-250, NH-16003, and sunobinop.
  • the opioid is a biased opioid ligand, such as oliceridine or SHR8554.
  • the opioid is kappa-opioid agonists, such as CR845 (Difelikefalin).
  • the nonsteroidal anti-inflammatory drug is chosen from aspirin, diflunisal, ibuprofen, naproxen, indomethacin, diclofenac, meclofenamate, mefenamic acid, meloxicam, piroxicam, nabumetone, celecoxib, etoricoxib, biphenyl acetic acid, aceclofenac, ketorolac, etodolac, ketotifen, loxoprofen, tenoxicam, zaltoprofen, zaltoprofen, esflurbiprofen, valdecoxib, lumiracoxib, imidazole salicylate, dexketoprofen, nimesulide, otenaproxes
  • the antiepileptic medication is a calcium channel blockers (CCB), sodium channel blocker, or GABA modulator.
  • CB calcium channel blockers
  • the antiepileptic medication is chosen from gabapentin, pregabalin, enacarbil, mirogabalin, crisugabalin, phenibut, baclofen, 4-fluorophenibut, 4- methylpregabalin, atagabalin (PD-200,390), imagabalin, PD-217,014, tolibut, oxcarbazepine, ziconotide, lomerizine, carbamazepine, oxcarbazepine, lamotrigine, lidocaine, ropivacaine, bupivacaine, valproic acid, topiramate, and divalproex sodium.
  • the antiepileptic medication is non- ⁇ 2 ⁇ antiepileptics, such as carbamazepine or oxcarbazepine.
  • the antiepileptic medication is chosen from pregabalin naproxencarbil, XG-004, RAP-219, AFA-281, priralfinamide, laflunimus, and CPP-115.
  • the antidepressant medication is a selective serotonin and norepinephrine reuptake inhibitor (SNRI), a tricyclic antidepressant (TCA), or a monoamine oxidases inhibitor (MAOI).
  • the selective serotonin and norepinephrine reuptake inhibitor is duloxetine, venlafaxine, desvenlafaxine, bupropion, or milnacipran.
  • the tricyclic antidepressant is amitriptyline, nortriptyline, desipramine, clomipramine, or imipramine.
  • the topical analgesic agent is lidocaine (e.g., 5%) patch or capsaicin (e.g., 8%) patch.
  • the NMDA antagonist is chosen from ketamine, methadone, memantine, amantadine, dextromethorphan, and L-4-chlorokynurenine.
  • the neurotoxin is Botulinum toxin A.
  • the cannabinoid is chosen from nabilone, THC, cannabidiol (CBD), nabiximols (Sativex), AP-707, ONO-1110, CMX-020, CNTX-6016, and OCT-461201.
  • the voltage-gated sodium channel inhibitor is the inhibitor of NaV1.3, NaV1.7, NaV1.8 and / or NaV1.9.
  • the selective NaV1.8 inhibitor is chosen from suzetrigine (VX-548), ANP-230, VX-150, JMK-000623, HBW-004285, DSP-2230, VX-993, VX-973, LTG- 001, kindolor, FZ-008, HRS-4800, HRS-2129, MK-5661, and STC-004.
  • the selective NaV1.7 inhibitor is chosen from tetrodotoxin, OLP-1002, aneratrigine, CC-8464, DSP-3905, S-151128, and ST-2427.
  • the voltage-gated sodium channel inhibitor is relutrigine or ANP- 230.
  • the transient receptor potential (TRP) channel blocker is TRPV1 agonist, TRPV1 antagonist, TRPA1 inhibitor, TRPM3 inhibitor, TRPM8 inhibitor, and TRPC5 antagonist.
  • the transient receptor potential (TRP) channel blocker is chosen from capsaicin, vocacapsaicin, resiniferatoxin (RTX), ACD-440, libvatrep, tivanisiran, JNJ- 39439335, SB-705498, AJH-2947, SER-014, SRP-001, XEN-D0501, QP-5113, LD-04185, LD- 2020, BHV-2100, Elismetrep, MT-8554, RQ-00434739, and QR-060127.
  • TRP transient receptor potential
  • the NGF inhibitor is chosen from Tanezumab, Fulranumab, Fasinumab, EP-9001A, MEDI-7352, SSS-40, DS-002, luvagrobart, TNM-009, and STSA-1001.
  • the purinergic receptor antagonist is P2X purinoreceptor antagonist, such as NC-2600, or LY-3857210, P2X3 antagonists, such as Gefapixant (Merck), Sivopixant (Shionogi), or VNUT Inhibitor such as Clodronate.
  • the adenosinergic pathway modulator is a modulator for Adenosine A1, A2A, A3 receptors; CD39/73 enzymes; or Adenosine kinase.
  • the adenosinergic pathway modulator is caffeine.
  • the sigma-1 receptor antagonist is ketamine, memantine, or amantadine
  • the KCC2 enhancer is CLP257 or kenpaullone.
  • the glutamate transport enhancer is ceftriaxone.
  • the TrkA receptor antagonist is AK-1830 or BR-01T.
  • the somatostatin receptor ligand is pasireotide, LY-3556050, or FZ002-037.
  • the Human Adenylyl Cyclase Associated Protein 1 (CAP1) modulator is FM-888 or NB-001.
  • the Angiotensin II receptor antagonist is CFTX-1554, TRD-205, or WXSH-0024.
  • the corticosteroid is chosen from dexamethasone, clobetasol, triamcinolone acetonide, difluprednate, loteprednol, and fluticasone.
  • the second therapeutic agent is chosen from GGG tri-agonist (e.g., retatrutide), metabotropic glutamate receptors (mGluR) modulator (e.g., basimglurant), TGF beta-1 Inhibitor (e.g., Vicatertide (SB-01)), gonadotropin-releasing hormone receptor(GnRH) antagonist (e.g., linzagolix), 11 ⁇ -HSD1 inhibitor (e.g., clofutriben), 17-HSD-1 inhibitor (e.g., OG-6219), Dual ENKephalinase Inhibitor (DENKI) (e.g., PL37), CCR2 receptor antagonist (e.g., CNTX-6970), IL- 10 receptor agonist (e.g., XT-150), nAChR dual alpha 4/beta 2 subunit stimulator (e.g., ATA-104), allosteric NEK7/NLRP
  • GGG tri-agonist
  • the second therapeutic agent is chosen from IRX-101, MR-107A- 02, 3-VM-1001, lysergide assisted therapy, SIL-1002, TRN-261, HR-1405-01, HRF-2105, TTAX- 03, KP-910, LYT-503, LL-50, ZeP-3, YR-1702, YZJ-1495, FB-1003, MK-4318, PZH-2108, HEC- 137076MsOH, VVZ-2471, SYNP-101, Pudafensine, BIOS-0618, and HSK-36357.
  • kits for treating or preventing pain in a subject in need thereof comprising administering to the subject an effective amount of a peripherally selective ⁇ 2AR agonist, wherein treating with the peripherally selective ⁇ 2AR agonist causes less side effects than treating with a non-peripherally selective ⁇ 2AR agonist, such as at similar or comparable dosage.
  • the peripherally selective ⁇ 2AR agonist comprises an ⁇ 2AR activation moiety covalently linked to a peripheral distribution moiety.
  • kits for treating or preventing a disease in a subject in need thereof comprising administering to the subject an effective amount of a peripherally selective ⁇ 2AR agonist, wherein the peripherally selective ⁇ 2AR agonist comprises an ⁇ 2AR activation moiety covalently linked to a peripheral distribution moiety.
  • the present disclosure relates to a method of treating or preventing a disease in a subject in need thereof, the method comprising administering to the 90 Attorney Docket No.: 071741.11025/5WO1 subject an effective amount of the pharmaceutical combination described herein, or administering to the subject (i) an effective amount of a first therapeutic agent, and (ii) an effective amount of a second therapeutic agent, wherein the first therapeutic agent is a peripherally selective ⁇ 2AR agonist, such as the peripherally selective ⁇ 2AR agonist described herein.
  • the term “effective amount” regarding a pharmaceutical combination includes the effective amount of the first therapeutical agent and the effective amount of the second therapeutical agent, and these two effective amounts can be identical or different, fixed or varying dependent on upon a variety of factors as described above.
  • the disease is chosen from glaucoma, analgesia, spasticity, nasal congestion, rosacea, rhinitis, anesthesia, menopausal hot flashes, dysmenorrhea, presbyopia, acute kidney injury, insomnia, inflammatory disease, and cancer.
  • the disease is pain.
  • the second therapeutic agent is any therapeutic agent other than a peripherally selective ⁇ 2AR agonist.
  • the present disclosure relates to a method for treating or preventing pain in a subject in need thereof, the method comprising administering to the subject an effective amount of a first analgesic agent, and an effective amount of a second analgesic agent, wherein the first analgesic agent is a peripherally selective alpha-2 adrenergic receptor ( ⁇ 2AR) agonist, such as the peripherally selective ⁇ 2AR agonist described herein.
  • the method for treating or preventing pain further comprising administering to the subject an effective amount of a third analgesic agent.
  • the present disclosure relates to a method for treating or preventing pain in a subject in need thereof, the method comprising administering to the subject an effective amount of a codrug that comprises a first analgesic agent and a second analgesic agent, wherein the first analgesic agent is a peripherally selective alpha-2 adrenergic receptor ( ⁇ 2AR) agonist, such as the peripherally selective ⁇ 2AR agonist described herein.
  • a codrug that comprises a first analgesic agent and a second analgesic agent
  • the first analgesic agent is a peripherally selective alpha-2 adrenergic receptor ( ⁇ 2AR) agonist, such as the peripherally selective ⁇ 2AR agonist described herein.
  • ⁇ 2AR peripherally selective alpha-2 adrenergic receptor
  • the body After administration, the body (through enzymatic cleavage or hydrolysis) breaks the linkage, releasing the two active drugs at the site of action or in systemic circulation.
  • 91 Attorney Docket No.: 071741.11025/5WO1
  • the peripherally selective ⁇ 2AR agonist comprises an ⁇ 2AR activation moiety covalently linked to a peripheral distribution moiety.
  • the pain is neuropathic pain, nociceptive pain, nociplastic pain, or mixed pain.
  • the neuropathic pain is chosen from diabetic peripheral neuropathy, postherpetic neuralgia, chemotherapy-induced peripheral neuropathy, Fabry disease– associated pain, trigeminal neuralgia, glossopharyngeal neuralgia, occipital neuralgia, HIV- associated neuropathy, hereditary sensory and autonomic neuropathy, Charcot-Marie-Tooth disease–related pain, small fiber neuropathy, brachial plexus avulsion, phantom limb pain, post- surgical neuropathic pain, carpal tunnel syndrome, other compression or entrapment neuropathies, amyloid neuropathy, vasculitic neuropathy, leprosy-associated pain, radiation-induced neuropathy, drug-induced neuropathy, metabolic neuropathies, endocrine-related neuropathies, toxic neuropathies, immune-mediated neuropathies, traumatic peripheral nerve injury, and heritable neuropathies, as well as post-stroke pain syndrome, spinal cord injury–related pain, multiple sclerosis–associated pain,
  • the nociceptive pain is somatic (i.e., from skin, muscles, or joints (e.g., cuts, fractures)) or visceral (i.e., from internal organs (e.g., appendicitis)).
  • the nociceptive pain is osteoarthritis-related rheumatoid pain, arthritis-associated post-traumatic pain, musculoskeletal pain, postoperative pain, tendonitis, myofascial pain, visceral inflammatory pain (e.g., appendicitis, cystitis, pancreatitis), dysmenorrhea, or inflammatory bowel disease-related pain.
  • the mixed pain is cancer-related pain (e.g., bone metastasis pain, tumor infiltration pain, perineural invasion), complex regional pain syndrome type I, failed back surgery syndrome, low back pain with radiculopathy endometriosis-associated pelvic pain, chronic post-surgical pain, chronic pancreatitis-associated pain, interstitial cystitis/bladder pain syndrome, 92 Attorney Docket No.: 071741.11025/5WO1 fibromyalgia, temporomandibular disorder, vulvodynia, irritable bowel syndrome with visceral pain, and combinations thereof.
  • the pain is acute pain or chronic pain.
  • the term “chronic pain” refers to pain that is persistent or recurrent pain lasting longer than 3 months.
  • the chronic pain is chronic primary pain, chronic cancer pain, chronic postsurgical and posttraumatic pain, chronic neuropathic pain, chronic headache and orofacial pain, chronic visceral pain, chronic musculoskeletal pain.
  • the pain is cancer pain.
  • the pain is post-surgery pain.
  • the second therapeutic agent or the second analgesic agent is any therapeutic agent or analgesic agent other than a peripherally selective alpha-2 adrenergic receptor ( ⁇ 2AR) agonist.
  • the second analgesic agent is chosen from opioids, acetaminophen (paracetamol), local analgesics, nonsteroidal anti-inflammatory drugs (NSAIDs), antiepileptic medication, antidepressant medication, topical analgesic agents, NMDA receptor antagonists, neurotoxins, cannabinoids, voltage-gated sodium channel inhibitors, adenosine agonists, transient receptor potential (TRP) channel modulators, NGF inhibitors, purinergic receptor antagonists, adenosinergic pathway modulators, sigma-1 receptor antagonists, KCC2 enhancers, glutamate transport enhancers, TrkA receptor antagonists, somatostatin receptor ligands, Human Adenylyl Cyclase Associated Protein 1 (CAP1) modulators, and angiotensin II receptor antagonists, corticosteroids, and any combinations thereof.
  • opioids opioids
  • acetaminophen paracetamol
  • NSAIDs nonsteroidal anti-inflammatory drugs
  • the opioid is chosen from meperidine, tramadol, codeine, pentazocine, dihydrocodeine, tapentadol, hydrocodone, morphine, oxycodone, methadone, oxymorphone, hydromorphone, butorphanol, buprenorphine, fentanyl, sufentanil, remifentanil, alfentanil, thebaine, diacetyl morphine, vicodin, dinorbuprenorphine, nalbuphine, and heroin.
  • the opioid is chosen from anrikefon, cebranopadol, STC-007, MR-309, desmetramadol, LPM-3480392, YZJ-4729, RFUS-144, HY-1608, NES-100, PN-6047, Cyt-1010, TRV-250, DMX-101, ADV-502, MEB-1170, TRV-734, KUR-101, 18F-FTC-146, MUM0-1, RFUS-250, NH-16003, and sunobinop.
  • the opioid is a biased opioid ligand, such as oliceridine or SHR8554.
  • the opioid is kappa-opioid agonists, such as CR845 (Difelikefalin).
  • the nonsteroidal anti-inflammatory drug is chosen from aspirin, diflunisal, ibuprofen, naproxen, indomethacin, diclofenac, meclofenamate, mefenamic acid, meloxicam, piroxicam, nabumetone, celecoxib, etoricoxib, biphenyl acetic acid, aceclofenac, ketorolac, etodolac, ketotifen, loxoprofen, tenoxicam, zaltoprofen, zaltoprofen, esflurbiprofen, valdecoxib, lumiracoxib, imidazole salicylate, dexketopro
  • NSAID nonsteroidal anti-inflammatory drug
  • the antiepileptic medication is a calcium channel blockers (CCB), sodium channel blocker, or GABA modulator.
  • CB calcium channel blockers
  • the antiepileptic medication is chosen from gabapentin, pregabalin, enacarbil, mirogabalin, crisugabalin, phenibut, baclofen, 4-fluorophenibut, 4- methylpregabalin, atagabalin (PD-200,390), imagabalin, PD-217,014, tolibut, oxcarbazepine, ziconotide, lomerizine, carbamazepine, oxcarbazepine, lamotrigine, lidocaine, ropivacaine, bupivacaine, valproic acid, topiramate, and divalproex sodium.
  • the antiepileptic medication is non- ⁇ 2 ⁇ antiepileptics, such as carbamazepine or oxcarbazepine.
  • the antiepileptic medication is chosen from pregabalin naproxencarbil, XG-004, RAP-219, AFA-281, priralfinamide, laflunimus, and CPP-115.
  • the antidepressant medication is a selective serotonin and norepinephrine reuptake inhibitor (SNRI), a tricyclic antidepressant (TCA), or a monoamine oxidases inhibitor (MAOI).
  • the selective serotonin and norepinephrine reuptake inhibitor is duloxetine, venlafaxine, desvenlafaxine, bupropion, or milnacipran.
  • the tricyclic antidepressant is amitriptyline, nortriptyline, desipramine, clomipramine, or imipramine.
  • the topical analgesic agent is lidocaine (e.g., 5%) patch or capsaicin (e.g., 8%) patch.
  • the NMDA antagonist is chosen from ketamine, methadone, memantine, amantadine, dextromethorphan, and L-4-chlorokynurenine.
  • the neurotoxin is Botulinum toxin A. 94 Attorney Docket No.: 071741.11025/5WO1
  • the cannabinoid is chosen from nabilone, THC, cannabidiol (CBD), nabiximols (Sativex), AP-707, ONO-1110, CMX-020, CNTX-6016, and OCT-461201.
  • the voltage-gated sodium channel inhibitor is the inhibitor of NaV1.3, NaV1.7, NaV1.8 and / or NaV1.9.
  • the selective NaV1.8 inhibitor is chosen from suzetrigine (VX-548), ANP-230, VX-150, JMK-000623, HBW-004285, DSP-2230, VX-993, VX-973, LTG- 001, kindolor, FZ-008, HRS-4800, HRS-2129, MK-5661, and STC-004.
  • the selective NaV1.7 inhibitor is chosen from tetrodotoxin, OLP-1002, aneratrigine, CC-8464, DSP-3905, S-151128, and ST-2427.
  • the voltage-gated sodium channel inhibitor is relutrigine or ANP- 230.
  • the transient receptor potential (TRP) channel blocker is TRPV1 agonist, TRPV1 antagonist, TRPA1 inhibitor, TRPM3 inhibitor, TRPM8 inhibitor, and TRPC5 antagonist.
  • the transient receptor potential (TRP) channel blocker is chosen from capsaicin, vocacapsaicin, resiniferatoxin (RTX), ACD-440, libvatrep, tivanisiran, JNJ- 39439335, SB-705498, AJH-2947, SER-014, SRP-001, XEN-D0501, QP-5113, LD-04185, LD- 2020, BHV-2100, Elismetrep, MT-8554, RQ-00434739, and QR-060127.
  • TRP transient receptor potential
  • the NGF inhibitor is chosen from Tanezumab, Fulranumab, Fasinumab, EP-9001A, MEDI-7352, SSS-40, DS-002, luvagrobart, TNM-009, and STSA-1001.
  • the purinergic receptor antagonist is P2X purinoreceptor antagonist, such as NC-2600, or LY-3857210, P2X3 antagonists, such as Gefapixant (Merck), Sivopixant (Shionogi), or VNUT Inhibitor such as Clodronate.
  • the adenosinergic pathway modulator is a modulator for Adenosine A1, A2A, A3 receptors; CD39/73 enzymes; or Adenosine kinase.
  • the adenosinergic pathway modulator is caffeine.
  • the sigma-1 receptor antagonist is ketamine, memantine, or amantadine
  • the KCC2 enhancer is CLP257 or kenpaullone.
  • the glutamate transport enhancer is ceftriaxone.
  • the TrkA receptor antagonist is AK-1830 or BR-01T. 95 Attorney Docket No.: 071741.11025/5WO1 [0447]
  • the somatostatin receptor ligand is pasireotide, LY-3556050, or FZ002-037.
  • the Human Adenylyl Cyclase Associated Protein 1 (CAP1) modulator is FM-888 or NB-001.
  • the Angiotensin II receptor antagonist is CFTX-1554, TRD-205, or WXSH-0024.
  • the corticosteroid is chosen from dexamethasone, clobetasol, triamcinolone acetonide, difluprednate, loteprednol, and fluticasone.
  • the second analgesic agent is chosen from GGG tri-agonist (e.g., retatrutide), metabotropic glutamate receptors (mGluR) modulator (e.g., basimglurant), TGF beta-1 Inhibitor (e.g., Vicatertide (SB-01)), gonadotropin-releasing hormone receptor(GnRH) antagonist (e.g., linzagolix), 11 ⁇ -HSD1 inhibitor (e.g., clofutriben), 17-HSD-1 inhibitor (e.g., OG-6219), Dual ENKephalinase Inhibitor (DENKI) (e.g., PL37), CCR2 receptor antagonist (e.g., CNTX-6970), IL- 10 receptor agonist (e.g., XT-150), nAChR dual alpha 4/beta 2 subunit stimulator (e.g., ATA-104), allosteric NEK7
  • GGG tri-agonist
  • the second analgesic agent is chosen from IRX-101, MR-107A-02, 3-VM-1001, lysergide assisted therapy, SIL-1002, TRN-261, HR-1405-01, HRF-2105, TTAX-03, KP-910, LYT-503, LL-50, ZeP-3, YR-1702, YZJ-1495, FB-1003, MK-4318, PZH-2108, HEC- 137076MsOH, VVZ-2471, SYNP-101, Pudafensine, BIOS-0618, and HSK-36357.
  • the administration of the first therapeutic agent and the second therapeutic agent provides a synergistic effect in treating the disease.
  • the administration of the first analgestic agent and the second analgestic agent provides a synergistic effect in treating the pain.
  • the term "synergistic effect” refers to a pharmacological interaction between two or more agents wherein the combined effect is greater than the sum of their individual effects. In the context of analgesic or therapeutic efficacy, synergy indicates that the combination achieves enhanced results at equal or lower doses compared to each compound used separately.
  • the synergistic effect is potentiation.
  • the term “potentiation” refers to a specific type of synergy in which one agent, often inactive or weakly active alone, significantly enhances the effect of another active compound when used in combination.
  • the administration of the first therapeutic agent and the second therapeutic agent provides an additive effect in treating the disease.
  • the administration of the first analgestic agent and the second analgestic agent provides an additive effect in treating the pain.
  • additive effect refers to a combined pharmacological effect that is equal to the sum of the effects of individual agents, without enhancement or suppression.
  • Additivity is typically defined within an acceptable experimental range (e.g., CI between 0.95 and 1.1) that accounts for biological variability.
  • CI combination index
  • the synergistic affect and additive effect in combination therapy can be measured and/or determined by any methods known in the art.
  • combination index (CI) is a quantitative metric used to assess the nature of drug-drug interactions, and CI values can be calculated based on models such as Bliss independence or Loewe additivity.
  • the drug-drug interactions in combination therapy can also be characterized by terms other than synergistic affect or additive effect.
  • the term “sparing effect” is the ability of one drug in a combination to reduce the required dose of another drug while maintaining the same therapeutic effect.
  • the sparing effect is the ability of a peripherally selective alpha-2 adrenergic receptor ( ⁇ 2AR) agonist in a combination to reduce the required dose of another therapeutic agent (such as a analgesic agent) while maintaining the same therapeutic effect.
  • the first therapeutic agent has a sparing effect on the second threapeutic agent.
  • the first analgestic agent has a sparing effect on the second analgesic agent.
  • second therapeutic agents and/or analgestic agents have dose-dependent side effects, which can be severe.
  • some typical dose-dependent side effects of second analgesic agents include, addiction, sedation and drowsiness, respiratory depression, constipation, nausea and vomiting, hypotension, dizziness, somnolence (sleepiness), peripheral edema (swelling), weight gain, blurred vision, difficulty with concentration/attention, dry mouth, and fatigue.
  • Such side effects are often dose-limiting side effects, and sometimes even life-threating.
  • the present disclosure provides a combination with a peripherally selective alpha-2 adrenergic receptor ( ⁇ 2AR) agonist allows the dose of second therapeutic agent/analgesic agent to be reduced dramatically, e.g., by 66.67%, while maintaining the same therapeutic effect, and thereby mnimizes adverse events such as those dose-dependent side effects listed above.
  • ⁇ 2AR peripherally selective alpha-2 adrenergic receptor
  • first therapeutic agent and the second therapeutic agent can be administrered by way of simultaneous, sequential or separate administration.
  • the following embodiments apply to the above all general aspects of methods of use.
  • the ⁇ 2AR activation moiety has formula of u la ula [0473] In some embodiments, the ⁇ 2AR activation moiety has formula of Y , wherein Y, A, B, R2, R3, m, and n are defined as in formula N [0474] In some embodiments, the ⁇ 2AR activation moiety has formu , wherein X 1 and Y 1 are defined as in formula (I-D). 99 Attorney Docket No.: 071741.11025/5WO1 [0475] In some embodiments, the ⁇ 2AR activation moiety has formu A B , wherein A and B are defined as in formula (II).
  • the peripheral distribution moiety has formula of , wherein RT is defined as in formula (I-A).
  • R T [0477] In some embodiments, the peripheral distribution moiety has formula of , wherein R T is defined as in formula (II).
  • the compounds of formula (I-A), (I-B), (I-C), (I-D), or (II) can be useful for activating ⁇ 2AR.
  • a method of treating or preventing a disease in human or in animal comprising administering to the subject a compound or composition described herein, e.g., administering an effective amount of a compound or composition described herein.
  • the disease is glaucoma, pain, spasticity, nasal congestion, rosacea, rhinitis, anesthesia, presbyopia, acute kidney injury, insomnia, inflammatory disease, cancer, etc.
  • the disease is pain.
  • the pain is nociceptive pain, nociplatic pain, neuropathic pain such as peripheral neuropathic pain, or mixed pain. Examples of peripheral neuropathic pain include, but 100 Attorney Docket No.: 071741.11025/5WO1 not limited to diabetic neuropathy, postherpetic neuralgia, HIV-associated pain, chemotherapy- induced peripheral neuropathy, and post-surgical neuropathic pain.
  • the compounds and pharmaceutical compositions described herein cause less side effects when treating pain, such as sedation, decreasing heart rate, and decreasing blood pressure in the treated subject.
  • the compounds and pharmaceutical compositions described herein do not cause sedative response in the treated subject.
  • TAA triethylamine
  • TES triethyl silane
  • Trt trityl group or triphenylmethyl group
  • MeOH refers to methanol.
  • EtOH refers to ethanol.
  • t-BuXphos tert-butyl-Xantphos
  • TMAl trimethylaluminum
  • Xantphos refers to 9,9-dimethyl-4,5-bis(diphenylphosphino)xanthene
  • Pd(PPH3)4 refers to tetrakis(triphenylphosphine)palladium(0) [0490]
  • Example 1 Example 1
  • Step 1 400mL THF and 36g (0.18mol, 4.0eq) of 3-bromobenzoic acid were added into a 500mL reaction flask. Following cooling to -65 °C, 135mL (4mol/L, 0.428mol, 7.5eq) of n- Butyllithium was added. The mixture was stirred at -65 °C for 2 hours before 20g (0.057mol, 1.0eq) of compound 1-1 and an additional 400mL of THF were introduced.
  • Step 2 150 ml of 55% HI, 7.5g (13.3mmol, 1.0eq) of compound 1-2, and 4.1g (133mmol, 10.0eq) of red phosphorus were added into a 200 mL sealing tube The mixture was stirred at 160 °C for 16 hours until LC-MS indicated completion. Following vacuum concentration, the residue was collected to produce 7.3g of compound 1-3, achieving a 100% yield.
  • Step 4 52ml DCM, and then 1.3g (2.37mmol, 1.0eq) of compound 1-4, 594mg (7.11mmol, 3.0eq) of methoxyammonium chloride, 2.45g (18.96mmol, 8.0eq) of DIPEA, 640mg (4.74mmol, 2.0eq) of HOBT, and 999mg (5.21mmol, 2.2eq) of EDCI were added to a 100 mL reaction flask. The mixture was stirred at room temperature for 5 hours until LC-MS confirmed the reaction's completion.
  • Step 5 18mL DCM and 900mg (1.56mmol, 1.0eq) of compound 1-5, along with 9mL of TFA, were added to a 50mL reaction flask. The reaction was stirred at room temperature for 2 hours until LC-MS showed completion. Following vacuum concentration, the residue underwent column chromatography to yield 670 mg of compound 1, achieving a 98.5% yield.
  • Step 6 Compound 1 was separated by chiral HPLC to afford compound 1-A and compound 1-B.
  • the first eluting enantiomer (compound 1-A) 103 Attorney Docket No.: 071741.11025/5WO1 with a retention time of 4.18min was isolated from the eluent with an enantiomeric excess of 100% in 80% yield.
  • the naming convention for separated enantiomers is systematic. "A” denotes the first eluting product from the chromatography, and "B" indicates the second.
  • Step 2 A 250mL three-necked round bottom flask received 113ml of DCM, 11.3 g (18mmol, 1.0eq) of compound 8-2, HSiEt3 (21g, 180mmol, 10eq), and TFA (21g, 180mmol, 10eq) under nitrogen at 0°C.
  • Step 3 17 g (18mmol, 1.0eq) of compound 8-3, TrtCl (12.6g, 45mmol, 2.5eq), 170ml of DCM and Et3N (9.1 g, 90mmol, 5eq) were mixed in a 500ml three-necked round bottom flask under nitrogen. After stirring for 16 hours at 25°C, completion was confirmed by LC-MS. The product was processed similarly to previous steps to yield 7.1 g of compound 8-4 (64%).
  • Step 4 100ml of DMF, 6 g (9.8mmol, 1.0eq) of compound 8-4, Zn(CN)2 (1.26g, 10.8mmol, 1.1eq), and Pd(PPh 3 ) 4 (1.26g, 1.1mmol, 0.11eq) were added under nitrogen to a 250ml three-necked round bottom flask. After stirring at 120°C for 2 hours, LC-MS confirmed completion. Following the standard work-up, 5.1 g of compound 8-5 (93%) was obtained.
  • Step 5 60ml of EtOH, 2 g (3.6mmol, 1.0eq) of compound 8-5, and 12mL of 30% KOH were added under nitrogen into a 100ml single-mouth flask. The mixture was refluxed for 72 hours. After concentration under vacuum and subsequent work-up, 1.9 g of compound 8-6 (91%) was purified.
  • Step 6 20ml of DCM, 1g (1.73mmol, 1.0eq) of compound 8-6, EDCI (0.432g, 2.25mmol, 1.3eq), DIPEA (0.893g, 6.92mmol, 4eq), HOBt (0.234g, 1.73mmol, 1.0eq), and methoxyammonium chloride (0.174g, 2.08mmol, 1.2eq) were combined in a 100mL single-mouth flask under nitrogen. Stirred for 16 hours at 25°C and then processed as before, this yielded 0.43 g of compound 8-7 (41%).
  • Step 7 10ml of DCM, 0.430 g (0.71mmol, 1eq) of compound 8-7, and BBr3 (0.435 g, 1.775mmol, 2.5eq) were mixed in a 25ml single-mouth flask at 0°C under nitrogen. Stirring continued for 3 hours at 0°C until LC-MS indicated completion, proceeding directly to the next step.
  • Step 8 The mixture from Step 7 and 10ml of MeOH were added to a 50ml single-mouth flask under nitrogen. Heated to reflux for 16 hours, completion was verified by LC-MS.
  • Step 2 3ml of DCM, 150mg (0.25mmol, 1eq) of compound 17-2, and 1.5ml of TFA were added to a 10ml single-mouth flask under N2 at 25°C. After stirring for 2 hours and confirmation of completion by LC-MS, the reaction mixture was vacuum concentrated and subjected to purification through a fast silica gel column, yielding 33mg of compound 17, which corresponds to a 28% yield.
  • Example 4 Synthesis of Compound 22 O OH O O O O O Br N H O O O NH2 O N S O p - , g .
  • Step 2 300mg of compound 22-2, 10ml DCM, and 5ml TFA were added to a 50mL three-necked flask. This mixture was stirred at room temperature (25°C) for 12 hours. LC-MS confirmed completion; the mixture was then diluted with water, adjusted to pH 10, and extracted with DCM. The organic phase was dried over Na 2 SO 4 , vacuum concentrated, and the resulting residue was column chromatographed on silica gel to obtain 85mg of compound 22-3 as a yellow solid, with a yield of 45.9%.
  • Step 1 In a 50mL reaction flask, 10mL of THF and 860mg (3.39mmol, 1.5eq) of 1,3- dibromo-2-fluorobenzene were combined and cooled to -65°C. Next, 1.4mL (3.39mmol, 1.5eq) of n-butyllithium was added. The mixture was stirred at this temperature for 2 hours before adding 1g 107 Attorney Docket No.: 071741.11025/5WO1 (2.26mmol, 1eq) of compound 27-1 and another 10mL of THF. It was stirred for an additional 30 minutes at -65°C, then allowed to warm to room temperature over 16 hours.
  • Step 2 A 50mL three-necked flask was charged with 600mg (0.971mmol, 1eq) of compound 27-2, 1.1g (9.71mmol, 10eq) of triethylsilane, and 1.1g (9.71mmol, 10eq) of TFA. The mixture was stirred at 25°C for 1 hour.
  • Step 3 Into a 25mL reaction flask, 10ml of DMF, 170mg (0.473mmol, 1eq) of compound 27-3, 158g (0.568mmol, 1.2eq) of triphenylmethyl chloride, and 96mg (0.946mmol, 2eq) of TEA were added. The mixture was stirred at 25°C for 12 hours until LC-MS indicated the reaction had completed.
  • Step 4 A 25mL reaction flask was prepared with 10ml DMF, 170mg (0.283mmol, 1eq) of compound 27-4, 100mg (0.848mmol, 3eq) of ZnCN, and 98mg (0.0848mmol, 0.3eq) of tetrakis(triphenylphosphine)palladium. Stirring was conducted at 150°C for 30 minutes in a microwave.
  • Step 5 To a 25mL reaction flask, 10mL of DMSO and 110mg (0.201mmol, 1eq) of compound 27-5 were added and cooled to 0°C. Then, 3ml of 30% H2O2 was added, and the mixture was stirred at 0°C for 1 hour. Following LC-MS confirmation of completion, water was added, and the organic phase was separated and concentrated under vacuum. Column chromatography purification yielded 100mg of compound 27-6, an 88.1% yield.
  • Step 2 Into a 25ml reaction flask, 5ml THF, 185mg (7.6mmol, 2.1eq) of magnesium chips, and 2g (7.2mmol, 2.0eq) of compound 28-2 were added.
  • Step 3 A 100mL reaction flask was charged with 14mL DCM, 1.4g (2.18mmol, 1.0eq) of compound 28-3, and 2.53g (21.8mmol, 10eq) of TES. After cooling to 0°C, 2.48g (21.8mmol, 109 Attorney Docket No.: 071741.11025/5WO1 10eq) of TFA was added. The mixture was warmed to 25°C for 5 hours, then concentrated under vacuum after LC-MS confirmed completion. The residue was column chromatographed to yield 500mg of compound 28-4, a 68.4% yield.
  • Step 4 In a 5mL reaction flask, 2mL THF, 100mg (0.29mmol, 1.0eq) of compound 28-4, 7mg (0.06mmol, 0.2eq) of DMAP, 94mg (0.43mmol, 1.5eq) of Boc2O, and 44mg (0.43mmol, 1.5eq) of TEA were mixed. The reaction was held at 25°C for 4 hours, confirmed by LC-MS. After vacuum concentration, the residue was purified by column chromatography, yielding 120mg of compound 28-5, an 85.5% yield.
  • Step 5 A 10mL reaction flask was loaded with 1.5mL acetic acid, 0.5ml water, and 120mg (0.25mmol, 1.0eq) of compound 28-5. After chilling to 0°C, 165mg (1.24mmol, 5eq) of NCS was added. The mixture was stirred at 0°C for 2 hours until LC-MS confirmed completion, then moved to the next step without purification. The yield was recorded as 100%.
  • Step 6 To a 50mL reaction flask, 10mL of 2M NH 2 CH 3 /THF was added and cooled to 0°C before introducing the crude compound 28-6.
  • Step 2 A 50ml closed tank received 20ml of 57 wt.% HI, 2.35g (4.599mmol, 1.0eq) of compound 31-2, and 1.43g (45.99mmol, 10eq) of red phosphorus. Stirred at 160°C overnight and checked by LC-MS for completion, the mixture was cooled to room temperature and concentrated to yield 5g of crude compound 31-3 (100% yield).
  • Step 4 Into a 200mL high-pressure reactor, 174mg (0.586mmol, 1eq) of compound 31-4, 10 mL of MeOH, 296mg (2.93mmol, 5eq) of TEA, and 48mg (0.0586mmol, 0.1eq) of PdCl 2 (dppf) 111 Attorney Docket No.: 071741.11025/5WO1 were introduced. The reaction, under 5MPa of carbon monoxide at 120°C for 48 hours, left 5% of the starting material, as shown by LC-MS. After filtration and concentration, 270mg of compound 31-5 was isolated by column chromatography (100% yield).
  • Step 5 A 50mL closed tank was charged with 100mg (0.312mmol, 1eq) of compound 31-5 and 5mL of MeOH/NH3 (15M/L). The mixture was stirred at 68°C overnight, cooled to room temperature, concentrated under vacuum, and then purified to obtain 10mg of compound 31 through pre-HPLC (10% yield). [0537] Example 8.
  • Step 2 A 50mL single-mouth flask was charged with 200mg (0.35mmol, 1eq) of compound 32-2, 5mL of dichloromethane, and 1mL of trifluoroacetic acid. The mixture was stirred at room temperature for 1 hour, with TLC confirming the reaction's completion.
  • Step 2 To a 200mL high-pressure reactor, 1g (1.74mmol, 1.0eq) of 58-1, 40 mL of MeOH, 40mL of DMSO, 530mg (5.24mmol, 3.0eq) of TEA, and 148mg (0.17mmol, 0.1eq) of PdCl2 (dppf) were added. The mixture was reacted with carbon monoxide at 5MPa and 100 °C for 48 hours, with LC-MS indicating 5% remaining raw material. After concentration, the residue was purified by column chromatography to yield 53mg of compound 58-2, a 51.2% yield.
  • Step 3 A 50mL sealed tube received 25mL of 16M NH3/MeOH and 530mg (0.89mol, 1.0eq) of compound 58-2. Stirred at 30°C for 16 hours until LC-MS confirmed completion, the residue was then purified by column chromatography to yield 360mg of compound 58-3, a 67.8% yield.
  • Step 4 Into a 10mL reaction flask, 3mL DCM, 100mg (0.17mmol, 1.0eq) of compound 58-3, and 195mg (1.7mmol, 10eq) of TES were added. After cooling to 0°C, 191mg (1.7mmol, 10eq) of TFA was introduced.
  • Example 10 Synthesis of Compound 60 O O O O O O O O O OH HCl/Et2O O N + Zn/TiCl4/THF/4h DCM 2h P/HI/AcOH/16h N NH T FA of zinc were added. This mixture was cooled to -10°C under a nitrogen atmosphere while stirring.
  • Step 2 To a 25mL three-necked flask, 10mL of DCM and 0.5g (1.54mmol, 1eq) of compound 60-1, along with 10mL of HCl/Et2O, were added. The mixture was stirred at room temperature for 3 hours. LC-MS indicated the reaction was complete. Concentrating under reduced pressure yielded 400mg of compound 60-2 as crude, with a 100% yield.
  • Step 2 Into a 100mL reaction flask, 40mL THF, 2g (7.0mmol, 1.0eq) of compound 61-1, and 3.4g (7.7mmol, 1.1eq) of (2,3-dimethylphenyl)(1-trityl-4-imidazolyl)methanone were combined and cooled to 0°C before adding 2.35g (21mmol, 3.0eq) of potassium tert-butoxide.
  • Step 3 A 25mL reaction flask was charged with 10mL DCM and 500mg of compound 61-2, followed by the addition of 2.5mL TFA. The reaction mixture was stirred at 27°C for 1 hour. LC-MS indicated completion, and after concentration, the residue was purified by column chromatography to yield 240mg of compound 61-3, an 84.5% yield.
  • Step 4 In a 10mL reaction flask, 3mL THF, 240mg of compound 61-3, and 120mg (50%) of Pd/C were added.
  • Step 5 To a 10mL reaction flask, 3mL DMF, 80mg (0.25mmol, 1.0eq) of compound 61- 4, 209mg (2.5mmol, 10eq) of methoxyammonium chloride, cooled to 0°C, then 386mg (3mmol, 12eq) of DIPEA and 142mg (0.37mmol, 1.5eq) of HATU were added.
  • Step 6 A 10mL reaction flask was prepared with 1mL DCM and 160mg of compound 61-5, and 0.5mL TFA was added. Stirred at 27°C for 1 hour, completion was confirmed by LC- MS. After concentration under vacuum, the residue was purified by prep-HPLC to yield 25mg of compound 61, a 15.3% yield.Overall yield: 2.1%. [0556] Example 12.
  • Step 2 A 100mL three-necked flask received 60ml of ACN, 3.2g (7.01mmol, 1eq) of 139-1, 3.14g (14.02mmol, 2eq) of CAS 39684-80-5, and 3.42g (10.51mmol, 1.5eq) of Cs2CO3. Stirred at 60°C for 12 hours and confirmed complete by LC-MS, the reaction was worked up and purified to give 930mg of 139-2, a 22.1% yield.
  • Step 4 A 25mL three-necked flask was charged with 5ml of THF, 130mg (0.260mmol, 1eq) of 139-3, 40mg (0.390mmol, 1.5eq) of TEA, and 57mg (0.286mmol, 1.1eq) of (Tetrahydro- 2H-pyran-4-yl)methanesulfonyl chloride (CAS 264608-29-9). Stirred at 25°C for 18 hours and verified complete by LC-MS, the mixture was worked up and purified to give 80mg of 139-4 as a white solid, yielding 46.5%.
  • Step 1 A 50mL reaction flask was charged with 25mL of toluene, 5g (0.0188mmol, 1.0eq) of 3-fluoro-4-bromobenzyl bromide, and 3.44g (0.0207mmol, 1.1eq) of triethyl phosphite. Stirred at 110°C for 18 hours, completion was confirmed by LC-MS. The reaction mixture was concentrated and the residue was purified by column chromatography to yield 5.69g of compound 156-1, with a 93.4% yield.
  • Step 2 To a 50mL reaction flask, 20mL of THF, 1g (3.08mmol, 1.0eq) of compound 156-1, and 1.36g (3.08mmol, 1.0eq) of (2,3-dimethylphenyl)(1-trityl-4-imidazolyl)methanone were added and cooled to 0°C. Then, 1.04g (9.24mmol, 3.0eq) of potassium tert-butoxide was introduced.
  • Step 3 A 10mL reaction flask received 5mL of DMF, 500mg (0.817mmol, 1.0eq) of 156- 2, 192mg (1.634mmol, 2.0eq) of zinc cyanide, and 95mg (0.0817mmol, 0.1eq) of Pd(PPh3)4. Stirred at 120°C for 18 hours, TLC indicated 50% of the raw materials remained.
  • Step 4 Into a 5mL reaction flask, 1.5mL of THF, 1.5mL of MeOH, 110mg (0.197mmol, 1.0eq) of 156-3, 54mg (0.394mmol, 2.0eq) of K2CO3, and 45mg (0.394mmol, 2.0eq) of 30% H 2 O 2 were combined under nitrogen. The mixture was stirred overnight at 20°C for 18 hours.
  • Step 5 In a 5mL reaction flask, 3mL of MeOH:THF (1:1), 90mg (0.156mmol, 1.0eq) of 156-4, and 87mg of Pd(OH) 2 were stirred at 48°C overnight under a hydrogen environment. LC- MS confirmed the reaction's completion.
  • Step 2 In a 25mL three-necked flask, 598mg (2.53mmol, 2.0eq) of 182-1 was dissolved in 6mL THF and cooled to -80°C under nitrogen. n-BuLi (2.5M, 2.5mL, 6.33mmol, 5.0eq) was added, stirred for 40 minutes, then a mixture of 560mg (1.27mmol, 1.0eq) of (2,3- dimethylphenyl)(1-trityl-4-imidazolyl)methanone in 6mL THF was introduced. After stirring for 1 hour and quenching with water, the mixture was warmed and extracted with ethyl acetate.
  • Example 15 Example 15
  • Step 2 188-1 (20g, 79.27mmol, 1.0eq) was dissolved in methanol (350mL), and NaBH 4 (14.99g, 0.39mol, 5.0eq) was added at 0°C.
  • Step 3 DMSO (50mL), 188-2 (4g, 17.83mmol, 1.0eq), PySO 3 (7.1g, 44.58mmol, 2.5eq), and TEA (4.51g, 44.58mmol, 2.5eq) were added to a 250mL flask.
  • Step 4 In a 50mL flask, THF (20mL) was combined with Zn (1.1g, 16.87mmol, 7.5eq) and TiCl4 (1.58g, 8.32mmol, 3.7eq) at -70°C, followed by 188-3 (500mg, 2.25mmol, 1.0eq) and a specified compound in THF. Stirred at 80°C for 2 hours, the reaction was completed, worked up, and purified to yield 270mg of 188-4 as a yellow solid (19.1% yield).
  • Step 6 188-5 (60mg, 0.096mmol, 1.0eq) was mixed with oxalyl dichloride (36.92mg, 0.291mmol, 3.0eq) in DCM (1mL) at 0°C, stirred at 15°C for an hour, concentrated to yield 60mg of 188-6 as a white solid (100% yield), and used directly in the next step.
  • Step 7 188-6 (60mg) was dissolved in THF (0.5 mL) and treated with NH 3 ⁇ THF (8mL) at 0°C, stirred at 15°C for an hour, concentrated to yield 60mg of 188-7 as a white solid (100% yield), and used directly in the next step.
  • Example 16 Example 16
  • Step 1 In a 100mL flask, 50mL of dichloromethane and 4-Iodo-1-trityl-1H-imidazole (11.8g, 0.027mol, 1.0eq) were combined. After cooling the mixture to 0°C, iPrMgClLiCl (1.3mol/L, 20.7mL, 0.027mol, 1.0eq) was added.
  • Step 2 Into a 200mL high-pressure tube, 180mL of dichloromethane, compound 401-1 (6.1g, 12.35mmol, 1.0eq), and MnO2 (6.44g, 74.1mmol, 6.0eq) were added. The mixture was stirred at 72°C for 5 hours. Completion was verified by LC-MS, and the mixture was then filtered to yield 5.6g of compound 401-2. achieving an 83.9% yield.
  • Step 3 A 1L reaction vessel was charged with 500 mL of diethyl ether and 20g (82.67mmol, 1eq) of 3,4-Dibromothiophene (Cas: 3141-26-2). Upon cooling to -78°C, 36.37mL (90.94mmol, 1.1eq) of n-BuLi was added dropwise. The mixture was stirred at -78°C for 30 minutes before 14.02g (90.94mmol, 1.1eq) of diethyl sulfate was added dropwise. Stirring continued at 25°C for 5 hours until LC-MS confirmed the reaction's completion.
  • Step 4 Into a 50 mL reaction flask, 5mL of THF and 387.28mg (2.03mmol, 2eq) of 401- 3 were introduced. Cooled to -78°C, 0.81mL (2.03mmol, 2eq) of n-BuLi was added dropwise. After stirring at -78°C for 30 minutes, a solution of 500mg (1.01mmol, 1.0eq) of 401-2 in 5mL of THF was added.
  • Step 5 To a 25mL flask, 7mL of dioxane, 340mg (0.561mmol, 1.0eq) of 401-4, 64.08mg (0.67mmol, 1.2eq) of MsNH4, 10.28mg (0.011mmol, 0.02eq) of Pd2(dba)3, 9.54mg (0.022mmol, 0.04eq) of tBuxphos, and 365.85mg (1.12mmol, 2eq) of Cs2CO3 were added. The mixture was stirred at 100°C for 5 hours.
  • Example 18 Synthesis of Compound 502
  • Step 1 A 500mL three-necked flask was loaded with 250mL of ACN, 25g (0.15mol, leq) of 502-1, 31.5g (0.16mol, 1.05eq) of diethyl chloromalonate, and 43g (0.31mol, 2eq) of K2CO3.
  • the reaction mixture was refluxed at 80°C overnight. After completion was confirmed by LC-MS, the mixture was concentrated under vacuum and purified via silica gel column chromatography to yield 40g of 502-2. The yield was 83%.
  • Step 2 In a IL three-necked flask, 250mL of DMF and 8g (0.21mol, 1 5eq, 60%) of NaH were combined and cooled to 0°C. Then, 40g (0.125mol, leq) of 502-2 dissolved in lOOmL of DMF was added at 0°C and stirred for 1 hour. Next, 30g (0.154mol, l.leq) of 3- (Bromomethyl)benzonitrile in lOOmL of DMF was added at 0°C, and the mixture was stirred at 58°C overnight. After completion (confirmed by LC-MS), the reaction was quenched with water, extracted with EA, dried over Na2SC>4, and concentrated. Purification by silica gel column chromatography yielded 36g of 502-3 with a 64% yield.
  • Step 3 A 500mL three-necked flask received 300mL of DMSO, 36g (0.08mol, leq) of 502-3, 9g (0.15mol, 2eq) of NaCl, and 11g (0.3mol, 4eq) of H2O. The mixture was stirred at 150°C overnight. LC-MS indicated the reaction was incomplete. The mixture was worked up similarly to previous steps and purified to yield 25g of 502-4 with an 86% yield.
  • Step 4 To a 500mL three-necked flask, 200mL of DMSO, 20g (0.055mol, leq) of 502-4, 13g (0.1 Imol, 2eq) of H2O2, and 15g (0.1 Imol, 2eq) of K2CO3 were added. The mixture was stirred at room temperature overnight and purified after standard work-up to yield 8g of 502-05. The yield was 38%.
  • Step 5 A 100mL three-necked flask was charged with 40mL of EtOH, 8g (0.021mol, 1eq) of 502-5, and 12.6g (0.21mol, 10eq) of ethylenediamine.
  • Step 2 A solution of 1-bromo-2-methoxybenzene (28.99g, 155mmol) in dry THF (150mL) was cooled to -78°C, to which n-BuLi (2.5M in hexane, 62mL, 155mmol) was added dropwise. After stirring at -78°C for 30 minutes, a solution of 503-1 (20g, 77.5mmol) in dry THF 124 Attorney Docket No.: 071741.11025/5WO1 (100mL) was added dropwise. The solution was then allowed to warm to room temperature and stirred for 16 hours.
  • n-BuLi 2.5M in hexane, 62mL, 155mmol
  • Step 3 A solution of 503-2 (1.5g, 4.9mmol), PdCl2(dppf) (360mg, 0.49mmol), and sodium carbonate (1.04g, 9.8mmol) in toluene:MeOH (10mL, 1:1 ratio) was heated at 100°C for 3 days under CO atmosphere.
  • reaction was diluted with water, extracted with EtOAc, and purified via silica gel chromatography, eluting with EtOAc/PE from 1% to 10%, to yield 700mg of methyl 3-[2-(2-methoxyphenyl)-2-oxoethyl]benzoate (503-3) as a yellow oil, with a 46.94% yield.
  • Step 4 To a solution of methyl 3-[2-(2-methoxyphenyl)-2-oxoethyl]benzoate (600mg, 2.1mmol) and O-methylhydroxylamine hydrochloride (264.38mg, 3.16mmol) in toluene (8mL), LiHMDS (1M, 8.4mL, 8.441mmol) was added and stirred at 25°C for 3 hours.
  • Step 5 A solution of N-methoxy-3-[2-(2-methoxyphenyl)-2-oxoethyl]benzamide (400mg, 1.3364mmol) and NH 4 OAc (1.545g, 20.046mmol) in IPA (8.0mL) was stirred at 25°C for 30 minutes before adding NaBH3CN (335.92mg, 5.34mmol) and heated at 80°C for 3 hours.
  • Step 6 To a solution of 3-[2-amino-2-(2-methoxyphenyl)ethyl]-N-methoxybenzamide (300mg, 0.9988mmol, 1eq) in DCM:DMF (5.0mL, 10:1 ratio), 1-chloro-2-isocyanatoethane (421.59mg, 3.9952mmol) was stirred at 25°C for 6 hours. After dilution with water and extraction with DCM, the combined organic phases were washed with brine, dried over sodium sulfate, and concentrated under vacuum. Without further purification, the crude product (250mg, 46.26% yield) was obtained as a yellow oil.
  • Step 7 To a solution of 3-(2- ⁇ [(2-chloroethyl)carbamoyl]amino ⁇ -2-(2- methoxyphenyl)ethyl)-N-methoxybenzamide (200mg, 0.4927mmol) in water (5.0mL), the mixture was heated at 100°C for 3 hours.
  • Step 1 In a 250mL flask, 100mL of THF and 13.1g (0.056mol, 1.5eq) of 1,3- dibromobenzene were combined. After cooling to -78°C, 22.4mL (0.056mol, 1.5eq) of 2.5M n- BuLi was added.
  • Step 2 A 50mL flask was charged with 20mL DMF, 1g (3.45mmol, 1.0eq) of 504-1, 492mg (5.175mmol, 1.5eq) of MsNH2, 2.25g (6.9mmol, 2.0eq) of Cs2CO3, 316mg (0.345mmol, 0.1eq) of Pd2(dba)3, and 293mg (0.69mmol, 0.2eq) of tBuxphos.
  • Step 3 To a 5mL flask, 1.5mL DCM, 134mg (0.439mmol, 1.0eq) of 504-2, 76mg (0.659mmol, 1.5eq) of TMSN 3 , and 31mg (0.0878mmol, 0.2eq) of InBr 3 were added.
  • Step 4 A 5mL flask received 1.5mL THF, 0.3mL water, 144mg (0.436mmol, 1.0eq) of 504-3, and 229mg (0.87mmol, 2.0eq) of PPh3.
  • Step 5 To a 5mL flask, 1mL dioxane, 52mg (0.171mmol, 1.0eq) of 504-4, and 60mg (0.5mmol, 2.9eq) of 2-chloroethyl isothiocyanate were added. Stirred at 80°C for 16 hours, completion confirmed by LC-MS, concentrated under vacuum and purified by pre-HPLC to yield 3.3mg of compound 504, a 5% yield. Overall yield: 2.2% [0612] Example 21.
  • Step 2 A mixture of N- ⁇ 3-[2-(2-methoxyphenyl)-2- oxoethyl]phenyl ⁇ methanesulfonamide (400mg, 0.5323mmol) and NH4OAc (1.448g, 18.785mmol) in isopropanol (8.0ml) was stirred at 25°C for 0.5 hour, then NaBH 3 CN (314.80mg, 5.0096mmol) 127 Attorney Docket No.: 071741.11025/5WO1 was added and the mixture was heated at 80°C for 4.5 hours. After cooling, the mixture was filtered through celite and concentrated.
  • Step 3 A solution of N- ⁇ 3-[2-amino-2-(2- methoxyphenyl)ethyl]phenyl ⁇ methanesulfonamide (505-2, 160mg, 0.4994mmol) and 4,5-dihydro- 1H-imidazole-2-sulfonic acid (224.96mg, 1.4982mmol) in butanol:water (5:1 ratio, 3.0ml) was heated at 120°C for 2 hours in a microwave reactor. After cooling, the mixture was concentrated, diluted with water, and extracted with EtOAc.
  • Step 4 In a 25mL bottle, 10mL of THF, 497mg (2.81mmol, 1.0eq) of 510-3 were cooled to -80°C under nitrogen.
  • Step 5 A 50mL bottle received 10mL of DMF, 570mg (1.65mmol, 1.0eq) of 510-4, 1.07g (3.3mmol, 2.0eq) of Cs 2 CO 3 , 235mg (2.47mmol, 1.5eq) of methanesulfonamide, 151mg (0.165mmol, 0.1eq) of Pd2(dba)3, and 141mg (0.33mmol, 0.2eq) of t-BuXphos. Stirred at 105°C for 2 hours under nitrogen, the reaction was worked up and purified to yield 600mg of 510-5, an 85.7% yield.
  • Step 6 In a 10mL bottle, 5mL of toluene, 200mg (0.554mmol, 1.0eq) of 510-5, 166.5mg (2.77mmol, 5.0eq) of ethylenediamine, and TMAl (1.39mL, 2.77mmol, 5.0eq) were stirred at 110°C overnight. After cooling and working up, the crude was purified to yield 29mg of 510-6, a 13.5% yield. [0623] Step 7: A 5mL bottle received 1mL of toluene and 29mg (0.0745mmol, 1.0eq) of 510-6, then 57mg (0.37mmol, 5.0eq) of POCl 3 was added.
  • Example 23 Synthesis of Compound B24 Potassium ethenylt- rifluoroborate O TosMIC, t-BuOK, CN Pd(PPh3)4 , Na 2 CO 3 CN KOH, EtOH, H 2 O O MeI, K 2 CO 3 DCE, rt, 12h 1,4-Dioxane, H2 O 100o OH C, 4h DMF, rt, 12h bromoindanone (10 g, 47.38 mmol, 1.0 equiv), and TosMIC (13.88 g, 71.07 mmol, 1.5 equiv) were added.
  • Step 2 To a 100 mL single-necked flask, 60 mL of dioxane, 6 mL of water, compound B24-1 (3 g, 13.51 mmol, 1.0 equiv), potassium ethenyltrifluoroborate (2.71 g, 20.26 mmol, 1.5 equiv), Na2CO3 (2.15 g, 20.26 mmol, 1.5 equiv), and Pd(PPh3)4 (0.78 g, 0.68 mmol, 0.05 equiv) were added. The solution was stirred at 90°C under nitrogen for 12 hours. TLC analysis confirmed the reaction was complete.
  • Step 3 To a 100 mL single-necked flask, 50 mL of ethanol, compound B24-2 (5 g, 13.53 mmol, 1.0 equiv), 7.59 g of water, and KOH (7.59 g, 135.32 mmol, 10.0 equiv) were added.
  • Step 4 To a 100 mL single-necked flask, 40 mL of DMF, 40 mL of DMF, compound B24-3 (3.9 g, 13.55 mmol, 1.0 equiv), and K2CO3 (3.74 g, 27.10 mmol, 2.0 equiv) were added. Methyl iodide (3.85 g, 27.10 mmol, 2.0 equiv) was then added to the stirred solution. The mixture was stirred at ambient temperature for 12 hours. LC-MS analysis confirmed the reaction was complete. The mixture was poured into 300 mL of water and extracted with MTBE (100 mL ⁇ 2).
  • Step 5 To a 25 mL three-necked flask, 10 mL of DCM and compound B24-4 (2 g, 9.89 mmol, 1.0 equiv) were added. Ozone was bubbled into the reaction mixture for 10 minutes. TLC analysis confirmed the reaction was complete. The solution was dried over Na2SO4, filtered, and concentrated under vacuum to afford the crude oil.
  • Step 6 To a 25 mL three-necked flask, 100 mL of THF, (2,3-dimethylphenyl)(1-trityl-4- imidazolyl)methanone (300 mg, 0.68 mmol, 1.0 equiv), compound B24-5 (139 mg, 0.68 mmol, 1.0 equiv), and zinc powder (177 mg, 2.71 mmol, 4.0 equiv) were added.
  • Step 7 To a 100 mL single-necked flask, 8 mL of THF, 8 mL of methanol, compound B24-6 (150 mg, 0.40 mmol, 1.0 equiv), and Pd(OH)2/C (80 mg) were added. The solution was stirred at 40°C under a hydrogen atmosphere for 12 hours. LC-MS analysis confirmed the reaction was complete. The solution was filtered and concentrated under vacuum.
  • Step 8 To a 10 mL single-necked flask, compound B24-7 (72 mg, 0.19 mmol, 1.0 equiv), 0.1 mL of water, 1 mL of ethanol, and NaOH (23 mg, 0.58 mmol, 3.0 equiv) were added. The reaction mixture was stirred at room temperature for 3 hours. LC-MS analysis confirmed the reaction was complete. To the reaction mixture, 0.5 mL of 2 N HCl was added, and the mixture was stirred for 5 minutes.
  • Step 1 A mixture of pyrazole (60 g, 0.881 mol, 1.0 equiv) and 3-chloroperoxybenzoic acid (152.1 g, 0.881 mol, 1.0 equiv) in ethyl acetate (3 L) was stirred at room temperature for 10 days. LC-MS showed the reaction was incomplete. The reaction mixture was concentrated and quenched with saturated sodium sulfite solution.
  • Step 2 A mixture of compound 27-01 (4.5 g, 0.0536 mol, 1.0 equiv), benzyl bromide (9.2 g, 0.0541 mol, 1.01 equiv), and DIPEA (7 g, 0.0541 mol, 1.01 equiv) in DCM (54 mL) was stirred at room temperature for 16 hours. LC-MS showed the reaction was complete.
  • Step 3 To a mixture of compound 27-02 (1 g, 5.74 mmol, 1.0 equiv) in 15 mL of THF was added n-BuLi (2.5 M, 2.5 mL, 6.31 mmol, 1.1 equiv) at -78°C.
  • Step 4 A mixture of compound 27-03 (1.16 g, 1.95 mmol, 1.0 equiv), (2,3- dimethylphenyl)(1-trityl-4-imidazolyl)methanone (1 g, 2.15 mmol, 1.1 equiv), and Pd(PPh3)2Cl2 (68 mg, 0.0975 mmol, 0.05 equiv) in toluene (10 mL) was stirred at 95°C for 16 hours. LC-MS showed the reaction was complete. The reaction mixture was filtered, and the organic solution was concentrated under vacuum.
  • Step 8 To a mixture of compound B27-07 (110 mg, 0.249 mmol, 1.0 equiv) in 2.5 mL of THF, NaH (11 mg, 0.274 mmol, 1.1 equiv) was added at 0°C. The reaction was stirred at room temperature for 0.5 hours under nitrogen, then BPO (6 mg, 0.0249 mmol, 0.1 equiv) was added. The mixture was stirred at room temperature for 5 days. LC-MS showed the reaction was incomplete. The reaction mixture was quenched with ice water (3 mL) and adjusted to pH 6 with 1 M HCl.
  • Step 2 A 50 mL reaction flask containing NaH (407.61 mg, 10.19 mmol) in DMF (5 mL) was cooled to 0°C under a nitrogen atmosphere. Ethyl 2-(diethoxyphosphoryl)propanoate (2.43 g, 10.19 mmol) in DMF (7 mL) was added dropwise. The mixture was stirred at 0°C for 1 hour. A solution of B34-1 (300 mg, 1.0191 mmol) in DMF (3 mL) was added dropwise at 0°C, and the reaction mixture was stirred at 90°C for 2 hours. LC-MS confirmed the reaction was complete.
  • Step 3 A sealed tube containing a solution of B34-2 (350 mg, 0.9247 mmol) in ethanol (10 mL), Pd/C (350 mg, 3.2889 mmol), and Pd(OH)2/C (350 mg, 2.4923 mmol) was evacuated and backfilled with hydrogen three times, then charged with hydrogen (3 MPa). The reaction mixture was stirred at 50°C for 16 hours under a hydrogen atmosphere. LC-MS confirmed the reaction was complete. The mixture was filtered, and the filter cake was washed with ethanol (30 mL). The filtrate was concentrated to dryness to afford B34-3 (300 mg) as a colorless oil. Yield: 78.03%.
  • Step 4 A 10 mL reaction flask containing a solution of B34-3 (200 mg, 0.5228 mmol) in THF (3 mL), H2O (1.5 mL), and methanol (3 mL) was stirred with NaOH (41.82 mg, 1.4056 mmol) at 25°C for 12 hours.
  • LC-MS confirmed the reaction was complete.
  • the solvent was removed, and the mixture was diluted with water (100 mL) and adjusted to pH 4 with 1 M HCl.
  • the mixture was extracted with EA (30 mL), and the organic phase was dried and concentrated.
  • Example 26 Synthesis of Compound B37 O N 3 N O HO OH O Trt C, 0 g, 140.7 mmol), ethane-1,2-diol (17.47 g, 281.4 mmol), and 4-methylbenzenesulfonic acid (2.42 g, 14 mmol) in toluene (200 mL) was heated at 110°C under a nitrogen atmosphere for 4 hours.
  • Step 2 A solution of B37-1 (3.7 g, 19.9 mmol) in THF (30 mL) was cooled to 0°C, and LAH (39.8 mL, 39.8 mmol) was added dropwise.
  • Step 3 A mixture of B37-2 (1 g, 6.3 mmol) in DCM (10 mL) and Dess-Martin periodinane (3.21 g, 7.5 mmol) was stirred at 25°C for 2 hours. TLC showed the reaction was complete.
  • Step 4 A mixture of 4-[(2,3-dimethylphenyl)carbonyl]-1-(triphenylmethyl)imidazole (1700.15 mg, 3.84 mmol), B37-3 (600 mg, 3.84 mmol), and Zn (1507.69 mg, 23.05 mmol) in THF (20 mL) was cooled to 0°C. TiCl4 (4372.16 mg, 23.05 mmol) was added dropwise. The mixture was stirred for 1 hour at 0°C and then heated to 70°C for 3 hours. LC-MS showed the reaction was complete.
  • Step 5 A mixture of TosMIC (208.91 mg, 1.07 mmol) and t-BuOK (350.20 mg, 3.12 mmol) in DMSO (3 mL) was stirred at 25°C for 30 minutes. LC-MS showed the reaction was complete.
  • Step 6 A mixture of B37-5 (120 mg, 0.4118 mmol) in 5 N KOH (5 mL) was heated at 80°C for 16 hours. LC-MS showed the reaction was complete. The mixture was quenched with water (20 mL) and extracted with EtOAc (3 ⁇ 20 mL). The combined organic layers were washed with water and brine, dried with sodium sulfate, and concentrated under vacuum to afford B37-6 (90 mg, 0.2755 mmol) as a white solid. Yield: 67%.
  • Step 7 A mixture of B37-6 (90 mg, 0.29 mmol), Pd/C (92.59 mg, 0.87 mmol), and Pd(OH)2/C (122.17 mg, 0.87 mmol) in methanol (3 mL) was heated at 50°C under a hydrogen atmosphere (3 MPa) for 16 hours in a high-pressure reactor. After cooling to ambient temperature, the mixture was filtered through Celite, and the filtrate was concentrated under vacuum. The residue was purified on a Biotage Isolera One (C18 column, eluting with 10–90% MeCN/H2O containing 0.1% TFA) to provide B37 (33.01 mg, 0.1046 mmol) as a white solid. Yield: 36%.
  • Example 27 Synthesis of Compound B40-A and B40-B Pd(OAc)2,dppp O Py,Tf2O,DCM TEA,CO,DMSO O Pd(PPh3)4,dioxane O DMP,DCM OH rt,2h OTf MeOH,78°C,7h 85°C,16h rt,1H O O ep2 s O step4 O step1 st tep3 Br Br B HO O OH HCl ers1 OH Cl rs2 pyridine (11.87 g, 0.15 mol, 3.0 eq) in DCM (100 mL) was cooled to 0°C.
  • Trifluoromethanesulfonic anhydride (23.97 g, 0.085 mol, 1.7 eq) was added at 0°C, and the mixture was stirred at room temperature for 2 hours under a nitrogen atmosphere. LC-MS analysis confirmed the reaction was complete. The reaction mixture was poured into 100 mL of ice water and extracted with DCM. The organic phase was washed with brine, dried over Na2SO4, and concentrated under vacuum. The residue was purified by silica gel column chromatography to afford compound B40-01 (15.53 g) as a colorless liquid. Yield: 93.6%.
  • Step 2 A mixture of B40-01 (8 g, 0.024 mol, 1.0 eq), TEA (4.62 g, 0.0461 mol, 1.9 eq), Pd(OAc)2 (1.1 g, 0.00482 mmol, 0.2 eq), and dppp (1.88 g, 0.0058 mmol, 0.24 eq) in a 1:1 mixture of DMSO and MeOH (260 mL) was stirred at 78°C under a 5 MPa CO atmosphere for 7 hours. LC-MS analysis confirmed the reaction was complete. The reaction mixture was poured into 1.2 L of ice water and extracted with DCM. The organic phase was dried over Na2SO4 and concentrated under vacuum.
  • Step 3 A mixture of B40-02 (3.357 g, 0.0139 mol, 1.0 eq), (tributylstannyl)methanol (6.68 g, 0.0208 mol, 1.5 eq), and Pd(PPh3)4 (1.2 g, 0.104 mmol, 0.075 eq) in dioxane (40 mL) was stirred at 85°C for 16 hours under a nitrogen atmosphere. LC-MS analysis showed the reaction was 138 Attorney Docket No.: 071741.11025/5WO1 incomplete.
  • Step 4 A mixture of B40-03 (610 mg, 3.14 mmol, 1.0 eq) and Dess-Martin periodinane (DMP) (2 g, 4.71 mmol, 1.5 eq) in DCM (6 mL) was stirred at room temperature for 1 hour under a nitrogen atmosphere. LC-MS analysis confirmed the reaction was complete. The reaction mixture was poured into 6 mL of saturated sodium bicarbonate solution, filtered, and extracted with DCM. The organic phase was dried over Na2SO4 and concentrated under vacuum.
  • DMP Dess-Martin periodinane
  • Step 5 A mixture of B40-04 (496 mg, 2.58 mmol, 1.0 eq), (2,3-dimethylphenyl)(1-trityl- 4-imidazolyl)methanone (1.14 g, 2.58 mmol, 1.0 eq), and Zn (1.26 g, 19.35 mmol, 7.5 eq) in THF (10 mL) was cooled to 0°C, and TiCl4 (1.81 g, 9.55 mmol, 3.7 eq) was added dropwise.
  • Step 6 A mixture of B40-05 (210 mg, 1 eq) and Pd(OH)2 (210 mg, 1.0 wt) in a 1:1 mixture of MeOH and THF (6 mL) was stirred at 45°C under a hydrogen atmosphere overnight.
  • Step 7 A total of 285 mg of B40-06 was separated on a column with dimensions 30 ⁇ 250 mm packed with CHIRALPAK® IB-10 (10 ⁇ m particle size). A mixture of 70% mobile phase A (CO2) and 30% mobile phase B (0.2% ammonia in MeOH/EtOH) was used. The operational conditions were: 35°C, flow rate 3 g/min, detection at 214 nm and 254 nm.
  • Step 8 A mixture of B40-06-P1 (125 mg, 0.345 mmol, 1.0 eq) and NaOH (55 mg, 1.38 mmol, 4.0 eq) in a 2:2:1 mixture of MeOH, THF, and H2O (2 mL) was stirred at 45°C for 16 hours under a nitrogen atmosphere.
  • Step 1 To a 250 mL reaction flask was added 60 mL THF, Zn (6.4 g, 98.74 mmol, 19 eq), cooled to 0°C, slowly added TiCl4 (9.5 g, 49.89 mmol, 9.6 eq) while controlling the temperature at 0-10°C.
  • Step 2 To a 100 mL reaction flask was added 36 mL DMF, B43-03 (1.85 g, 5.03 mmol, 1.0 eq), triphenylmethyl chloride (1.68 g, 6.04 mmol, 1.2 eq), and TEA (1.02 g, 10.07 mmol, 2 eq). The reaction mixture was stirred at 25°C for 15 hours. LC-MS showed the reaction was completed. The mixture was poured into water and extracted with EA.
  • Step 3 To a 50 mL reaction flask was added 20 mL toluene, B43-04 (1 g, 1.64 mmol, 1.0 eq), DIPEA (424 mg, 3.28 mmol, 2 eq), benzyl mercaptan (224 mg, 1.80 mmol, 1.1 eq), Pd2(dba)3 (150 mg, 0.164 mmol, 0.1 eq), and xantphos (190 mg, 0.328 mmol, 0.2 eq).
  • Step 4 To a 25 mL reaction flask was added 6 mL acetic acid and 2 mL water, B43-05 (400 mg, 0.612 mmol, 1 eq). After the mixture was cooled to 0°C, NCS (409 mg, 3.06 mmol, 5 eq) was added. The reaction mixture was stirred at 20°C for 2 hours. LC-MS showed the reaction was completed.
  • Step 5 To a 25 mL three-necked flask was added 5 mL DCM, B43-06 (theoretical yield: 385 mg, 0.612 mmol, 1 eq), TEA (620 mg, 6.12 mmol, 10 eq), and methyl 3-aminopropionate hydrochloride (427 mg, 3.06 mmol, 5 eq). The reaction mixture was stirred at 25°C for 2 hours. LC-MS showed the reaction was completed. The mixture was poured into water and extracted with DCM.
  • Step 6 A mixture of B43-07 (130 mg), Pd(OH)2/C (50 mg), CH3OH (7 mL), and THF (7 mL) was stirred at 40°C for 4 days under a hydrogen atmosphere. The catalyst was removed by filtration, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to afford B43-08 (50 mg). Yield: 58.7%.
  • Step 7 A mixture of B43-08 (50 mg, 0.109 mmol, 1 eq), NaOH (9 mg, 0.219 mmol, 2 eq), CH3OH (4 mL), THF (4 mL), and H2O (2 mL) was stirred at 40°C for 18 hours. LC-MS showed the reaction was completed. The mixture was concentrated under vacuum. Then 10 mL DCM and 5 mL TFA were added, and the solution was stirred for 10 minutes before evaporating the solvent. The residue was purified by preparative HPLC to give B43 (25 mg) as a white solid. Yield: 41.0%. Overall yield: 3.37%.
  • Example 29 Synthesis of Compound B50 O O ydride h eOH , , , , g, 63 mmol, 3 eq), and MeI (6 g, 42 mmol, 2 eq) in DMF (50 mL) was stirred at 30°C overnight under N2. LC-MS showed the reaction was completed. The reaction mixture was diluted with water (100 mL) and extracted with EA (100 mL ⁇ 2). The organic phase was dried over Na2SO4 and concentrated under vacuum. The residue was purified by column chromatography on silica gel to afford B50-01 (5 g). Yield: 95.24%.
  • Step 2 A mixture of B50-01 (5 g, 20 mmol, 1 eq), 2,4-Dimethoxyaniline (3.7 g, 22 mmol, 1.1 eq), and K2CO3 (4 g, 30 mmol, 1.5 eq) in DMF (50 mL) was stirred at 110°C overnight under N2. LC-MS showed the reaction was completed. The reaction mixture was diluted with water (50 mL) and extracted with EA (50 mL ⁇ 2). The organic phase was dried over Na2SO4 and concentrated under vacuum. The residue was purified by column chromatography on silica gel to afford B50-02 (6 g). Yield: 70%.
  • Step 3 A mixture of B50-02 (5 g, 13 mmol, 1 eq) and TFA (40 mL) in DCM (50 mL) was stirred at 30°C for 2 hours. LC-MS showed the reaction was completed. The reaction mixture was concentrated under vacuum. The mixture was diluted with saturated sodium carbonate solution and extracted with DCM (50 mL ⁇ 1). The organic phase was dried over Na2SO4 and concentrated under vacuum. The residue was purified by column chromatography on silica gel to afford B50-03 (3.6 g). Yield: 100%.
  • Step 4 A mixture of B50-03 (3.6 g, 15 mmol, 1 eq) and acetic anhydride (2.2 g, 21 mmol, 1.5 eq) in toluene (40 mL) was stirred at 90°C for 16 hours under N2. LC-MS showed the reaction was completed. The reaction mixture was diluted with saturated sodium bicarbonate solution (20 mL) and extracted with EA (20 mL ⁇ 2). The organic phase was dried over Na2SO4 and concentrated under vacuum. The residue was purified by column chromatography on silica gel to afford B50-04 (3.7 g). Yield: 84.7%.
  • Step 5 A mixture of B50-04 (3.7 g, 12.7 mmol, 1 eq), (tributylstannyl)methanol (6 g, 19 mmol, 1.5 eq), and Pd(PPh3)4 (733 mg, 0.635 mmol, 0.05 eq) in 1,4-dioxane (40 mL) was refluxed at 90°C overnight under N2. LC-MS showed the reaction was incomplete. The mixture was filtered and concentrated under vacuum. The residue was purified by column chromatography on silica gel to afford B50-05 (3.8 g). Yield: 77.6%.
  • Step 6 To a solution of B50-05 (3.8 g, 15.8 mmol, 1 eq) in DCM (25 mL), Dess-Martin periodinane (8 g, 19 mmol, 1.2 eq) was added at 0°C. The reaction mixture was stirred for 2 hours at room temperature. LC-MS showed the reaction was completed. The mixture was filtered and concentrated under vacuum. The residue was purified by column chromatography on silica gel to afford B50-06 (2.4 g). Yield: 63.6%.
  • Step 7 To a solution of B50-06 (2.2 g, 9.2 mmol, 2.4 eq), Zn (1.7 g, 26.6 mmol, 7 eq), and (2,3-Dimethylphenyl)(1-trityl-4-imidazolyl)methanone (1.7 g, 3.8 mmol, 1 eq) in THF (170 mL), titanium tetrachloride (2.5 g, 13.3 mmol, 3.5 eq) was added at 0°C. The reaction mixture was 143 Attorney Docket No.: 071741.11025/5WO1 stirred for 3 hours at 80°C. LC-MS showed the reaction was completed.
  • Step 8 A mixture of B50-07 (800 mg, 1.96 mmol, 1 eq), Pd(OH)2/C (700 mg), TFA (200 mg), and water (10 drops) in MeOH (16 mL) was stirred at 50°C for 16 hours under H2. LC-MS showed the reaction was completed. The mixture was filtered and concentrated under vacuum to afford B50-08 (848 mg) as a crude product.
  • Step 9 A mixture of B50-08 (848 mg, 2.07 mmol, 1 eq) and SOCl2 (400 mg, 4.1 mmol, 2 eq) in MeOH (16 mL) was stirred at 80°C for 16 hours under N2. LC-MS showed the reaction was completed. The reaction mixture was concentrated under vacuum. The mixture was diluted with saturated sodium bicarbonate solution and extracted with EA (20 mL ⁇ 2). The organic phase was dried over Na2SO4 and concentrated under vacuum. The residue was purified by column chromatography on silica gel to afford B50-09 (330 mg). Yield: 41.3%.
  • Step 10 A mixture of B50-09 (200 mg, 0.54 mmol, 1 eq) in 4 M HCl (6 mL) was stirred at 45°C for 0.5 hours. NaNO2 (45 mg, 0.65 mmol, 1.2 eq) in water (0.5 mL) was added at 0°C. The reaction mixture was stirred for 1 hour at 0°C. CuCl (270 mg, 2.7 mmol, 5 eq) in water (0.5 mL) was added at 0°C. The reaction mixture was stirred for 2 hours at 30°C. LC-MS showed the reaction was completed. The mixture was diluted with saturated sodium bicarbonate solution and extracted with EA (20 mL ⁇ 2).
  • Example 30 Synthesis of Compound B53 O O O KSC(S)OEt, acetone DLP, Vinyl pivalate, Dichloroethane DLP, Dichloroethane, CSA B 0-10o C 16h S O 80o Br opiv TFA - bromophenyl)propan-1-one (10 g, 0.0343 mol, 1 eq) was cooled to 0°C. Potassium O-ethyl xanthate (6.05 g, 0.03773 mol, 1.1 eq) was added. The reaction mixture was stirred until the starting material disappeared.
  • Step 2 A 100 mL three-necked flask containing 45 mL dichloroethane, B53-01 (9 g, 0.0271 mol, 1 eq), and vinyl pivalate (6.95 g, 0.0542 mol, 2 eq) was refluxed under nitrogen.
  • Step 3 A 500 mL three-necked flask containing 140 mL dichloroethane, B53-02 (14 g, 0.0271 mol, 1 eq), and camphorsulfonic acid (629 mg, 0.00271 mol, 0.1 eq) was refluxed under nitrogen. Lauroyl peroxide (DLP) was then added to the refluxing solution followed by additional portions (2.16 g, 5.42 mmol, 0.2 eq, every 1-1.5 hours). When TLC monitoring showed that the starting material was consumed (after 1 eq of DLP), the solution was cooled to room temperature. The organic phase was evaporated.
  • DLP Lauroyl peroxide
  • Step 4 A 100 mL three-necked flask containing 19 mL toluene, B53-03 (1.1 g, 4.66 mmol, 1 eq), and p-TsOH ⁇ H2O (3.1 g, 16.3 mmol, 2.9 eq) was refluxed for 4 hours. When the starting material was totally consumed, the mixture was allowed to cool to room temperature, neutralized with saturated Na2CO3, extracted with DCM, dried, and evaporated under reduced pressure. The residue was purified by a fast silica gel column to give 1.1 g of B53-04.
  • Step 5 A 100 mL three-necked flask containing 11 mL DCM, B53-04 (1.1 g, 4.66 mmol, 1 eq), pyridine (1.11 g, 13.98 mmol, 3 eq), and trifluoromethanesulfonic anhydride (Tf2O) (2.23 g, 7.92 mmol, 1.7 eq) was stirred for 1.5 hours at room temperature. TLC showed the reaction was complete. The organic phase was washed with water, and the aqueous phase was extracted with DCM. The organic phases were combined, dried over Na2SO4, and concentrated under reduced pressure.
  • Tf2O trifluoromethanesulfonic anhydride
  • Step 6 A 200 mL autoclave containing 22 mL MeOH, 22 mL DMSO, B53-05 (1.3 g, 3.53 mmol, 1 eq), TEA (679.2 mg, 6.71 mmol, 1.9 eq), dppp (274.75 mg, 0.847 mmol, 0.24 eq), and Pd(OAc)2 (159.92 mg, 0.706 mmol, 0.2 eq) was stirred for 6 hours at 78°C under CO (5 MPa). GC-MS showed the reaction was complete.
  • Step 7 A 25 mL three-necked flask containing 7 mL 1,4-dioxane, B53-06 (683 mg, 2.46 mmol, 1 eq), tri-n-butylstannylmethanol (1.183 g, 3.69 mmol, 1.5 eq), and Pd(PPh3)4 (142.14 mg, 0.123 mmol, 0.05 eq) was stirred for 16 hours at 80°C under nitrogen.
  • Step 8 A 25 mL three-necked flask containing 7.5 mL DCM, B53-07 (370 mg, 1.61 mmol, 1 eq), and MnO2 (1.4 g, 16.1 mmol, 10 eq) was stirred for 16 hours at 45°C. The mixture was filtered and evaporated to dryness.
  • Step 9 A 50 mL three-necked flask containing 14 mL THF and powdered Zn (763 mg, 11.67 mmol, 19 eq) was cooled to 0°C under nitrogen. TiCl4 (1.12 g, 5.89 mmol, 9.6 eq) was added dropwise, and the mixture was stirred for 1 hour at 70°C. The mixture was cooled to 0°C.
  • Step 10 A 10 mL single-necked flask containing 1 mL MeOH, 4 drops of water, 4 drops of TFA, B53-09 (100 mg, 0.252 mmol, 1 eq), and 10% Pd/C (100 mg) was stirred for 6 hours at 50°C under a hydrogen atmosphere. LC-MS showed the reaction was complete. The mixture was evaporated to dryness to give 154 mg of B53-10. Yield: 100%.
  • ⁇ 2AAR FLIPR assay This experimental protocol involved cell seeding and a FLIPR assay using the ⁇ 2AAR ( ⁇ 2A-adrenergic receptor) cell line hosted in HEK293 cells.
  • the growth media used is DMEM (11965-092, Gibco) supplemented with 10% FBS (FSP500, Excell), 300 ⁇ g/mL G418 (10131-027, Gibco), and 2 ⁇ g/mL Blasticidin S HCl (BS) (A11139-03, Gibco).
  • FBS FSP500, Excell
  • BS Blasticidin S HCl
  • Cells were then treated with 0.05% EDTA-Trypsin (25300-062, Gibco), incubated at 37°C for 1-2 minutes, and monitored under an inverted microscope. The cells were detached, resuspended in growth media, and centrifuged at room temperature at 1000 rpm for 5 minutes. After discarding the supernatant, the cell pellet was resuspended in growth media to a concentration of 10 ⁇ 10 5 cells per mL. This suspension was added to 384-well plates (19-Jul-38, Greiner) at 20 ⁇ L per well and incubated overnight at 37°C in 5% CO 2 .
  • EDTA-Trypsin 25300-062, Gibco
  • the FLIPR assay began with the preparation of the assay buffer comprising 20 mM HEPES (15630-106, Invitrogen), 1 ⁇ HBSS (14025-076, Invitrogen), and 0.5% BSA (B2064, Sigma). A 250 mM Probenecid solution was prepared in this buffer.
  • the Fluo-4 DirectTM Loading Buffer was made by dissolving Fluo-4 DirectTM crystals (F10471, Invitrogen) in the FLIPR Assay Buffer and adding Probenecid. The buffer was then vortexed and allowed to stand for over 5 minutes, shielded from light.
  • testing compounds for agonist activite were serially diluted and transferred to a 384-well compound plate (25-Jan-39, Greiner). The cell plate was then treated with 2 ⁇ Fluo-4 DirectTM loading buffer and incubated for 50 minutes at 37 oC in a 5% CO2 atmosphere, followed by 10 minutes at room temperature. Subsequently, the FLIPR assay buffer was added to the compound plate, which is then centrifuged. 169 Attorney Docket No.: 071741.11025/5WO1 [0702] The cell plate was analyzed in the FLIPR Tetra+ System for fluorescence signals. For the agonist test, reference compounds were added to the cell plates, and fluorescence was measured.
  • the “Max-Min” calculation began from Read 1 to the maximum allowed. The data were analyzed using Prism software to calculate activation percentage for agonists and inhibition percentage for antagonists. The results were then fitted using specific models to determine EC50 for agonists.
  • the experimental protocol utilized various reagents and apparatus, including Penicillin/Streptomycin (100x) (SV30010, Hyclone), Poly-L-lysine hydrobromide (P1399, Sigma), and different types of 384-well plates such as the 384-Well PP 2.0 Microplate (PP-0200, LABCYTE) and 384 well Low Dead Volume Microplate (LP-0200, LABCYTE). The use of specific reference compounds like UK14304 was also integral to the assay. [0704] Example 2.
  • ⁇ 2AAR binding assay [0705] The ⁇ 2AR Binding Assay was conducted using a stable HEK293 cell line, specifically constructed by WuXi AppTec for targeting ⁇ 2AAR. This assay primarily focused on the binding activity of the radioligand [3H]-RX 821002 (PerkinElmer, NET1153250UC) to ⁇ 2AAR, with the membrane concentration set at 0.5 ⁇ g/well and the radioligand concentration at 0.5 nM.
  • Essential equipment for this assay includes Unifilter-96 GF/C filter plates (Perkin Elmer, 6005174), 96 well conical polypropylene plates (Agilent, 5042-1385), TopSeal-A sealing film (Perkin Elmer, 6050185), a MicroBeta2 reader (CNLL0153, Perkin Elemer, 1310887), and a cell harvester (UNIFILTER-96, Perkin Elemer, 1951369), all procured from Perkin Elmer. Both the assay and wash buffers consist of 50 mM Tris-HCl at a pH of 7.4 (Tris base, Sigma, T1503-1KG).
  • the plates were dried at 50 °C for one hour.
  • the next step involved sealing the bottom of the filter plate wells with Perkin Elmer Unifilter-96 backing seal tape and adding 50 ⁇ L of MicroScint-O cocktail (PerkinElmer, 6013611) to each well.
  • the top of the plates was then sealed with TopSeal-A sealing 170 Attorney Docket No.: 071741.11025/5WO1 film.
  • the trapped 3H was quantified using a Perkin Elmer MicroBeta2 Reader.
  • MDR1-MDCK Permeability Assay [0711] MDR1-MDCK II cells (obtained from Piet Borst at the Netherlands Cancer Institute) were seeded onto Polycarbonate membranes (PC) in 96-well insert systems at 3.33 x 10 5 cells/ mL until to 4-7 days for confluent cell monolayer formation. Selected ⁇ 2AR agonist from Table 3 and Table 4 were diluted with the transport buffer (HBSS with 10.0 mM Hepes, pH7.4) from DMSO stock solution to a concentration of 2 ⁇ M (DMSO ⁇ 1%) and applied to the apical or basolateral side of the cell monolayer.
  • transport buffer HBSS with 10.0 mM Hepes, pH7.4
  • Digoxin was used as a positive control for the P-glycoprotein (P-gp) substrate, while clonidine, dexmedetomidine, faldomidine and brimonidine were used as negative control.
  • Permeation of the test compounds from A to B direction and/or B to A direction was determined in duplicate.
  • Digoxin was tested at 10.0 ⁇ M from A to B direction and B to A direction in duplicate. The plate was incubated for 2.5 hours in CO2 incubator at 37.0 ⁇ 1.0°C, with 5.0% CO 2 at saturated humidity without shaking. In addition, the efflux ratio of each compound was also determined. Test and reference compounds were quantified by LC/MS/MS analysis based on the peak area ratio of analyte/IS.
  • lucifer yellow rejection assay was applied to determine the cell monolayer integrity. Buffers were removed from both apical and basolateral chambers, followed by the addition of 75 ⁇ L of 100 ⁇ M lucifer yellow in transport buffer and 250 ⁇ L transport buffer in apical and basolateral chambers, respectively. The plate was incubated for 30 minutes at 37.0°C 176 Attorney Docket No.: 071741.11025/5WO1 with 5.0% CO2 and 95.0% relative humidity without shaking. After 30 minutes incubation, 20 ⁇ L of lucifer yellow samples were taken from the apical sides, followed by the addition of 60 ⁇ L of transport Buffer.
  • the apparent permeability coefficient Papp (dCr/dt) ⁇ Vr / (A x C0) wherein dCr/dt is the cumulative concentration of compound in the receiver chamber as a function of time ( ⁇ M/s); Vr is the solution volume in the receiver chamber (0.075 mL on the apical side, 0.25 mL on the basolateral side); A is the surface area for the transport, i.e.0.143 cm2 for the area of the monolayer; and C0 is the initial concentration in the donor chamber ( ⁇ M).
  • the binding affinity of various compounds to plasma proteins was evaluated, including clonidine HCl, dexmedetomidine HCl, 1-B HCl, and 44-B HCl, with warfarin serving as a control.
  • the experiment utilized a HT-Dialysis plate (HTD 96 b) and a dialysis membrane with a molecular weight cutoff of 12-14 kDa.
  • the plasma was derived from male C57BL/6J mice, treated with EDTA-K2 as an anticoagulant.
  • the experimental procedure commenced with plasma thawing under cold tap water, followed by centrifugation at 3220 ⁇ g for 5 minutes to eliminate clots, and pH adjustment to 7.4 ⁇ 0.1.
  • Dialysis membranes were initially hydrated in ultra-pure water for about one hour and then treated in a 20:80 ethanol-water mixture for 20 minutes. These prepared membranes could be used immediately or stored at 2-8°C for up to a month. Membranes were rinsed in ultra-pure water before use.
  • Test and control compounds were prepared at a 400 ⁇ M concentration by diluting stock solutions with DMSO. Working solutions were further diluted to create 2 ⁇ M loading matrix solutions, which were thoroughly mixed. In the assay, 50 ⁇ L aliquots of these solutions were dispensed in triplicate into a Sample Collection Plate, balanced with blank PBS to a final volume of 100 ⁇ L per well.
  • a stop solution containing acetonitrile, tolbutamide, and labetalol was added, and samples were mixed and cooled at 2 to 8°C.
  • 100 ⁇ L aliquots from the loading matrix were placed in the dialysis well's donor side, matched with an equal volume of PBS on the receiver side, and incubated at 37°C for 4 hours.
  • Post-dialysis samples from both sides were collected, balanced to 100 ⁇ L with corresponding blank fluids, treated with stop solution, vortexed, and centrifuged to prepare for LC- MS/MS analysis.
  • Data analysis involved calculating the percentages of Unbound, Bound, and Recovery of the compounds post-dialysis.
  • %Unbound was calculated as the ratio of the compound's peak area 178 Attorney Docket No.: 071741.11025/5WO1 on the receiver side to its internal standard, reflecting the fraction that crossed the membrane.
  • %Bound was the complement of %Unbound, representing the fraction retained on the donor side.
  • %Recovery was determined from the peak area ratios on both sides of the membrane, assessing the dialysis efficiency in retaining the compound.
  • the initial preparation of the dialysis membrane involved thawing brain homogenate in a water bath at room temperature and subsequently heating it at 37°C for 10 minutes.
  • the dialysis setup utilized was from HT Dialysis LLC, featuring a HT-Dialysis plate (Model HTD 96 b) and a dialysis membrane with a molecular weight cutoff of 12-14 kDa.
  • the membrane underwent a comprehensive pretreatment which included hydration in ultra-pure water at room temperature for approximately one hour. This was followed by separation and immersion in a 20:80 ethanol:water solution for about 20 minutes. After this treatment, the membranes were either used immediately or stored at 2-8°C for up to one month, with a final rinse in ultra-pure water prior to experimental use.
  • test and control substances were first dissolved to create 400 ⁇ M working solutions by mixing 4 ⁇ L of stock solution with 96 ⁇ L of DMSO. These working solutions were then further diluted to 2 ⁇ M in a blank matrix by combining 3 ⁇ L of the prepared solution with 597 ⁇ L of matrix, ensuring thorough mixing.
  • 50 ⁇ L aliquots of the 2 ⁇ M compound-matrix mixture were dispensed in triplicate into a Sample Collection Plate. Each aliquot was paired with an equal volume of blank 179 Attorney Docket No.: 071741.11025/5WO1 PBS to standardize the total volume to 100 ⁇ L per well at a 1:1 matrix to PBS ratio.
  • a stop solution comprising 500 ⁇ L of acetonitrile with tolbutamide and labetalol at 250nM each was added to stabilize the samples at T0. The samples were then shaken at 800 rpm for 10 minutes and stored at 2-8°C.
  • the dialysis procedure included assembling the dialysis device according to the manufacturer's specifications, loading the matrix aliquots into the donor side of the dialysis wells, and conducting the dialysis under a humidified atmosphere with 5% CO 2 at 37°C for 4 hours.
  • Post-dialysis 50 ⁇ L samples were collected from both the receiver and donor sides into new 96-well plates. Volumes were adjusted to 100 ⁇ L by adding an equivalent amount of the opposite blank matrix or PBS.
  • %Undiluted Unbound 100 ⁇ 1/D / ((1 / (F/T) - 1) + 1/D), where D is the dilution factor (10).
  • %Undiluted Bound was derived as 100 - %Undiluted Unbound.
  • %Recovery 100 ⁇ (F + T) / T0, with F and T representing the peak area ratios of the compound to the internal standard on the receiver and donor sides respectively, after 4 hours of incubation.
  • the brain protein binding result is shown in Table 7.
  • mice were acclimated to the test facility for at least 3 days. During this period, their general health was assessed by veterinary staff or other authorized personnel. The mice were housed in groups of up to four per cage in polysulfone cages, using either certified aspen shaving bedding or corncob bedding. This bedding was regularly tested for environmental contaminants by the manufacturer. The facility's environment was carefully controlled to maintain a temperature range of 20-26°C, relative humidity between 40 to 70%, and a 12-hour light/12-hour dark cycle, although this cycle can be interrupted for study-related activities. Temperature and humidity were continuously monitored by the Vawasala ViewLinc Monitoring system.
  • Compounds such as 5 mg/kg Clonidine HCl, 5 mg/kg dexmedetomidine HCl, 5 mg/kg and 80 mg/kg compound 1-B, and 5 mg/kg and 80 mg/kg compound 44-B were administered in a 20% HP- ⁇ -CD solution in water, with sample collections scheduled at 0.5, 1, 2, and 8 hours post-dosing.
  • Blood collections were performed from the saphenous vein or another suitable site, with approximately 0.1 mL collected per time point into pre-chilled commercial EDTA-K2 tubes. The samples were kept on wet ice until centrifugation at 4°C and 3,200 g for 10 minutes.
  • the plasma was then transferred into pre-labeled 96-well plates or polypropylene tubes, quick-frozen over dry ice, and stored at -60°C or lower until LC-MS/MS analysis.
  • Table 8 The in vivo drug distribution result is shown in Table 8. [0739] Table 8: the in vivo drug distribution Brain Spinal cord Kp, uu, Compounds dosage logBB logSB /plasma, Kp /plasma brain Clonidine HCl 5 mg/kg 2.27 1.78 0.356 0.252 0.739 Dexmedetomidine HCl 5 mg/kg 1.26 1.14 0.101 0.133 1.17 5 mg/kg below detection limit* 1-B HCl 80 mg/kg 0.0598 0.0153 -1.22 -0.732 0.032 5 mg/kg below detection limit* 44-B HCl 80 mg/kg 0.0334 0.0511 -1.39 -1.29 0.013 * drug in brain and spinal cord is below the detection limit (DL).
  • DL detection limit
  • the animals were anesthetized with Zoletil 50 (50 mg/kg, 2.5 mL/kg, i.p.) and Xylazine Hydrochloride (8 mg/kg, 2.5 mL/kg, i.p.), with a toe pinch used to ensure full anesthesia before incision, and ophthalmic ointment applied to the rodents' eyes to prevent drying of the corneas.
  • Zoletil 50 50 mg/kg, 2.5 mL/kg, i.p.
  • Xylazine Hydrochloride 8 mg/kg, 2.5 mL/kg, i.p.
  • mice were individually placed in plastic enclosures with mesh bottoms, allowing full paw access. For three consecutive days, mice were acclimated for 15 minutes each day. Mechanical allodynia baseline measurements were performed on day 14. Animals not exhibiting allodynia (PWT>0.6 g) were excluded, leaving 24 qualified animals (PWT ⁇ 0.6 g) who were then randomly divided into three groups based on their baseline PWT, in addition to 6 sham mice forming a Sham group, totaling four groups with 6-8 mice each.
  • the administration route for the therapeutic intervention for compounds 1-B with a dosage from 1 mg/mL to 20 mg/mL, and 10-B, 44-B, 45-B, 46-B, 47-B, 121, 136, 118, 156 and 175 was oral (p.o.) with a dosage of 1 mg/mL, while the ones for 1 mg/kg morphine via s.c. and 3mg/kg pregabalin via p.o. as positive control, which were prepared in a 20% HP- ⁇ -CD solution.
  • mice 1-B, 10-B, 44-B 45-B, 46-B and 47-B are the active enantiomers of 1, 10, 44, 45, 46, 47, respectively, while 121, 136, 118, and 156 are racemate.
  • the solution was vortexed to ensure thorough mixing until homogeneous.
  • the dosage administered to the mice was 10 ml/kg.
  • Mechanical allodynia tests were conducted on the left hind paw of mice, which were individually placed in plastic enclosures with mesh bottoms for full paw access and acclimated for 15 minutes prior to testing.
  • the mid-plantar hind paw was probed using a series of eight Von Frey filaments with logarithmically incremental stiffness: 0.02 g (2.36), 0.04 g (2.44), 0.07 g (2.83), 0.16 g (3.22), 0.4 g (3.61), 0.6 g (3.84), 1 g (4.08), and 1.4 g (4.17).
  • the filaments were applied perpendicularly to the paw's plantar surface with enough force to slightly buckle against it, maintaining contact for 6-8 seconds. Tests were spaced by 5-second intervals to ensure clear resolution of any response to the prior stimulus, with a sharp withdrawal or flinching upon filament removal indicating a positive response.
  • EB indicates the analgesic ratio of the other analgesia drugs, such as morphine, pregabalin, suzetrigine, duloxetine, amitriptyline, and ketorolac.
  • EAB denotes the analgesic ratio resulting from a combination treatment involving compound 1-B or compound 44-B and one of the following drugs: morphine, pregabalin, suzetrigine, duloxetine, amitriptyline, and ketorolac.
  • mice Male C3H/He mice were anesthetized with a combination of Zoletil 50 (50 mg/kg) and Xylazine 185 Attorney Docket No.: 071741.11025/5WO1 Hydrochloride (8 mg/kg) administered via intraperitoneal injection, and positioned supinely. The right hind limb was shaved and sterilized. A minimal incision was made on the right hind leg to sever the patellar ligaments and expose the condyles of the distal femur. The proximal femur was perforated using a 0.3 mL syringe needle.
  • the remaining qualified animals were then randomly assigned into four groups based on their baseline PWT values.
  • the animals received a single injection of test compounds, including pregabalin 3mg/kg p.o., morphine 1mg/kg s.c., 44-B 1mg/kg p.o., as well as a group for 1-B 20mg/kg p.o. and 44-B 20mg/kg p.o. at a dose of 10 mL/kg based on body weight, and mechanical allodynia tests were performed at various time points post- administration as dictated by different experimental requirements with sham group and vehicle group.
  • the plantar aspect of the left hind paw was cleansed with three rounds of alternating Betadine and 70% ethanol applications, allowing the surface to air-dry. A 0.5-mm longitudinal incision was then made through the skin and fascia from 2 mm proximal to the heel towards the toes. The plantar muscle was longitudinally incised while preserving the origin and insertion points. Hemostasis was achieved with gentle pressure, and the skin was closed with two mattress sutures. Post-surgery, all surgical instruments were cleaned and re-sterilized using a glass bead sterilizer.
  • the objective of this study is to evaluate the in vivo efficacy study of test srticles in the Treatment of Subcutaneous Colorectal Cancer Syngeneic Model MC38 in Female C57BL6/J mice.
  • the mice are Mus musculus C57BL6/J, female, supplied by Beijing HFK Bioscience Co. LTD, with an average age of 6-8 weeks.
  • the cage is polysulfone IVC cage, with a temperature 20-26°C and humidity 40 – 70%.
  • the light cycle is 12 hours light and 12 hours dark.
  • mice is feed by a diet of standard rodent chow, irradiated, ad libitum.
  • the water is autoclaved filtered RO (reverse osmosis) softened, filtered water, ad libitum.
  • RO reverse osmosis
  • the MC38 cancer cells were maintained in vitro with DMEM medium supplemented with 10% fetal bovine serum and 50 ⁇ g/mL Hygromycin B at 37oC in an atmosphere of 5% CO 2 in air. The cells in exponential growth phase were harvested and quantitated by cell counter before tumor inoculation. Each mouse was inoculated subcutaneously at the right rear flank region with MC38 tumor cells (1x 10 6 ) in 0.1 mL of PBS mixed with PBS for tumor development. The randomization started when the mean tumor size reached approximately 121.36 mm 3 .30 mice were enrolled in the study. All animals were randomly allocated to 5 study groups, 6 mice in each group.
  • Randomization was performed based on “Matched distribution” method. The date of randomization was denoted as day 0. [0771] The treatment was initiated on the same day of randomization (day 0) per study design. After tumor cells inoculation, the animals were checked daily for morbidity and mortality. During routine monitoring, the animals were checked for any effects of tumor growth and treatments on behavior such as mobility, food and water consumption, body weight gain/loss (Body weights were measured twice per week after randomization), eye/hair matting and any other abnormalities. Mortality and observed clinical signs were recorded for individual animals in detail.
  • Dosing as well as tumor and body weight measurements 189 Attorney Docket No.: 071741.11025/5WO1 were conducted in a Laminar Flow Cabinet. The body weights and tumor volumes were measured by using StudyDirectorTM software (version 3.1.399.19). [0772] The body weights of all animals were monitored throughout the study and animals were euthanized if they lost over 20% of their body weight relative to the weight on the day of randomization.
  • the individual mouse was euthanized if its tumor volume exceeds 3000 mm3.
  • any animal exhibiting an ulcerated or necrotic tumor were separated immediately and singly housed and monitored daily before the animal was euthanized or until tumor regression was completed.
  • the mouse was euthanized rapidly if a) tumor ulcerates, and the ulceration diameter was greater than 5 mm, or pus or necrosis observed, and b) tumor burden, including metastasis, compromises animal’s normal physiologic performances, e.g., orientation, access to food or water, etc.
  • the body weight between randomization grouping is shown in FIG.5A.
  • the tumor growth of each treatment group and control group is shown in FIG.5B.
  • TGI tumor growth inhibition
  • T/C tumor growth inhibition
  • Table 12 Pharmacodynamic analysis of each group in Subcutaneous Colorectal Cancer Syngeneic Model MC38 D17 Group Tumor volume (mm 3 ) M ) TGI (% P Value ( mean ⁇ SE ) T/C (%) (Compared with control group) G1 2478.07 ⁇ 439.10 -- -- -- G2 766.52 ⁇ 147.17 72.62% 27.38 ⁇ 0.05 G3 1009.16 ⁇ 258.31 62.32% 37.68 ⁇ 0.05 G4 996.54 ⁇ 270.05 62.86% 37.14 ⁇ 0.05 [0775] Another in vivo efficacy study of clonidine, 44-B and their combination therapy with Anti-mPD-1 in the treatment of subcutaneous colorectal cancer syngeneic model MC38 in female C57BL6/J mice was carried out with same protocol but different treatment design.10mg/kg Anti- mPD-1 i.p.
  • BIWx5 does, 2mg/kg clonidine p.o. QDx15days, and 2mg/mL 44-B HCl p.o. 190 Attorney Docket No.: 071741.11025/5WO1 QD ⁇ 15days were administrated as the single therapy, while the 10mg/kg Anti-mPD-1 i.p. BIWx5 does and 2mg/kg clonidine p.o. QD ⁇ 15days, as well as the combination of 10mg/kg Anti-mPD-1 i.p. BIWx5 does and 5mg/kg 44-B p.o. QDx15days were administrated as the combination therapy.
  • the experiment design is in Table 13. [0776] Table 13.
  • Dosing Volume ROA Dosing Frequency & mice (mg/kg) (mg/ml) ( ⁇ L/g) Duration 1 6 Vehicle -- -- * 2 6 Anti-mPD-1 10 1 3 6 Clonidine 2 0.2 p. . y 4 6 44-B HCl 2 0.2 10 p.o. QD*15days 5 6 44-B HCl 5 0.5 10 p.o. QD*15days 6 6 Clonidine 2 0.2 10 p.o. QD*15days Anti-mPD-1 10 1 10 i.p. BIW ⁇ 5 doses 7 6 44-B HCl 2 0.2 10 p.o.
  • the TGI% (1/T/C) ⁇ 100%.
  • EA represents the TGI of compound 44-B.
  • EB indicates the TGI of An notes the analgesic ratio resulting from a combined treatment involving compound 44-B and Anti-mPD-1.
  • the data is shown in Table 14.
  • the combination group of 10mg/kg Anti-mPD-1 i.p. BIW ⁇ 5 does and 5mg/kg 44-B p.o.
  • mice Spontaneous Locomotor Activity in Mice
  • the study evaluated the effects of clonidine, brimonidine tartrate, compound 1-B HCl, compound 44-B HCl on spontaneous locomotor activity in male C57BL/6 mice. Initially, mice were acclimatized to the testing environment for 8 hours the day before the experiment, followed by at least 2 hours of habituation on the day of the test. The mice were then grouped randomly based on their body weight into six per group, ensuring a balanced distribution for the administration of the drug, which was dissolved in 20% HP- ⁇ -CD in water.
  • clonidine at a dose of 1mg/kg and compound 1-B HCl at concentrations of 1mg/kg, 10mg/kg, and 20mg/kg were freshly prepared and administered orally at a volume of 10mL/kg.
  • clonidine at a dose of 1mg/kg, brimonidine tartrate at a dose of 1mg/kg and compound 44-B HCl at a dose of 1mg/kg were freshly prepared and administered orally at a volume of 10mL/kg.
  • the locomotor activity was monitored by placing the mice in the center of a test box, with a video tracking system measuring the distance traveled every 5 minutes for 60 minutes.
  • mice were randomly assigned to groups based on their body weight to ensure homogeneity across the groups in terms of weight before any treatment was administered.
  • Rotarod training occurred two days prior to the testing phase.
  • the mice On the first training day, the mice underwent three trials on the rotarod at a speed of 6 rpm, each lasting 120 seconds, with 30-minute intervals between trials. If a mouse fell off before completing 120 seconds, it was immediately placed back on the rotarod to complete the training duration.
  • the training consisted of a single trial at the same speed of 6 rpm but extended to 300 seconds. Mice that fell before the 300-second mark were similarly returned to the rotarod to ensure they reached the full training time.
  • mice On the test day, treatments were administered orally to the mice at a dosage volume of 10 mL/kg based on their body weight. The treatments included a vehicle, clonidine (1 mg/kg), and 44- B HCl at three dosages (1 mg/kg, 10 mg/kg, and 20 mg/kg). The time of compound administration was designated as time zero.
  • the rotarod test was conducted at 30, 60, and 120 minutes post-administration, with each session lasting 300 seconds at a speed of 6 rpm. The primary measure was the latency time until a mouse fell from the rotarod, which served as an indicator of the compounds' effects on motor function. [0787] Data were recorded in Microsoft Excel and subsequently analyzed using GraphPad Prism.

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Abstract

La présente invention concerne de nouveaux agonistes du récepteur adrénergique alpha-2 (α2AR) et leurs utilisations. En particulier, la présente divulgation concerne des composés contenant de l'imidazole, en particulier, de formule (I-A), de formule (I-B), de formule (I-C), de formule (I-D) ou de formule (II). Ces composés peuvent être utiles en tant qu'agonistes de α2AR sélectifs en périphérie pour le traitement ou la prévention de maladies liées à ceux-ci, utilisés seuls ou en combinaison avec un second agent thérapeutique.
PCT/US2025/027331 2024-05-01 2025-05-01 Combinaison comprenant un agoniste du récepteur adrénergique alpha-2 Pending WO2025231265A1 (fr)

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