WO2008011478A2 - Methods for treating chronic pain using 3-aryl-3-hydroxy-2-amino-propionic acid amides, 3-heteroaryl-3-hydroxy-2-amino-propionic acid amides and related compounds - Google Patents

Methods for treating chronic pain using 3-aryl-3-hydroxy-2-amino-propionic acid amides, 3-heteroaryl-3-hydroxy-2-amino-propionic acid amides and related compounds Download PDF

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WO2008011478A2
WO2008011478A2 PCT/US2007/073806 US2007073806W WO2008011478A2 WO 2008011478 A2 WO2008011478 A2 WO 2008011478A2 US 2007073806 W US2007073806 W US 2007073806W WO 2008011478 A2 WO2008011478 A2 WO 2008011478A2
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compound
carbons
alkyl
pain
groups
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PCT/US2007/073806
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French (fr)
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WO2008011478A3 (en
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John E. Donello
Fabien J. Schweighoffer
Bertrand Leblond
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Allergan, Inc.
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    • 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
    • A61K31/40—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with one nitrogen as the only ring hetero atom, e.g. sulpiride, succinimide, tolmetin, buflomedil
    • 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
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/4409—Non condensed pyridines; Hydrogenated derivatives thereof only substituted in position 4, e.g. isoniazid, iproniazid
    • 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
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/445—Non condensed piperidines, e.g. piperocaine
    • 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
    • A61K31/535—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with at least one nitrogen and one oxygen as the ring hetero atoms, e.g. 1,2-oxazines
    • A61K31/5355—Non-condensed oxazines and containing further heterocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]

Definitions

  • the present invention is directed to methods of treating a patient suffering from one or more types of chronic pain using using derivatives of 3- aryl-3-hydroxy-2-amino-propionic acid amides, 3-heteroaryl-3-hydroxy-2- amino-propionic acid amides, and related compounds.
  • United States Patent Application Publication Nos. 2003/0153768 and 2003/0050299 disclose several examples of the above- mentioned known compounds.
  • the ⁇ /-acyl compounds of these references are said to be useful as ⁇ /-acylsphingosine glucosyltransferase inhibitors, the amide and the reduced compounds are described as intermediates in their preparations.
  • Illustrative specific examples of compounds of these references are shown below:
  • L-f/?reo-PDMP and some other known compounds used in the methods of this invention are commercially available, in pure enantiomeric and racemic forms, as applicable, from Matreya, LLC Pleasant Gap, Pennsylvania.
  • Serija (1965) (4), 474-7 disclose the following compound.
  • the present invention is directed to methods of treating a patient suffering from one or more types of chronic pain using compounds of Formula 1
  • R 2 is H, alkyl of 1 to 6 carbons or the Ri and R 2 groups together with the nitrogen form a saturated or unsaturated 4, 5, 6 or 7 membered ring that optionally includes one or two heteroatoms independently selected from N, O and S, said 4, 5, 6 or 7 membered ring optionally being substituted with one or two COOH, CH 2 OH, OH, B(OH) 2 , cyano or halogen groups or with one or two alkyl groups having 1 to 6 carbons, or one or two carbons of said rings being attached to an oxygen to form keto groups and said 4, 5, 6 or 7 membered ring optionally being condensed with an aromatic or non-aromatic 5 or 6 membered ring that optionally includes 1 or heteroatoms selected from N, O and S;
  • R 3 is independently selected from H, alkyl of 1 to 20 carbons, cycloalkyl of 3 to 6 carbons, aryl or heteroaryl, aryl-alkyl, aryl-(hydroxy)alkyl, heteroaryl-alkyl or hetero-(hydroxy)alkyl where the alkyl moiety has 1 to 4 carbons, said aryl or heteroaryl groups being optionally substituted with 1 to 3 groups independently selected from the group consisting of halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 6 carbons, or R 3 is CO- R 7 , SO 2 R 7 or CO-O-R 7 where R 7 is H, alkyl of 1 to 1 to 20 carbons, alkyl of 1 to 20 carbons substituted with and NH 2 group or with an NH-COalkyl group where the alkyl group has one to 6 carbons, aryl or heteroaryl, aryl-alkyl or heteroaryl-alkyl where the
  • R5 and R ⁇ are independently selected from H, halogen, alkyl of 1 to 6 carbons, halogen substituted alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 6 carbons, phenyl, or
  • R5 and R ⁇ together with the atoms to which they are attached jointly form a carbocyclic or a heterocyclic ring, the carbocyclic ring having 5 or 6 atoms in the ring, the heterocyclic ring having 5 or 6 atoms in the ring and 1 to 3 heteroatoms independently selected from N, O and S; said carbocyclic or heterocyclic ring jointly formed by R5 and R& being optionally substituted with 1 to 6 Rg groups where Rg is independently selected from halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 6 carbons or a pharmaceutically acceptable salt of said compound with the proviso that Formula 1 does not cover compounds where R 4 is H, Ri and R 2 jointly with the nitrogen form a pyrrolidino or morpholino ring, the sum of m, n and q is 3, and none of W, X and Y represent a heteroatom with the further proviso that the formula does not cover the compounds of the formula
  • the present invention is also directed to methods of treating types of pain using the compounds of Formula 2
  • Ri is H or alkyl of 1 to 6 carbons
  • R 2 is H, alkyl of 1 to 6 carbons or the Ri and R 2 groups together with the nitrogen form a saturated or unsaturated 4, 5, 6 or 7 membered ring that optionally includes one or two heteroatoms independently selected from N, O and S, said 4, 5, 6 or 7 membered ring optionally being substituted with one or two COOH, CH 2 OH, OH, B(OH) 2 , cyano or halogen groups or with one or two alkyl groups having 1 to 6 carbons, or one or two carbons of said rings being attached to an oxygen to form keto groups and said 4, 5, 6 or 7 membered ring optionally being condensed with an aromatic or non-aromatic 5 or 6 membered ring that optionally includes 1 or heteroatoms selected from N, O and S; R 3 is independently selected from H, alkyl of 1 to 20 carbons, cycloalkyl of 3 to 6 carbons, aryl or heteroaryl, aryl-alkyl, aryl-(hydroxy)alkyl, heteroary
  • R-io is not hydrogen;
  • Rn is H, alkyl of 1 to 6 carbons or CO-R 12 where R 12 is alkyl of 1 to 6 carbons;
  • m, n and q are integers independently selected from O, 1 , 2 or 3 with the proviso that the sum of m, n and q is 2 or 3;
  • s is zero (O) or when X is N then s is zero (O) or 1 ;
  • W, X and Y independently represent a CH, CR 5 , CR ⁇ or a heteroatom selected independently of N, O and S, and
  • R5 and R ⁇ are independently selected from H, halogen, alkyl of 1 to 6 carbons, halogen substituted alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 6 carbons, phenyl, or
  • R5 and R ⁇ together with the atoms to which they are attached jointly form a carbocyclic or a heterocyclic ring, the carbocyclic ring having 5 or 6 atoms in the ring, the heterocyclic ring having 5 or 6 atoms in the ring and 1 to 3 heteroatoms independently selected from N, O and S; said carbocyclic or heterocyclic ring jointly formed by R5 and Re being optionally substituted with 1 to 6 Rg groups where Rg is independently selected from halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carb ' ons and thioxy of 1 to 6 carbons or a pharmaceutically acceptable salt of said compound.
  • Any of the compounds described here may be used to treat a patient suffering from one or more types of chronic pain including neuropathic pain, inflammatory pain, headache pain, somatic pain, visceral pain, and referred pain.
  • Most compounds that are useful in the method of the invention contain one or more asymmetric centers, such that the compounds may exist in enantiomeric as well as in diastereomeric forms.
  • most of the compounds of the present invention have two asymmetric carbons adjacent to one another and therefore can exist in erythro or threo form, with each of these two forms having dextrorotatory (D) or levorotary (L) enantiomers.
  • D dextrorotatory
  • L levorotary
  • the threo form is generally preferred in accordance with the present invention for analgesic activity, unless it is specifically noted otherwise, the scope of the present invention includes all enantiomers, diastereomers and diastereomeric or racemic mixtures.
  • the designation "DL” or “(+/-)” or “( ⁇ )” in this application includes the pure dextrorotatory enantiomer, the pure levorotatory enantiomer and all racemic mixtures, including mixtures where the two enantiomers are present in equal or in unequal proportions.
  • only one of the enantiomers is actually shown but when the designation "DL” or "(+/-)” or or “( ⁇ )” appears it also includes the enantiomeric form (mirror image) of the structure actually shown in the formula. For example:
  • DL-threo (the other enantiomer shown) and all racemic mixtures of the two optical isomers are also included.
  • one enantiomer of the threo, and in some cases of the erythro, enantiomers is significantly more active as an analgesic than the other enantiomer of the same pair.
  • the isolated enantiomer which is significantly more active than the other is considered a novel and inventive composition even if the racemic mixture or the other opposite enantiomer of the same compound have already been described in the prior art.
  • Some of the compounds that are useful in the method of the present invention contain three or more asymmetric centers.
  • An example is the following compound
  • Compound 214 named Compound 214 in the description.
  • the formula shown in the description for Compound 214 indicates two compounds of the threo isomer, but the two compounds indicated are not mirror images of each other, they are diastereomers. Another isomer pair is shown and described as Compound 215. (2S,3R) & (2R,3S)
  • alkyl in the general description and definition of the compounds includes straight chain as well as branch-chained alkyl groups.
  • the compounds of the invention may form salts with pharmaceutically acceptable acids or bases, and such pharmaceutically acceptable salts of the compounds of Formula 1 and of Formula 2 are also within the scope of the invention.
  • none of the W, X and Y groups is a heteroatom.
  • compounds are preferred where the sum of m, n and q is 3 and the aromatic group is unsubstituted or substituted with one or more halogen, alkyl of 1 to 6 carbons, or halogen substituted alkyl of 1 to 6 carbons.
  • Compounds within this class are also preferred where the R5 and Re groups form a carbocyclic ring, or a heterocyclic ring.
  • one of the variables W, X and Y represents a heteroatom, preferably nitrogen and the sum of m, n and q is 3.
  • one or two of the variables W, X and Y represent a heteroatom, selected from N, O or S and the sum of m, n and q is 2.
  • R 4 is H or an acyl group, more preferably H.
  • R 3 compounds in accordance with Formula 1 are preferred where both R 3 groups are Hand where one R 3 group is H and the other is benzyl, monohalogeno, dihalogeno, methyl or methoxy substituted benzyl, cyclohexyl, an alkyl of 1 to 7 carbons, COR 7 , COOR 7 where R 7 is alkyl of 1 to 15 carbons, benzyloxy, phenyl, methoxyphenyl, monohalogen or dihalogeno substituted phenyl, a 2-hydroxy-1-phenylethyl group or an alkyl group of 1 to 20 carbons itself substituted with an NH 2 , NHCOR 7 , or NHCOOR 7 group.
  • Ri and R 2 in the compounds of Formula 1 compounds are preferred in accordance with the invention where Ri and R2 jointly form a pyrrolidine, a 3-fluoro or a 3,3-difluoro or an 3-hydroxy substituted pyrrolidine, a morpholine, a thiomorpholine, a piperazine, an alkyl substituted piperazine where the alkyl group has 1 to 6 carbons, an azetidine, a tetrahydrothiazole, an indoline, or a 2/-/-pyrrol ring or Ri and R 2 are two alkyl groups of 1 to 3 carbons.
  • Rg and R 1 O compounds are generally preferred where Rg and R 1 O are both hydrogen, where one of these two variables is hydroxy and the other is alkyl of 1 to 6 carbons, where the Rg and R 1 O groups jointly form an NOR 11 group, and where Rg is hydrogen, the dashed line between carbons 2 and 3 represent a double bond and R 1 O does not exist.
  • R 11 compounds of Formula 2 are preferred where R 11 is H, or COR-12 where R 12 is alkyl of 1 to 3 carbons.
  • R 1 and R 2 jointly with the nitrogen form a five-membered ring, where both R 3 groups are hydrogen and where one of the R3 groups is hydrogen and the other is formyl.
  • the presently most preferred novel compounds of the invention are disclosed with their structural formulas in the ensuing Tables and or description, showing activity of exemplary compounds relevant to their ability to act as analgesics.
  • BIOLOGICAL ACTIVITY, MODES OF ADMINISTRATION The compounds described here may be used to treat a patient suffering from one or more types of chronic pain, including neuropathic pain, inflammatory pain, headache pain, somatic pain, visceral pain, and referred pain.
  • treat means to deal with medically. It includes, for example, administering a compound of the invention to prevent a pain, to alleviate its severity, and to prevent its reoccurance.
  • Pain means any unpleasant sensory experience, usually associated with a physical disorder.
  • the physical disorder may or may not be apparent to a clinician.
  • Pain is of two types: chronic and acute.
  • An "acute pain” is a pain of short duration having a sudden onset.
  • One type of acute pain for example, is cutaneous pain felt on injury to the skin or other superficial tissues, such as caused by a cut or a burn. Cutaneous nociceptors terminate just below the skin, and due to the high concentration of nerve endings, produce a well-defined, localized pain of short duration.
  • Chronic pain includes neuropathic pain, inflammatory pain, headache pain, somatic pain visceral pain and referred pain.
  • Neuroopathic pain means abnormal sensory input, resulting in discomfort, from the peripheral nervous system, central nervous systems, or both. A. Symptoms of neuropathic pain
  • neuropathic pain can involve persistent, spontaneous pain, as well as allodynia (a painful response to a stimulus that normally is not painful), hyperalgesia (an accentuated response to a painful stimulus that usually causes only a mild discomfort, such as a pin prick), or hyperpathia (where a short discomfort becomes a prolonged severe pain).
  • allodynia a painful response to a stimulus that normally is not painful
  • hyperalgesia an accentuated response to a painful stimulus that usually causes only a mild discomfort, such as a pin prick
  • hyperpathia where a short discomfort becomes a prolonged severe pain.
  • Neuropathic pain may be caused by any of the following.
  • a traumatic insult such as, for example, a nerve compression injury (e.g., a nerve crush, a nerve stretch, a nerve entrapment or an incomplete nerve transsection); a spinal cord injury (e.g., a hemisection of the spinal cord); a limb amputation; a contusion; an inflammation (e.g., an inflammation of the spinal cord); or a surgical procedure.
  • a nerve compression injury e.g., a nerve crush, a nerve stretch, a nerve entrapment or an incomplete nerve transsection
  • a spinal cord injury e.g., a hemisection of the spinal cord
  • a limb amputation e.g., a contusion
  • an inflammation e.g., an inflammation of the spinal cord
  • surgical procedure e.g., a surgical procedure.
  • An ischemic event including, for example, a stroke and heart attack.
  • An infectious agent Exposure to a toxin, including, for example, a drug, an alcohol, a heavy metal (e.g., lead, arsenic, mercury), an industrial agent (e.g., a solvent, fumes from a glue) or nitrous oxide.
  • a heavy metal e.g., lead, arsenic, mercury
  • an industrial agent e.g., a solvent, fumes from a glue
  • a disease including, for example, an inflammatory disorder, a neoplastic tumor, an acquired immune deficiency syndrome (AIDS), Lymes disease, a leprosy, a metabolic disease, a neurodegenerative disease, a spinal stenosis, a mononeuropathy, a polyneuropathy, and a peripheral nerve disorder, such as a neuroma.
  • AIDS acquired immune deficiency syndrome
  • Neuralgia is a pain that radiates along the course of one or more specific nerves usually without any demonstrable pathological change in the nerve structure. The causes of neuralgia are varied. Chemical irritation, inflammation, trauma (including surgery), compression by nearby structures (for instance, tumors), and infections may all lead to neuralgia. In many cases, however, the cause is unknown or unidentifiable. Neuralgia is most common in elderly persons, but it may occur at any age.
  • a neuralgia includes, without limitation, a trigeminal neuralgia, a spinal stenosis, a post-herpetic neuralgia, a postherpetic neuralgia, a glossopharyngeal neuralgia, pain associated with nerve entrapment disorders, a sciatica and an atypical facial pain.
  • Neuralgia is a painful disorder of the cranial nerves. Falling under the category of neuralgia are trigeminal neuralgia (TN), atypical facial pain, and postherpetic neuralgia (caused by shingles or herpes). The affected nerves are responsible for sensing touch, temperature and pressure in the facial area from the jaw to the forehead.
  • the disorder generally causes short episodes of excruciating pain, usually for less than two minutes and on only one side of the face.
  • the pain can be described in a variety of ways such as “stabbing,” “sharp,” “like lightning,” “burning,” and even “itchy”.
  • the pain can also present as severe or merely aching and last for extended periods.
  • the pain associated with TN is recognized as one the most excruciating pains that can be experienced.
  • Simple stimuli such as eating, talking, washing the face, or any light touch or sensation can trigger an attack (even the sensation of a gentle breeze).
  • the attacks can occur in clusters or as an isolated attack.
  • Trigeminal neuralgia is the most common form of neuralgia. It affects the main sensory nerve of the face, the trigeminal nerve ("trigeminal” literally means “three origins”, referring to the division of the nerve into 3 branches).
  • This condition involves sudden and short attacks of severe pain on the side of the face, along the area supplied by the trigeminal nerve on that side.
  • the pain attacks may be severe enough to cause a facial grimace, which is classically referred to as a painful tic (tic douloureux).
  • the cause of trigeminal neuralgia is a blood vessel or small tumor pressing on the nerve.
  • disorders such as multiple sclerosis (an inflammatory disease affecting the brain and spinal cord), certain forms of arthritis, and diabetes (high blood sugar) may also cause trigeminal neuralgia, but a cause is not always identified. In this condition, certain movements such as chewing, talking, swallowing, or touching an area of the face may trigger a spasm of excruciating pain.
  • a related but rather uncommon neuralgia affects the glosso-pharyngeal nerve, which provides sensation to the throat. Symptoms of this neuralgia are short, shock-like episodes of pain located in the throat.
  • Neuralgia may occur after infections such as shingles, which is caused by the varicella-zoster virus, a type of herpesvirus. This neuralgia produces a constant burning pain after the shingles rash has healed. The pain is worsened by movement of or contact with the affected area. Not all of those diagnosed with shingles go on to experience postherpetic neuralgia, which can be more painful than shingles. The pain and sensitivity can last for months or even years. The pain is usually in the form of an intolerable sensitivity to any touch but especially light touch. Postherpetic neuralgia is not restricted to the face; it can occur anywhere on the body but usually occurs at the location of the shingles rash. Depression is not uncommon due to the pain and social isolation during the illness.
  • Postherpetic neuralgia may be debilitating long after signs of the original herpes infection have disappeared.
  • Other infectious diseases that may cause neuralgia are syphilis and Lyme disease.
  • Diabetes is another common cause of neuralgia. This very common medical problem affects almost 1 out of every 20 Americans during adulthood. Diabetes damages the tiny arteries that supply circulation to the nerves, resulting in nerve fiber malfunction and sometimes nerve loss. Diabetes can produce almost any neuralgia, including trigeminal neuralgia, carpal tunnel syndrome (pain and numbness of the hand and wrist), and meralgia paresthetica (numbness and pain in the thigh due to damage to the lateral femoral cutaneous nerve).
  • Strict control of blood sugar may prevent diabetic nerve damage and may accelerate recovery in patients who do develop neuralgia.
  • Other medical conditions that may be associated with neuralgias are chronic renal insufficiency and porphyria - a hereditary disease in which the body cannot rid itself of certain substances produced after the normal breakdown of blood in the body. Certain drugs may also cause this problem.
  • Deafferentation indicates a loss of the sensory input from a portion of the body, and can be caused by interruption of either peripheral sensory fibres or nerves from the central nervous system.
  • a deafferentation pain syndrome includes, without limitation, an injury to the brain or spinal cord, a post-stroke pain, a phantom pain, a paraplegia, a brachial plexus avulsion injuries, lumbar radiculopathies. 3.
  • CRPSs Complex regional pain syndromes
  • CRPS is a chronic pain syndrome with two forms.
  • CRPS 1 currently replaces the term "reflex sympathetic dystrophy syndrome". It is a chronic nerve disorder that occurs most often in the arms or legs after a minor or major injury.
  • CRPS 1 is associated with severe pain; changes in the nails, bone, and skin; and an increased sensitivity to touch in the affected limb.
  • CRPS 2 replaces the term causalgia, and results from an identified injury to the nerve.
  • a CRPS includes, without limitation, a CRPS Type I (reflex sympathetic dystrophy) and a CRPS Type Il (causalgia).
  • a neuropathy is a functional or pathological change in a nerve and is characterized clinically by sensory or motor neuron abnormalities.
  • Central neuropathy is a functional or pathological change in the central nervous system.
  • Peripheral neuropathy is a functional or pathological change in one or more peripheral nerves.
  • the peripheral nerves relay information from your central nervous system (brain and spinal cord) to muscles and other organs and from your skin, joints, and other organs back to your brain.
  • Peripheral neuropathy occurs when these nerves fail to carry information to and from the brain and spinal cord, resulting in pain, loss of sensation, or inability to control muscles.
  • the failure of nerves that control blood vessels, intestines, and other organs results in abnormal blood pressure, digestion problems, and loss of other basic body processes.
  • Risk factors for neuropathy include diabetes, heavy alcohol use, and exposure to certain chemicals and drugs. Some people have a hereditary predisposition for neuropathy.
  • Prolonged pressure on a nerve is another risk for developing a nerve injury.
  • Pressure injury may be caused by prolonged immobility (such as a long surgical procedure or lengthy illness) or compression of a nerve by casts, splints, braces, crutches, or other devices.
  • Polyneuropathy implies a widespread process that usually affects both sides of the body equally. The symptoms depend on which type of nerve is affected. The three main types of nerves are sensory, motor, and autonomic. Neuropathy can affect any one or a combination of all three types of nerves. Symptoms also depend on whether the condition affects the whole body or just one nerve (as from an injury). The cause of chronic inflammatory polyneuropathy is an abnormal immune response.
  • the specific antigens, immune processes, and triggering factors are variable and in many cases are unknown. It may occur in association with other conditions such as HIV, inflammatory bowel disease, lupus erythematosis, chronic active hepatitis, and blood cell abnormalities.
  • Peripheral neuropathy may involve a function or pathological change to a single nerve or nerve group (monneuropathy) or a function or pathological change affecting multiple nerves (polyneuropathy).
  • Diabetes diabetes (diabetic neuropathy )
  • Uremia from kidney failure
  • Cancer including bone cancer and other cancers
  • Polyneuropathy is a peripheral neuropathy involving the loss of movement or sensation to an area caused by damage or destruction to multiple peripheral nerves.
  • Polyneuropathic pain includes, without limitation, post-polio syndrome, postmastectomy syndrome, diabetic neuropathy, alcohol neuropathy, amyloidosis, toxin exposure, AIDS, hypothyroidism, uremia, vitamin deficiencies, chemotherapy-induced pain, 2',3'-didexoycytidine (ddC) treatment, exposure to the anticonvulsant phenytoin, exposure to antibiotics including chloramphenicol, nitrofurantoin and sulfonamineds, exposure to sedatives including barbital and hexobarbital, Guillain-Barre syndrome, Fabry's disease or polyneuropathy secondary to cancers such as multiple myeloma.
  • ddC 2',3'-didexoycytidine
  • Mononeuropathy is a peripheral neuropathy involving loss of movement or sensation to an area caused by damage or destruction to a single peripheral nerve or nerve group. Mononeuropathy is most often caused by damage to a local area resulting from injury or trauma, although occasionally systemic disorders may cause isolated nerve damage (as with mononeuritis multiplex). The usual causes are direct trauma, prolonged pressure on the nerve, and compression of the nerve by swelling or injury to nearby body structures. The damage includes destruction of the myelin sheath (covering) of the nerve or of part of the nerve cell (the axon). This damage slows or prevents conduction of impulses through the nerve. Mononeuropathy may involve any part of the body.
  • Mononeuropathic pain includes, without limitation, a sciatic nerve dysfunction, a common peroneal nerve dysfunction, a radial nerve dysfunction, an ulnar nerve dysfunction, a cranial mononeuropathy Vl, a cranial mononeuropathy VII, a cranial mononeuropathy III (compression type), a cranial mononeuropathy III (diabetic type), an axillary nerve dysfunction, a carpal tunnel syndrome, a femoral nerve dysfunction, a tibial nerve dysfunction, a Bell's palsy, a thoracic outlet syndrome, a carpal tunnel syndrome, and a sixth (abducent) nerve palsy.
  • a sciatic nerve dysfunction includes, without limitation, a sciatic nerve dysfunction, a common peroneal nerve dysfunction, a radial nerve dysfunction, an ulnar nerve dysfunction, a cranial mononeuropathy Vl, a cranial mononeuropathy VII, a cranial mononeuropathy III (compression type),
  • Distal axonopathies are the result of some metabolic or toxic derangement of neurons. They may be caused by metabolic diseases such as diabetes, renal failure, deficiency syndromes such as malnutrition and alcoholism, or the effects of toxins or drugs. Distal axonopathy (aka dying back neuropathy) is a type of peripheral neuropathy that results from some metabolic or toxic derangement of peripheral nervous system (PNS) neurons.
  • PNS peripheral nervous system
  • Myelinopathy is due to primary destruction of myelin or the myelinating Schwann cells, which leaves the axon intact, but causes an acute failure of impulse conduction. This demyelination slows down or completely blocks the conduction of electical impulses through the nerve.
  • the most common cause is acute inflammatory demyelinating polyneuropathy (AIDP, better known as Guillain-Barre syndrome), though other causes include chronic inflammatory demyelinating polyneuropathy (CIDP), genetic metabolic disorders (e.g., leukodystrophy or Charcot-Marie-Tooth disease), or toxins.
  • CIDP chronic inflammatory demyelinating polyneuropathy
  • Neuronopathies are the result of destruction of peripheral nervous system (PNS) neurons.
  • Neurotoxins may cause neuronopathies, such as the chemotherapy agent vincristine.
  • Neuronopathy is dysfunction due to damage to neurons of the peripheral nervous system (PNS), resulting in a peripheral neuropathy. It may be caused by motor neurone diseases, sensory neuronopathies (e.g., Herpes zoster), toxic substances or autonomic dysfunction.
  • PNS peripheral nervous system
  • a person with neuronopathy may present in different ways, depending on the cause, the way it affects the nerve cells, and the type of nerve cell that is most affected.
  • Focal entrapment neuropathies e.g., carpal tunnel syndrome
  • the compounds of the invention may be used to treat pain caused by or otherwise associated with any of the following inflammatory conditions.
  • Arthritic disorders include, for example, a rheumatoid arthritis; a juvenile rheumatoid arthritis; a systemic lupus erythematosus (SLE); a gouty arthritis; a scleroderma; an osteoarthritis; a psoriatic arthritis; an ankylosing spondylitis; a Reiter's syndrome (reactive arthritis); an adult Still's disease; an arthritis from a viral infection; an arthritis from a bacterial infection, such as, e.g., a gonococcal arthritis and a non-gonococcal bacterial arthritis (septic arthritis); a Tertiary Lyme disease; a tuberculous arthritis; and an arthritis from a fungal infection, such as, e,g,, a blastomycosis
  • Autoimmune diseases include, for example, a Guillain-Barre syndrome, a Hashimoto's thyroiditis, a pernicious anemia, an Addison's disease, a type I diabetes, a systemic lupus erythematosus, a dermatomyositis, Sjogren's syndrome, a lupus erythematosus, a multiple sclerosis, a myasthenia gravis, a Reiter's syndrome, a Grave's disease, and a rheumatoid arthritis.
  • Connective tissue disorders include, for example, a spondylarthritis a dermatomyositis, and a fibromyalgia syndrome .
  • Inflammation caused by injury including, for example, a crush, puncture, stretch of a tissue or joint, may cause chronic inflammatory pain.
  • Inflammation caused by infection including, for example, a tuberculosis or an interstitial keratitis may cause chronic inflammatory pain. Infection may also result in inflammatory bowel diseases and irritable bowel syndromes.
  • Neuritis is an inflammatory process affecting a nerve or group of nerves. Symptoms depend on the nerves involved, but may include pain, paresthesias, paresis, or hypesthesia (numbness).
  • Examples include: a. Brachial neuritis b. Retrobulbar neuropathy, an inflammatory process affecting the part of the optic nerve lying immediately behind the eyeball. c. Optic neuropathy, an inflammatory process affecting the optic nerve causing sudden, reduced vision in the affected eye. The cause of optic neuritis is unknown. The sudden inflammation of the optic nerve (the nerve connecting the eye and the brain) leads to swelling and destruction of the myelin sheath. The inflammation may occasionally be the result of a viral infection, or it may be caused by autoimmune diseases such as multiple sclerosis. Risk factors are related to the possible causes. d. Vestibular neuritis, a viral infection causing an inflammatory process affecting the vestibular nerve. G. Joint inflammation
  • Inflammation of the joint such as that caused by bursitis or tendonitis, for example, may cause chronic inflammatory pain. ///. Headache Pain
  • the compounds of the invention may be used to treat pain caused by or otherwise associated with any of the following headache conditions.
  • a headache (medically known as cephalgia) is a condition of mild to severe pain in the head; sometimes neck or upper back pain may also be interpreted as a headache. It may indicate an underlying local or systemic disease or be a disorder in itself.
  • A. Muscular/myogenic headache Muscular/myogenic headaches appear to involve the tightening or tensing of facial and neck muscles; they may radiate to the forehead. Tension headache is the most common form of myogenic headache.
  • a tension headache is a condition involving pain or discomfort in the head, scalp, or neck, usually associated with muscle tightness in these areas. Tension headaches result from the contraction of neck and scalp muscles. One cause of this muscle contraction is a response to stress, depression or anxiety. Any activity that causes the head to be held in one position for a long time without moving can cause a headache. Such activities include typing or use of computers, fine work with the hands, and use of a microscope.
  • a tension-type headache includes, without limitation, an episodic tension headache and a chronic tension headache.
  • B. vascular headache The most common type of vascular headache is migraine. Other kinds of vascular headaches include cluster headaches, which cause repeated episodes of intense pain, and headaches resulting from high blood pressure 1.
  • Migraine A migraine is a heterogeneous disorder that generally involves recurring headaches. Migraines are different from other headaches because they occur with other symptoms, such as, e.g., nausea, vomiting, or sensitivity to light. In most people, a throbbing pain is felt only on one side of the head.
  • a migraine headache includes, without limitation, a migraine without aura (common migraine), a migraine with aura (classic migraine), a menstrual migraine, a migraine equivalent (acephalic headache), a complicated migraine, an abdominal migraine and a mixed tension migraine. 2.
  • Cluster headache includes, without limitation, a migraine without aura (common migraine), a migraine with aura (classic migraine), a menstrual migraine, a migraine equivalent (acephalic headache), a complicated migraine, an abdominal migraine and a mixed tension migraine.
  • Cluster headaches affect one side of the head (unilateral) and may be associated with tearing of the eyes and nasal congestion. They occurs in clusters, happening repeatedly every day at the same time for several weeks and then remitting.
  • Traction and inflammatory headache are usually symptoms of other disorders, ranging from stroke to sinus infection.
  • Rebound headaches also known as medication overuse headaches, occur when medication is taken too frequently to relieve headache. Rebound headaches frequently occur daily and can be very painful. H. Chronic sinusitis headache
  • Sinusitis is inflammation, either bacterial, fungal, viral, allergic or autoimmune, of the paranasal sinuses.
  • Chronic sinusitis is one of the most common complications of the common cold. Symptoms include: Nasal congestion; facial pain; headache; fever; general malaise; thick green or yellow discharge; feeling of facial 'fullness' worsening on bending over. In a small number of cases, chronic maxillary sinusitis can also be brought on by the spreading of bacteria from a dental infection.
  • Chronic hyperplastic eosinophilic sinusitis is a noninfective form of chronic sinusitis.
  • J. lctal headaches lctal headaches are headaches associated with seizure activity.
  • the compounds of the invention may be used to treat pain caused by or otherwise associated with any of the following somatic pain conditions.
  • Somatic pain originates from ligaments, tendons, bones, blood vessels, and even nerves themselves. It is detected with somatic nociceptors.
  • the scarcity of pain receptors in these areas produces a dull, poorly-localized pain of longer duration than cutaneous pain; examples include sprains and broken bones. Additional examples include the following.
  • Excessive muclse tension can be caused, for example, by a sprain or a strain.
  • Repetitive motion disorders can result from overuse of the hands, wrists, elbows, shoulders, neck, back, hips, knees, feet, legs, or ankles.
  • Muscle disorders causing somatic pain include, for example, a polymyositis, a dermatomyositis, a lupus, a fibromyalgia, a polymyalgia rheumatica, a macrophagic myofasciitis, and a rhabdomyolysis. Muscle pain can also be secondary to neurological and neuromuscular disorders including without limitation Parkinson's disease, Huntington's chorea, dystonias, tardive dyskinesias, drug-induced dyskinesias and dystonias, dyskinesias
  • Myalgia is muscle pain and is a symptom of many diseases and disorders. The most common cause for myalgia is either overuse or overstretching of a muscle or group of muscles. Myalgia without a traumatic history is often due to viral infections. Longer-term myalgias may be indicative of a metabolic myopathy, some nutritional deficiencies or chronic fatigue syndrome. E. Infection
  • Infection can cause somatic pain.
  • infections include, for example, an abscess in the muscle, a trichinosis, an influenza, a Lyme disease, a malaria, a Rocky Mountain spotted fever, Avian influenza, the common cold, community-acquired pneumonia, meningitis, monkeypox, Severe Acute Respiratory Syndrome, toxic shock syndrome, trichinosis, typhoid fever, and upper respiratory tract infection.
  • F. Drugs include, for example, an abscess in the muscle, a trichinosis, an influenza, a Lyme disease, a malaria, a Rocky Mountain spotted fever, Avian influenza, the common cold, community-acquired pneumonia, meningitis, monkeypox, Severe Acute Respiratory Syndrome, toxic shock syndrome, trichinosis, typhoid fever, and upper respiratory tract infection.
  • Drugs can cause somatic pain.
  • Such drugs include, for example, cocaine, statins for lowering cholesterol (such as atorvastatin, simvastatin, and lovastatin), and ACE inhibitors for lowering blood pressure (such as enalaphl and captophl).
  • statins for lowering cholesterol such as atorvastatin, simvastatin, and lovastatin
  • ACE inhibitors for lowering blood pressure (such as enalaphl and captophl).
  • G. Prolonged nociceptive pain including without limitation to bone fracture pain, spinal stenosis, and post-surgical pain.
  • the compounds of the invention may be used to treat pain caused by or otherwise associated with any of the following visceral pain conditions.
  • Visceral pain originates from body's viscera, or organs.
  • Visceral nociceptors are located within body organs and internal cavities. The even greater scarcity of nociceptors in these areas produces pain that is usually more aching and of a longer duration than somatic pain.
  • Visceral pain is extremely difficult to localise, and several injuries to visceral tissue exhibit "referred" pain, where the sensation is localised to an area completely unrelated to the site of injury. Examples of visceral pain include the following.
  • Functional visceral pain includes, for example, an irritable bowel syndrome and a chronic functional abdominal pain (CFAP), a functional constipation and a functional dyspepsia, a non-cardiac chest pain (NCCP) and a chronic abdominal pain.
  • CFAP chronic functional abdominal pain
  • NCCP non-cardiac chest pain
  • Chronic gastrointestinal inflammation includes, for example, a gastritis, an inflammatory bowel disease, e.g., a Crohn's disease, an ulcerative colitis, a microscopic colitis, a diverticulitis and a gastroenteritis; an interstitial cystitis; an intestinal ischemia; a cholecystitis; an appendicitis; a gastroesophageal reflux; an ulcer, a nephrolithiasis, an urinary tract infection, a pancreatitis and a hernia.
  • a gastritis an inflammatory bowel disease, e.g., a Crohn's disease, an ulcerative colitis, a microscopic colitis, a diverticulitis and a gastroenteritis
  • an interstitial cystitis an intestinal ischemia
  • a cholecystitis cholecystitis
  • an appendicitis a gastroesophageal reflux
  • Autoimmune pain includes, for example, a sarcoidosis and a vasculitis.
  • Organic visceral pain includes, for example, pain resulting from a traumatic, inflammatory or degenerative lesion of the gut or produced by a tumor impinging on sensory innervation.
  • Treatment-induced visceral pain includes, for example, a pain attendant to chemotherapy therapy or a pain attendant to radiation therapy.
  • the compounds of the invention may be used to treat pain caused by or otherwise associated with any of the following referred pain conditions.
  • Referred pain arises from pain localized to an area separate from the site of pain stimulation. Often, referred pain arises when a nerve is compressed or damaged at or near its origin. In this circumstance, the sensation of pain will generally be felt in the territory that the nerve serves, even though the damage originates elsewhere.
  • a common example occurs in intervertebral disc herniation, in which a nerve root arising from the spinal cord is compressed by adjacent disc material. Although pain may arise from the damaged disc itself, pain will also be felt in the region served by the compressed nerve (for example, the thigh, knee, or foot). Relieving the pressure on the nerve root may ameliorate the referred pain, provided that permanent nerve damage has not occurred.
  • Myocardial ischaemia (the loss of blood flow to a part of the heart muscle tissue) is possibly the best known example of referred pain; the sensation can occur in the upper chest as a restricted feeling, or as an ache in the left shoulder, arm or even hand. Pain Reversal
  • An art-accepted model or assay for measuring an analgesic effect of a compound in chronic pain is the model known as Kim and Chung 1992, Pain 150, pp 355-363 ⁇ Chung model).
  • This model involves the surgical ligation of the L5 (and optionally the L6) spinal nerves on one side in experimental animals. Rats recovering from the surgery gain weight and display a level of general activity similar to that of normal rats. However, these rats develop abnormalities of the foot, wherein the hindpaw is moderately everted and the toes are held together. More importantly, the hindpaw on the side affected by the surgery appears to become sensitive to low-threshold mechanical stimuli and will perceive pain instead of the faint sensation of touch.
  • the surgical site is shaved and prepared either with betadine or Novacaine. Incision is made from the thoracic vertebra XIII down toward the sacrum. Muscle tissue is separated from the spinal vertebra (left side) at the L4 - S2 levels. The L6 vertebra is located and the transverse process is carefully removed with a small rongeur to expose the L4 - L6 spinal nerves. The L5 and L6 spinal nerves are isolated and tightly ligated with 6-0 silk thread. The same procedure is done on the right side as a control, except no ligation of the spinal nerves is performed.
  • the wounds are sutured.
  • a small amount of antibiotic ointment is applied to the incised area, and the rat is transferred to the recovery plastic cage under a regulated heat-temperature lamp.
  • the test drugs are administered by intraperitoneal (i.p.) injection or oral gavage (p.o.).
  • i.p. administration the compounds are formulated in H 2 O and given in a volume of 1 ml/kg body weight by injecting into the intraperitoneal cavity.
  • p.o. administration the compounds are formulated in H2O and given in a volume of 1 ml/kg body weight using an 18-gauge, 3 inch gavage needle that is slowly inserted through the esophagus into the stomach.
  • Tactile allodynia is assessed via von Frey hairs, which are a series of fine hairs with incremental differences in stiffness. Rats are placed in a plastic cage with a wire mesh bottom and allowed to acclimate for approximately 30 minutes. To establish the pre-drug baseline, the von Frey hairs are applied perpendicularly through the mesh to the mid-plantar region of the rats' hindpaw with sufficient force to cause slight buckling and held for 6-8 seconds. The applied force has been calculated to range from 0.41 to 15.1 grams. If the paw is sharply withdrawn, it is considered a positive response. A normal animal will not respond to stimuli in this range, but a surgically ligated paw will be withdrawn in response to a 1-2 gram hair.
  • the 50% paw withdrawal threshold is determined using the method of Dixon, W.J., Ann. Rev. Pharmacol. Toxicol. 20:441-462 (1980) hereby incorporated by reference.
  • Tactile allodynia is measured prior to and 15, 30, and 60 minutes after drug administration.
  • the post-drug threshold is compared to the pre-drug threshold and the percent reversal of tactile sensitivity is calculated based on a normal threshold of 15.1 grams. Table 1 below indicates the degree of pain reversal obtained in the
  • the intraperitonial (i.p.) and/or intravenous (iv) administration of the compounds was in doses ranging from 1 ⁇ g/kg to 300 ⁇ g/kg or 3 mg/kg PO and the peak percentage of reversal of allodynia was measured at 15, 30 or 60 minutes after administration, as is indicated in the table. Data are expressed as the highest % allodynia reversal (out of 3 time points: 15 min, 30 min, or 60 min. post- drug) with a minimum of a 20% allodynia reversal in the rat Chung model. Comparisons between groups (drug treated vs.
  • the compounds of the invention may be administered at pharmaceutically effective dosages.
  • dosages are normally the minimum dose necessary to achieve the desired therapeutic effect; in the treatment of chronic pain, this amount would be roughly that necessary to reduce the discomfort caused by the pain to tolerable levels.
  • doses generally will be in the range of 0.1-5,000 mg/day; more preferably in the range of 1 to 3,000 mg/day, 10 mg to 500 mg/day, 500 to 1 ,000 mg/day, 1 ,000 to 1 ,500 mg/day, 1 ,500 to 2,000 mg/day, 2,000 to 2,500 mg/day, or 2,500 to 3,000 mg/day.
  • the actual amount of the compound to be administered in any given case will be determined by a physician taking into account the relevant circumstances, such as the severity of the pain, the age and weight of the patient, the patient's general physical condition, the cause of the pain, and the route of administration.
  • the compounds are useful in the treatment of pain in a mammal; particularly a human being.
  • the patient will be given the compound orally in any acceptable form, such as a tablet, liquid, capsule, powder and the like.
  • other routes may be desirable or necessary, particularly if the patient suffers from nausea.
  • Such other routes may include, without exception, transdermal, intrapehtonial, parenteral, subcutaneous, intranasal, intrathecal, intramuscular, intravenous and intrarectal modes of delivery.
  • Compositions useful in the method of the invention may further include an excipient.
  • Such an excipient may be a carrier or a diluent; this is usually mixed with the active compound, or permitted to dilute or enclose the active compound.
  • the carrier may be solid, semi-solid, or liquid material that acts as an excipient or vehicle for the active compound.
  • the formulations may also include wetting agents, emulsifying agents, preserving agents, sweetening agents, and/or flavoring agents. If used as in an ophthalmic or infusion format, the formulation will usually contain one or more salt to influence the osmotic pressure of the formulation.
  • Absolute configuration of compounds of the invention can generally speaking be determined in accordance with methods known in the state of the art, such as X-ray chhstallography.
  • Compounds 203 and 204 are mentioned as examples for which the absolute configurations were determined by X-ray chhstallography analysis of the corresponding (1 S)- camphanylamide D-(10) camphorsulfonic acid salt.
  • Compound 204 was assigned (2S, 3R). Its enantiomer, Compound 203 was assigned by default the (2R, 3S) absolute configuration.
  • R * substantially corresponds to the 5, 6, or 7 membered ring structure on the left side of Formula 1 (as the formula is depicted in the Summary and in the instant claims) and R ** substantially corresponds to the Ri groups in Formula 1.
  • (+l-)-threo ft* «»s-oxazoline
  • methyl isocyanoacetate (or ethyl isocyanoacetate available commercially) is reacted with an "amine” which includes the R ** groups to provide the 2-isocyanoacetic acid amide derivative shown in the general scheme.
  • amines used in the reaction are pyrrolidine, pipehdine, azetidine, morpholine, 2,5- dihydro-1 H-pyrrole, dialkylamines such as diethylamine, 3-fluoro-, 3,3-difluoro or 3-hydroxy substituted pyrrolidines. Specific examples of these "amines" abound in the experimental description.
  • the 2-isocyanoacetic acid amide derivative is then reacted in the presence of base (such as KOH) with an "aldehyde” which includes the R * group to provide a trans "oxazoline” with high diastereoselectivity (transxis ratios generally > 97:3) as shown in the general reaction scheme 1.
  • the trans oxazoline is then treated with a strong acid, such as HCI, to open the ring and to provide the f/?reo-3-substituted-3- hydroxy-2-amino-propionic acid amides (with threo:erythro ratios generally > 97:3) of the invention as shown in General Scheme 1.
  • EDCI stands for 1-(3-dimethylaminopropyl)- ethylcarbodiimide hydrochloride
  • HOBT stands for 1-hydroxybenzothazole
  • BOC 2 O stands for di-f-butyl-dicarbonate
  • TEA stands for triethylamine.
  • the A- pyridyl group can be substituted with an R * group (as defined in connection with Scheme 1 ) and the indoline can be susbstituted with other weak nucleophilic amines of the formula NH(R ** ) 2 (R ** defined as in connection with Reaction Scheme 1 ) to provide other compounds of Formula 1 analogous to compounds 242 and 243.
  • threo and erythro isomers when both are formed in the reactions leading to the compounds of the invention, can typically be separated by chromatographic methods.
  • the more abundantly formed threo isomers can also be converted into the erythro isomers by oxidizing to the ketone level the hydroxyl group in the 3 position of the propanoic acid moiety and subsequently reducing the resulting ketone to the hydroxyl level.
  • Separation of enantiomeric mixtures can be performed on Chiralpack columns which are well known in the art. (See, for example, the preparation of Compound 204).
  • acylated derivatives of the 2- amino function can be prepared by using acyl chlorides such as acetyl chloride and hexanoyl chloride. (See, Method G and the preparation of Compound 51 ).
  • Carbamate derivatives of the 2-amino function can be obtained by using chloroformates, such as benzylchloroformate. (See, for example, the preparation of Compound 58).
  • the tertiary butyl carbamoyl function can also serve as a removable protecting group of the 2-amino function, (see for example the preparation of Compounds 219 and 224).
  • 2-amino function of the compounds of the invention is already acylated or bears a carbamoyl group, then the 3-hydroxy group of the propanoic acid moiety can be subjected to acylation by reagents such as acetic anhydride. (See for example the preparation of Compound 217).
  • Alkylation of the 2-amino function is readily performed by condensing the compound bearing the 2-NH 2 group with an aldehyde to obtain a Schiff base intermediate which can be reduced, without isolation, to provide the N- alkyl, arylalkyl or heteroaryl-alkyl compound.
  • the procedure described for preparing Compound 234 can be generalized to make compounds of the invention where the 2-amino function bears an aryl( hydroxy)alkyl or heteroaryl(hydroxy)alkyl group.
  • Compounds of Formula 2 where the Rg and R-m groups jointly form an oxime (NOH) group can be obtained by oxidizing the 3-hydroxyl group of the propanoic acid moiety to the ketone stage and reacting the resulting ketone with hydroxylamine.
  • NOH oxime
  • the 4-pyhdyl group can be substituted with an R * group (as defined in connection with Scheme 1 ) and the pyrrolidine can be substituted with amines of the formula NH(R ** ) 2 (R ** defined as in connection with Reaction Scheme 1 ) to provide other compounds of Formula 2 analogous to compound 236 or to its enantiomer (S)-2-amino-3-(pyridin-4-yl)-1-(pyrrolidin-1-yl)propan-1-one dihydrochlohde Compound 240.
  • BLE 041 10B Preparation of oxazolines: BLE 041 10B. SLA 07122A. SLA 07124A. SLA 07124B. SLA 07132. BLE 041 10A. Compound 19. BLE 04124A. BLE 04124B. BLE 04124C, BLE 04124D, BLE 04130B, BLE 04130C, BLE 04130D, BLE 04136B. BLE 04136C, BAL 01016, BLE 04136D, BAL 01014. SLA 07194A. SLA 07174, BAL 01028A, BLA 01028B, SLA 07158 and SLA 07180.
  • SLA 07122A was prepared in accordance with method method D using 2-isocyano-1-(pipehdin-1 -yl)ethanone (0.4 g, 26.3 mmol), potassium hydroxide (0.15 g, 26.7 mmol) in methanol (5 mL) and pyridine-4- carbaldehyde (0.37 mL, 40.9 mmol). The solution was stirred 20 h at 0 0 C. frans-(4,5-Dihydro-5-(pyridin-4-yl)oxazol-4-yl)(piperidin-1 -yl)methanone SLA 07122A was obtained as a yellow solid (0.353 g, 52 % yield).
  • SLA 07124 A SLA 071 18 was prepared in accordance with method D using 2- isocyano-1-morpholinoethanone (0.40 g, 25.95 mmol), potassium hydroxide (0.146 g, 26.0 mmol) in methanol (5 mL) and pyhdine-4-carbaldehyde (0.36 mL, 40.4 mmol). The solution was stirred 22 h at 0 0 C. frans-(4,5-Dihydro-5- (pyhdin-4-yl)oxazol-4-yl)(morpholino)methanone SLA 07124A was obtained as a yellow solid (0.168 g, 25 % yield).
  • SLA 07124B was prepared in accordance with method D using tert- butyl 4-(2-isocyanoacetyl)piperazine-1-carboxylate SLA 071 16C (0.41 g, 16.20 mmol), potassium hydroxide (0.91 g, 16.2 mmol) in methanol (5 mL) and pyhdine-4-carbaldehyde (0.227 mL, 25.2 mmol). The solution was stirred 22 h at 0 0 C.
  • SLA 07132 was prepared in accordance with method D using 2- lsocyano-1 -thiomorpholinoethanone SLA 0713OA (0.752 g, 4.41 mmol), potassium hydroxide (0.250 g, 4.45 mmol) in methanol (10 mL) and pyridine- 4-carbaldehyde (0.436 mL, 4.85 mmol). The solution was stirred 24 h at 0 0 C. frans-(4,5-Dihydro-5-(pyridin-4-yl)oxazol-4-yl)(thiomorpholino)methanone SLA 07132 was obtained as a yellow foam (1.01 g, 83 %).
  • BLE 041 10A was prepared in accordance with method D using 2- pyridine carboxaldehyde (1.02 mL, 10.84 mmol). 7ra/7s-(4,5-dihydro-5- (pyhdin-2-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 041 1 OA was obtained as a yellow pale oil (0.45 g, 19 % yield).
  • BLE 04124A was prepared in accordance with method D using thiophen-3-carboxaldehyde (0.475 mL, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol).
  • BLE 04124B was prepared in accordance with method D using thiophen-2-carboxaldehyde (0.507 mL, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol).
  • BLE 04124C was prepared in accordance with method D using 2- thiazolecarboxaldehyde (0.476 mL, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1-(pyrrolidin-1 -yl)ethanone BLE 04098 (0.75 g, 5.43 mmol).
  • BLE 04124D was prepared in accordance with method D using thianaphtene-3-carboxaldehyde (0.88 g, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol). After work-up the residue obtained was purified by column chromatography (EtOAc) to led after evaporation to trans- ⁇ 5-
  • BLE 0413OB was prepared in accordance with method D using 3- furaldehyde (0.453 g, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol).
  • BLE 0413OC was prepared in accordance with method D using 2- naphtaldehyde (0.847 g, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol).
  • BLE 0413OD was prepared in accordance with method D using 1- naphtaldehyde (0.736 mL, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol).
  • BLE 04136B was prepared in accordance with method D using 2- quinoline carbaldehyde (0.852 g, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol).
  • BLE 04136C was prepared in accordance with method D using 4- quinoline carbaldehyde (0.852 g, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol).
  • BLE 04136C was prepared in accordance with method D using 2- furaldehyde (0.449 ml_, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 ml.) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol).
  • BAL 01014 was prepared in accordance with method D using 2- methoxy-3-pyridinecarboxaldehyde (0.64 ml, 5.43 mmol), KOH (0.305 mg, 5.43 mmol) in methanol (5 mL) and 2-isocyano-1 -(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol). After work-up frans-(4,5-dihydro-5-(2- methoxypyhdin-3-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BAL 01014 was obtained (0.74 mg, 50 % yield) as a white solid.
  • SLA 07194A was prepared in accordance with method D using pyridine-4-carbaldehyde (1.14 mL, 9.52 mmol), KOH (0.54 g, 9.60 mmol) in methanol (5 mL) and ⁇ /, ⁇ /-diethyl-2-isocyanoacetamide SLA 07184A (1.21 g, 8.65 mmol). After work-up and column chromatography on florisil (ethyl acetate) frans- ⁇ /, ⁇ /-diethyl-4,5-dihydro-5-(pyhdin-4-yl)oxazole-4-carboxamide SLA 07194A was obtained as a brown oil (0.25 g, 12 % yield).
  • Methyl 2-chloropyridine-4-carboxylate (2.50 g, 14.60 mmol) was dissolved in anhydrous THF (50 mL) and this solution was cooled to -78 0 C under N 2 atmosphere.
  • Diisobutylaluminium hydride 1.0 M in hexanes (63.3 mL, 63.30 mmol) was added dropwise stabilizing the temperature between - 50 0 C and -70 0 C.
  • the reaction mixture was stirred 1 .5 h at -78 0 C and allowed to stand at room temperature for 3 h.
  • a solution of aqueous 10 % NH 4 CI was slowly added and the mixture was extracted with ethyl acetate (3 x 300 ml_).
  • the dropping funnel was washed with dichloromethane (2 x 5 mL) and charged with a solution of thethylamine (480 ⁇ l_, 6.51 mmol) in dichloromethane (4 mL) which was added (10 min) to the reaction mixture and finally the reaction flask was allowed to warm to 0 0 C over 10 min.
  • the reaction solution was transferred to a 500 mL separatory funnel charged with 130 mL of a 5 % aqueous NH 4 CI solution.
  • SLA 07174 was prepared in accordance with method D using 2- chloropyridine-4-carbaldehyde SLA 07156 (0.12 g, 1.05 mmol), KOH (0.06 g, 1.05 mmol) in methanol (10 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.146 g, 1.05 mmol). The solution was stirred 24 h with continued cooling.
  • BAL 01028A was prepared in accordance with method D using 3- bromo-4-pyhdinecarboxaldehyde (1.010 g, 5.43 mmol), KOH (0.305 g, 5.43 mmol) in methanol (5 mL) and 2-isocyano-1-pyrrolidin-1-yl-ethanone BLE 04134 (0.75 g, 5.43 mmol). The mixture was stirred at 0 0 C until precipitation and concentrated. The mixture was partitioned between EtOAc (50 ml) and H 2 O (25 ml). The aqueous layer was extracted twice with EtOAc (25 mL).
  • BAL 01028B was prepared in accordance with method D using 2- isocyano-1-pyrrolidin-1 -yl-ethanone BLE 04134 (0.75 g, 5.43 mmol), KOH (0.305 g, 5.43 mmol) in methanol (5 mL) and 3-chloro-isonicotinaldehyde (0.769 g, 5.43 mmol). The solution was stirred 3 h at 0 0 C. trans- ⁇ 5- ⁇ 3- Chloropyridin-4-yl)-4,5-dihydrooxazol-4-yl)(pyrrolidin-1 -yl)methanone BAL 01028B (1.20 g, 65% yield) was obtained as a white solid.
  • SLA 07158 was prepared in accordance with method D using 2- chloropyridine-4-carbaldehyde SLA 07156 (0.47 g, 3.31 mmol), KOH (0.184 g, 3.33 mmol) in methanol (10 mL) and 2-lsocyano-1-(2/-/-pyrrol-1 (5/-/)- yl)ethanone SLA 07178 (0.410 g, 3.01 mmol). The solution was stirred 2 h with continued cooling.
  • SLA 07158 was prepared in accordance with method D using pyridine- 4-carbaldehyde (0.293 mL, 2.40 mmol), KOH (0.13 g, 2.32 mmol) in methanol (10 mL) and 2-isocyano-1 -(2H-pyrrol-1 (5H)-yl)ethanone SLA 07178 (0.301 g, 2.20 mmol). The solution was stirred 2 h with continued cooling.
  • Compound 21 was prepared following method E with trans- ⁇ 4,5- dihydro-5-(pyridin-2-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 041 10 B (0.44 g, 1 .79 mmol), hydrochloric acid 37 % (1 .0 mL) and methanol (10 mL). After 2.5 h at 50 0 C and work-up DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-2-yl)- 1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochloride Compound 21 was obtained as a yellow solid (0.44 g, 84 % yield).
  • Compound 23 was prepared following method E with trans- ⁇ 4,5- dihydro-5-(thiophen-3-yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone BLE 04124A (0.486 g, 1.94 mmol), hydrochloric acid 37 % (0.6 mL) and methanol (10 ml_). After 3.5 h at 50 0 C and work-up DL-f/?reo-2-amino-3-hydroxy-1 -(pyrrolidin-1 - yl)-3-(thiophen-3-yl)propan-1-one hydrochloride Compound 23 was obtained as a white solid (0.480 g, 89.5 % yield).
  • Compound 27 was prepared following method E with frans-(5-(furan-3- yl)-4,5-dihydrooxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 0413OB (0.800 g, 3.41 mmol), hydrochloric acid 37 % (0.6 mL) and methanol (10 mL). After 3.5 h at 50 0 C and work-up DL-f/?reo-2-amino-3-(furan-3-yl)-3-hydroxy-1- (pyrrolidin-1-yl)propan-1-one hydrochloride Compound 27 was obtained as a white solid (0.738 g, 83 % yield).
  • Compound 28 was prepared following method E with trans- ⁇ 4,5- dihydro-5-(naphthalen-3-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 0413OC (0.745 g, 2.53 mmol), hydrochloric acid 37 % (0.6 ml.) and methanol (10 ml_). After 3.5 h at 50 0 C and work-up DL-f/?reo-2-amino-3-hydroxy-3- (naphthalen-2-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one hydrochloride Compound 28 was obtained as a white solid (0.706 g, 87 % yield).
  • Compound 30 was prepared following method E with trans- ⁇ 4,5- dihydro-5-(quinolin-2-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 04136B (0.923 g, 3.13 mmol), hydrochloric acid 37 % (0.6 mL) and methanol (15 mL). After 3.5 h at 50 0 C and work-up DL-f/?reo-2-amino-3-hydroxy-1 -(pyrrolidin-1 - yl)-3-(quinolin-2-yl)propan-1-one dihydrochlohde Compound 30 was obtained as a yellow solid (1 .098 g, 98 % yield).
  • Compound 32 was prepared following method E with trans- ⁇ 4,5- dihydro-5-(quinolin-3-yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone BAL 01016 (0.905 g, 3.41 mmol), hydrochloric acid 37 % (0.6 ml.) and methanol (10 ml_). After 2 h at RT and work-up ⁇ /-(DL-f/?reo-1-hydroxy-3-oxo-3-(pyrrolidin-1-yl)-1- (quinolin-3-yl)propan-2-yl)formamide hydrochloride Compound 32 was obtained as a white solid (240 mg, 20.0 % yield).
  • Compound 33 was prepared following method E with trans- ⁇ 4,5- dihydro-5-(quinolin-3-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BAL 01016 (0.91 g, 3.41 mmol), hydrochloric acid 37% (0.6 mL) and methanol (10 ml_). After 3 h at 50 0 C and work-up DL-f/?reo-2-amino-3-hydroxy-1 -(pyrrolidin-1 -yl)- 3-(quinolin-3-yl)propan-1-one dihydrochlohde Compound 33 (678 mg, 55 % yield) was obtained as a white solid.
  • Compound 38 was prepared following method E with trans- ⁇ 4,5- dihydro-5-(pyridin-4-yl)oxazol-4-yl)(thiomorpholino)methanone SLA 07132 (0.926 g, 3.36 mmol), hydrochloric acid 37 % (1.1 ml.) and methanol (10 ml_). After 3 h at 50 0 C and work-up DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-4-yl)-1 - thiomorpholinopropan-1 -one dihydrochloride Compound 38 was obtained as a pale yellow solid (1 .1 g, 99 % yield).
  • Compound 40 was prepared following method E with frans- ⁇ /, ⁇ /-diethyl- 4,5-dihydro-5-(pyridin-4-yl)oxazole-4-carboxamide diethylamide SLA 07194A (254 mg, 1.03 mmol), hydrochloric acid 37 % (1.0 mL) and methanol (10 mL). After 2 h at RT and work-up DL-f/?reo-2-amino- ⁇ /, ⁇ /-diethyl-3-hydroxy-3- (pyhdin-4-yl)propanamide dihydrochloride Compound 40 was obtained (212 mg, 67 % yield) as a pale yellow solid. (+/-) 2. HCI
  • Compound 42 was prepared following method E with trans- ⁇ 5- ⁇ 3- bromopyridin-4-yl)-4,5-dihydrooxazol-4-yl)(pyrrolidin-1 -yl)methanone BAL 01028A (1.141 g, 3.52 mmol), hydrochloric acid 37 % (0.6 ml.) and methanol (15 ml_). After 3 h at 50 0 C and work-up DL-f/?reo-2-amino-3-(3-bromopyridin- 4-yl)-3-hydroxy-1-(pyrrolidin-1-yl)propan-1-one dihydrochloride Compound 42 was obtained as a white solid (667 mg, 49 % yield).
  • the reaction mixture was allowed to reach room temperature, stirred for 16 h and partitioned with H 2 O (3 x 4 ml_), washed with brine (3 x 4 ml_), NaOH (0.5 M, 3 x 4 ml.) and the organic layer was evaporated, adsorbed on silica gel (0.3 g) with EtOAc.
  • the desired product was isolated by column chromatography using a gradient O to 8 % [v/v] MeOH in EtOAc.
  • the solid obtained was dissolved in ethanol (1 mL) and a solution of HCI (0.8 M, 1 mL) in EtOH was added. Evaporation of the volatiles led to the corresponding hydrochloride salt.
  • the compound was prepared according to method F with hexanoyl chloride (59 mg, 0.484 mmol) and DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-4- yl)-1-(pyrrolidin-1-yl)propan-1 -one dihydrochlohde Compound 22. /V-(DL- f/?reo-1 -Hydroxy-3-oxo-1 -(pyhdin-4-yl)-3-(pyrrolidin-1 -yl)propan-2- yl)hexanamide hydrochloride Compound 51 was obtained as an off white solid. (56 mg, 34 % yield).
  • the compound was prepared according to method F with heptanoyl chloride (72 mg, 0.484 mmol) and DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-4- yl)-1-(pyrrolidin-1-yl)propan-1 -one dihydrochloride Compound 22. /V-(DL- f/?reo-1 -Hydroxy-3-oxo-1 -(pyhdin-4-yl)-3-(pyrrolidin-1 -yl)propan-2- yl)heptanamide hydrochloride Compound 52 was obtained as an off white solid. (192 mg, 66 % yield).
  • the compound was prepared according to method F with octanoyl chloride (78 mg, 0.484 mmol) and DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-4- yl)-1-(pyrrolidin-1-yl)propan-1 -one dihydrochloride Compound 22. /V-(DL- f/?reo-1 -Hydroxy-3-oxo-1 -(pyridin-4-yl)-3-(pyrrolidin-1 -yl)propan-2- yl)octanamide hydrochloride Compound 53 was obtained as an off white solid. (131 mg, 75 % yield).
  • the compound was prepared according to method F with benzoyl chloride (141 mg, 0.484 mmol) and DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-4- yl)-1-(pyrrolidin-1-yl)propan-1 -one dihydrochlohde Compound 22. /V-(DL- f/?reo-1 -Hydroxy-3-oxo-1 -(pyhdin-4-yl)-3-(pyrrolidin-1 -yl)propan-2- yl)benzamide hydrochloride Compound 55 was obtained as an off white solid. (67 mg, 34 % yield).
  • the compound was prepared according to method F with 4-methoxybenzoyl chloride (82 mg, 0.484 mmol) and DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-4- yl)-1-(pyrrolidin-1-yl)propan-1 -one dihydrochlohde Compound 22. /V-(DL- f/?reo-1 -Hydroxy-3-oxo-1 -(pyridin-4-yl)-3-(pyrrolidin-1 -yl)propan-2-yl)-4- methoxy-benzamide hydrochloride Compound 56 was obtained as an off white solid. (105 mg, 58 % yield).
  • the compound was prepared according to method G with decanoyl chloride (1 1 1 ⁇ l_, 0.53 mmol). After work-up ⁇ /-(DL-f/?reo-3-hydroxy-1 -oxo-3- (pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-2-yl)decanamide hydrochloride Compound 59 was obtained as a white solid (1 15 mg, 55 % yield).
  • the compound was prepared according to method H with paraformaldehyde (21 mg, 0.65 mmol). After colummn chromatography
  • the compound was prepared according to method H with heptaldehyde (82 mg, 0.68 mmol). After column chromatography with
  • the compound was prepared according to method H with 4- chlorobenzaldehyde (98 mg, 0.70 mmol). After column chromatography
  • the compound was prepared according to method H with 4- methoxybenzaldehyde (95 mg, 0.70 mmol). After column chromatography
  • BLE 04136D was prepared in accordance with method D using 2- furaldehyde (0.449 mL, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1 -(pyrrolidin-1 -yl)ethanone BLE 04098 (0.75 g, 5.42 mmol).
  • Compound 201 was prepared following method E with frans-(5-(furan- 2-yl)-4,5-dihydrooxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 04136D (0.30 g, 1.28 mmol), hydrochloric acid 37 % (0.3 mL) and methanol (10 ml_). After overnight at RT and work-up ( ⁇ )-f/?reo-2-amino-3-(furan-2-yl)-3-hydroxy-1 - (pyrrolidin-1-yl)propan-1-one hydrochloride (0.22 g, 66 % yield) was obtained as a pale brown solid.

Abstract

Disclosed herein are methods of treating a patient suffering from one or more types of chronic pain using compounds of Formula 1 wherein the variables have the meaning disclosed in the specification.

Description

METHODS FOR TREATING CHRONIC PAIN USING 3-ARYL-3- HYD ROXY-
2-AMINO-PROPIONIC ACID AMIDES, 3-HETEROARYL-3-HYDROXY-2-
AMINO-PROPIONIC ACID AMIDES AND RELATED COMPOUNDS
BACKGROUND OF THE INVENTION The present invention is directed to methods of treating a patient suffering from one or more types of chronic pain using using derivatives of 3- aryl-3-hydroxy-2-amino-propionic acid amides, 3-heteroaryl-3-hydroxy-2- amino-propionic acid amides, and related compounds.
Several compounds falling within one or more of the general definitions as "derivatives of 3-aryl-3-hydroxy-2-amino-propionic acid amides, of 3- heteroaryl-3-hydroxy-2-amino-propionic acid amides, of 1 -aryl-1-hydroxy-2,3- diamino-propyl amines, 1-heteroaryl-1-hydroxy-2,3-diamino-propyl amines" are known in the patent and scientific literature.
For example, United States Patent Application Publication Nos. 2003/0153768 and 2003/0050299 disclose several examples of the above- mentioned known compounds. The Λ/-acyl compounds of these references are said to be useful as Λ/-acylsphingosine glucosyltransferase inhibitors, the amide and the reduced compounds are described as intermediates in their preparations. Illustrative specific examples of compounds of these references are shown below:
Figure imgf000004_0001
X=H1H X = O
Figure imgf000004_0002
X=H1H X = O
The publication Shin etal. Tetrahedron Asymmetry, 2000, 11, 3293-3301 discloses the following compounds:
Figure imgf000005_0001
(1R,2R)-2-((S)-1-phenylethylamino)-3- (1R,2R)-2-amino-3-morpholino-1- morpholino-1-phenylpropan-1-ol phenylpropan-1-ol
Figure imgf000005_0002
D-thmo-PDMP
L-f/?reo-PDMP and some other known compounds used in the methods of this invention are commercially available, in pure enantiomeric and racemic forms, as applicable, from Matreya, LLC Pleasant Gap, Pennsylvania.
Figure imgf000005_0003
United States Patent Nos. 5,945,442; 5,952,370; 6,030,995 and 6,051 ,598, which are all related to each other as being based on same or related disclosures, describe compounds which are structurally similar to the known compounds shown above. The compounds of these U.S. patent references are said to be inhibitors of the enzyme glucosylceramide (GlcCer) synthethase. A publication in Journal of Labelled Compounds &
Radiopharmaceuticals (1996), 38(3), 285-97 discloses the compound of the formula
Figure imgf000006_0001
Published PCT application WO 01/38228 discloses
Figure imgf000006_0002
in connection with a chromatographic method. Kastron et al. in Latvijas PSR Zinatnu Akademijas Vestis, Kimijas
Serija (1965) (4), 474-7 disclose the following compound.
Figure imgf000006_0003
SUMMARY OF THE INVENTION The present invention is directed to methods of treating a patient suffering from one or more types of chronic pain using compounds of Formula 1
Formula 1 where Ri is H or alkyl of 1 to 6 carbons,
R2 is H, alkyl of 1 to 6 carbons or the Ri and R2 groups together with the nitrogen form a saturated or unsaturated 4, 5, 6 or 7 membered ring that optionally includes one or two heteroatoms independently selected from N, O and S, said 4, 5, 6 or 7 membered ring optionally being substituted with one or two COOH, CH2OH, OH, B(OH)2, cyano or halogen groups or with one or two alkyl groups having 1 to 6 carbons, or one or two carbons of said rings being attached to an oxygen to form keto groups and said 4, 5, 6 or 7 membered ring optionally being condensed with an aromatic or non-aromatic 5 or 6 membered ring that optionally includes 1 or heteroatoms selected from N, O and S;
R3 is independently selected from H, alkyl of 1 to 20 carbons, cycloalkyl of 3 to 6 carbons, aryl or heteroaryl, aryl-alkyl, aryl-(hydroxy)alkyl, heteroaryl-alkyl or hetero-(hydroxy)alkyl where the alkyl moiety has 1 to 4 carbons, said aryl or heteroaryl groups being optionally substituted with 1 to 3 groups independently selected from the group consisting of halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 6 carbons, or R3 is CO- R7, SO2R7 or CO-O-R7 where R7 is H, alkyl of 1 to 1 to 20 carbons, alkyl of 1 to 20 carbons substituted with and NH2 group or with an NH-COalkyl group where the alkyl group has one to 6 carbons, aryl or heteroaryl, aryl-alkyl or heteroaryl-alkyl where the alkyl moiety has 1 to 4 carbons, said aryl or heteroaryl groups being optionally substituted with 1 to 3 groups independently selected from the group consisting of halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 6 carbons; R4 is H, alkyl of 1 to 6 carbons or CO-Rs where Rs is alkyl of 1 to 6 carbons; the wavy lines represent bonds connected to carbons having R or S configuration; the dashed lines represent a bond or absence of a bond with the proviso that the ring containing the dashed lines is aromatic; m, n and q are integers independently selected from O, 1 , 2 or 3 with the proviso that the sum of m, n and q is 2 or 3; s is zero (O) or when X is N then s is zero (O) or 1 ; W, X and Y independently represent a CH, CR5, CRε or a heteroatom selected independently of N, O and S, and
R5 and Rε are independently selected from H, halogen, alkyl of 1 to 6 carbons, halogen substituted alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 6 carbons, phenyl, or
R5 and Rε together with the atoms to which they are attached jointly form a carbocyclic or a heterocyclic ring, the carbocyclic ring having 5 or 6 atoms in the ring, the heterocyclic ring having 5 or 6 atoms in the ring and 1 to 3 heteroatoms independently selected from N, O and S; said carbocyclic or heterocyclic ring jointly formed by R5 and R& being optionally substituted with 1 to 6 Rg groups where Rg is independently selected from halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 6 carbons or a pharmaceutically acceptable salt of said compound with the proviso that Formula 1 does not cover compounds where R4 is H, Ri and R2 jointly with the nitrogen form a pyrrolidino or morpholino ring, the sum of m, n and q is 3, and none of W, X and Y represent a heteroatom with the further proviso that the formula does not cover the compounds of the formula below:
Figure imgf000008_0001
DL-erythro
The present invention is also directed to methods of treating types of pain using the compounds of Formula 2
Figure imgf000009_0001
Formula 1
where Ri is H or alkyl of 1 to 6 carbons,
R2 is H, alkyl of 1 to 6 carbons or the Ri and R2 groups together with the nitrogen form a saturated or unsaturated 4, 5, 6 or 7 membered ring that optionally includes one or two heteroatoms independently selected from N, O and S, said 4, 5, 6 or 7 membered ring optionally being substituted with one or two COOH, CH2OH, OH, B(OH)2, cyano or halogen groups or with one or two alkyl groups having 1 to 6 carbons, or one or two carbons of said rings being attached to an oxygen to form keto groups and said 4, 5, 6 or 7 membered ring optionally being condensed with an aromatic or non-aromatic 5 or 6 membered ring that optionally includes 1 or heteroatoms selected from N, O and S; R3 is independently selected from H, alkyl of 1 to 20 carbons, cycloalkyl of 3 to 6 carbons, aryl or heteroaryl, aryl-alkyl, aryl-(hydroxy)alkyl, heteroaryl-alkyl or hetero-(hydroxy)alkyl where the alkyl moiety has 1 to 4 carbons, said aryl or heteroaryl groups being optionally substituted with 1 to 3 groups independently selected from the group consisting of halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 6 carbons, or R3 is CO- R7, SO2R7 or CO-O-R7 where R7 is H, alkyl of 1 to 1 to 20 carbons, alkyl of 1 to 20 carbons substituted with an NH2, NHCOR7 or NHCOOR7 group, aryl or heteroaryl, aryl-alkyl or heteroaryl-alkyl where the alkyl moiety has 1 to 4 carbons, said aryl or heteroaryl groups being optionally substituted with 1 to 3 groups independently selected from the group consisting of halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 6 carbons; the wavy lines represent bonds connected to carbons having R or S configuration; the dashed lines represent a bond or absence of a bond with the proviso that the ring containing the dashed lines is aromatic; Rg and R-io are independently H, alkyl of 1 to 6 carbons or ORn, or Rg and R-io jointly represent NORn with the proviso that when the dashed lines between carbons 2 and 3 of the propionic acid moiety represents a bond then Ri0 does not exist and Rg is not ORn with the further proviso that when Rg is ORn then
R-io is not hydrogen; Rn is H, alkyl of 1 to 6 carbons or CO-R12 where R12 is alkyl of 1 to 6 carbons; m, n and q are integers independently selected from O, 1 , 2 or 3 with the proviso that the sum of m, n and q is 2 or 3; s is zero (O) or when X is N then s is zero (O) or 1 ;
W, X and Y independently represent a CH, CR5, CRε or a heteroatom selected independently of N, O and S, and
R5 and Rε are independently selected from H, halogen, alkyl of 1 to 6 carbons, halogen substituted alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 6 carbons, phenyl, or
R5 and Rε together with the atoms to which they are attached jointly form a carbocyclic or a heterocyclic ring, the carbocyclic ring having 5 or 6 atoms in the ring, the heterocyclic ring having 5 or 6 atoms in the ring and 1 to 3 heteroatoms independently selected from N, O and S; said carbocyclic or heterocyclic ring jointly formed by R5 and Re being optionally substituted with 1 to 6 Rg groups where Rg is independently selected from halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carb'ons and thioxy of 1 to 6 carbons or a pharmaceutically acceptable salt of said compound.
Any of the compounds described here may be used to treat a patient suffering from one or more types of chronic pain including neuropathic pain, inflammatory pain, headache pain, somatic pain, visceral pain, and referred pain.
DETAILED DESCRIPTION OF THE INVENTION Most compounds that are useful in the method of the invention contain one or more asymmetric centers, such that the compounds may exist in enantiomeric as well as in diastereomeric forms. In fact, most of the compounds of the present invention have two asymmetric carbons adjacent to one another and therefore can exist in erythro or threo form, with each of these two forms having dextrorotatory (D) or levorotary (L) enantiomers. Although the threo form is generally preferred in accordance with the present invention for analgesic activity, unless it is specifically noted otherwise, the scope of the present invention includes all enantiomers, diastereomers and diastereomeric or racemic mixtures. In light of the foregoing, it should be clearly understood that the designation "DL" or "(+/-)" or "(±)" in this application includes the pure dextrorotatory enantiomer, the pure levorotatory enantiomer and all racemic mixtures, including mixtures where the two enantiomers are present in equal or in unequal proportions. Moreover, for simplicity sake in many of the structural formulas, such as in the example below, only one of the enantiomers is actually shown but when the designation "DL" or "(+/-)" or or "(±)" appears it also includes the enantiomeric form (mirror image) of the structure actually shown in the formula. For example:
Figure imgf000011_0001
HCI
DL-threo (Only one enantiomer shown)
Thus, in the example above, only one enantiomer is shown, but because the designation "DL" (or or "(+/-)" or "(±)") appears below the formula, its optical isomer
Figure imgf000012_0001
HCI
DL-threo (the other enantiomer shown) and all racemic mixtures of the two optical isomers are also included. In the case of some compounds of the present invention one enantiomer of the threo, and in some cases of the erythro, enantiomers is significantly more active as an analgesic than the other enantiomer of the same pair. For this reason the isolated enantiomer which is significantly more active than the other is considered a novel and inventive composition even if the racemic mixture or the other opposite enantiomer of the same compound have already been described in the prior art.
Some of the compounds that are useful in the method of the present invention contain three or more asymmetric centers. An example is the following compound
Figure imgf000012_0002
(2S.3R) & (2R.3S)
Compound 214 named Compound 214 in the description. The formula shown in the description for Compound 214 indicates two compounds of the threo isomer, but the two compounds indicated are not mirror images of each other, they are diastereomers. Another isomer pair is shown and described as Compound 215.
Figure imgf000013_0001
(2S,3R) & (2R,3S)
Compound 215
Keeping the foregoing examples in mind the reader one of ordinary skill in the art should readily understand the scope of each described example, although in a broad sense all isomers, enantiomers and racemic mixtures are within the scope of the invention.
The term "alkyl" in the general description and definition of the compounds includes straight chain as well as branch-chained alkyl groups.
Generally speaking the compounds of the invention may form salts with pharmaceutically acceptable acids or bases, and such pharmaceutically acceptable salts of the compounds of Formula 1 and of Formula 2 are also within the scope of the invention.
Referring now to the novel compounds of Formula 1 , in a class of preferred compounds of the invention none of the W, X and Y groups is a heteroatom. Within this class, compounds are preferred where the sum of m, n and q is 3 and the aromatic group is unsubstituted or substituted with one or more halogen, alkyl of 1 to 6 carbons, or halogen substituted alkyl of 1 to 6 carbons. Compounds within this class are also preferred where the R5 and Re groups form a carbocyclic ring, or a heterocyclic ring.
In another class of preferred compounds in accordance with Formula 1 one of the variables W, X and Y represents a heteroatom, preferably nitrogen and the sum of m, n and q is 3.
In still another class of preferred compounds in accordance with Formula 1 one or two of the variables W, X and Y represent a heteroatom, selected from N, O or S and the sum of m, n and q is 2. Referring still to the compounds of Formula 1 , compounds are preferred where R4 is H or an acyl group, more preferably H.
With reference to the variables R3, compounds in accordance with Formula 1 are preferred where both R3 groups are Hand where one R3 group is H and the other is benzyl, monohalogeno, dihalogeno, methyl or methoxy substituted benzyl, cyclohexyl, an alkyl of 1 to 7 carbons, COR7, COOR7 where R7 is alkyl of 1 to 15 carbons, benzyloxy, phenyl, methoxyphenyl, monohalogen or dihalogeno substituted phenyl, a 2-hydroxy-1-phenylethyl group or an alkyl group of 1 to 20 carbons itself substituted with an NH2, NHCOR7, or NHCOOR7 group.
Referring now to the variables Ri and R2 in the compounds of Formula 1 , compounds are preferred in accordance with the invention where Ri and R2 jointly form a pyrrolidine, a 3-fluoro or a 3,3-difluoro or an 3-hydroxy substituted pyrrolidine, a morpholine, a thiomorpholine, a piperazine, an alkyl substituted piperazine where the alkyl group has 1 to 6 carbons, an azetidine, a tetrahydrothiazole, an indoline, or a 2/-/-pyrrol ring or Ri and R2 are two alkyl groups of 1 to 3 carbons.
Referring now to the novel compounds of Formula 2, with respect to the variables W, X, Y, m, n, q, R1, R2, R5, Re, R3 compounds are gerenerally preferred in which these variables have the same preferences as in compounds of Formula 1.
With respect to Rg and R1O, compounds are generally preferred where Rg and R1O are both hydrogen, where one of these two variables is hydroxy and the other is alkyl of 1 to 6 carbons, where the Rg and R1O groups jointly form an NOR11 group, and where Rg is hydrogen, the dashed line between carbons 2 and 3 represent a double bond and R1O does not exist. With respect to R11 compounds of Formula 2 are preferred where R11 is H, or COR-12 where R12 is alkyl of 1 to 3 carbons. Presently still more preferred are Compounds of Formula 2 where R1 and R2 jointly with the nitrogen form a five-membered ring, where both R3 groups are hydrogen and where one of the R3 groups is hydrogen and the other is formyl.
The presently most preferred novel compounds of the invention are disclosed with their structural formulas in the ensuing Tables and or description, showing activity of exemplary compounds relevant to their ability to act as analgesics. BIOLOGICAL ACTIVITY, MODES OF ADMINISTRATION The compounds described here may be used to treat a patient suffering from one or more types of chronic pain, including neuropathic pain, inflammatory pain, headache pain, somatic pain, visceral pain, and referred pain.
To "treat," as used here, means to deal with medically. It includes, for example, administering a compound of the invention to prevent a pain, to alleviate its severity, and to prevent its reoccurance.
The term "pain," as used here, means any unpleasant sensory experience, usually associated with a physical disorder. The physical disorder may or may not be apparent to a clinician. Pain is of two types: chronic and acute. An "acute pain" is a pain of short duration having a sudden onset. One type of acute pain, for example, is cutaneous pain felt on injury to the skin or other superficial tissues, such as caused by a cut or a burn. Cutaneous nociceptors terminate just below the skin, and due to the high concentration of nerve endings, produce a well-defined, localized pain of short duration. "Chronic pain" is a pain other than an acute pain. Chronic pain includes neuropathic pain, inflammatory pain, headache pain, somatic pain visceral pain and referred pain.
/. Neuropathic Pain
The compounds of the invention may be used to treat pain caused by or otherwise associated with any of the following neuropathic pain conditions. "Neuropathic pain" means abnormal sensory input, resulting in discomfort, from the peripheral nervous system, central nervous systems, or both. A. Symptoms of neuropathic pain
Symptoms of neuropathic pain can involve persistent, spontaneous pain, as well as allodynia (a painful response to a stimulus that normally is not painful), hyperalgesia (an accentuated response to a painful stimulus that usually causes only a mild discomfort, such as a pin prick), or hyperpathia (where a short discomfort becomes a prolonged severe pain). B. Causes of neuropathic pain
Neuropathic pain may be caused by any of the following.
1. A traumatic insult, such as, for example, a nerve compression injury (e.g., a nerve crush, a nerve stretch, a nerve entrapment or an incomplete nerve transsection); a spinal cord injury (e.g., a hemisection of the spinal cord); a limb amputation; a contusion; an inflammation (e.g., an inflammation of the spinal cord); or a surgical procedure.
2. An ischemic event, including, for example, a stroke and heart attack.
3. An infectious agent 4. Exposure to a toxin, including, for example, a drug, an alcohol, a heavy metal (e.g., lead, arsenic, mercury), an industrial agent (e.g., a solvent, fumes from a glue) or nitrous oxide.
5. A disease, including, for example, an inflammatory disorder, a neoplastic tumor, an acquired immune deficiency syndrome (AIDS), Lymes disease, a leprosy, a metabolic disease, a neurodegenerative disease, a spinal stenosis, a mononeuropathy, a polyneuropathy, and a peripheral nerve disorder, such as a neuroma.
C. Types of neuropathic pain 1. Neuralgia. A neuralgia is a pain that radiates along the course of one or more specific nerves usually without any demonstrable pathological change in the nerve structure. The causes of neuralgia are varied. Chemical irritation, inflammation, trauma (including surgery), compression by nearby structures (for instance, tumors), and infections may all lead to neuralgia. In many cases, however, the cause is unknown or unidentifiable. Neuralgia is most common in elderly persons, but it may occur at any age. A neuralgia, includes, without limitation, a trigeminal neuralgia, a spinal stenosis, a post-herpetic neuralgia, a postherpetic neuralgia, a glossopharyngeal neuralgia, pain associated with nerve entrapment disorders, a sciatica and an atypical facial pain. Neuralgia is a painful disorder of the cranial nerves. Falling under the category of neuralgia are trigeminal neuralgia (TN), atypical facial pain, and postherpetic neuralgia (caused by shingles or herpes). The affected nerves are responsible for sensing touch, temperature and pressure in the facial area from the jaw to the forehead. The disorder generally causes short episodes of excruciating pain, usually for less than two minutes and on only one side of the face. The pain can be described in a variety of ways such as "stabbing," "sharp," "like lightning," "burning," and even "itchy". In the atypical form of TN, the pain can also present as severe or merely aching and last for extended periods. The pain associated with TN is recognized as one the most excruciating pains that can be experienced.
Simple stimuli such as eating, talking, washing the face, or any light touch or sensation can trigger an attack (even the sensation of a gentle breeze). The attacks can occur in clusters or as an isolated attack.
Symptoms include sharp, stabbing pain or constant, burning pain located anywhere, usually on or near the surface of the body, in the same location for each episode; pain along the path of a specific nerve; impaired function of affected body part due to pain, or muscle weakness due to concomitant motor nerve damage; increased sensitivity of the skin or numbness of the affected skin area (feeling similar to a local anesthetic such as a Novacaine shot); and any touch or pressure is interpreted as pain. Movement may also be painful. Trigeminal neuralgia is the most common form of neuralgia. It affects the main sensory nerve of the face, the trigeminal nerve ("trigeminal" literally means "three origins", referring to the division of the nerve into 3 branches). This condition involves sudden and short attacks of severe pain on the side of the face, along the area supplied by the trigeminal nerve on that side. The pain attacks may be severe enough to cause a facial grimace, which is classically referred to as a painful tic (tic douloureux). Sometimes, the cause of trigeminal neuralgia is a blood vessel or small tumor pressing on the nerve. Disorders such as multiple sclerosis (an inflammatory disease affecting the brain and spinal cord), certain forms of arthritis, and diabetes (high blood sugar) may also cause trigeminal neuralgia, but a cause is not always identified. In this condition, certain movements such as chewing, talking, swallowing, or touching an area of the face may trigger a spasm of excruciating pain.
A related but rather uncommon neuralgia affects the glosso-pharyngeal nerve, which provides sensation to the throat. Symptoms of this neuralgia are short, shock-like episodes of pain located in the throat.
Neuralgia may occur after infections such as shingles, which is caused by the varicella-zoster virus, a type of herpesvirus. This neuralgia produces a constant burning pain after the shingles rash has healed. The pain is worsened by movement of or contact with the affected area. Not all of those diagnosed with shingles go on to experience postherpetic neuralgia, which can be more painful than shingles. The pain and sensitivity can last for months or even years. The pain is usually in the form of an intolerable sensitivity to any touch but especially light touch. Postherpetic neuralgia is not restricted to the face; it can occur anywhere on the body but usually occurs at the location of the shingles rash. Depression is not uncommon due to the pain and social isolation during the illness.
Postherpetic neuralgia may be debilitating long after signs of the original herpes infection have disappeared. Other infectious diseases that may cause neuralgia are syphilis and Lyme disease. Diabetes is another common cause of neuralgia. This very common medical problem affects almost 1 out of every 20 Americans during adulthood. Diabetes damages the tiny arteries that supply circulation to the nerves, resulting in nerve fiber malfunction and sometimes nerve loss. Diabetes can produce almost any neuralgia, including trigeminal neuralgia, carpal tunnel syndrome (pain and numbness of the hand and wrist), and meralgia paresthetica (numbness and pain in the thigh due to damage to the lateral femoral cutaneous nerve). Strict control of blood sugar may prevent diabetic nerve damage and may accelerate recovery in patients who do develop neuralgia. Other medical conditions that may be associated with neuralgias are chronic renal insufficiency and porphyria - a hereditary disease in which the body cannot rid itself of certain substances produced after the normal breakdown of blood in the body. Certain drugs may also cause this problem.
2. Deafferentation.
Deafferentation indicates a loss of the sensory input from a portion of the body, and can be caused by interruption of either peripheral sensory fibres or nerves from the central nervous system. A deafferentation pain syndrome, includes, without limitation, an injury to the brain or spinal cord, a post-stroke pain, a phantom pain, a paraplegia, a brachial plexus avulsion injuries, lumbar radiculopathies. 3. Complex regional pain syndromes (CRPSs)
CRPS is a chronic pain syndrome with two forms. CRPS 1 currently replaces the term "reflex sympathetic dystrophy syndrome". It is a chronic nerve disorder that occurs most often in the arms or legs after a minor or major injury. CRPS 1 is associated with severe pain; changes in the nails, bone, and skin; and an increased sensitivity to touch in the affected limb. CRPS 2 replaces the term causalgia, and results from an identified injury to the nerve. A CRPS, includes, without limitation, a CRPS Type I (reflex sympathetic dystrophy) and a CRPS Type Il (causalgia).
4. Neuropathy. A neuropathy is a functional or pathological change in a nerve and is characterized clinically by sensory or motor neuron abnormalities.
Central neuropathy is a functional or pathological change in the central nervous system.
Peripheral neuropathy is a functional or pathological change in one or more peripheral nerves. The peripheral nerves relay information from your central nervous system (brain and spinal cord) to muscles and other organs and from your skin, joints, and other organs back to your brain. Peripheral neuropathy occurs when these nerves fail to carry information to and from the brain and spinal cord, resulting in pain, loss of sensation, or inability to control muscles. In some cases, the failure of nerves that control blood vessels, intestines, and other organs results in abnormal blood pressure, digestion problems, and loss of other basic body processes. Risk factors for neuropathy include diabetes, heavy alcohol use, and exposure to certain chemicals and drugs. Some people have a hereditary predisposition for neuropathy. Prolonged pressure on a nerve is another risk for developing a nerve injury. Pressure injury may be caused by prolonged immobility (such as a long surgical procedure or lengthy illness) or compression of a nerve by casts, splints, braces, crutches, or other devices. Polyneuropathy implies a widespread process that usually affects both sides of the body equally. The symptoms depend on which type of nerve is affected. The three main types of nerves are sensory, motor, and autonomic. Neuropathy can affect any one or a combination of all three types of nerves. Symptoms also depend on whether the condition affects the whole body or just one nerve (as from an injury). The cause of chronic inflammatory polyneuropathy is an abnormal immune response. The specific antigens, immune processes, and triggering factors are variable and in many cases are unknown. It may occur in association with other conditions such as HIV, inflammatory bowel disease, lupus erythematosis, chronic active hepatitis, and blood cell abnormalities.
Peripheral neuropathy may involve a function or pathological change to a single nerve or nerve group (monneuropathy) or a function or pathological change affecting multiple nerves (polyneuropathy).
Peripheral neuropathies Hereditary disorders
Charcot-Marie-Tooth disease
Friedreich's ataxia Systemic or metabolic disorders
Diabetes (diabetic neuropathy )
Dietary deficiencies (especially vitamin B-12)
Excessive alcohol use (alcoholic neuropathy )
Uremia (from kidney failure ) Cancer (including bone cancer and other cancers)
Infectious or inflammatory conditions
AIDS Hepatitis
Colorado tick fever
Diphtheria
Guillain-Barre syndrome HIV infection without development of AIDS
Leprosy
Lyme disease
Polyarteritis nodosa
Rheumatoid arthritis Sarcoidosis
Sjogren's syndrome
Syphilis
Systemic Lupus erythematosus amyloid Exposure to toxic compounds
Sniffing glue or other toxic compounds
Nitrous oxide
Industrial agents - especially solvents
Heavy metals (lead, arsenic, mercury, etc.) Neuropathy secondary to drugs like analgesic nephropathy
Rhabdomyolysis
Macrohagic myofasciitis
Highly Active Anti-Retrviral Therapy (HAART)-induced neuropathy Chemotherapy lncuced Neuropathy
Miscellaneous causes
Ischemia (decreased oxygen/decreased blood flow)
Prolonged exposure to cold temperature a. Polyneuropathy Polyneuropathy is a peripheral neuropathy involving the loss of movement or sensation to an area caused by damage or destruction to multiple peripheral nerves. Polyneuropathic pain, includes, without limitation, post-polio syndrome, postmastectomy syndrome, diabetic neuropathy, alcohol neuropathy, amyloidosis, toxin exposure, AIDS, hypothyroidism, uremia, vitamin deficiencies, chemotherapy-induced pain, 2',3'-didexoycytidine (ddC) treatment, exposure to the anticonvulsant phenytoin, exposure to antibiotics including chloramphenicol, nitrofurantoin and sulfonamineds, exposure to sedatives including barbital and hexobarbital, Guillain-Barre syndrome, Fabry's disease or polyneuropathy secondary to cancers such as multiple myeloma. b. Mononeuropathy Mononeuropathy is a peripheral neuropathy involving loss of movement or sensation to an area caused by damage or destruction to a single peripheral nerve or nerve group. Mononeuropathy is most often caused by damage to a local area resulting from injury or trauma, although occasionally systemic disorders may cause isolated nerve damage (as with mononeuritis multiplex). The usual causes are direct trauma, prolonged pressure on the nerve, and compression of the nerve by swelling or injury to nearby body structures. The damage includes destruction of the myelin sheath (covering) of the nerve or of part of the nerve cell (the axon). This damage slows or prevents conduction of impulses through the nerve. Mononeuropathy may involve any part of the body. Mononeuropathic pain, includes, without limitation, a sciatic nerve dysfunction, a common peroneal nerve dysfunction, a radial nerve dysfunction, an ulnar nerve dysfunction, a cranial mononeuropathy Vl, a cranial mononeuropathy VII, a cranial mononeuropathy III (compression type), a cranial mononeuropathy III (diabetic type), an axillary nerve dysfunction, a carpal tunnel syndrome, a femoral nerve dysfunction, a tibial nerve dysfunction, a Bell's palsy, a thoracic outlet syndrome, a carpal tunnel syndrome, and a sixth (abducent) nerve palsy. c. Generalized peripheral neuropathies Generalized peripheral neuropathis are symmetrical, and usually due to various systematic illnesses and disease processes that affect the peripheral nervous system in its entirety. They are further subdivided into several categories: i. Distal axonopathies are the result of some metabolic or toxic derangement of neurons. They may be caused by metabolic diseases such as diabetes, renal failure, deficiency syndromes such as malnutrition and alcoholism, or the effects of toxins or drugs. Distal axonopathy (aka dying back neuropathy) is a type of peripheral neuropathy that results from some metabolic or toxic derangement of peripheral nervous system (PNS) neurons. It is the most common response of nerves to metabolic or toxic disturbances, and as such may be caused by metabolic diseases such as diabetes, renal failure, deficiency syndromes such as malnutrition and alcoholism, or the effects of toxins or drugs. The most common cause of distal axonopathy is diabetes, and the most common distal axonopathy is diabetic neuropathy. ii. Myelinopathies are due to a primary attack on myelin causing an acute failure of impulse conduction. The most common cause is acute inflammatory demyelinating polyneuropathy (AIDP; aka Guillain-Barre syndrome), though other causes include chronic inflammatory demyelinating syndrome (CIDP), genetic metabolic disorders (e.g., leukodystrophy), or toxins. Myelinopathy is due to primary destruction of myelin or the myelinating Schwann cells, which leaves the axon intact, but causes an acute failure of impulse conduction. This demyelination slows down or completely blocks the conduction of electical impulses through the nerve. The most common cause is acute inflammatory demyelinating polyneuropathy (AIDP, better known as Guillain-Barre syndrome), though other causes include chronic inflammatory demyelinating polyneuropathy (CIDP), genetic metabolic disorders (e.g., leukodystrophy or Charcot-Marie-Tooth disease), or toxins. iii. Neuronopathies are the result of destruction of peripheral nervous system (PNS) neurons. They may be caused by motor neurone diseases, sensory neuronopathies (e.g., Herpes zoster), toxins or autonomic dysfunction. Neurotoxins may cause neuronopathies, such as the chemotherapy agent vincristine. Neuronopathy is dysfunction due to damage to neurons of the peripheral nervous system (PNS), resulting in a peripheral neuropathy. It may be caused by motor neurone diseases, sensory neuronopathies (e.g., Herpes zoster), toxic substances or autonomic dysfunction. A person with neuronopathy may present in different ways, depending on the cause, the way it affects the nerve cells, and the type of nerve cell that is most affected. iv. Focal entrapment neuropathies (e.g., carpal tunnel syndrome) represent an additional category of generalized peripheral neuropathies.
//. Inflammatory pain The compounds of the invention may be used to treat pain caused by or otherwise associated with any of the following inflammatory conditions.
A. Arthritic disorder
Arthritic disorders include, for example, a rheumatoid arthritis; a juvenile rheumatoid arthritis; a systemic lupus erythematosus (SLE); a gouty arthritis; a scleroderma; an osteoarthritis; a psoriatic arthritis; an ankylosing spondylitis; a Reiter's syndrome (reactive arthritis); an adult Still's disease; an arthritis from a viral infection; an arthritis from a bacterial infection, such as, e.g., a gonococcal arthritis and a non-gonococcal bacterial arthritis (septic arthritis); a Tertiary Lyme disease; a tuberculous arthritis; and an arthritis from a fungal infection, such as, e,g,, a blastomycosis
B. Autoimmune diseases
Autoimmune diseases include, for example, a Guillain-Barre syndrome, a Hashimoto's thyroiditis, a pernicious anemia, an Addison's disease, a type I diabetes, a systemic lupus erythematosus, a dermatomyositis, Sjogren's syndrome, a lupus erythematosus, a multiple sclerosis, a myasthenia gravis, a Reiter's syndrome, a Grave's disease, and a rheumatoid arthritis.
C. Connective tissue disorder
Connective tissue disorders include, for example, a spondylarthritis a dermatomyositis, and a fibromyalgia syndrome . D. Injury
Inflammation caused by injury, including, for example, a crush, puncture, stretch of a tissue or joint, may cause chronic inflammatory pain. E. Infection
Inflammation caused by infection, including, for example, a tuberculosis or an interstitial keratitis may cause chronic inflammatory pain. Infection may also result in inflammatory bowel diseases and irritable bowel syndromes. F. Neuritis
Neuritis is an inflammatory process affecting a nerve or group of nerves. Symptoms depend on the nerves involved, but may include pain, paresthesias, paresis, or hypesthesia (numbness).
Examples include: a. Brachial neuritis b. Retrobulbar neuropathy, an inflammatory process affecting the part of the optic nerve lying immediately behind the eyeball. c. Optic neuropathy, an inflammatory process affecting the optic nerve causing sudden, reduced vision in the affected eye. The cause of optic neuritis is unknown. The sudden inflammation of the optic nerve (the nerve connecting the eye and the brain) leads to swelling and destruction of the myelin sheath. The inflammation may occasionally be the result of a viral infection, or it may be caused by autoimmune diseases such as multiple sclerosis. Risk factors are related to the possible causes. d. Vestibular neuritis, a viral infection causing an inflammatory process affecting the vestibular nerve. G. Joint inflammation
Inflammation of the joint, such as that caused by bursitis or tendonitis, for example, may cause chronic inflammatory pain. ///. Headache Pain
The compounds of the invention may be used to treat pain caused by or otherwise associated with any of the following headache conditions. A headache (medically known as cephalgia) is a condition of mild to severe pain in the head; sometimes neck or upper back pain may also be interpreted as a headache. It may indicate an underlying local or systemic disease or be a disorder in itself. A. Muscular/myogenic headache Muscular/myogenic headaches appear to involve the tightening or tensing of facial and neck muscles; they may radiate to the forehead. Tension headache is the most common form of myogenic headache.
A tension headache is a condition involving pain or discomfort in the head, scalp, or neck, usually associated with muscle tightness in these areas. Tension headaches result from the contraction of neck and scalp muscles. One cause of this muscle contraction is a response to stress, depression or anxiety. Any activity that causes the head to be held in one position for a long time without moving can cause a headache. Such activities include typing or use of computers, fine work with the hands, and use of a microscope.
Sleeping in a cold room or sleeping with the neck in an abnormal position may also trigger this type of headache. A tension-type headache, includes, without limitation, an episodic tension headache and a chronic tension headache. B. Vascular headache The most common type of vascular headache is migraine. Other kinds of vascular headaches include cluster headaches, which cause repeated episodes of intense pain, and headaches resulting from high blood pressure 1. Migraine A migraine is a heterogeneous disorder that generally involves recurring headaches. Migraines are different from other headaches because they occur with other symptoms, such as, e.g., nausea, vomiting, or sensitivity to light. In most people, a throbbing pain is felt only on one side of the head. Clinical features such as type of aura symptoms, presence of prodromes, or associated symptoms such as vertigo, may be seen in subgroups of patients with different underlying pathophysiological and genetic mechanisms. A migraine headache, includes, without limitation, a migraine without aura (common migraine), a migraine with aura (classic migraine), a menstrual migraine, a migraine equivalent (acephalic headache), a complicated migraine, an abdominal migraine and a mixed tension migraine. 2. Cluster headache
Cluster headaches affect one side of the head (unilateral) and may be associated with tearing of the eyes and nasal congestion. They occurs in clusters, happening repeatedly every day at the same time for several weeks and then remitting.
D. High blood pressure headache
E. Traction and inflammatory headache Traction and inflammatory headaches are usually symptoms of other disorders, ranging from stroke to sinus infection.
F. Hormone headache
G. Rebound headache
Rebound headaches, also known as medication overuse headaches, occur when medication is taken too frequently to relieve headache. Rebound headaches frequently occur daily and can be very painful. H. Chronic sinusitis headache
Sinusitis is inflammation, either bacterial, fungal, viral, allergic or autoimmune, of the paranasal sinuses. Chronic sinusitis is one of the most common complications of the common cold. Symptoms include: Nasal congestion; facial pain; headache; fever; general malaise; thick green or yellow discharge; feeling of facial 'fullness' worsening on bending over. In a small number of cases, chronic maxillary sinusitis can also be brought on by the spreading of bacteria from a dental infection. Chronic hyperplastic eosinophilic sinusitis is a noninfective form of chronic sinusitis. I. An organic headache J. lctal headaches lctal headaches are headaches associated with seizure activity.
IV. Somatic pain
The compounds of the invention may be used to treat pain caused by or otherwise associated with any of the following somatic pain conditions. Somatic pain originates from ligaments, tendons, bones, blood vessels, and even nerves themselves. It is detected with somatic nociceptors. The scarcity of pain receptors in these areas produces a dull, poorly-localized pain of longer duration than cutaneous pain; examples include sprains and broken bones. Additional examples include the following.
A. Excessive muscle tension
Excessive muclse tension can be caused, for example, by a sprain or a strain.
B. Repetitive motion disorders
Repetitive motion disorders can result from overuse of the hands, wrists, elbows, shoulders, neck, back, hips, knees, feet, legs, or ankles.
C. Muscle disorders Muscle disorders causing somatic pain include, for example, a polymyositis, a dermatomyositis, a lupus, a fibromyalgia, a polymyalgia rheumatica, a macrophagic myofasciitis, and a rhabdomyolysis. Muscle pain can also be secondary to neurological and neuromuscular disorders including without limitation Parkinson's disease, Huntington's chorea, dystonias, tardive dyskinesias, drug-induced dyskinesias and dystonias, dyskinesias
(paroxysmal), amyotrophic lateral sclerosis, multiple sclerosis, myoclonus, progressive supranuclear palsy, corticobasal degeneration, choreoathetosis, spasticity, Wilson disease, multiple system atrophy (including Shy-Drager syndrome, striatonigral degeneration and olivopontocerebellar atrophy), and hereditary spastic paraplegia (including familial spastic paraparesis, familial spastic paraplegia, hereditary spastic paraparesis, Strumpell-Lorraine syndrome, and Strumpell's disease).
D. Myalgia
Myalgia is muscle pain and is a symptom of many diseases and disorders. The most common cause for myalgia is either overuse or overstretching of a muscle or group of muscles. Myalgia without a traumatic history is often due to viral infections. Longer-term myalgias may be indicative of a metabolic myopathy, some nutritional deficiencies or chronic fatigue syndrome. E. Infection
Infection can cause somatic pain. Examples of such infection include, for example, an abscess in the muscle, a trichinosis, an influenza, a Lyme disease, a malaria, a Rocky Mountain spotted fever, Avian influenza, the common cold, community-acquired pneumonia, meningitis, monkeypox, Severe Acute Respiratory Syndrome, toxic shock syndrome, trichinosis, typhoid fever, and upper respiratory tract infection. F. Drugs
Drugs can cause somatic pain. Such drugs include, for example, cocaine, statins for lowering cholesterol (such as atorvastatin, simvastatin, and lovastatin), and ACE inhibitors for lowering blood pressure (such as enalaphl and captophl). G. Prolonged nociceptive pain including without limitation to bone fracture pain, spinal stenosis, and post-surgical pain.
V. Visceral pain
The compounds of the invention may be used to treat pain caused by or otherwise associated with any of the following visceral pain conditions. Visceral pain originates from body's viscera, or organs. Visceral nociceptors are located within body organs and internal cavities. The even greater scarcity of nociceptors in these areas produces pain that is usually more aching and of a longer duration than somatic pain. Visceral pain is extremely difficult to localise, and several injuries to visceral tissue exhibit "referred" pain, where the sensation is localised to an area completely unrelated to the site of injury. Examples of visceral pain include the following.
A. Functional visceral pain
Functional visceral pain includes, for example, an irritable bowel syndrome and a chronic functional abdominal pain (CFAP), a functional constipation and a functional dyspepsia, a non-cardiac chest pain (NCCP) and a chronic abdominal pain.
B. Chronic gastrointestinal inflammation
Chronic gastrointestinal inflammation includes, for example, a gastritis, an inflammatory bowel disease, e.g., a Crohn's disease, an ulcerative colitis, a microscopic colitis, a diverticulitis and a gastroenteritis; an interstitial cystitis; an intestinal ischemia; a cholecystitis; an appendicitis; a gastroesophageal reflux; an ulcer, a nephrolithiasis, an urinary tract infection, a pancreatitis and a hernia.
C. Autoimmune pain
Autoimmune pain includes, for example, a sarcoidosis and a vasculitis. D. Organic visceral pain
Organic visceral pain includes, for example, pain resulting from a traumatic, inflammatory or degenerative lesion of the gut or produced by a tumor impinging on sensory innervation.
E. Treatment-induced visceral pain Treatment-induced visceral pain includes, for example, a pain attendant to chemotherapy therapy or a pain attendant to radiation therapy.
Vl. Referred pain The compounds of the invention may be used to treat pain caused by or otherwise associated with any of the following referred pain conditions.
Referred pain arises from pain localized to an area separate from the site of pain stimulation. Often, referred pain arises when a nerve is compressed or damaged at or near its origin. In this circumstance, the sensation of pain will generally be felt in the territory that the nerve serves, even though the damage originates elsewhere. A common example occurs in intervertebral disc herniation, in which a nerve root arising from the spinal cord is compressed by adjacent disc material. Although pain may arise from the damaged disc itself, pain will also be felt in the region served by the compressed nerve (for example, the thigh, knee, or foot). Relieving the pressure on the nerve root may ameliorate the referred pain, provided that permanent nerve damage has not occurred. Myocardial ischaemia (the loss of blood flow to a part of the heart muscle tissue) is possibly the best known example of referred pain; the sensation can occur in the upper chest as a restricted feeling, or as an ache in the left shoulder, arm or even hand. Pain Reversal
An art-accepted model or assay for measuring an analgesic effect of a compound in chronic pain (in particular peripheral neuropathy) is the model known as Kim and Chung 1992, Pain 150, pp 355-363 {Chung model). This model involves the surgical ligation of the L5 (and optionally the L6) spinal nerves on one side in experimental animals. Rats recovering from the surgery gain weight and display a level of general activity similar to that of normal rats. However, these rats develop abnormalities of the foot, wherein the hindpaw is moderately everted and the toes are held together. More importantly, the hindpaw on the side affected by the surgery appears to become sensitive to low-threshold mechanical stimuli and will perceive pain instead of the faint sensation of touch. This sensitivity to normally non-painful touch, called "tactile allodynia", develops within the first week after surgery and lasts for at least two months. The allodynia response includes lifting the affected hindpaw to escape from the stimulus, licking the paw and holding it in the air for many seconds. None of these responses is normally seen in the control group. To produce the tactile allodynia, rats are anesthetized before surgery.
The surgical site is shaved and prepared either with betadine or Novacaine. Incision is made from the thoracic vertebra XIII down toward the sacrum. Muscle tissue is separated from the spinal vertebra (left side) at the L4 - S2 levels. The L6 vertebra is located and the transverse process is carefully removed with a small rongeur to expose the L4 - L6 spinal nerves. The L5 and L6 spinal nerves are isolated and tightly ligated with 6-0 silk thread. The same procedure is done on the right side as a control, except no ligation of the spinal nerves is performed.
After a complete hemostasis is confirmed, the wounds are sutured. A small amount of antibiotic ointment is applied to the incised area, and the rat is transferred to the recovery plastic cage under a regulated heat-temperature lamp.
On the day of the experiment, at least seven days after the surgery, typically six rats per test group are administered the test drugs by intraperitoneal (i.p.) injection or oral gavage (p.o.). For i.p. administration, the compounds are formulated in H2O and given in a volume of 1 ml/kg body weight by injecting into the intraperitoneal cavity. For p.o. administration, the compounds are formulated in H2O and given in a volume of 1 ml/kg body weight using an 18-gauge, 3 inch gavage needle that is slowly inserted through the esophagus into the stomach.
Tactile allodynia is assessed via von Frey hairs, which are a series of fine hairs with incremental differences in stiffness. Rats are placed in a plastic cage with a wire mesh bottom and allowed to acclimate for approximately 30 minutes. To establish the pre-drug baseline, the von Frey hairs are applied perpendicularly through the mesh to the mid-plantar region of the rats' hindpaw with sufficient force to cause slight buckling and held for 6-8 seconds. The applied force has been calculated to range from 0.41 to 15.1 grams. If the paw is sharply withdrawn, it is considered a positive response. A normal animal will not respond to stimuli in this range, but a surgically ligated paw will be withdrawn in response to a 1-2 gram hair. The 50% paw withdrawal threshold is determined using the method of Dixon, W.J., Ann. Rev. Pharmacol. Toxicol. 20:441-462 (1980) hereby incorporated by reference. Tactile allodynia is measured prior to and 15, 30, and 60 minutes after drug administration. The post-drug threshold is compared to the pre-drug threshold and the percent reversal of tactile sensitivity is calculated based on a normal threshold of 15.1 grams. Table 1 below indicates the degree of pain reversal obtained in the
Chung model with exemplary compounds of the invention. The intraperitonial (i.p.) and/or intravenous (iv) administration of the compounds was in doses ranging from 1 μg/kg to 300 μg/kg or 3 mg/kg PO and the peak percentage of reversal of allodynia was measured at 15, 30 or 60 minutes after administration, as is indicated in the table. Data are expressed as the highest % allodynia reversal (out of 3 time points: 15 min, 30 min, or 60 min. post- drug) with a minimum of a 20% allodynia reversal in the rat Chung model. Comparisons between groups (drug treated vs. saline treated) were made using a two-tailed, 2-sample, unpaired t-test. Compounds that are not shown which were not statistically analgesic following an IP dose of 300 ug/kg, but may still be analgesic. Compounds that do not exhibit significant analgesia at 100 mg/kg are not considered to be analgesic.
Figure imgf000033_0001
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Figure imgf000044_0001
Modes of Administration:
The compounds of the invention may be administered at pharmaceutically effective dosages. Such dosages are normally the minimum dose necessary to achieve the desired therapeutic effect; in the treatment of chronic pain, this amount would be roughly that necessary to reduce the discomfort caused by the pain to tolerable levels. For human adults such doses generally will be in the range of 0.1-5,000 mg/day; more preferably in the range of 1 to 3,000 mg/day, 10 mg to 500 mg/day, 500 to 1 ,000 mg/day, 1 ,000 to 1 ,500 mg/day, 1 ,500 to 2,000 mg/day, 2,000 to 2,500 mg/day, or 2,500 to 3,000 mg/day. However, the actual amount of the compound to be administered in any given case will be determined by a physician taking into account the relevant circumstances, such as the severity of the pain, the age and weight of the patient, the patient's general physical condition, the cause of the pain, and the route of administration.
The compounds are useful in the treatment of pain in a mammal; particularly a human being. Preferably, the patient will be given the compound orally in any acceptable form, such as a tablet, liquid, capsule, powder and the like. However, other routes may be desirable or necessary, particularly if the patient suffers from nausea. Such other routes may include, without exception, transdermal, intrapehtonial, parenteral, subcutaneous, intranasal, intrathecal, intramuscular, intravenous and intrarectal modes of delivery. Compositions useful in the method of the invention may further include an excipient. Such an excipient may be a carrier or a diluent; this is usually mixed with the active compound, or permitted to dilute or enclose the active compound. If a diluent, the carrier may be solid, semi-solid, or liquid material that acts as an excipient or vehicle for the active compound. The formulations may also include wetting agents, emulsifying agents, preserving agents, sweetening agents, and/or flavoring agents. If used as in an ophthalmic or infusion format, the formulation will usually contain one or more salt to influence the osmotic pressure of the formulation. SYNTHETIC METHODS FOR OBTAINING THE COMPOUNDS OF THE
INVENTION, EXPERIMENTAL
The compound of the invention can be synthesized by utilizing the synthetic methods described in the experimental below, or such modifications of the below described experimental methods which will become readily apparent to those skilled in the art in light of the present disclosure. GENERAL
1H NMR spectra were recorded at ambient temperature with an Avance 300 (Bruker) spectrometer. The compounds were analyzed by reverse phase high performance liquid chromatography (HPLC) using a Waters
Autopurification System equipped with a Waters 2525 Pump, a Waters 2696 photodiode array detector, and a XTerra column (Part. No. 186000482, 5 μm, C18, 4.5 x 50 mm). The HPLC method used was a gradient of 5 % solvent B to 100 % in 7 min. Solvent A was H2O with 0.05 % TFA and solvent B was CH3CN with 0.05 % TFA (Method A).
Melting points were measured with a Buchi B-545 melting point apparatus and were uncorrected. To isolate reaction products the solvent were removed by evaporation using a vacuum rotatory evaporator, the water bath temperature not exceeding 40 0C.
Absolute configuration of compounds of the invention, where applicable, can generally speaking be determined in accordance with methods known in the state of the art, such as X-ray chhstallography. Compounds 203 and 204 are mentioned as examples for which the absolute configurations were determined by X-ray chhstallography analysis of the corresponding (1 S)- camphanylamide D-(10) camphorsulfonic acid salt. As a result Compound 204 was assigned (2S, 3R). Its enantiomer, Compound 203 was assigned by default the (2R, 3S) absolute configuration. GENERAL SYNTHETIC ROUTES The compound of the invention can be synthesized by utilizing the synthetic methods described in a general sense immediately below and in more detail in the experimental section of the present application, or by such modifications of the below described experimental methods which will become readily apparent to those skilled in the art in light of the present disclosure.
A general synthetic route to the novel compounds of the invention which are amides of substituted (+/-)-f/?reo-3-hydroxy-2-aminopropionic acid of the Generalized Structure 1 is described below.
OH O
NH2 R**
General Structure 1
In General Structure 1 , for the sake of simplicity of description R* substantially corresponds to the 5, 6, or 7 membered ring structure on the left side of Formula 1 (as the formula is depicted in the Summary and in the instant claims) and R** substantially corresponds to the Ri groups in Formula 1.
Figure imgf000048_0001
methylisocyanoacetate 'amine" 2-Isocyano-l-(subst.-amino)ethanone
R* -CHO strong base "aldehyde" (KOH)
Figure imgf000048_0002
(+l-)-threo ft*««s-oxazoline General Structure 1
General Reaction Scheme 1
Thus, in accordance with General Scheme 1 , methyl isocyanoacetate (or ethyl isocyanoacetate available commercially) is reacted with an "amine" which includes the R** groups to provide the 2-isocyanoacetic acid amide derivative shown in the general scheme. Typical examples for the amines used in the reaction are pyrrolidine, pipehdine, azetidine, morpholine, 2,5- dihydro-1 H-pyrrole, dialkylamines such as diethylamine, 3-fluoro-, 3,3-difluoro or 3-hydroxy substituted pyrrolidines. Specific examples of these "amines" abound in the experimental description. The 2-isocyanoacetic acid amide derivative is then reacted in the presence of base (such as KOH) with an "aldehyde" which includes the R* group to provide a trans "oxazoline" with high diastereoselectivity (transxis ratios generally > 97:3) as shown in the general reaction scheme 1. The trans oxazoline is then treated with a strong acid, such as HCI, to open the ring and to provide the f/?reo-3-substituted-3- hydroxy-2-amino-propionic acid amides (with threo:erythro ratios generally > 97:3) of the invention as shown in General Scheme 1.
Compounds of Formula 1 where the amino group of formula NH(R**)2 is a weaker nucleophile, such as indoline, thiomorpholine and the like, can be made as illustrated in Reaction Scheme 2 for the synthesis of (±)-threo-2- amino-3-hydroxy-1 -(indolin-1 -yl)-3-(pyhdin-4-yl)propan-1 -one dihydrochlohde Compound 243 and (±)-f/?reo-2-amino-3-hydroxy-1-(thiazolidin-3-yl)-3- (pyhdin-4-yl)propan-1-one dihydrochloride Compound 242.
Figure imgf000050_0001
R = Me, terf-butyl a)
% yield
Figure imgf000050_0002
(±) fΛreo
Compound 242
a) KOH, MeOH; b) Indoline, EDCI, TEA, HOBT, CH2CI2. c) HCI (1 M) in MeOH d) i. Silica Gel Chromatography.ii HCI (0.1 M) in /-PrOH e) BOC2O, NaOH, Dioxane. f) Thiazolidine, EDCI, TEA, HOBT, CH2CI2; g) Silica Gel Chromatography g) HCI (1 M) in MeOH.
Reaction Scheme 2
In Reaction Scheme 2 EDCI stands for 1-(3-dimethylaminopropyl)- ethylcarbodiimide hydrochloride; HOBT stands for 1-hydroxybenzothazole; BOC2O stands for di-f-butyl-dicarbonate and TEA stands for triethylamine. As it will be readily understood by those skilled in the art, for a more general synthetic route, such as the one shown in Reaction Scheme 2, the A- pyridyl group can be substituted with an R* group (as defined in connection with Scheme 1 ) and the indoline can be susbstituted with other weak nucleophilic amines of the formula NH(R**)2 (R** defined as in connection with Reaction Scheme 1 ) to provide other compounds of Formula 1 analogous to compounds 242 and 243. lsomehcally pure and/or enantiomerically pure compounds and further derivatives of the 3-substituted-3-hydroxy-2-amino-propionic acid amides are obtained by separation techniques and reactions which, per se, are well known to the synthetic chemist. The experimental section of the present invention abounds in examples of such separation techniques and reactions. Some of the typical separation techniques and reactions are generally described below.
Separation of threo and erythro isomers, when both are formed in the reactions leading to the compounds of the invention, can typically be separated by chromatographic methods. The more abundantly formed threo isomers can also be converted into the erythro isomers by oxidizing to the ketone level the hydroxyl group in the 3 position of the propanoic acid moiety and subsequently reducing the resulting ketone to the hydroxyl level. (See, for example, the preparation of Compound 219). Separation of enantiomeric mixtures can be performed on Chiralpack columns which are well known in the art. (See, for example, the preparation of Compound 204).
The amino function in the 2-position of the propanoic acid moiety is, generally speaking, more reactive towards acylation and carbamoylation than the hydroxyl group in the 3 position. Therefore, acylated derivatives of the 2- amino function can be prepared by using acyl chlorides such as acetyl chloride and hexanoyl chloride. (See, Method G and the preparation of Compound 51 ). Carbamate derivatives of the 2-amino function can be obtained by using chloroformates, such as benzylchloroformate. (See, for example, the preparation of Compound 58). The tertiary butyl carbamoyl function can also serve as a removable protecting group of the 2-amino function, (see for example the preparation of Compounds 219 and 224). When the 2-amino function of the compounds of the invention is already acylated or bears a carbamoyl group, then the 3-hydroxy group of the propanoic acid moiety can be subjected to acylation by reagents such as acetic anhydride. (See for example the preparation of Compound 217). Alkylation of the 2-amino function is readily performed by condensing the compound bearing the 2-NH2 group with an aldehyde to obtain a Schiff base intermediate which can be reduced, without isolation, to provide the N- alkyl, arylalkyl or heteroaryl-alkyl compound. The procedure described for preparing Compound 234 can be generalized to make compounds of the invention where the 2-amino function bears an aryl( hydroxy)alkyl or heteroaryl(hydroxy)alkyl group.
Several compounds of the invention of Formula 2 can be obtained by derivatization of compounds of Formula 1 , or by such modification of the synthetic routes leading to compounds of Formula 1 which will become readily apparent to those skilled in the art in light of the present disclosure. For example, compounds of Formula 2 where Rg is OH or ORn and R-m is alkyl can be made by using a "ketone" bearing the R-io group, instead of the "aldehyde" in General Reaction Scheme 1.
Compounds of Formula 2 where the Rg and R-m groups jointly form an oxime (NOH) group can be obtained by oxidizing the 3-hydroxyl group of the propanoic acid moiety to the ketone stage and reacting the resulting ketone with hydroxylamine.
Another general synthetic route for making several compounds of Formula 2 is illustrated in Synthetic Scheme 3 adapted for synthesizing (R)- 2-amino-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochlohde, Compound 236 of the present invention.
Figure imgf000053_0001
Synthetic Scheme 3
Figure imgf000053_0002
2 HCI
Compound 236
As it will be readily understood by those skilled in the art, for a more general synthetic route, such as the one shown in Reaction Shyntetic Scheme 3, the 4-pyhdyl group can be substituted with an R* group (as defined in connection with Scheme 1 ) and the pyrrolidine can be substituted with amines of the formula NH(R**)2 (R** defined as in connection with Reaction Scheme 1 ) to provide other compounds of Formula 2 analogous to compound 236 or to its enantiomer (S)-2-amino-3-(pyridin-4-yl)-1-(pyrrolidin-1-yl)propan-1-one dihydrochlohde Compound 240.
DETAILED DESCRIPTION OF THE SYNTHESIS OF PREFERRED COMPOUNDS (EXPERIMENTAL) Preparation of compound 12
2-lsocvano-1-(pyrrolidin-1 -yl)ethanone BLE 04098.
To stirred and cooled (0 0C) methyl isocyanoacetate (96 % technical grade, 5.0 g, 47.8 mmol) was slowly added in 0.75 h pyrrolidine (6.5 mL, 78 mmol). The mixture was stirred for 1.5 h with continued cooling and then concentrated. The resulting oil was co-evaporated twice from Ch^C^hexane to remove residual pyrrolidine. 2-lsocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 was obtained as a yellow solid (6.85 g, 98 % yield) and used in the next step without purification.
Figure imgf000054_0001
BLE 04098
MW: 138.17; Yield: 98 %; yellow solid; Mp (0C) = 73.9. 1H-NMR (CDCI3, δ): 1 .81-2.08 (m, 4H, 2xCH2), 3.35-3.45 (m, 2H, -NCH2), 3.50-3.60 (m, 2H, -NCH2;, 4.23 (s, 2H, CH2CO).
frans-(4,5-Dihvdro-5-(2,3-dihydrobenzo[ά1[1 ,41dioxin-6-yl)oxazol-4- yl)(pyrrolidin-1 -yl)methanone BLE 04100. To a stirred and cooled (00C) solution of potassium hydroxide (0.43 mg,
7.60 mmol) in MeOH (6.5 mL) were added successively 1 ,4-benzodioxan-6- carboxaldehyde (1 .31 g, 7.96 mmol) and 2-isocyano-1-(pyrrolidin-1- yl)ethanone BLE 04098 (1.0 g, 6.57 mmol). The solution was stirred 3 h at 00C and then concentrated. The residue was partitioned between EtOAc (100 mL) and water. The organic layer was combined with 2 additional EtOAc extracts (2 x 100 mL), washed with brine, dried over MgSO4, filtered and evaporated. Concentration afford to a crude product which was purified by column chromatography on silica (EtOAc) to yield, after evaporation and drying, to frans-4,5-dihydro-5-(2,3-dihydrobenzo[ό][1 ,4]dioxin-6-yl)oxazol-4- yl)(pyrrolidin-1 -yl)methanone BLE 04100 as a colourless oil (1.76 g, 89 % yield).
Figure imgf000054_0002
BLE 04100 MW: 440.49; Yield: 89 %; colorless oil.
1H-NMR (CDCI3, δ): 1 .75-2.10 (m, 4H, 2xCH2), 3.40-3.59 (m, 3H, 1 .5xCH2N), 3.85-4.00 (m, 1 H, 0.5xCH2N), 4.26 (s, 4H, CH2O), 4.59 (dd, 1 H, J = 7.5 Hz, J = 2.2 Hz, CH-N), 6.00 (d, 1 H, J = 7.5 Hz, CH-O), 6.75-6.90 (m, 3H, ArH), 7.00 (d, 1 H, J = 2.2 Hz, CH=N).
frans-(4,5-Dihvdro-5-(4-methoxyphenyl)oxazol-4-yl)(pyrrolidin-1-yl)methanone SLA 07074.
To a stirred and cooled (00C) solution of potassium hydroxide (0.37 g, 6.57 mmol) in methanol (30 mL) was added a mixture of 4-methoxy- benzaldehyde (0.88 mL, 7.23 mmol) and 2-isocyano-1-(pyrrolidin-1- yl)ethanone BLE 04098 (1.0 g, 6.57 mmol). The solution was stirred 4 h with continued cooling and then concentrated. The residue was partitioned between ethyl acetate and water. The organic layer was combined with additional ethyl acetate extracts, washed with aqueous sodium chloride and dried over MgSO4. Concentration afforded a crude product as a glassy solid. Flash chromatography over silica (ethyl acetate) yielded to frans-(4,5-dihydro- 5-(4-methoxyphenyl)oxazol-4-yl)(pyrrolidin-1-yl)methanone SLA 07074 as a pale yellow solid (1.2 g, 90.5 %).
Figure imgf000055_0001
SLA 07074
MW: 274.32; Yield: 90.5 %; pale yellow solid; Mp (0C): 91.2. Rf:0.30 (EtOAc). 1H-NMR (CDCI3, δ): 1 .75-2.08 (m, 4H, 2xCH2), 3.40-3.58 (m, 3H, CH2N), 3.52 (s, 3H, CH3O), 3.88-3.98 (m, 1 H, CH2N), 4.59 (dd, 1 H, J = 7.6 Hz, J = 2.2 Hz, CH-N), 6.06 (d, 1 H, J = 7.6 Hz, CH-O), 6.90 (d, 2H, J = 8.7 Hz, ArH), 7.01 (d, 1 H, J = 2.2 Hz, CH=N), 7.25 (d, 2H, J = 8.7 Hz, ArH). MS-ESI m/z (% rel. Int.): 275.1 ([MH]+, 10), 247.1 (100). HPLC: Method A, detection UV 280 nm, SLA 07074 RT = 5.2 min, peak area 92 %.
DL-f/?reo-2-Amino-3-hvdroxy-3-(4-niethoxyphenyl)-1 -(pyrrolidin-1 -yl)propan-1 - one hydrochloride SLA 07078.
To a stirred solution of frans-(4,5-dihydro-5-(4-methoxyphenyl)oxazol- 4-yl)(pyrrolidin-1-yl)methanone SLA 07074 (1.61 g, 5.93 mmol) in methanol (13 mL) was added hydrochloric acid (1 mL). After heating at 50 0C for 3h the mixture reaction was concentrated and the resulting yellow oil was co- evaporated twice with ethyl acetate before solidifying. Trituration (ethyl acetate) and drying afforded DL-f/?reo-2-amino-3-hydroxy-3-(4- methoxyphenyl)-1 -(pyrrolidin-1 -yl)propan-1 -one hydrochloride SLA 07078 as a white solid (1 .64 g, 93 %).
Figure imgf000056_0001
SLA 07078
MW: 300.78; Yield: 93 %; white Solid; Mp (0C): 177.0.
1H-NMR (CD3OD, δ): 1.32-1.50 (m, 1 H, 0.5xCH2), 1.50-1.88 (m, 3H,
1.5xCH2), 2.15-2.28 (m, 1 H, CH2N), 3.15-3.42 (m, 3H, 1.5xCH2N), 3.79 (s,
3H, CH3O), 4.06 (d, 1 H, J = 9.2 Hz, CH-N), 4.78 (d, 1 H, J = 9.2 Hz, CHO), 6.94 (d, 2H, J = 8.5 Hz, ArH), 7.34 (d, 2H, J = 8.5 Hz, ArH).
13C-NMR (CD3OD, δ): 24.8, 26.6, 47.2, 47.6, 55.9, 59.6, 73.9, 1 15.0 (2xC), 128.9 (2xC), 132.5, 161.7, 166.4.
DL-f/?reo-2-amino-3-(2,3-dihvdrobenzorάiπ ,41dioxin-6-yl)-3-hvdroxy-1- (pyrrolidin-1-yl)propan-1-one hydrochloride Compound 12.
To a stirred solution of frans-4,5-dihydro-5-(2,3- dihydrobenzo[ό][1 ,4]dioxin-6-yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone BLE 04100 (1.74 g, 5.77 mmol) in methanol (15 mL) was added hydrochloric acid (1 ml_). After heating at 50 0C for 3 h the mixture reaction was concentrated and the resulting yellow oil was co-evaporated twice with ethyl acetate before solidifying. Trituration (ethyl acetate) and drying afforded DL-f/?reo-2-amino-3- (2,3-dihydrobenzo[ό][1 ,4]dioxin-6-yl)-3-hydroxy-1 -(pyrrolidin-1 -yl)propan-1 - one hydrochloride Compound 12 as a white solid (1.85 g, 95 %).
Figure imgf000057_0001
(+/-)
Compound 12
MW: 328.79; Yield: 95.0 %; White Solid; Mp (0C): 176.2.
1H-NMR (CD3OD, δ): 1.42-1.58 (m, 1 H, 0.5xCH2), 1.58-1.70 (m, 1 H, 0.5xCH2), 1.70-1.88 (m, 2H, CH2), 3.20-3.45 (m, 4H, 2xN-CH2), 4.06 (d, 1 H, J
= 9.1 Hz, CH-N), 4.25 (s, 2H, OCH2), 4.75 (d, 1 H, J = 9.2 Hz, CH-O), 4.89 (s,
2H, OCH2), 6.82-6.95 (m, 3H, ArH).
13C-NMR (CD3OD, δ): 24.9, 26.7, 47.3, 47.6, 59.5, 65.7, 73.6, 1 16.4, 1 18.3,
120.3, 133.7, 145.1 , 145.6, 166.4.
Preparation of Compound 18.
Method B:
To a stirred and cooled (0 0C) solution of potassium hydroxide (380 mg, 5.80 mmol) in MeOH (5 mL) were added successively aldehyde (5.80 mmol) and 2-isocyano-1 -(pyrrolidin-1 -yl)ethanone BLE 04098 (0.8 g, 5.8 mmol). The solution was stirred 3 h at 0 0C and then concentrated. The residue was partitioned between CH2CI2 (100 mL) and water. The organic layer was washed with brine, dried over MgSO4, filtered and evaporated. Concentration afford to a crude product which was purified by column chromatography on silica (cyclohexane: EtOAc = 70:30 to 0:100) to yield, after evaporation and drying, to an intermediate oxazoline. To a stirred solution of oxazoline in methanol (15 mL) was added hydrochloric acid (1 mL, 12 mmol). After heating at 60 0C for 2 h, the mixture reaction was then concentrated and the resulting yellow oil was coevaporated twice with MeOH before solidifying. Trituration in EtOAc:MeOH = 10:1 followed by filtration gave title compound as a white solid.
DL-f/7reo-2-Amino-3-(biphenyl-4-yl)-3-hvdroxy-1 -(pyrrolidin-1 -yl)propan-1 -one hydrochloride Compound 18.
The compound was prepared according to method B with 4- phenylbenzaldehyde (1.05 g, 5.78 mmol). DL-f/?reo-2-Amino-3-(biphenyl-4- yl)-3-hydroxy-1 -(pyrrolidin-1 -yl)propan-1 -one hydrochloride Compound 18 was obtained as a pale brown solid (0.55 g, 28 % yield).
Figure imgf000058_0001
Compound 18
MW: 346.85; Yield: 28 %; Pale Brown Solid; Mp (0C): 197.3.
1H-NMR (CD3OD, δ): 1 .25-1.42 (m, 1 H, 0.5xCH2), 1.50-1.60 (m, 1 H, 0.5xCH2), 1.60-1.80 (m, 1 H, 0.5xCH2), 2.20-2.30 (m, 2H, N-CH2), 3.15-3.30 (m, 2H, N-CH2), 3.30-3.45 (1 H, m, N-CH2), 4.13 (d, 1 H, J = 9.2 Hz, CH-N), 4.85-4.95 (m, 1 H, CH-O), 7.32 -7.38 (m, 1 H, ArH), 7.46 (dd, 2H, J = 7.1 Hz, J = 7.8Hz, ArH), 7.52 (d, 2H, J = 8.3 Hz, ArH), 7.58-7.70 (m, 4H, ArH).
13C-NMR (CD3OD, δ): 24.8, 26.5, 47.2, 47.6, 59.5, 78.7, 127.9, 128.1 , 128.2,
128.8, 130.0, 139.7, 141.6, 143.3, 166.3.
MS-ESI m/z (% rel. Int.): 31 1.2 ([MH]+, 60).
HPLC: Method A, detection UV 254 nm, Compound 18 RT = 4.50 min, peak area 99.9 %.
Preparation of 2-isocvano derivatives: SLA 071 16B, SLA 071 16C, SLA 071 18, SLA 0713OA, SLA 07178, and SLA 07184A. 2-lsocvano-1-(piperidin-1 -yl)ethanone SLA 071 16B.
Prepared in accordance with Method B with methyl isocyanoacetate (2.46 g, 24.63 mmol) and piperidine (3.22 ml_, 37.85 mmol). The reaction mixture was stirred 1 h at RT and then concentrated. The residue was dissolved in dichloromethane (50 mL) and the organic layer was washed with 10 % aqueous citric acid (2 x 25 mL), dried over MgSO4, filtered and evaporated. 2-lsocyano-1-(pipehdin-1-yl)ethanone SLA 071 16B was obtained as an orange solid (3.13 g, 83 % yield).
Figure imgf000059_0001
SLA 071 16B
MW: 152.19; Yield: 83 %; Orange Solid; Mp (0C): 81.6. 1H-NMR (CDCI3, δ): 1 .56-1.74 (m, 6H, CH2C), 3.33 (t, 2H, J = 5.7 Hz, CH2N), 3.58 (t, 2H, J = 5.7 Hz, CH2N), 4.29 (s, 2H, CH2CO).
ferf-Butyl 4-(2-isocvanoacetyl)piperazine-1-carboxylate SLA 071 16C.
Prepared in accordance with Method B with methyl isocyanoacetate (2.51 g, 25.29 mmol) and piperazine-1-carboxylic acid tert-butyl ester (6.28 g, 33.85 mmol. The reaction mixture was stirred 1 h at RT and then concentrated. The residue was dissolved in dichloromethane (50 mL) and the organic layer was washed with 10 % aqueous citric acid (2 x 25 mL), dried over MgSO4, filtered and evaporated. ferf-Butyl 4-(2- isocyanoacetyl)piperazine-1-carboxylate SLA 071 16C was obtained as a colorless oil (0.41 g, 6.5 % yield).
Figure imgf000059_0002
SLA 071 16C MW: 253.14; Yield: 6.5 %; Colorless oil. 1H-NMR (CDCI3, δ): 1 .47 (s, 9H, tBu), 3.38 (t, 2H, J = 5.3 Hz, CH2N), 3.45- 3.53 (m, 4H, CH2N), 3.62 (t, 2H, J = 5.5 Hz, CH2N), 4.32 (s, 2H, CH2CO).
2-lsocvano-1-morpholinoethanone SLA 071 18. Prepared in accordance with Method B with methyl isocyanoacetate
(2.51 g, 25.30 mmol) and morpholine (3.30 ml_, 38.05 mmol). The reaction mixture was stirred 24 h at RT and then concentrated. The residue was dissolved in dichloromethane (50 mL) and the organic layer was washed with 10 % aqueous citric acid (2 x 25 mL), dried over MgSO4, filtered and evaporated. 2-lsocyano-1-morpholinoethanone SLA 071 18 was obtained as a brown oil (2.28 g, 58 % yield).
Figure imgf000060_0001
SLA 071 18 MW: 154.17; Yield: 58 %; Brown Oil. Rf\ 0.20 (EtOAc:cyclohexane = 50:50).
1H-NMR (CDCI3, δ): 3.42 (t, 2H, J = 4.9 Hz, CH2N), 3.65 (t, 2H, J = 5.1 Hz, CH2N), 3.73 (t, 4H, J = 5.0 Hz, CH2O), 4.31 (s, 2H, CH2CO).
2-lsocvano-1-thiomorpholinoethanone SLA 0713OA.
Prepared in accordance with Method B with methyl isocyanoacetate (2.50 g, 25.28 mmol) and thiomorpholine (4.25 mL, 37.85 mmol). The reaction mixture was stirred 22 h at RT and then concentrated. The residue was dissolved in dichloromethane (50 mL) and the organic layer was washed with 10 % aqueous citric acid (2 x 25 mL), dried over MgSO4, filtered and evaporated. 2-lsocyano-1-thiomorpholinoethanone SLA 0713OA was obtained as a yellow solid (3.05 g, 71 % yield).
Figure imgf000060_0002
SLA 0713OA
MW: 170.23; Yield: 71 %; Yellow Solid; Mp (0C): 144.4. Rf : 0.35 (EtOAσcyclohexane = 50:50). 1H-NMR (CDCI3, δ): 2.68 (m, 4H, 2xCH2S), 3.67 (m, 2H, N-CH2), 3.90 (m, 2H, N-CH2), 4.31 (s, 2H, COCH2).
2-lsocvano-1-(2H-pyrrol-1 (5H)-yl)ethanone SLA 07178.
Prepared in accordance with Method B with methyl isocyanoacetate
(LOO g, 10.10 mmol) and dihydro-1 /-/-pyrrole (1 .01 mL, 15.15 mmol). The reaction mixture was stirred 5 h at 50 0C and concentrated. The residue was dissolved in dichloromethane (50 mL) and the organic layer was washed with 10 % aqueous citric acid (2 x 25 mL), dried over MgSO4, filtered and evaporated. 2-lsocyano-1 -(2/-/-pyrrol-1 (5/-/)-yl)ethanone SLA 07178 was obtained (1.0 g, 73 % yield) as a yellow solid.
Figure imgf000061_0001
SLA 07178
MW: 136.15; Yield: 73 %; Yellow Solid. Rf\ 0.35 (EtOAσcyclohexane = 50:50).
1H-NMR (CDCI3, δ): 4.23 (s, 4H, 2xCH2N), 4.31 (s, 2H, CH2N), 5.80-5.86 (m, 1 H, CH=C), 5.90-5.95 (m, 1 H, CH=C).
Λ/.Λ/-Diethyl-2-isocvanoacetamide SLA 07184A.
Prepared in accordance with Method B with methyl isocyanoacetate (2.50 g, 25.29 mmol) and diethylamine (1.96 mL, 37.94 mmol). The reaction mixture was stirred 5 h at 50 0C and concentrated. The residue was dissolved in dichloromethane (50 mL) and the organic layer was washed with 10 % aqueous citric acid (2 x 25 mL), dried over MgSO4, filtered and evaporated. Λ/,Λ/-Diethyl-2-isocyanoacetamide SLA 07184A was obtained (1.213 g, 34 % yield) as a brown oil.
Figure imgf000062_0001
SLA 07184A
MW: 140.18; Yield: 34 %; Brown Oil.
Rf : 0.35 (EtOAc:cyclohexane = 50:50).
1H-NMR (CDCI3, δ): 1 .15-1.26 (m, 6H, CH3), 3.21-3.30 (m, 2H, CH2N), 3.38-
3.45 (m, 2H, CH2N), 4.26 (s, 2H, CH2CO).
Preparation of oxazolines: BLE 041 10B. SLA 07122A. SLA 07124A. SLA 07124B. SLA 07132. BLE 041 10A. Compound 19. BLE 04124A. BLE 04124B. BLE 04124C, BLE 04124D, BLE 04130B, BLE 04130C, BLE 04130D, BLE 04136B. BLE 04136C, BAL 01016, BLE 04136D, BAL 01014. SLA 07194A. SLA 07174, BAL 01028A, BLA 01028B, SLA 07158 and SLA 07180.
frans-(4,5-Dihvdro-5-(pyridin-3-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 041 10B.
General Method D for oxazolines formation:
To a stirred and cooled (0 0C) solution of potassium hydroxide (0.55 g, 9.80 mmol) in methanol (10 mL) were added a mixture of 3-pyhdine carboxaldehyde (1 .03 mL, 10.84 mmol) and 2-isocyano-1-(pyrrolidin-1 - yl)ethanone BLE 04098 (1.50 g, 10.86 mmol). The solution was stirred 3 h at 0 0C and then concentrated. The residue was partitioned between ethyl acetate (100 mL) and water. The organic layer was combined with two additional ethyl acetate extracts (2x100 mL), washed with aqueous sodium chloride and dried over MgSO4, filtered and evaporated. Concentration afforded a crude product which was purified by column chromatography on silica (CH2CI2:Me0H = 98:2) to yield to fra/7S-(4,5-dihydro-5-(pyridin-3- yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone BLE 041 1 OB (0.95 g, 39 % yield) as a pale yellow pale solid.
Figure imgf000063_0001
BLE 041 10B
MW: 245.28; Yield: 39 %; Yellow Pale Solid; Mp (0C): 107.0. 1H-NMR (CDCI3, δ): 1 .78-2.10 (m, 4H, 2xCH2), 3.40-3.61 (m, 3H, CH2N), 3.90-4.04 (m, 1 H, CH2N), 4.59 (dd, 1 H, J = 7.7 Hz, J = 2.2 Hz, CH-N), 6.21 (d, 1 H, J = 7.7 Hz, CH-O), 7.04 (d, 1 H, J = 2.2 Hz, 0-CH=N), 7.33 (m, 1 H, ArH), 7.64 (m, 1 H, ArH), 8.59 (d, 2H, J = 2.8 Hz, ArH).
13C-NMR (CDCI3, δ): 24.2, 26.0, 46.4, 46.6, 75.7, 79.3, 123.7, 133.5, 135.3, 147.6, 149.9, 155.2, 166.2.
frans-(4,5-Dihvdro-5-(pyridin-4-yl)oxazol-4-yl)(piperidin-1 -yl)methanone SLA 07122 A.
SLA 07122A was prepared in accordance with method method D using 2-isocyano-1-(pipehdin-1 -yl)ethanone (0.4 g, 26.3 mmol), potassium hydroxide (0.15 g, 26.7 mmol) in methanol (5 mL) and pyridine-4- carbaldehyde (0.37 mL, 40.9 mmol). The solution was stirred 20 h at 0 0C. frans-(4,5-Dihydro-5-(pyridin-4-yl)oxazol-4-yl)(piperidin-1 -yl)methanone SLA 07122A was obtained as a yellow solid (0.353 g, 52 % yield).
Figure imgf000063_0002
SLA 07122A MW: 259.30; Yield: 52 %; Yellow Solid; Mp (0C): 1 1 1.7. Rf\ 0.80 (MeOH:CH2CI2 = 10:90).
1H-NMR (CDCI3, δ): 1 .55-1.78 (m, 6H, 3xCH2), 3.45-3.60 (m, 2H, CH2N), 3.70-3.85 (m, 2H, CH2N), 4.60 (dd, 1 H, J = 7.8 Hz, J = 2.3 Hz, CH-N), 6.27 (d, 1 H, J = 7.8 Hz, CH-O), 7.01 (d, 1 H, J = 2.3 Hz, CH=N), 7.23 (dd, 2H, J = 4.5 Hz, J = 1.6 Hz, ArH), 8.61 (dd, 2H, J = 4.5 Hz, J = 1 .5 Hz, ArH).
frans-(4,5-Dihvdro-5-(pyridin-4-yl)oxazol-4-yl)(morpholino)methanone SLA 07124 A. SLA 071 18 was prepared in accordance with method D using 2- isocyano-1-morpholinoethanone (0.40 g, 25.95 mmol), potassium hydroxide (0.146 g, 26.0 mmol) in methanol (5 mL) and pyhdine-4-carbaldehyde (0.36 mL, 40.4 mmol). The solution was stirred 22 h at 0 0C. frans-(4,5-Dihydro-5- (pyhdin-4-yl)oxazol-4-yl)(morpholino)methanone SLA 07124A was obtained as a yellow solid (0.168 g, 25 % yield).
Figure imgf000064_0001
SLA 07124A
MW: 261.28; Yield: 25 %; Yellow Solid; Mp (0C): 90.5. Rf: 0.30 (EtOAσcyclohexane = 20:80). 1H-NMR (CDCI3, δ): 3.46-4.02 (m, 8H, 2xCH2O, 2xCH2N), 4.56 (dd, 1 H, J = 7.8 Hz, J = 2.3 Hz, CH-N), 6.27 (d, 1 H, J = 7.9 Hz, CH-O), 7.02 (d, 1 H, J = 2.3 Hz, CH=N), 7.24 (dd, 2H, J = 4.6 Hz, J = 1 .4 Hz, ArH), 8.63 (dd, 2H, J = 4.5 Hz, J = 1.6 Hz, ArH).
frans-(4,5-Dihvdro-5-(pyridin-4-yl)oxazol-4-yl)(4-te/f-butyloxycarbonyl- piperazin-1-yl)methanone SLA 07124B.
SLA 07124B was prepared in accordance with method D using tert- butyl 4-(2-isocyanoacetyl)piperazine-1-carboxylate SLA 071 16C (0.41 g, 16.20 mmol), potassium hydroxide (0.91 g, 16.2 mmol) in methanol (5 mL) and pyhdine-4-carbaldehyde (0.227 mL, 25.2 mmol). The solution was stirred 22 h at 0 0C. fra/7S-(4,5-Dihydro-5-(pyridin-4-yl)oxazol-4-yl)(4-terf- butyloxycarbonyl-piperazin-1-yl)methanone SLA 07124B was obtained as a pale yellow solid (0.335 g, 58 % yield).
Figure imgf000065_0001
SLA 07124B
MW: 360.41 ; Yield: 58 %; Pale Yellow Solid; Mp (0C): 157.2°C. 1H-NMR (CDCI3, δ): 1 .47 (s, 9H, tBu), 3.25-4.02 (m, 8H, CH2N), 4.58 (dd, 1 H, J = 7.8 Hz, J = 2.3 Hz, CH-N), 6.27 (d, 1 H, J = 7.8 Hz, CH-O), 7.01 (d, 1 H, J = 2.3 Hz, CH=N), 7.24 (dd, 2H, J = 4.6 Hz, J = 1.4 Hz, ArH), 8.62 (dd, 2H, J = 4.5 Hz, J = 1.6 Hz, ArH).
frans-(4,5-Dihvdro-5-(pyridin-4-yl)oxazol-4-yl)(thiomorpholino)methanone SLA 07132.
SLA 07132 was prepared in accordance with method D using 2- lsocyano-1 -thiomorpholinoethanone SLA 0713OA (0.752 g, 4.41 mmol), potassium hydroxide (0.250 g, 4.45 mmol) in methanol (10 mL) and pyridine- 4-carbaldehyde (0.436 mL, 4.85 mmol). The solution was stirred 24 h at 0 0C. frans-(4,5-Dihydro-5-(pyridin-4-yl)oxazol-4-yl)(thiomorpholino)methanone SLA 07132 was obtained as a yellow foam (1.01 g, 83 %).
Figure imgf000065_0002
SLA 07132
MW: 277.35; Yield: 83 % ι; Yellow Foam. Rf: 0.80 (MeOHiCH2CI2 = 10:90).
1H-NMR (CDCI3, δ): 2.53-2.92 (m, 4H, 2xCH2), 3.58-3.70 (m, 1 H, CH2N), 3.78-3.88 (m, 1 H, CH2N), 4.15-4.30 (m, 2H, CH2N), 4.56 (dd, J = 7.8 Hz, J = 2.3 Hz, 2H, CH-N), 6.27 (d, 1 H, J = 7.8 Hz, CH-O), 7.02 (d, 1 H, J = 2.3 Hz, N=CH-O), 7.22 (d, 2H, J = 6.1 Hz, ArH), 8.61 (dd, 2H, J = 6.1 Hz, ArH). 13C-NMR (CDCI3, δ): 27.3, 28.0, 45.4, 48.6, 74.9, 79.6, 120.0 (2xC), 148.5, 150.3 (2xC), 154.8, 166.2.
frans-(4,5-Dihvdro-5-(pyridin-2-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 041 10A.
BLE 041 10A was prepared in accordance with method D using 2- pyridine carboxaldehyde (1.02 mL, 10.84 mmol). 7ra/7s-(4,5-dihydro-5- (pyhdin-2-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 041 1 OA was obtained as a yellow pale oil (0.45 g, 19 % yield).
Figure imgf000066_0001
BLE 041 1 OA
MW: 245.28; Yield: 19 %; Yellow Pale Oil.
1H-NMR (CDCI3, δ): 1 .73-2.08 (m, 4H, 2xCH2), 3.35-3.70 (m, 3H, CH2N), 3.85-4.00 (m, 1 H, CH2N), 5.05 (dd, 1 H, J = 6.9 Hz, J = 2.2 Hz, CH-N), 6.18
(d, 1 H, J = 6.9 Hz, CH-O), 7.02 (d, 1 H, J = 2.1 Hz, 0-CH=N), 7.25 (m, 1 H,
ArH), 7.43 (d, 1 H, J = 7.8 Hz, ArH), 7.69 (dt, 1 H, J = 7.8 Hz, J = 1 .8 Hz ArH),
8.62 (m, 1 H1 ArH).
13C-NMR (CDCI3, δ): 24.2, 25.9, 46.3, 46.5, 73.4, 81.3, 121.5, 123.2, 136.8, 149.8, 154.8, 158.0, 166.9.
frans-(4,5-Dihvdro-5-(pyridin-4-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone Compound 19. SLA 07092 was prepared in accordance with method D using pyridine- 4-carbaldehyde (1.88 ml_, 19.76 mmol), KOH (1.01 g, 18.00 mmol) in methanol (18 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (2.73 g, 19.76 mmol). The residue was partitioned between ethyl acetate (200 mL) and water (150 mL). The organic layer was combined with additional ethyl acetate extracts (2 x 150 mL), washed with aqueous sodium chloride (2 x 150 mL) and dried over MgSO4, filtered and evaporated. 7ra/7s-(4,5-dihydro- 5-(pyhdin-4-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone Compound 19 was obtained as a white solid (4.32 g, 98 % yield).
Figure imgf000067_0001
Compound 19
MW: 245.28; Yield: 98 %; White Solid; Mp (0C) = 69.2. Rf. 0.65 (MeOH:CH2CI2 = 10:90). 1H-NMR (CDCI3, δ): 1 .78-2.06 (m, 4H, 2xCH2), 3.44-3.60 (m, 3H, CH2N),
3.90-4.01 (m, 1 H, CH2N;, 4.52 (dd, 1 H, J = 7.9 Hz, J = 2.2 Hz, CH-N), 6.19 (d, J = 7.9 Hz, 1 H, CH-O), 7.03 (d, 1 H, J = 2.2 Hz, N=CH-O), 7.24 (dd, 2H, J = 4.5 Hz, J = 1.5 Hz, ArH), 8.61 (dd, 2H, J = 4.5 Hz, J = 1.5 Hz, ArH).
frans-(4,5-Dihvdro-5-(thiophen-3-yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone BLE 04124A.
BLE 04124A was prepared in accordance with method D using thiophen-3-carboxaldehyde (0.475 mL, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol). After work-up the residue obtained was rechstallized from ethyl acetate to obtain after filtration frans-(4,5-dihydro-5- (thiophen-3-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 041 1 OA as a yellow pale solid (0.498 g, 40 % yield).
Figure imgf000068_0001
BLE 04124A
MW: 250.32; Yield: 40.5 %; Yellow Pale Solid; Mp (0C): 105.9. 1H-NMR (CDCI3, δ): 1 .78-2.10 (m, 4H, CH2), 3.42-3.61 (m, 3H, CH2N), 3.90- 4.02 (m, 1 H, CH2N), 4.63 (dd, 1 H, J = 7.4 Hz, J = 2.2 Hz, CH-N), 6.20 (d, 1 H, J = 7.4 Hz, CH-O), 6.98 (d, 1 H, J = 2.2 Hz, 0-CH=N), 7.03 (dd, 1 H, J = 5.0 Hz, J = 1.3 Hz, CH=C), 7.30 (dt, 1 H, J = 3.0 Hz, J = 1.3 Hz, CH=C), 7.36 (dd, 1 H, J = 5.0 Hz, J = 3.0 Hz, CH=C).
13C-NMR (CDCI3, δ): 24.2, 26.0, 46.4, 46.6, 74.6, 77.9, 122.7, 125.1 , 127.3, 140.4, 155.3, 166.7.
MS-ESI m/z (% rel. Int.): 251.0 ([MH]+, 17), 223 (40), 179.9 (60), 151.9 (63),
123.9 (100).
HPLC: Method A, detection UV 254 nm, BLE 04124A, RT = 4.4 min, peak area 98.0 %.
frans-(4,5-Dihvdro-5-(thiophen-2-yl)oxazol-4-yl)(pyrrolidin-1 -yl)niethanone BLE 04124B.
BLE 04124B was prepared in accordance with method D using thiophen-2-carboxaldehyde (0.507 mL, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol). After work-up the residue obtained was purified by column chromatography (EtOAc) to led after evaporation to trans-{4,5- dihydro-5-( thiophen-2-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 04124B as a yellow pale solid (0.713 g, 58 % yield).
Figure imgf000068_0002
BLE 04124B MW: 250.32; Yield: 58 %; Yellow Pale Solid; Mp (0C): 71 .3.
1H-NMR (CDCI3, δ): 1 .78-2.10 (m, 4H, CH2), 3.42-3.62 (m, 3H, CH2N), 3.90-
4.03 (m, 1 H, CH2N), 4.76 (dd, 1 H, J = 7.3 Hz, J = 2.2 Hz, CH-N), 6.37 (d, 1 H,
J = 7.3 Hz, CH-O), 6.96 (d, 1 H, J = 2.2 Hz, 0-CH=N), 7.00 (dd, 1 H, J = 5.0 Hz, J = 3.5 Hz, CH=C), 7.1 1 (d, 1 H, J = 3.1 Hz, CH=C), 7.33 (dd, 1 H, J = 5.0
Hz, J = 0.7 Hz, CH=C).
13C-NMR (CDCI3, δ): 24.2, 26.0, 46.4, 46.6, 75.5, 77.6, 126.3 (2xC), 127.1 ,
142.0, 154.9, 166.3.
MS-ESI m/z (% rel. Int.): 251.0 ([MH]+, 15), 223 (100). HPLC: Method A, detection UV 254 nm, BLE 04124B, RT = 3.8 min, peak area > 90 %.
frans-(4,5-Dihvdro-5-(thiazol-2-yl)oxazol-4-yl)(pyrrolidin-1-yl)niethanone BLE 04124C. BLE 04124C was prepared in accordance with method D using 2- thiazolecarboxaldehyde (0.476 mL, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1-(pyrrolidin-1 -yl)ethanone BLE 04098 (0.75 g, 5.43 mmol). After work-up the residue obtained was purified by column chromatography (EtOAc) to led after evaporation to trans-{4,5- dihydro-5-(thiazol-2-yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone BLE 04124C as a colourless oil (0.564 g, 45.5 % yield).
Figure imgf000069_0001
BLE 04124C
MW: 251.3; Yield: 45.5 %; colourless Oil. 1H-NMR (CDCI3, δ): 1 .80-2.10 (m, 4H, CH2), 3.47-3.70 (m, 3H, CH2N), 3.91 - 4.02 (m, 1 H1 CH2N), 5.18 (dd, 1 H, J = 6.4 Hz, J = 2.2 Hz, CH-N), 6.40 (d, 1 H, J = 6.4 Hz, CH-O), 6.97 (d, 1 H, J = 2.2 Hz, 0-CH=N), 7.38 (d, 1 H, J = 3.3 Hz, CH=C), 7.81 (d, 1 H, J = 3.3 Hz, CH=C). 13C-NMR (CDCI3, δ): 24.2, 26.0, 46.4, 46.5, 73.7, 78.2, 120.1 , 143.3, 154.3, 166.1 , 168.2.
MS-ESI m/z (% rel. Int.): 252.0 ([MH]+, 18), 225 (30), 198.9 (37), 153.9 (48), 143.0 (100). HPLC: Method A, detection UV 254 nm, BLE 04124C, RT = 3.5 min, peak area > 90 %.
frans-(5-(Benzo[ά1thiophen-3-yl)-4,5-dihvdrooxazol-4-yl)(pyrrolidin-1- vDmethanone BLE 04124D.
BLE 04124D was prepared in accordance with method D using thianaphtene-3-carboxaldehyde (0.88 g, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol). After work-up the residue obtained was purified by column chromatography (EtOAc) to led after evaporation to trans-{5-
(benzo[ό]thiophen-3-yl)-4,5-dihydrooxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 04124D as a white solid (1 .12 g, 75.5 % yield).
Figure imgf000070_0001
BLE 04124D
MW: 300.38; Yield: 75.5 %; White Solid; Mp (0C): 92.2.
1H-NMR (CDCI3, δ): 1 .75-2.08 (m, 4H, CH2), 3.36-3.49 (m, 1 H, CH2N), 3.50- 3.62 (m, 1 H, CH2N), 3.89-4.00 (m, 1 H, CH2N), 4.75 (dd, 1 H, J = 7.6 Hz, J =
2.2 Hz, CH-N), 6.54 (d, 1 H, J = 7.6 Hz, CH-O), 7.08 (d, 1 H, J = 2.2 Hz, O-
CH=N), 7.35 (m, 2H, ArH), 7.45 (s, 1 H, C=CH-S), 7.67-7.75 (m, 1 H, ArH),
7.84-7.92 (m, 1 H1 ArH).
13C-NMR (CDCI3, δ): 24.2, 26.0, 46.5, 46.6, 73.3, 77.7, 121.8, 123.1 , 124.1 , 124.6, 124.8, 134.0, 136.4, 141.0, 155.4, 166.6.
MS-ESI m/z (% rel. Int.): 301.0 ([MH]+, 30), 273.0 (100). HPLC: Method A, detection UV 254 nm, BLE 04124D, RT = 4.2 min, peak area 92.0 %.
frans-(5-(Furan-3-yl)-4,5-dihvdrooxazol-4-yl)(pyrrolidin-1 -yl)niethanone BLE 0413OB.
BLE 0413OB was prepared in accordance with method D using 3- furaldehyde (0.453 g, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol). After work-up the residue was washed with a minimum of ethyl acetate to led, after filtration and drying, to frans-(5-(furan-3-yl)-4,5-dihydrooxazol-4- yl)(pyrrolidin-1 -yl)methanone BLE 0413OB as a white solid (0.837 g, 72.5 % yield).
Figure imgf000071_0001
BLE 04130B MW: 234.25; Yield: 72.5 %; White Solid; Mp (0C): 136.7.
1H-NMR (CDCI3, δ): 1 .80-2.10 (m, 4H, CH2), 3.47-3.58 (m, 3H, CH2N), 3.91- 4.02 (m, 1 H, CH2N), 4.61 (dd, 1 H, J = 7.3 Hz, J = 2.1 Hz, CH-N), 6.10 (d, 1 H, J = 7.3 Hz, CH-O), 6.36 (dd, 1 H, J = 1.6 Hz, J = 0.6 Hz, CH=C), 6.95 (d, 1 H, J = 2.1 Hz, 0-CH=N), 7.44 (t, 1 H, J = 1 .6 Hz, OCH=C); 7.50 (d, 1 H, J = 0.6 Hz, OCH=C).
13C-NMR (CDCI3, δ): 24.2, 26.0, 46.4, 46.6, 70.0, 74.0, 108.1 , 124.0, 140.4, 144.2, 155.3, 166.6.
frans-(4,5-Dihvdro-5-(naphthalen-3-yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone BLE 0413OC.
BLE 0413OC was prepared in accordance with method D using 2- naphtaldehyde (0.847 g, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol). After work-up the residue was washed with a minimum of ethyl acetate to led, after filtration and drying, to trans-(4,5-dihydro-5-(naphthalen-3- yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 0413OC as a white solid (0.791 g, 54.5 % yield).
Figure imgf000072_0001
BLE 0413OC
MW: 294.35; Yield: 54.5 %; White Solid; Mp (0C): 1 17.9.
1H-NMR (CDCI3, δ): 1 .78-2.07 (m, 4H, CH2), 3.37-3.49 (m, 1 H, CH2N), 3.49-
3.61 (m, 2H, CH2N), 3.88-3.99 (m, 1 H, CH2N), 4.67 (dd, 1 H, J = 7.7 Hz, J = 2.2 Hz, CH-N), 6.31 (d, 1 H, J = 7.7 Hz, CH-O), 7.10 (d, 1 H, J = 2.2 Hz, O-
CH=N), 7.38 (dd, 1 H, J = 8.5 Hz, J = 1.7 Hz, ArH); 7.45-7.54 (m, 2H, ArH),
7.79-7.90 (m, 4H, ArH).
13C-NMR (CDCI3, δ): 23.8, 25.7, 46.1 , 46.2, 75.3, 81.4, 122.7, 124.9, 126.1 ,
126.2, 127.4, 127.7, 128.7, 132.8, 132.9, 136.5, 155.2,166.4. MS-ESI m/z (% rel. Int.): 295.1 ([MH]+, 40), 267.1 (100).
HPLC: Method A, detection UV 254 nm, BLE 0413OC, RT = 4.2 min, peak area 92.0 %.
frans-(4,5-Dihvdro-5-(naphthalen-4-yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone BLE 04130D.
BLE 0413OD was prepared in accordance with method D using 1- naphtaldehyde (0.736 mL, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol). After work-up the residue was purified by column chromatography on silica (EtOAc:cyclohexane = 80:20 to 90:10) to led, after evaporation, to trans-(4,5-dihydro-5-(naphthalen-4-yl)oxazol-4-yl)(pyrrolidin-1- yl)methanone BLE 0413OD as a colorless gum (0.850 g, 58.5 % yield).
Figure imgf000073_0001
BLE 04130D
MW: 294.35; Yield: 58.5 %; Colorless gum.
1H-NMR (CDCI3, δ): 1 .75-2.02 (m, 4H, CH2), 3.25-3.37 (m, 1 H, CH2N), 3.52- 3.67 (m, 2H, CH2N), 3.82 -3.93 (m, 1 H, CH2N), 4.62 (dd, 1 H, J = 7.0 Hz, J = 2.0 Hz, CH-N), 6.89 (d, 1 H, J = 7.0 Hz, CH-O), 7.16 (d, 1 H, J = 2.0 Hz, O- CH=N), 7.44-7.58 (m, 4H, ArH), 7.80-7.90 (m, 3H, ArH). 13C-NMR (CDCI3, δ): 24.2, 25.9, 46.5 (2xC), 75.3, 79.2, 122.5, 123.0, 125.4, 126.0, 126.8, 128.7, 129.0, 129.9, 133.9, 135.5, 155.5,166.9. MS-ESI m/z (% rel. Int.): 295.1 ([MH]+, 50), 267.1 (100).
HPLC: Method A, detection UV 254 nm, BLE 0413OD, RT = 4.2 min, peak area 95.0 %.
frans-(4,5-Dihvdro-5-(αuinolin-2-yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone BLE 04136B.
BLE 04136B was prepared in accordance with method D using 2- quinoline carbaldehyde (0.852 g, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol). After work-up the residue was purified by column chromatography on silica (EtOAc) to led, after evaporation, to trans-{4,5- dihydro-5-(quinolin-2-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 04136B as a yellow pale solid (0.966 g, 60.3 % yield).
Figure imgf000073_0002
BLE 04136B
MW: 295.34; Yield: 60.3 %; Yellow Pale Solid; Mp (0C): 93.8. 1H-NMR (CDCI3, δ): 1 .85-2.10 (m, 4H, CH2), 3.50-3.66 (m, 2H, CH2N), 3.67- 3.80 (m, 1 H, CH2N), 3.92-4.03 (m, 1 H, CH2N), 5.32 (dd, 1 H, J = 7.8 Hz, J = 2.1 Hz, CH-N), 6.31 (d, 1 H, J = 7.8 Hz, CH-O), 7.06 (d, 1 H, J = 2.1 Hz, O- CH=N), 7.51 -7.60 (m, 2H, ArH); 7.72 (t, 1 H, J = 8.4 Hz, ArH), 7.83 (t, 1 H, J = 8.1 Hz, ArH), 8.07 (d, 1 H, J = 8.4 Hz, ArH), 8.20 (d, 1 H, J = 8.4 Hz, ArH). 13C-NMR (CDCI3, δ): 24.3, 26.2, 46.5, 46.7, 73.0, 82.1 , 1 19.2, 126.9, 127.7, 127.8, 129.5, 129.9, 137.2, 147.7, 155.0, 158.2, 167.3.
frans-(4,5-Dihvdro-5-(isoαuinolin-4-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 04136C.
BLE 04136C was prepared in accordance with method D using 4- quinoline carbaldehyde (0.852 g, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol). After work-up the residue was washed with a minimum of EtOAc to led, after filtration and drying, to frans-(4,5-dihydro-5-(isoquinolin-4- yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone BLE 04136C as a white solid (0.640 g, 40 % yield).
Figure imgf000074_0001
BLE 04136C
MW: 295.34; Yield: %; White Solid; Mp (0C): 152.0.
1H-NMR (CDCI3, δ): 1 .85-2.08 (m, 4H, CH2), 3.28-3.40 (m, 1 H, CH2N), 3.54-
3.69 (m, 2H, CH2N), 3.84-3.95 (m, 1 H, CH2N), 4.57 (dd, 1 H, J = 6.8 Hz, J =
2.1 Hz, CH-N), 6.93 (d, 1 H, J = 6.8 Hz, CH-O), 7.15 (d, 1 H, J = 2.1 Hz, O- CH=N), 7.41 (d, 1 H, J = 4.5 Hz, ArH); 7.59 (m, 1 H, ArH), 7.76 (m, 1 H, ArH),
7.91 (d, 1 H, J = 8.3 Hz, ArH), 8.16 (d, 1 H, J = 8.3 Hz, ArH), 8.92 (d, 1 H, J =
4.5 Hz, ArH).
13C-NMR (CDCI3, δ): 24.2, 25.9, 46.7 (2xC), 75.6, 77.8, 1 16.4, 123.1 , 124.8,
127.5, 129.6, 130.5, 145.5, 148.4, 150.3, 155.0, 166.0. frans-(4,5-Dihvdro-5-(αuinolin-3-yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone BAL 01016.
To a stirred and cooled (0 0C) solution of KOH (0.31 g, 5.43 mmol) in 5 mL MeOH were added successively quinoline-3-carboxaldehyde (0.85 g, 5.43 mmol) and 2-isocyano-1-pyrrolidin-1-yl-ethanone BLE 04134 (0.75 g, 5.43 mmol). The mixture was stirred at 0 0C until precipitation and concentrated. The mixture was partitioned between EtOAc (50 mL) and H2O (25 mL). The aqueous layer was extracted twice with EtOAc (25 ml). The EtOAc fractions were combined, washed twice with brine (2x25 mL), dried over MgSO4 and filtered. After evaporation and drying frans-(4,5-dihydro-5-(quinolin-3- yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone BAL 01016 was obtained (0.96 g, 60 % yield) as a white solid.
Figure imgf000075_0001
BAL 01016
MW: 295.34; Yield: 60 %; White Solid; Mp (0C): 144.4.
Rf : 0.15 (EtOAc).
1H NMR (CDCI3, δ): 1.75-2.10 (m, 4H, 2xCH2), 3.40-3.62 (m, 3 H, CH2N),
3.90-4.05 (m, 1 H, CH2N), 4.70 (dd, 1 H, J = 7.8 Hz, J = 2.2 Hz, CH-N), 6.40 (d, 1 H, J = 7.8 Hz, CH-O), 7.10 (d, 1 H, J = 2.2 Hz, OCH=N), 7.58 (dt, 1 H, J =
1.1 Hz, J = 8.0 Hz, ArH) ), 7.73 (dt, 1 H, J = 1.4 Hz, J = 6.9 Hz, ArH), 7.83 (dd,
1 H, J = 1.2 Hz, J = 8.2 Hz, ArH), 8.12 (m, 2H, ArH), 8.87 (d, 1 H, J = 2.2 Hz,
ArH).
13C-NMR (CDCI3, δ): 24.2, 26.0, 46.6, 46.6, 75.8, 79.7, 127.3, 127.5, 127.9, 129.4, 130.0, 132.3, 133.2, 148.1 , 148.4, 155.3, 166.2.
MS-ESI m/z (% rel. Int.): 296.1 ([MH]+, 5), 314.1 (100).
frans-(5-(Furan-2-yl)-4,5-dihvdrooxazol-4-yl)(pyrrolidin-1 -yl)methanone BLE 04136D. BLE 04136C was prepared in accordance with method D using 2- furaldehyde (0.449 ml_, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 ml.) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol). After work-up the residue was purified by column chromatography on silica (cyclohexane: EtOAc = 100:0 to 0:100) to led, after evaporation, to trans- (5-(furan-2-yl)-4,5-dihydrooxazol-4-yl)(pyrrolidin-1 -yl)methanone BLE 04136D as a yellow pale oil (0.742 g, 58.5 % yield).
Figure imgf000076_0001
BLE 04136D MW: 234.25; Yield: 58.5 %; Yellow Pale Oil.
1H-NMR (CDCI3, δ): 1 .80-2.10 (m, 4H, CH2), 3.47-3.60 (m, 3H, CH2N), 3.94- 4.06 (m, 1 H, CH2N), 4.94 (dd, 1 H, J = 7.4 Hz, J = 2.2 Hz, CH-N), 6.14 (d, 1 H, J = 7.4 Hz, CH-O), 6.37 (dd, 1 H, J = 3.3 Hz, J = 1.8 Hz, CH=C), 6.48 (d, 1 H, J = 3.3 Hz, CH=C), 6.93 (d, 1 H, J = 2.2 Hz, 0-CH=N), 7.44 (d, 1 H, J = 1.8 Hz, OCH=C).
13C-NMR (CDCI3, δ): 24.2, 26.0, 46.4, 46.5, 71.3, 74.5, 1 10.2, 1 10.5, 143.6, 150.4, 155.0, 166.3.
frans-(4,5-Dihvdro-5-(2-methoxypyhdin-3-yl)oxazol-4-yl)(pyrrolidin-1- vDmethanone BAL 01014.
BAL 01014 was prepared in accordance with method D using 2- methoxy-3-pyridinecarboxaldehyde (0.64 ml, 5.43 mmol), KOH (0.305 mg, 5.43 mmol) in methanol (5 mL) and 2-isocyano-1 -(pyrrolidin-1-yl)ethanone BLE 04098 (0.75 g, 5.43 mmol). After work-up frans-(4,5-dihydro-5-(2- methoxypyhdin-3-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BAL 01014 was obtained (0.74 mg, 50 % yield) as a white solid.
Figure imgf000077_0001
BAL 01014
MW: 275.30; Yield: 50 %; White Solid; Mp (0C): 1 10.1.
Rf : 0.25 (EtOAc). 1H NMR (CDCI3, δ): 1.82-2.10 (m, 4H, 2xCH2), 3.40-3.62 (m, 3 H, CH2N),
3.80-3.90 (m, 3 H, CH2N), 3.93 (s, 3H, OMe), 4.61 (dd, 1 H, J = 7 Hz, J = 2 Hz,
CH-N), 6.14 (d, 1 H, J = 7 Hz, CH-O), 6.90 (dd, 1 H, J = 7.3 Hz, J = 5 Hz, ArH),
7.02 (d, 1 H, J = 2 Hz, OCH=N), 7.60 (dd, 1 H, J = 7.3 Hz, J = 1 .7 Hz, ArH) ),
8.13 (dd, 1 H, , J = 5 Hz, J = 1.8 Hz1 ArH). 13C-NMR (CDCI3, δ): 24.3, 26.1 , 46.3, 46.6, 53.5, 73.5, 78.1 , 1 16.8, 122.2,
135.2, 146.5, 155.3, 160.5 and 167.4.
MS-ESI m/z (% rel. Int.): 276.1 ([MH]+, 42).
HPLC: Method A, detection UV 254 nm, BAL 01014 RT = 3.63 min, peak area
97.2 %.
frans-Λ/,Λ/-Diethyl-4,5-dihvdro-5-(pyridin-4-yl)oxazole-4-carboxamide SLA
07194 A.
SLA 07194A was prepared in accordance with method D using pyridine-4-carbaldehyde (1.14 mL, 9.52 mmol), KOH (0.54 g, 9.60 mmol) in methanol (5 mL) and Λ/,Λ/-diethyl-2-isocyanoacetamide SLA 07184A (1.21 g, 8.65 mmol). After work-up and column chromatography on florisil (ethyl acetate) frans-Λ/,Λ/-diethyl-4,5-dihydro-5-(pyhdin-4-yl)oxazole-4-carboxamide SLA 07194A was obtained as a brown oil (0.25 g, 12 % yield).
Figure imgf000077_0002
SLA 07194 A MW: 247.29; Yield: 12 %; Brown Oil. Rf\ 0.15 (AcOEt = 100).
1H-NMR (CDCI3, δ): 1 .16-1.34 (m, 6H, CH3), 3.30-3.80 (m, 4H, CH2N), 4.60 (dd, 1 H, J = 7.7 Hz, J = 2.2 Hz, CH-N), 6.22 (d, 1 H, J = 7.7 Hz, CH-O), 7.06 (d, J = 2.2 Hz, CH=N), 7.23 (d, 2H, J = 5.8 Hz, ArH), 8.61 (d, 2H, J = 6.0 Hz, ArH).
Preparation of 2-chloropyridine-4-carbaldehvde SLA 07156.
Methyl 2-chloropyridine-4-carboxylate SLA 07150.
2-Chloro-isonicotinic acid (5.10 g, 32.38 mmol) was dissolved in methanol (150 mL). Thionyl chloride (12 mL) was added. This suspension was stirred 5 h at 70 0C and concentrated in vacuo. The residue was dissolved in dichloromethane (250 mL) washed with a solution of 10 % aqueous K2CO3 (2 x 150 mL) dried with MgSO4, filtered and evaporated. Methyl 2-chloropyridine-4-carboxylate SLA 07150 was obtained as a yellow solid (5.06 g, 91 %).
Figure imgf000078_0001
SLA 07150 MW: 171.58; Yield: 91 %; Yellow Solid; Mp (0C): 33.0. Rf\ 0.80 (MeOH:CH2CI2 = 10:90).
1H-NMR (CDCI3, δ): 3.98 (s, 3H, CH3), 7.78 (dd, 1 H, J = 5.1 Hz, J = 1.3 Hz, ArH), 7.89 (d, 1 H, J = 0.6 Hz ArH), 8.55 (dd, 1 H, J = 5.1 Hz, J = 0.6 Hz, ArH).
(2-Chloropyhdin-4-yl)methanol SLA 07152.
Methyl 2-chloropyridine-4-carboxylate (2.50 g, 14.60 mmol) was dissolved in anhydrous THF (50 mL) and this solution was cooled to -78 0C under N2 atmosphere. Diisobutylaluminium hydride 1.0 M in hexanes (63.3 mL, 63.30 mmol) was added dropwise stabilizing the temperature between - 50 0C and -70 0C. The reaction mixture was stirred 1 .5 h at -78 0C and allowed to stand at room temperature for 3 h. A solution of aqueous 10 % NH4CI was slowly added and the mixture was extracted with ethyl acetate (3 x 300 ml_). The combined organic layers were washed with water (3 x 20 ml_), brine (2 x 20 ml_), dried over MgSO4, filtered and evaporated. (2- Chloropyridin-4-yl)methanol SLA 07152 was obtained as a yellow oil (1 .97 g, 94 % yield).
Figure imgf000079_0001
SLA 07152
MW: 143.71 ; Yield: 94 %; Yellow Oil. Rf : 0.35 (EtOAc:cyclohexane = 30:70).
1H-NMR (CDCI3, δ): 2.95 (s broad, 1 H, OH), 4.75 (s, 2H, CH2O), 7.21 (dd, 1 H, J = 5.1 Hz, J = 1.2 Hz, ArH), 7.37 (d, 1 H, J = 1.2 Hz ArH), 8.29 (d, 1 H, J = 5.1 Hz, ArH).
MS-ESI m/z (rel. int.): 144.0 ([MH]+, 100).
HP LC: Method A, detection UV 254 nm, SLA 07152 RT = 3.45 min, peak area 99.9 %.
2-Chloropyridine-4-carbaldehvde SLA 07156.
In a 250 mL tricol equipped with a low temperature thermometer and two equalizing dropping funnels was charged oxalyl dichloride (1.24 g, 9.81 mmol) in dichloromethane (15 mL) and this solution was stirred under N2 at - 78 0C. The first equalizing dropping funnel was connected to a nitrogen flow line and was charged with a solution of (2-chloropyridin-4-yl)methanol SLA 07152 (0.94 g, 6.54 mmol) with dichloromethane (15 mL). The other was charged with a solution of dimethyl sulfoxide anhydrous (1 .7 mL, 19.63 mmol) in dichloromethane (2 mL) and this solution was added dropwise (25 min) in order to stabilize the temperature between -60 0C and -70 0C. At the end of the addition the reaction solution was warmed to -60 0C over a period of 20 min then the solution of (2-chloropyhdin-4-yl)methanol SLA 07152 was added dropwise (50 min) keeping the temperature between -50 0C and -60 0C in the reactor then the mixture reaction was warmed to -45 0C over a period of 30 min. The dropping funnel was washed with dichloromethane (2 x 5 mL) and charged with a solution of thethylamine (480 μl_, 6.51 mmol) in dichloromethane (4 mL) which was added (10 min) to the reaction mixture and finally the reaction flask was allowed to warm to 0 0C over 10 min. The reaction solution was transferred to a 500 mL separatory funnel charged with 130 mL of a 5 % aqueous NH4CI solution. The two phases were separated the aqueous phase was extracted with dichloromethane (3 x 50 mL) and the combined organic phases were washed with 1 M aqueous phosphate buffer (pH = 7; 4x10OmL), then dried over MgSO4, filtered and evaporated. 2- Chloropyridine-4-carbaldehyde SLA 07156 was obtained as an orange solid (0.740 g, 76 % yield).
Figure imgf000080_0001
SLA 07156
MW: 141.57; Yield: 76 %; Orange Solid. Rf: 0.35 (EtOAc:cyclohexane = 30:70).
1H-NMR (CDCI3, δ): 7.65 (dd, 1 H, J = 5.0 Hz, J = 1 .3 Hz, ArH), 7.75 (d, 1 H, J = 1.3 Hz, ArH) 8.66 (d, 1 H, J = 5.0 Hz, ArH), 10.05 (s, 1 H, CHO).
frans-(5-(2-Chloropyridin-4-yl)-4,5-dihvdrooxazol-4-yl)(pyrrolidin-1 - vDmethanone SLA 07174.
SLA 07174 was prepared in accordance with method D using 2- chloropyridine-4-carbaldehyde SLA 07156 (0.12 g, 1.05 mmol), KOH (0.06 g, 1.05 mmol) in methanol (10 mL) and 2-isocyano-1-(pyrrolidin-1-yl)ethanone BLE 04098 (0.146 g, 1.05 mmol). The solution was stirred 24 h with continued cooling. After work-up frans-(5-(2-chloropyridin-4-yl)-4,5-dihydrooxazol-4- yl)(pyrrolidin-1 -yl)methanone SLA 07174 was obtained as a yellow solid (0.19 g, 66 % yield).
Figure imgf000081_0001
(+/-) SLA 07174
MW: 279.72; Yield: 66 %; Yellow Solid; Mp (0C): 1 16.3.
1H-NMR (CDCI3, δ): 1 .86-2.07 (m, 4H, CH2), 3.45-3.62 (m, 3H, CH2N), 3.93-
4.01 (m, 1 H1 CH2N), 4.50 (dd, J = 8.0 Hz, J = 2.3 Hz, 1 H, CH-N), 6.19 ( d, 1 H,
J = 8.0 Hz, CH-O), 7.02 (d, 1 H, J = 2.3 Hz, CH=N), 7.17 (td, 1 H, J = 5.1 Hz J = 0.9 Hz, J = 0.4 Hz, ArH), 7.29 (d, 1 H, J = 0.7 Hz, ArH), 8.38 (d, 1 H, J = 5.1
Hz, ArH).
13C-NMR (CDCI3, δ): 22.5, 24.4, 44.9, 45.0, 74.3, 77.3, 1 17.2, 1 19.0, 148.6,
150.4, 150.6, 153.1 , 164.0.
frans-(5-(3-Bromopyridin-4-yl)-4,5-dihvdrooxazol-4-yl)(pyrrolidin-1 - vDmethanone BAL 01028A.
BAL 01028A was prepared in accordance with method D using 3- bromo-4-pyhdinecarboxaldehyde (1.010 g, 5.43 mmol), KOH (0.305 g, 5.43 mmol) in methanol (5 mL) and 2-isocyano-1-pyrrolidin-1-yl-ethanone BLE 04134 (0.75 g, 5.43 mmol). The mixture was stirred at 0 0C until precipitation and concentrated. The mixture was partitioned between EtOAc (50 ml) and H2O (25 ml). The aqueous layer was extracted twice with EtOAc (25 mL). The EtOAc fractions were combined, washed twice with brine (2x25 mL), dried over MgSO4 and filtered. After evaporation and frans-(5-(3-bromopyhdin-4-yl)- 4,5-dihydrooxazol-4-yl)(pyrrolidin-1 -yl)methanone BAL 01028A was obtained (1 .20 g, 68 % yield) as a white solid.
Figure imgf000081_0002
BAL 01028A
MW: 324.17; Yield: 68 %; White Solid; Mp (0C): 160.8. Rf : 0.25 (EtOAc = 100).
1H NMR (CDCI3, δ): 1.82-2.08 (m, 4H, 2xCH2), 3.45-3.65 (m, 3H, CH2N), 3.80- 3.92 (m, 1 H, CH2N), 4.60 (dd, 1 H, J = 2.1 Hz, J = 6.1 Hz, CH-N), 6.30 (d, 1 H, J = 6.1 Hz, CH-O), 7.10 (d, 1 H, J = 2.1 Hz, OCH=N), 7.30(d, 1 H, J = 5.0 Hz, ArH) ), 8.55 (d, 1 H, J = 5.0 Hz, ArH), 8.72 (s, 1 H, ArH). 13C-NMR (CDCI3, δ): 24.3, 26.0, 46.4, 46.6, 74.5, 79.6, 1 18.6, 121 .1 , 148.3, 148.8, 152.1 , 155.1 , 166.2. MS-ESI m/z (% rel. Int.): 324.1/326.1 ([MH]+, 50/50), 239.0 (100).
HPLC: Method A, detection UV 254 nm, BAL 01028A RT = 3.50 min, peak area 96.8 %.
frans-(5-(3-Chloropyridin-4-yl)-4,5-dihvdrooxazol-4-yl)(pyrrolidin-1 - vDmethanone BAL 01028B.
BAL 01028B was prepared in accordance with method D using 2- isocyano-1-pyrrolidin-1 -yl-ethanone BLE 04134 (0.75 g, 5.43 mmol), KOH (0.305 g, 5.43 mmol) in methanol (5 mL) and 3-chloro-isonicotinaldehyde (0.769 g, 5.43 mmol). The solution was stirred 3 h at 0 0C. trans-{5-{3- Chloropyridin-4-yl)-4,5-dihydrooxazol-4-yl)(pyrrolidin-1 -yl)methanone BAL 01028B (1.20 g, 65% yield) was obtained as a white solid.
Figure imgf000082_0001
BAL 01028B
MW: 279.72; Yield: 65 %; White Solid; Mp (0C): 162. Rf : 0.25 (EtOAc = 100).
1H NMR (CDCI3, δ): 1.82-2.08 (m, 4H, CH2), 3.45-3.65 (m, 3H, CH2N), 3.82- 3.93 (m, 1 H, CH2N), 4.62 (dd, 1 H, J = 2.1 Hz, J = 6.1 Hz, CH-N), 6.38 (d, 1 H, J = 6.1 Hz, CH-O), 7.08 (d, 1 H, J = 2.1 Hz, OCH=N), 7.33 (d, 1 H, J = 5.0 Hz, ArH), 8.52 (d, 1 H, J = 5.0 Hz, ArH), 8.59 (s, 1 H, ArH). 13C-NMR (CD3OD, δ): 24.3, 26.0, 46.4, 46.6, 74.4, 77.9, 120.6, 128.8, 146.6, 148.3, 149.7, 155.0, 166.1.
MS-ESI m/z (% rel. Int.): 280.1/282.1 ([MH]+, 39/14). HPLC: Method A, detection UV 254 nm, BAL 01028B RT = 3.47 min, peak area 97.2 %.
frans-(5-(2-Chloropyhdin-4-yl)-4,5-dihvdrooxazol-4-yl)(2/-/-pyrrol-1 (5/-/)- vDmethanone SLA 07158.
SLA 07158 was prepared in accordance with method D using 2- chloropyridine-4-carbaldehyde SLA 07156 (0.47 g, 3.31 mmol), KOH (0.184 g, 3.33 mmol) in methanol (10 mL) and 2-lsocyano-1-(2/-/-pyrrol-1 (5/-/)- yl)ethanone SLA 07178 (0.410 g, 3.01 mmol). The solution was stirred 2 h with continued cooling. After work-up and column chromatography on florisil (EtOAc), frans-(5-(2-chloropyhdin-4-yl)-4,5-dihydrooxazol-4-yl)(2/-/-pyrrol- 1 (5/-/)-yl)methanone SLA 07158 was obtained as a yellow solid (0.597 g, 84 %).
SLA 07158 MW: 277.71 ; Yield: 84 %; Yellow Solid; Mp (0C): 90.2. Rf\ 0.10 (EtOAc).
1H-NMR (CDCI3, δ): 4.26-4.37 (m, 3H, CH2N), 4.48-4.52 (dd, 1 H, J = 2.3 Hz, J = 8.0 Hz, CH-N), 4.75-4.85 (m, 1 H, CH2N), 5.80-5.95 (m, 2H, CH=CH), 6.20 (d, 1 H, J = 8 Hz, CH-O), 7.02 (d, J = 2.3 Hz, CH=N), 7.17 (td, 1 H, J = 5.1 Hz, J = 0.8 Hz, J = 0.6 Hz, ArH), 7.30 (t, 1 H, J = 0.6 Hz, ArH), 8.38 (d, 1 H, J = 5.1 Hz, ArH).
13C-NMR (CDCI3, δ): 53.4, 53.9, 75.8, 78.9, 1 18.9, 120.7, 125.2, 125.4, 150.3, 151.9, 152.3, 154. frans-(4,5-Dihvdro-5-(pyridin-4-yl)oxazol-4-yl)(2/-/-pyrrol-1 (5/-/)-yl)methanone SLA 07180.
SLA 07158 was prepared in accordance with method D using pyridine- 4-carbaldehyde (0.293 mL, 2.40 mmol), KOH (0.13 g, 2.32 mmol) in methanol (10 mL) and 2-isocyano-1 -(2H-pyrrol-1 (5H)-yl)ethanone SLA 07178 (0.301 g, 2.20 mmol). The solution was stirred 2 h with continued cooling. After work-up and column chromatography on florisil (EtOAc), frans-(4,5-dihydro-5-(pyridin- 4-yl)oxazol-4-yl)(2H-pyrrol-1 (5H)-yl)methanone SLA 07180 was obtained (0.284 g, 53 % yield) as a yellow oil.
Figure imgf000084_0001
(+/-)
SLA 07180
MW: 243.26; Yield: 53 %; Yellow Oil.
Rf. 0.15 (AcOEt). 1H-NMR (CDCI3, δ): 4.28-4.33 (m, 3H, CH2N), 4.52-4.56 (dd, 1 H, J = 7.8 Hz, J
= 2.2 Hz, CH-N), 4.73-4.82 (m, 1 H, CH2N), 5.80-5.93(m, 2H, CH=CH), 6.18
(d, 1 H, J = 7.8 Hz, CH-O), 7.08 (d, J = 2.2 Hz, CH=N), 7.27 (d, 2H, J = 6.0 Hz,
ArH), 8.59 (d, 2H, J = 6.0 Hz, ArH).
13C-NMR (CDCI3, δ): 53.6, 53.9, 75.8, 79.7, 120.2, 125.3, 125.6, 148.7, 150.5, 155.3, 166.1.
Preparation of Compound 20, Compound 21 , Compound 22, Compound 23, Compound 24, Compound 25, Compound 26, Compound 27, Compound 28, Compound 29, Compound 30, Compound 31 , Compound 32, Compound 34, Compound 35, Compound 36, Compound 37, Compound 38, Compound 39, Compound 40, Compound 41 , Compound 42, Compound 43, Compound 44, Compound 45, Compound 46, Compound 48, Compound 49 and Compound 5_α
General method for oxazolines acidic hydrolysis: method E: DL-f/?reo-2-Amino-3-hvdroxy-3-(pyridin-3-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochloride Compound 20.
To a solution of frans-(4,5-dihydro-5-(pyridin-3-yl)oxazol-4- yl)(pyrrolidin-1 -yl)methanone BLE 041 1 OB (0.932 g, 3.80 mmol) in methanol (10 ml.) was added hydrochloric acid 37 % (1.2 ml_). After heating (50 0C) the mixture for 2.25 h the reaction mixture was concentrated and the crude product was coevaporated twice with ethyl acetate. After trituration with ethyl acetate, filtration and drying DL-f/?reo-2-amino-3-hydroxy-3-(pyhdin-3-yl)-1- (pyrrolidin-1-yl)propan-1-one dihydrochloride Compound 20 was obtained as a white solid (1 .10 g, 94 % yield).
Figure imgf000085_0001
Compound 20
MW: 308.2; Yield: 94 %; White Solid; Mp (0C): 123.4.
1H-NMR (CD3OD, δ): 1.65-2.00 (m, 4H, 2xCH2), 2.82-3.1 1 (m, 1 H, -CH2N), 3.30-3.57 (m, 2H, CH2N), 3.57-3.77 (m, 1 H, CH2N), 4.54 (d, 1 H, J = 5.3 Hz,
CH-N), 5.38 (d, 1 H, J = 5.3 Hz, CH-O), 8.15 (dd, 1 H, J = 7.6 Hz, J = 5.0 Hz,
ArH), 8.68 (d, 1 H, J = 7.6 Hz, ArH), 8.89 (d, 1 H, J = 7.6 Hz, ArH), 8.96 (s, 1 H,
ArH).
13C-NMR (CD3OD, δ): 24.9, 26.9, 47.7, 48.2, 58.1 , 69.6, 128.7, 141.5, 141.6, 143.1 , 146.5, 165.4.
DL-f/?reo-2-Amino-3-hvdroxy-3-(pyridin-2-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihvdrochlohde Compound 21 .
Compound 21 was prepared following method E with trans-{4,5- dihydro-5-(pyridin-2-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 041 10 B (0.44 g, 1 .79 mmol), hydrochloric acid 37 % (1 .0 mL) and methanol (10 mL). After 2.5 h at 50 0C and work-up DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-2-yl)- 1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochloride Compound 21 was obtained as a yellow solid (0.44 g, 84 % yield).
Figure imgf000086_0001
Compound 21
MW: 308.28; Yield: 84%; Yellow Solid.
1H-NMR (CD3OD, δ): 1.75-2.01 (m, 4H, 2xCH2), 3.10-3.22 (m, 1 H, CH2N), 3.39-3.60 (m, 2H, CH2N), 3.63-3.75 (m, 1 H, CH2N), 4.71 (d, 1 H, J = 5.0 Hz, CH1N), 5.55 (d, 1 H1 J = 5.0 Hz, CH-O), 8.05 (t, 1 H, J = 6.4 Hz, ArH), 8.13 (d, 1 H, J = 8.0 Hz, ArH), 8.61 (t, 1 H, J = 8.0 Hz, ArH), 8.84 (d, 1 H, J = 5.6 Hz, ArH).
DL-f/?reo-2-Amino-3-hvdroxy-3-(pyridin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochlohde Compound 22.
Compound 22 was prepared following method E with trans-{4,5- dihydro-5-(pyridin-4-yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone Compound 19
(0.750 g, 3.07 mmol), hydrochloric acid 37 % (1.0 mL) and methanol (10 ml_). After 3.0 h at 50 0C and work-up DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-4-yl)-
1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochlohde Compound 22 was obtained as a white solid (0.935 g, 99 % yield).
Figure imgf000086_0002
Compound 22 MW: 308.28; Yield: 99 %; White Solid; Mp (0C): 1 17.0.
1H-NMR (CD3OD, δ): 1.75-2.03 (m, 4H, 2xCH2), 2.93-3.08 (m, 1 H, CHN), 3.32-3.75 (m, 3H, 2xCH2), 4.54 (d, 1 H, J = 5.9 Hz, CH1N), 5.40 (d, 1 H, J = 5.9 Hz, CH-O), 8.21 (d, 2H, J = 5.8 Hz, ArH), 8.94 (d, 2H, J = 5.8 Hz, ArH). MS-ESI m/z (% rel. int.): 236.1 ([MH]+, 17), 219 (25), 148 (100). HPLC: Method A, detection UV 254 nm, Compound 22 RT = 0.8 min, peak area 96.3 %. DL-f/?reo-2-Amino-3-hvdroxy-1 -(pyrrolidin-1 -yl)-3-(thiophen-3-yl)propan-1 -one hydrochloride Compound 23.
Compound 23 was prepared following method E with trans-{4,5- dihydro-5-(thiophen-3-yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone BLE 04124A (0.486 g, 1.94 mmol), hydrochloric acid 37 % (0.6 mL) and methanol (10 ml_). After 3.5 h at 50 0C and work-up DL-f/?reo-2-amino-3-hydroxy-1 -(pyrrolidin-1 - yl)-3-(thiophen-3-yl)propan-1-one hydrochloride Compound 23 was obtained as a white solid (0.480 g, 89.5 % yield).
Figure imgf000087_0001
HCl (+/")
Compound 23
MW: 276.7; Yield: 89.5 %; White Solid; Mp (0C): 227.4. 1H-NMR (CD3OD, δ): 1.47-1.88 (m, 4H, 2xCH2), 2.31-2.46 (m, 1 H, CH2N), 3.18-3.46 (m, 3H, CH2N), 4.16 (d, 1 H, J = 9.0 Hz, CH-N), 4.97 (d, 1 H, J = 9.0 Hz, CH-O), 7.14 (dd, 1 H, J = 4.9 Hz, J = 1 .1 Hz, ArH), 7.40-7.50 (m, 2H, ArH).
13C-NMR (CD3OD, δ): 24.9, 26.7, 47.3, 47.6, 59.2, 70.6, 124.1 , 127.1 , 127.7, 142.3, 166.3.
DL-f/?reo-2-Amino-3-hvdroxy-1 -(pyrrolidin-1 -yl)-3-(thiophen-2-yl)propan-1 -one hydrochloride Compound 24.
Compound 24 was prepared following method E with trans-{4,5- dihydro-5-(thiophen-2-yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone BLE 04124B
(0.677 g, 2.70 mmol), hydrochloric acid 37 % (0.6 mL) and methanol (10 mL). After 3.5 h at 50 0C and work-up DL-f/?reo-2-amino-3-hydroxy-1 -(pyrrolidin-1 - yl)-3-(thiophen-2-yl)propan-1-one hydrochloride Compound 24 was obtained as a white solid (0.630 g, 84.5 % yield).
Figure imgf000088_0001
Compound 24
MW: 276.7; Yield: 84.5 %; White Solid; Mp (0C): 183.2. 1H-NMR (CD3OD, δ): 1.49-1.90 (m, 4H, 2xCH2), 2.36-2.48 (m, 1 H, CH2N), 3.20-3.48 (m, 3H1 CH2N), 4.18 (d, 1 H, J = 9.1 Hz, CH-N), 5.14 (d, 1 H1 J = 9.1 Hz, CH-O), 7.00-7.08 (m, 2H, ArH), 7.45 (dd, 1 H, J = 4.9 Hz, J = 1 .6 Hz, ArH). 13C-NMR (CD3OD, δ): 24.9, 26.8, 47.3, 47.7, 59.6, 70.5, 126.3, 127.0, 128.2, 144.5, 166.1.
DL-f/?reo-2-amino-3-hvdroxy-1 -(pyrrolidin-1 -yl)-3-(thiazol-2-yl)propan-1 -one dihvdrochlohde Compound 25.
Compound 25 was prepared following method E with trans-{4,5- dihydro-5-(thiazol-2-yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone BLE 04124C
(0.558 g, 2.22 mmol), hydrochloric acid 37 % (0.6 ml.) and methanol (10 ml_). After 3.5 h at 50 0C and work-up DL-f/?reo-2-amino-3-hydroxy-1-(pyrrolidin-1 - yl)-3-(thiazol-2-yl)propan-1-one dihydrochloride Compound 25 was obtained as a pale yellow solid (0.532 g, 76.5 % yield).
Figure imgf000088_0002
.HCI (+/-)
Compound 25 MW: 276.7; Yield: 76.5 %; Pale Yellow Solid; Mp (0C): 145.8.
1H-NMR (CD3OD, δ): 1.75-2.00 (m, 4H, 2xCH2), 3.05-3.17 (m, 1 H, -CH2N), 3.36-3.58 (m, 2H, CH2N), 3.58-3.70 (m, 1 H, CH2N), 4.67 (d, 1 H, J = 5.4 Hz, CH-N), 5.49 (d, 1 H, J = 5.4 Hz, CH-O), 7.84 (d, 1 H, J = 3.4 Hz ArH), 7.99 (d, 1 H, J = 3.4 Hz, ArH). 13C-NMR (CD3OD, δ): 24.9, 27.0, 47.7, 48.0, 57.5, 69.9, 123.6, 142.1 , 165.3, 173.3. DL-f/?reo-2-Amino-3-(3a,7a-dihvdrobenzo[ά1thiophen-3-yl)-3-hvdroxy-1 - (pyrrolidin-1-yl)propan-1-one hydrochloride Compound 26.
Compound 26 was prepared following method E with trans-{5-
(benzo[ό]thiophen-3-yl)-4,5-dihydrooxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 04124D (1 .050 g, 3.49 mmol), hydrochloric acid 37 % (1 .2 ml.) and methanol (10 ml_). After 3.5 h at 50 0C and work-up DL-f/?reo-2-amino-3- (3a,7a-dihydrobenzo[ό]thiophen-3-yl)-3-hydroxy-1 -(pyrrolidin-1 -yl)propan-1 - one hydrochloride Compound 26 was obtained as a white solid (0.970 g, 85 % yield).
Figure imgf000089_0001
Compound 26 MW: 326.84; Yield: 85 %; White Solid; Mp (0C): 207.0. 1H-NMR (CD3OD, δ): 0.92-1.09 (m, 2H, 2xCH2), 1.42-1.60 (m, 2H, 2xCH2), 1.83-1.98 (m, 1 H, CH2N), 2.76-2.91 (m, 1 H, CH2N), 3.06-3.25 (m, 2H, - CH2N), 4.30 (d, 1 H, J = 9.5 Hz, CH-N), 5.29 (d, 1 H, J = 9.5 Hz, CH-O), 7.35- 7.43 (m, 2H, ArH), 7.78-7.89 (m, 2H, ArH), 7.90-7.97 (m, 1 H, ArH). 13C-NMR (CD3OD, δ): 24.5, 26.4, 47.3, 47.4, 59.0, 69.5, 123.1 , 124.0, 125.4, 126.1 , 126.8, 136.6, 138.3, 141.9, 166.1 .
DL-f/?reo-2-Amino-3-(furan-3-yl)-3-hvdroxy-1 -(pyrrolidin-1 -yl)propan-1 -one hydrochloride Compound 27.
Compound 27 was prepared following method E with frans-(5-(furan-3- yl)-4,5-dihydrooxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 0413OB (0.800 g, 3.41 mmol), hydrochloric acid 37 % (0.6 mL) and methanol (10 mL). After 3.5 h at 50 0C and work-up DL-f/?reo-2-amino-3-(furan-3-yl)-3-hydroxy-1- (pyrrolidin-1-yl)propan-1-one hydrochloride Compound 27 was obtained as a white solid (0.738 g, 83 % yield).
Figure imgf000090_0001
Compound 27
MW: 260.72; Yield: 83 %; White Solid; Mp (0C): 218.0.
1H-NMR (CD3OD, δ): 1.62-1.95 (m, 4H, 2xCH2), 2.82-2.95 (m, 1 H, CH2N), 3.22-3.38 (m, 1 H, CH2N), 3.39-3.55 (m, 2H, CH2N), 4.19 (d, 1 H, J = 8.4 Hz,
CH-N), 4.90 (d, 1 H, J = 8.4 Hz, CH-O), 6.49 (m, 1 H, ArH), 7.52-7.57 (m, 2H,
ArH).
13C-NMR (CD3OD, δ): 24.9, 26.7, 47.4, 48.0, 58.7, 67.2, 109.8, 125.9, 142.0,
145.2, 166.3.
DL-f/?reo-2-Amino-3-hydroxy-3-(naphthalen-2-yl)-1 -(pyrrolidin-1 -yl)propan-1 - one hydrochloride Compound 28.
Compound 28 was prepared following method E with trans-{4,5- dihydro-5-(naphthalen-3-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 0413OC (0.745 g, 2.53 mmol), hydrochloric acid 37 % (0.6 ml.) and methanol (10 ml_). After 3.5 h at 50 0C and work-up DL-f/?reo-2-amino-3-hydroxy-3- (naphthalen-2-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one hydrochloride Compound 28 was obtained as a white solid (0.706 g, 87 % yield).
Figure imgf000090_0002
HCI
Compound 28
MW: 320.81 ; Yield: 87 %; White Solid; Mp (0C): 173.8. 1H-NMR (CD3OD, δ): 0.93-1.10 (m, 1 H, CH2), 1.20-1 .37 (m, 1 H, CH2), 1.44- 1.71 (m, 2H, CH2), 1.99-2.10 (m, 1 H, CH2N), 3.1 1-3.26 (m, 2H, CH2N), 3.31- 3.41 (m, 1 H, CH2N), 4.23 (d, 1 H, J = 9.1 Hz, CH-N), 5.06 (d, 1 H, J = 9.1 Hz, CH-O), 7.50-7.63 (m, 3H, ArH), 7.87-7.97 (m, 4H, ArH). 13C-NMR (CD3OD, δ): 24.6, 26.3, 47.2, 47.5, 59.4, 74.3, 125.1 , 126.9, 127.7, 127.8,128.8, 129.0, 129.4, 134.5, 135.0, 138.0, 166.4.
DL-f/?reo-2-Amino-3-hvdroxy-3-(naphthalen-1 -yl)-1 -(pyrrolidin-1 -yl)propan-1 - one hydrochloride Compound 29.
Compound 29 was prepared following method E with trans-{4,5- dihydro-5-(naphthalen-4-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 0413OD (0.794 g, 2.69 mmol), hydrochloric acid 37 % (0.6 ml.) and methanol (10 ml_). After 3.5 h at 50 0C and work-up DL-f/?reo-2-amino-3-hydroxy-3- (naphthalen-1-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one hydrochloride Compound 29 was obtained as a white solid (0.768 g, 89 % yield).
Figure imgf000091_0001
HCI
Compound 29
MW: 320.81 ; Yield: 89 %; White Solid; Mp (0C): 177.8. 1H-NMR (CD3OD, δ): 0.71 -0.91 (m, 2H, CH2), 1.29-1 .51 (m, 3H, CH2), 2.54-
2.67 (m, 1 H, CH2N), 2.88-3.02 (m, 1 H, CH2N), 3.02-3.16 (m, 1 H, CH2N), 4.27
(d, 1 H, J = 9.8 Hz, CH-N), 5.67 (d, 1 H, J = 9.8 Hz, CH-O), 7.50-7.61 (m, 3H,
ArH), 7.90-7.98 (m, 3H, ArH), 8.08-8.14 (m, 1 H, ArH).
13C-NMR (CD3OD, δ): 24.4, 26.2, 47.1 , 47.3, 59.5, 70.3, 124.0, 126.5 (2xC), 127.2, 127.4, 129.9, 130.4, 132.1 , 135.0, 137.1 , 166.1.
DL-f/?reo-2-Amino-3-hvdroxy-1 -(pyrrolidin-1 -yl)-3-(αuinolin-2-yl)propan-1 -one dihydrochlohde Compound 30.
Compound 30 was prepared following method E with trans-{4,5- dihydro-5-(quinolin-2-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 04136B (0.923 g, 3.13 mmol), hydrochloric acid 37 % (0.6 mL) and methanol (15 mL). After 3.5 h at 50 0C and work-up DL-f/?reo-2-amino-3-hydroxy-1 -(pyrrolidin-1 - yl)-3-(quinolin-2-yl)propan-1-one dihydrochlohde Compound 30 was obtained as a yellow solid (1 .098 g, 98 % yield).
Figure imgf000092_0001
Compound 30
MW: 358.26; Yield: 98 %; Yellow Solid; Mp (0C): 131.5.
1H-NMR (CD3OD, δ): 1.69-2.07 (m, 4H, CH2), 3.16-3.34 (m, 3H, CH2), 3.37- 3.60 (m, 2H, CH2N), 3.77-3.88 (m, 1 H, CH2-N), 5.85 (d, 1 H, J = 4.9 Hz, CH-
O), 8.03 (t, 1 H, J = 7.6 Hz, ArH), 8.17-8.30 (m, 2H, ArH), 8.40 (d, 1 H, J = 8.3
Hz, ArH), 8.56 (d, 1 H, J = 8.6 Hz, ArH), 9.25 (d, 1 H, J = 8.6 Hz, ArH), not seen under H2O (d, 1 H, CH-NH2).
13C-NMR (CD3OD, δ): 24.9, 27.0, 47.9, 48.2, 57.3, 70.3, 121.5, 122.5, 130.4, 130.5, 131.5, 136.5, 140.2, 148.5, 157.8, 164.8.
DL-f/?reo-2-Amino-3-hvdroxy-3-(isoαuinolin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 - one dihydrochloride Compound 31.
Compound 31 was prepared following method E with trans-{4,5- dihydro-5-(isoquinolin-4-yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone BLE
04136C (0.597 g, 2.02 mmol), hydrochloric acid 37 % (0.4 mL) and methanol (10 mL). After 3.5 h at 50 0C and work-up DL-f/?reo-2-amino-3-hydroxy-3- (isoquinolin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochloride Compound 31 was obtained as a off white solid (0.716 g, 99 % yield).
Figure imgf000092_0002
2.HCI (+/")
Compound 31
MW: 358.27; Yield: 99 %; Off White Solid; Mp (0C): 158.5. 1H-NMR (CD3OD, δ): 1.04-1.32 (m, 2H, CH2), 1.51-1 .72 (m, 2H, CH2), 2.05- 2.20 (m, 1 H, CH2N), 2.68-2.80 (m, 1 H, CH2N), 3.20-2.47 (m, 1 H, CH2N), 4.57 (d, 1 H, J = 8.5 Hz, CH-NH2), 5.99 (d, 1 H, J = 8.5 Hz, CH-OH), 8.09 (t, 1 H, J = 8.6 Hz, ArH), 8.27 (t, 1 H, J = 8.6 Hz, ArH), 8.38 (d, 1 H, J = 8.6 Hz, ArH), 8.45 (d, 1 H, J = 8.6 Hz, ArH), 8.55 (d, 1 H, J = 5.6 Hz, ArH), 9.35 (d, 1 H, J = 5.6 Hz, ArH).
13 C-NMR (CD3OD, δ): 24.6, 26.5, 47.5, 48.0, 58.2, 69.1 , 122.3, 122.6, 125.9, 127.7, 131.7, 136.7, 138.9, 146.1 , 159.4, 165.2.
Λ/-(DL-f/?reo-1 -hydroxy-3-oxo-3-(pyrrolidin-1 -yl)-1 -(quinolin-3-yl)propan-2- vDformamide hydrochloride Compound 32.
Compound 32 was prepared following method E with trans-{4,5- dihydro-5-(quinolin-3-yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone BAL 01016 (0.905 g, 3.41 mmol), hydrochloric acid 37 % (0.6 ml.) and methanol (10 ml_). After 2 h at RT and work-up Λ/-(DL-f/?reo-1-hydroxy-3-oxo-3-(pyrrolidin-1-yl)-1- (quinolin-3-yl)propan-2-yl)formamide hydrochloride Compound 32 was obtained as a white solid (240 mg, 20.0 % yield).
Figure imgf000093_0001
Compound 32
MW: 349.81 ; Yield: 20.0 %; White Solid; Mp (0C): 203.2.
1H NMR (CD3OD, δ): 1.78-2.09 (m, 4H, CH2), 3.35-3.58 (m, 2 H, CH2N), 3.58-
3.80 (m, 2 H, CH2N), 5.28 (d, 1 H, J = 4 Hz, CH-N), 5.51 (d, 1 H, J = 4 Hz, CH- O), 8.00 (t, 2H1 J = 7.1 Hz, ArH) ), 8.18 (t, 1 H, J = 6.9 Hz, ArH), 8.26 (d, 1 H, J
= 8.6 Hz, ArH) , 8.36 (d, 1 H, J = 8.3 Hz, ArH), 9.18 (s, 1 H, CHO), 9.26 (s, 1 H,
ArH).
13C-NMR (CD3OD, δ): 25.1 , 27.0, 47.5, 48.3, 55.5, 71.2, 121.4, 129.9, 130.6,
131.5, 136.2, 137.3, 138.5, 145.3, 145.8, 163.4, 168.7. MS-ESI m/z (% rel. Int.): 314 ([MH]+, 50), 158.1 (100).
HPLC: Method A, detection UV 254nm, Compound 32 RT = 3.36 min, peak area 99.9 %. DL-f/?reo-2-Amino-3-hvdroxy-1 -(pyrrolidin-1 -yl)-3-(αuinolin-3-yl)propan-1 -one dihvdrochloride Compound 33.
Compound 33 was prepared following method E with trans-{4,5- dihydro-5-(quinolin-3-yl)oxazol-4-yl)(pyrrolidin-1-yl)methanone BAL 01016 (0.91 g, 3.41 mmol), hydrochloric acid 37% (0.6 mL) and methanol (10 ml_). After 3 h at 50 0C and work-up DL-f/?reo-2-amino-3-hydroxy-1 -(pyrrolidin-1 -yl)- 3-(quinolin-3-yl)propan-1-one dihydrochlohde Compound 33 (678 mg, 55 % yield) was obtained as a white solid.
Figure imgf000094_0001
2 HCI (+/-) Compound 33
MW: 358.26; Yield: 55 %; White Solid; Mp (0C): 190.9.
1H NMR (CD3OD, δ): 1.57-1 .80 (m, 2H, CH2), 1.80-1.99 (m, 2H, CH2), 3.01-
3.20 (m, 1 H, CH2N), 3.35-3.61 (m, 2H, CH2N), 3.61-3.82(m, 1 H, CH2N), 4.70
(d, 1 H, J = 5.0 Hz, CH-N), 5.58 (d, 1 H, J = 5.0 Hz, CH-O), 7.96-8.1 1 (m, 1 H, ArH), 8.18-8.29 (m, 1 H, ArH), 8.29-8.38 (m, 1 H, ArH), 8.38-8.49 (m, 1 H, ArH),
9.28 (s, 1 H, ArH), 9.34 (s, 1 H, ArH).
13C-NMR (CD3OD, δ): 24.9, 26.9, 47.8, 48.3, 58.2, 69.8, 121.8, 130.0, 130.8,
131.9, 135.4, 136.9, 139.3, 145.1 , 146.2, 165.6.
MS-ESI m/z (% rel. Int.): 286.2 ([MH]M OO). HPLC: Method A, detection UV 254 nm, Compound 33 RT = 3.15 min, peak area 97.0 %.
DL-f/?reo-2-Amino-3-(2-chloropyhdin-4-yl)-3-hvdroxy-1-(2H-pyrrol-1 (5H)- yl)propan-1 -one dihvdrochloride Compound 34. Compound 34 was prepared following method E with trans-{5-{2- chloropyridin-4-yl)-4,5-dihydrooxazol-4-yl)(2/-/-pyrrol-1 (5/-/)-yl)methanone SLA 07158 (0.597 g, 2.02 mmol), hydrochloric acid 37 % (1 .0 mL) and methanol (10 mL). After 2 h at room temperature and work-up DL-f/?reo-2-amino-3-(2- chloropyridin-4-yl)-3-hydroxy-1 -(2/-/-pyrrol-1 (5/-/)-yl)propan-1 -one dihydrochloride Compound 34 (0.656 mg, 91 % yield) was obtained as a pale yellow solid.
Figure imgf000095_0001
Compound 34 MW: 340.63; Yield: 91 %; Pale Yellow Solid; Mp (0C): 196.2.
1H-NMR (CD3OD, δ): 3.45-3.50 (m, 1 H, CH2N), 4.04-4.15 (m, 1 H, CH2N), 4.22-4.36 (m, 3H, CH2N & CHNH2), 5.05 (d, 1 H, J = 7.1 Hz, -CHO), 5.71 (d, 1 H, J = 4.3 Hz, CH=CH), 5.84 (d, 1 H, J = 4.3 Hz, CH=CH), 7.47 (d, 1 H, J = 5.0 Hz, ArH), 7.57 (s, 1 H, ArH), 8.39 (d, 1 H, J = 5.0 Hz, ArH). 13C-NMR (CD3OD, δ): 54.3, 54.5, 58.2, 71 .7, 122.0, 123.5, 125.7, 126.3, 151.0, 152.8, 154.0, 165.8.
DL-f/?reo-2-Amino-3-hvdroxy-3-(pyhdin-4-yl)-1 -(piperidin-1 -yl)propan-1 -one dihydrochlohde Compound 35. Compound 35 was prepared following method E with trans-{4,5- dihydro-5-(pyridin-4-yl)oxazol-4-yl)(pipehdin-1-yl)methanone SLA 07122A (0.33 g, 1 .27 mmol), hydrochloric acid 37 % (1 .0 ml.) and methanol (10 ml_). After 3 h at 50 0C and work-up DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-4-yl)-1 - (pipehdin-1 -yl)propan-1-one dihydrochloride Compound 35 was obtained as a yellow solid (0.375 g, 91 % yield).
Figure imgf000095_0002
(+/-) 2. HCl
Compound 35
MW: 322.31 ; Yield: 91 %; Yellow Solid; Mp (0C): 145. 1H-NMR (CD3OD, δ): 1.05-1.17 (m, 1 H, CH2) 1.28-1 .65 (m, 5H, CH2), 2.75- 3.00 (m, 1 H, CH2N), 3.10-3.22 (m, 1 H, CH2N), 3.23-3.38 (m, 1 H, CH2N), 3.53- 3.65 (m, 1 H, CH2N), 4.68 (d, 1 H, J = 5.8 Hz, CHNH2), 5.14 (d, 1 H, J = 5.8 Hz, CHO), 8.06 (d, 2H, J = 6.0 Hz, ArH), 8.79 (d, 2H, J = 6.5 Hz, ArH).
DL-f/?reo-2-Amino-3-hvdroxy-3-(pyridin-4-yl)-1 -(morpholin-1 -yl)propan-1 -one dihydrochloride Compound 36.
Compound 36 was prepared following method E with trans-{4,5- dihydro-5-(pyridin-4-yl)oxazol-4-yl)(morpholino)methanone SLA 07124A
(0.146 g, 0.56 mmol), hydrochloric acid 37 % (1.0 mL) and methanol (10 ml_).
After 3 h at 50 0C and work-up DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-4-yl)-1 - (morpholin-1 -yl)propan-1 -one dihydrochloride Compound 36 was obtained as a pale yellow solid (0.143 g, 89 % yield).
Figure imgf000096_0001
Compound 36
MW: 287.78; Yield: 89 %; Pale Yellow Solid; Mp (0C): 1 15.9. 1H-NMR (CD3OD, δ): 3.32-3.82 (m, 8H, 4xCH2), 5.41 (d, 1 H, J = 5.0 Hz, CHO- ), 8.28 (d, 2H, J = 5.9 Hz, ArH), 8.97 (d, 2H, J = 5.8 Hz, ArH), CHNH2 not seen.
DL-f/?reo-2-Amino-3-hvdroxy-3-(pyhdin-4-yl)-1 -(piperazin-1 -yl)propan-1 -one thhydrochlohde Compound 37.
Compound 37 was prepared following method E with trans-{4,5- dihydro-5-(pyridin-4-yl)oxazol-4-yl)(4-ferf-butyloxycarbonyl-piperazin-1- yl)methanone SLA 07124B (0.31 g, 0.86 mmol), hydrochloric acid 37 % (1 .0 mL) and methanol (10 mL). After 3 h at 50 0C and work-up DL-f/?reo-2-amino- 3-hydroxy-3-(pyhdin-4-yl)-1 -(piperazin-1 -yl)propan-1 -one thhydrochloride Compound 37 was obtained as a yellow solid (0.303 g, 71 %).
Figure imgf000097_0001
(+/-) 3. HCI
Compound 37
MW: 359.8; Yield: 71 %; Yellow Solid; Mp (0C): 201 .4. Rf. 0.20 (CH2CI2:Me0H = 90:10), free base. 1H-NMR (CD3OD, δ): 3.31-3.48 (m, 4H, 2xCH2), 3.63-3.90 (m, 2H, CH2N), 4.00-4.35 (m, 2H, CH2N), 5.15 (d, 1 H, J = 4.5 Hz, CHNH2), 5.58 (d, 1 H, J = 4.5 Hz, CHO), 8.38 (d, 2H, J = 6.4 Hz, ArH), 9.04 (d, 2H, J = 6.5 Hz, ArH).
DL-f/?reo-2-amino-3-hvdroxy-3-(pyridin-4-yl)-1-thiomorpholinopropan-1-one dihydrochloride Compound 38.
Compound 38 was prepared following method E with trans-{4,5- dihydro-5-(pyridin-4-yl)oxazol-4-yl)(thiomorpholino)methanone SLA 07132 (0.926 g, 3.36 mmol), hydrochloric acid 37 % (1.1 ml.) and methanol (10 ml_). After 3 h at 50 0C and work-up DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-4-yl)-1 - thiomorpholinopropan-1 -one dihydrochloride Compound 38 was obtained as a pale yellow solid (1 .1 g, 99 % yield).
Figure imgf000097_0002
Compound 38 MW: 340.35; Yield: 99 %; Pale Yellow Solid; Mp (0C): 200.6.
1H-NMR (CD3OD, δ): 2.42-2.52 (m, 1 H, CH2), 2.53-2.70 (m, 1 H, CH2), 2.70- 2.90 (m, 2H, CH2), 3.45-3.71 (m, 2H, CH2N), 3.87-4.00 (m, 1 H, CH2N), 4.18- 4.28 (m, 1 H, CH2N), 5.44 (d, 1 H, J = 5.1 Hz, CHO), 8.34 (d, 2H, J = 5.9 Hz, ArH), 9.03 (d, 2H, J = 5.6 Hz, ArH), -CHNH2 not seen (under H2O). 13C-NMR (CD3OD, δ): 27.9, 28.7, 46.6, 50.0, 56.0, 71 .3, 126.9 (2xC), 143.2 (2xC), 161.3, 165.7. DL-f/?reo-2-Amino-3-hvdroxy-3-(pyridin-4-yl)-1 -(2/-/-pyrrol-1 (5/-/)-yl)propan-1 - one dihvdrochloride Compound 39.
Compound 39 was prepared following method E with trans-{4,5- dihydro-5-(pyridin-4-yl)oxazol-4-yl)(2H-pyrrol-1 (5H)-yl)methanone SLA 07180 (0.276 g, 1.14 mmol), hydrochloric acid 37 % (1.0 ml.) and methanol (10 ml_). After 2.5 h at RT and work-up DL-f/?reo-2-amino-3-hydroxy-3-(pyhdin-4-yl)-1- (2H-pyrrol-1 (5H)-yl)propan-1-one dihydrochloride Compound 39 was obtained (343 mg, 99 % yield) as a white solid.
Figure imgf000098_0001
Compound 39
MW: 306.27; Yield: 99 %; White Solid; Mp (0C): 186.3.
1H-NMR (CD3OD, δ): 3.91-4.02 (m, 1 H, CH-NH2), 4.09-4.21 (m, 1 H, CH2), 4.27-4.41 (m, 1 H, CH2), 4.44-4.59 (m, 1 H, CH2), 4.55 (d, 1 H, J = 5.7 Hz, CH2N), 5.46 (d, 1 H, J = 5.7 Hz, CHO), 5.80-5.90 (m, 2H, CH=CH), 8.24 (d, 1 H, J = 6.3 Hz, ArH), 8.93 (d, 1 H, J = 5.7 Hz, ArH).
13C-NMR (CD3OD, δ): 54.6, 54.7, 57.6, 71.0, 125.9, 126.4, 126.8 (2xC), 143.1 (2xC), 161.6, 165.5. MS-ESI m/z (% rel. Int.): 234.1 ([MH]+, 5), 137.1 (100).
DL-f/?reo-2-Amino-Λ/,Λ/-diethyl-3-hvdroxy-3-(pyridin-4-yl)propanamide dihvdrochloride Compound 40.
Compound 40 was prepared following method E with frans-Λ/,Λ/-diethyl- 4,5-dihydro-5-(pyridin-4-yl)oxazole-4-carboxamide diethylamide SLA 07194A (254 mg, 1.03 mmol), hydrochloric acid 37 % (1.0 mL) and methanol (10 mL). After 2 h at RT and work-up DL-f/?reo-2-amino-Λ/,Λ/-diethyl-3-hydroxy-3- (pyhdin-4-yl)propanamide dihydrochloride Compound 40 was obtained (212 mg, 67 % yield) as a pale yellow solid.
Figure imgf000099_0001
(+/-) 2. HCI
Compound 40
MW: 310.30; Yield: 67 %; Pale Yellow Solid; Mp (0C): 159.6°C. Rf. 0.10 (CH2CI2:Me0H = 90:10), free base.
1H-NMR (CD3OD, δ): 1.01-1.12 (m, 6H, 2xCH3), 3.01-3.31 (m, 3H, CH2), 3.40- 3.52 (m, 1 H, CH2), 4.64 (d, 1 H, J = 6.8 Hz, CHN), 5.31 (d, 1 H, J = 6.8 Hz, CHO), 8.22 (d, 1 H, J = 6.4 Hz, ArH), 8.94 (d, 1 H, J = 6.4 Hz, ArH). 13C-NMR (CD3OD, δ): 12.9, 14.4, 42.1 , 43.4, 55.9, 72.1 , 126.9 (2xC), 143.3 (2xC), 161.5, 166.1.
MS-ESI m/z (% rel. Int.): 238.1 ([MH]+, 5), 137.1 (100).
DL-f/?reo-2-Amino-3-(2-chloropyridin-4-yl)-3-hvdroxy-1-(pyrrolidin-1-yl)propan- 1-one dihydrochloride Compound 41. Compound 41 was prepared following method E with trans-{5-{2- chloropyridin-4-yl)-4,5-dihydrooxazol-4-yl)(pyrrolidin-1 -yl)methanone SLA 07174 (0.179 g, 0.64 mmol), hydrochloric acid 37 % (1.0 ml.) and methanol (7 ml_). After 2 h at RT and work-up DL-f/?reo-2-amino-3-(2-chloropyridin-4-yl)-3- hydroxy-1 -(pyrrolidin-1-yl)propan-1 -one dihydrochloride Compound 41 was obtained (212 mg, 67 (142 mg, 65 % yield) as a pale yellow solid.
Figure imgf000099_0002
Compound 41
MW: 342.65; Yield: 65 %; Pale Yellow Solid; Mp (0C): 184.3. Rf. 0.15 (CH2CI2:Me0H = 90:10), free base. 1H-NMR (CD3OD, δ): 1.50-1.90 (m, 4H, 2xCH2), 2.50-2.61 (m, 1 H, CH2N), 3.25-3.38 (m, 1 H, CH2N), 3.40-3.53 (m, 2H, CH2N), 4.26 (d, 1 H, J = 7.7 Hz, CHN), 4.99 (d, 1 H, J = 7.7 Hz, CHO), 7.45 (d, 1 H, J = 4.5 Hz, ArH), 7.53 (s,
1 H, ArH), 8.41 (d, 1 H, J = 4.9 Hz, ArH).
13C-NMR (CD3OD, δ): 24.9, 26.7, 47.5, 48.0, 58.4, 71.9, 122.0, 123.5, 151.1 ,
152.7, 154.0, 165.7. MS-ESI m/z (% rel. Int.): 270.1/272.1 ([MH]+, 40/13), 171.0/172.0 (100/32).
DL-f/?reo-2-Amino-3-(3-bromopyridin-4-yl)-3-hvdroxy-1 -(pyrrolidin-1-yl)propan- 1-one dihydrochloride Compound 42.
Compound 42 was prepared following method E with trans-{5-{3- bromopyridin-4-yl)-4,5-dihydrooxazol-4-yl)(pyrrolidin-1 -yl)methanone BAL 01028A (1.141 g, 3.52 mmol), hydrochloric acid 37 % (0.6 ml.) and methanol (15 ml_). After 3 h at 50 0C and work-up DL-f/?reo-2-amino-3-(3-bromopyridin- 4-yl)-3-hydroxy-1-(pyrrolidin-1-yl)propan-1-one dihydrochloride Compound 42 was obtained as a white solid (667 mg, 49 % yield).
Figure imgf000100_0001
(+/-) 2. HCI
Compound 42
MW: 387.10; Yield: 49.0 %; White Solid; Mp (0C): 216.3.
1H NMR (CD3OD, δ): 1.73-1 .99 (m, 4H, 2xCH2), 3.01-3.05 (m, 1 H, CH2N),
3.44-3.51 (m, 2H, CH2N), 3.60-3.73(m, 1 H, CH2N), 4.60 (d, 1 H, J = 5.5 Hz, CH-N), 5.54 (d, 1 H, J = 5.5 Hz, CH-O), 8.26 (d, 1 H, J = 5.7 Hz, ArH) ), 8.86
(d, 1 H, J = 5.7 Hz1 ArH), 9.10 (s, 1 H1 ArH).
13C-NMR (CD3OD, δ): 24.8, 27.1 , 56.4, 70.1 , 122.7, 127.6, 145.1 , 148.4,
156.8, 165.0, 2xC not seen.
MS-ESI m/z (% rel. Int.): 314.1/316.1 ([MH]+, 35/35), 215.0/217 (50/50). HPLC: Method A, detection UV 254 nm, Compound 42 RT = 3.08 min, peak area 92.8 %.
DL-f^reo^-Amino-S-O-chloropyridin^-vD-S-hvdroxy-i-fpyrrolidin-i-vDpropan- 1-one dihydrochloride Compound 43. Compound 43 was prepared following method E with trans-{5-{3- chloropyridin-4-yl)-4,5-dihydrooxazol-4-yl)(pyrrolidin-1 -yl)methanone BAL 01028B (0.925 g, 3.31 mmol), hydrochloric acid 37 % (0.6 ml.) and methanol (15 ml_). After 2 h at 50 0C and work-up DL-f/?reo-2-amino-3-(3-chloropyridin- 4-yl)-3-hydroxy-1-(pyrrolidin-1-yl)propan-1-one dihydrochlohde Compound 43 was obtained as a white solid (599 mg, 53 % yield).
Figure imgf000101_0001
(+/-) 2. HCI
Compound 43
MW: 342.65; Yield: 53 %; White Solid; Mp (0C): 214.0. 1H NMR (CD3OD, δ): 1.75-2.02 (m, 4H, 2xCH2), 3.13-3.25 (m, 1 H, CH2N),
3.39-3.62(m, 2H, CH2N), 3.65-3.80 (m, 1 H, CH2N), 4.66 (d, 1 H, J = 4.5 Hz,
CH-N), 5.66 (d, 1 H, J = 4.6 Hz, CH-O), 8.40 (d, 1 H, J = 5.8 Hz, ArH), 8.93 (d,
1 H, J = 5.8 Hz1 ArH), 9.13 (s, 1 H1 ArH).
13C-NMR (CD3OD, δ): 23.3, 25.6, 46.6, 46.8, 54.6, 66.5, 126.5, 132.4, 141.7, 142.9, 155.6, 163.4.
MS-ESI m/z (% rel. Int.): 270/272 ([MH]+, 33/1 1 ), 171/173 (100/32).
HPLC: Method A, detection UV 254 nm, Compound 43 RT = 2.80 min, peak area 97.2 %.
DL-f/?reo-3-Hvdroxy-1-oxo-3-(1-oxy-pyridin-4-yl)-1-(pyrrolidin-1-yl)propan-2- ylcarbamate BAL 01060.
To a solution of DL-f/?reo-3-hydroxy-1 -oxo-3-(pyhdin-4-yl)-1-(pyrrolidin- 1-yl)propan-2-ylcarbamate (300 mg, 0.81 mmol, free base obtained from Compound 58 by K2CO3, CH2CI2 treatment) in dichloromethane (40 mL) was added MCPBA (350 mg, 2.03 mmol). The resulting mixture was stirred overnight at room temperature. The mixture was concentrated and the crude product was purified by column chromatography (EtOAc:MeOH = 70:30). DL- f/?reo-3-Hydroxy-1 -oxo-3-(1 -oxy-pyridin-4-yl)-1 -(pyrrolidin-1 -yl)propan-2- ylcarbamate BAL 01060 was obtained as a white solid (292 mg, 94 % yield).
Figure imgf000102_0001
BAL 01060
MW: 385.41 ; Yield: 94 %; White Solid.
1H NMR CDCI3, δ): 1 .71-2.00 (m, 4H, 2xCH2), 3.35-3.53 (m, 3H, CH2N), 3.54- 3.68 (m, 1 H, CH2N), 4.65 (dd, 1 H, J = 9.6 Hz, J = 2.0 Hz, CH-N), 4.90-5.12 (m, 3H, CH2O & OH), 5.20 (d, 1 H, J = 1.9 Hz, CH-O), 5.83 (d, 1 H, J = 9.6 Hz, NH), 7.20-7.40 (m, 7H, ArH), 8.08 (d, 2H, J = 7.1 Hz, ArH).
DL-f/?reo-2-Amino-3-hydroxy-3-(1 -oxy-pyridin-4-yl)-1 -pyrrolidin-1 -yl-propan-1 - one hydrochloride Compound 44.
[2-Hydroxy-2-(1 -oxy-pyridin-4-yl)-1 -(pyrrolidine-1 -carbonyl)-ethyl]- carbamic acid benzyl ester BAL 01060 (0.26 g, 0.67 mmol) was dissolved in a 6 N hydrochloric acid solution (10 mL). The solution was stirred for 0.75 h at 100 0C. The residue was concentrated, dissolved in MeOH:EtOAc = 50:50 and heated at reflux. After cooling, the mixture was evaporated, triturated in MeOH and filtered to obtain DL-f/?reo-2-amino-3-hydroxy-3-(1-oxy-pyhdin-4- yl)-1-pyrrolidin-1-yl-propan-1-one hydrochloride Compound 44 (65 mg, 33 % yield) as a white solid.
OH O
NH2
HCI
Compound 44
MW: 287.74; Yield: 33 %; White Solid; Mp (0C): 178.5. 1H NMR (D2O, δ): 1.55-1.93 (m, 4H, 2xCH2), 2.65-3.80 (m, 1 H, CH2N), 3.22- 3.56 (m, 3H, CH2N), 4.43 (d, 1 H, J = 7.6 Hz, CH-N), 5.19 (d, 1 H, J = 7.6 Hz, CH-O), 7.69 (d, 2H, J = 6.1 Hz, ArH), 8.39 (d, 2H, J = 6.9 Hz, ArH). 13C-NMR (D2O, δ): 24.1 , 25.8, 47.3, 48.0, 57.4, 70.5, 125.7 (2χC), 139.9
Figure imgf000103_0001
.
MS-ESI m/z (% rel. Int.): 252.1 ([MH]+, 18), 120.0 (100). HPLC: Method A, detection UV 254nm, Compound 44 RT = 0.8 min, peak area 99.9 %.
DL-f/?reo-2-(Diniethylamino)-3-hvdroxy-3-(pyhdin-4-yl)-1-(pyrrolidin-1- yl)propan-1 -one dihydrochloride Compound 45.
DL-f/?reo-2-Amino-3-hydroxy-3-(pyridin-4-yl)-1-(pyrrolidin-1-yl)propan- 1-one dihydrochloride Compound 22 (0.50 g, 1.62 mmol) and para formaldehyde (0.245 g, 8.1 1 mmol) were stirred in methanol (25 ml_) for 10 min. Sodium cyanoborohydride (0.612 g, 9.73 mmol) was added. The solution was stirred 19 h at 50 0C and then concentrated. The residue was partitioned between dichloromethane and water. The aqueous layer was basified with 1 N sodium hydroxyde (pH = 10). The organic layer was combined with additional dichloromethane extracts, washed with aqueous sodium chloride and dried with MgSO4. The crude product was purified by column chromatography on silica (CH2CI2:Me0H = 95:05). DL-f/?reo-2-(dimethylamino)-3-hydroxy-3- (pyhdin-4-yl)-1-(pyrrolidin-1-yl)propan-1-one SLA 07140 was obtained (187 mg, 44 %) as a yellow oil. To a stirred solution of DL-f/?reo-2-(dimethylamino)- 3-hydroxy-3-(pyhdin-4-yl)-1-(pyrrolidin-1-yl)propan-1-one SLA 07140 (0.142 g, 0.54 mmol) in ethyl acetate (5 mL) was added dropwise via syringe 4 mL of a solution of HCI in Et2O (0.3 M). The reaction mixture was stirred at 0 0C for 0.5 h. The precipitate was filtered, washed with Et2O and dried. DL-threo-2- (Dimethylamino)-3-hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochloride Compound 45 was obtained (0.077 g, 43 % yield) as a white solid.
Figure imgf000103_0002
(+/-) 2 HCI
Compound 45 MW: 336.34; Yield: 43 %; White Solid; Mp (0C): 201.0.
1H-NMR (CD3OD,.: 1 .48-1.64 (m, 2H, CH2), 1.65-1.83 (m, 2H, CH2), 2.60-2.72 (m, 1 H, CH2N), 3.15-3.33 (m, 1 H, CH2), 3.30-3.52 (m, 2H, CH2), 4.60 (d, 1 H, J = 8.4 Hz, CHNH2), 5. 41 (d, 1 H, J = 8.4 Hz, CHO), 8.10 (d, 2H, J = 6.6 Hz, ArH), 8.84 (d, 2H, J = 6.7 Hz, ArH).
DL-f/?reo-2-Amino-1 -(pyridin-4-yl)-3-(pyrrolidin-1 -yl)propan-1 -ol Compound 46. To a stirred suspension of DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-4-yl)- 1-(pyrrolidin-1-yl)propan-1 -one dihydrochlohde Compound 22 (0.86 g, 2.80 mmol) in tetrahydrofuran (108 mL) under nitrogen atmosphere was slowly added, in two portions, lithium aluminium hydride (0.64 g, 16.82 mmol) at 0 0C. The mixture reaction was stirred at RT for 20 h and quenched by a slow, dropwise addition of 2 N aqueous sodium hydroxyde (8.4 mL, 6 eq). The yellow precipitate was filtered. The organic layer was washed by water (80 mL) and the organic layer was removed and combined with additional ethyl acetate extracts (4 x 200 mL) and dried over MgSO4, filtered and evaporated. The crude product was purified by column chromatography on silica (CH2CI2:Me0H:NH3 = 94:05:01 ). After evaporation and drying Dl-threo-2- amino-1-(pyridin-4-yl)-3-(pyrrolidin-1-yl)propan-1 -ol Compound 46 was obtained (0. 075 g, 12 % yield) as a pale yellow solid.
OH
Figure imgf000104_0001
Compound 46
MW: 221.30; Yield: 12 %; Pale Yellow Solid. Rf. 0.35 (CH2CI2:Me0H:NH3 = 90:08:02). 1H-NMR (CD3OD, δ): 1.60-1.80 (m, 4H, 2xCH2), 2.30-2.80 (m, 6H, 3xCH2N), 3.14-3.19 (m, 1 H, CHNH2), 4.68 (d, 1 H, J = 3.0 Hz, CHO), 7.30 (d, 2H, J = 6.0 Hz, ArH), 8.55 (d, 2H, J = 6.0 Hz, ArH).
13C-NMR (CD3OD, δ): 23.5 (2xC), 54.1 , 54.7 (2xC), 60.1 , 74.5, 121.4 (2xC), 149.5 (2xC), 152.1. MS-ESI m/z (rel. int.): 222.1 ([MH]+, 100), 205.0 (80), 189.0 (45), 151.0 (70), 134.0 (42), 121 .9 (100), 107.9 (40).
DL-f/?reo-2-Amino-3-hvdroxy-3-(2-niethoxypyridin-3-yl)-1-(pyrrolidin-1- yl)propan-1 -one dihydrochloride Compound 48.
7rans-(4,5-Dihydro-5-(2-methoxypyridin-3-yl)oxazol-4-yl)(pyrrolidin-1- yl)methanone BAL 01014 (0.465 g, 1.69 mmol) was dissolved in methanol (6 ml_). The solution of hydrochloric acid (37 %, 0.3 mL) was added via a syringe at RT. The mixture was stirred for 3 h at RT. The residue was concentrated, dissolved in the minimum of MeOH, precipitated with EtOAc and filtered to obtain a white solid DL-f/?reo-2-amino-3-hydroxy-3-(2-methoxypyhdin-3-yl)-1- (pyrrolidin-1-yl)propan-1-one dihydrochloride Compound 48 (103 mg, 18.0 % yield).
Figure imgf000105_0001
(+/-) 2. HCI Compound 48
MW: 338.23; Yield: 18.0 %; White Solid; Mp (0C): 171.5.
1H NMR (CD3OD, δ): 1.85-2.10 (m, 4H, CH2), 3.30-3.82 (m, 4 H, CH2N), 4.26
(s, 3H, OCH3), 4.60 (d, 1 H, J = 3.7 Hz, CH-N), 5.45 (d, 1 H, J = 3.7 Hz, CH-O),
7.39 (dd, 1 H, J = 5.6 Hz, J = 7.3, ArH), 8.32 (dd, 1 H, J = 5.6 Hz, J = 7.3 Hz, ArH).
13C-NMR (CD3OD, δ): 24.9, 27.1 , 47.8, 47.9, 56.4, 56.7, 66.0, 1 19.2, 125.6,
141.5, 143.9, 160.6, 166.1.
MS-ESI m/z (% rel. Int.): 266.2 ([MH]+, 30), 248.2.0 (100).
HPLC: Method A, detection UV 254nm, Compound 48 RT = 3.31 min, peak area 97.9 %.
3-(DL-f/?reo-2-Amino-1 -hydroxy-3-oxo-3-pyrrolidin-1 -yl-propyD-1 /-/-pyhdin-2- one hydrochloride Compound 49. 7rans-(4,5-Dihydro-5-(2-methoxypyridin-3-yl)oxazol-4-yl)(pyrrolidin-1- yl)methanone BAL 01014 (0.684 g, 2.487 mmol) was dissolved in methanol (10 ml_). A solution of hydrochloric acid (37 %, 0.6 mL) was added via syringe at RT. The mixture was stirred for 22 h at reflux. The residue was concentrated, triturated with EtOAc and filtered to obtain a yellow pale solid 3- (DL-f/?reo-2-amino-1 -hydroxy-3-oxo-3-pyrrolidin-1 -yl-propyl)-1 /-/-pyhdin-2-one hydrochloride Compound 49 (136 mg, 19.0 % yield).
Figure imgf000106_0001
HCI (+/-) Compound 49
MW: 287.74; Yield: 19.0 %; Yellow Pale Solid; Mp (0C): 180.
1H NMR (CD3OD, δ): 1.82-2.09 (m, 4H, 2xCH2), 3.35-3.80 (m, 4 H, 2xCH2N),
4.63 (s, 1 H1 CH-N), 5.17 (s, 1 H, CH-O), 6.56 (t, 1 H, ArH) ), 7.5 (d, 1 H, J = 6.1
Hz, ArH), 7.86 (d, 1 H, J = 6.5 Hz, ArH). 13C-NMR (CD3OD, δ): 24.2, 26.0, 46.6, 46.6, 75.8, 79.7, 127.3, 127.5, 127.9,
129.4, 130.0, 132.3, 133.2, 148.1 , 148.4, 155.3, 166.2.
MS-ESI m/z (% rel. Int.): 252.1 ([MH]+, 18), 163.0 (100).
HPLC: Method A, detection UV 254nm, Compound 49 RT = 1.13 min, peak area 84.0 %.
Preparation of Compound 51 , Compound 52, Compound 53, Compound 54 Compound 55, Compound 56 and Compound 57.
General procedures: Method F:
To a suspension of DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-4-yl)-1- (pyrrolidin-1-yl)propan-1-one dihydrochloride Compound 22 (0.150 g, 0.44 mmol) in CH2CI2 (4 mL) was added TEA (0.185 mL, 1 .32 mmol) and the reaction mixture was stirred for 10 min and cooled in an ice bath with continuous stirring. The acyl chloride (0.484 mmol) was dissolved in CH2CI2 (1 ml.) and added dropwise to the reaction mixture. The reaction mixture was allowed to reach room temperature, stirred for 16 h and partitioned with H2O (3 x 4 ml_), washed with brine (3 x 4 ml_), NaOH (0.5 M, 3 x 4 ml.) and the organic layer was evaporated, adsorbed on silica gel (0.3 g) with EtOAc. The desired product was isolated by column chromatography using a gradient O to 8 % [v/v] MeOH in EtOAc. The solid obtained was dissolved in ethanol (1 mL) and a solution of HCI (0.8 M, 1 mL) in EtOH was added. Evaporation of the volatiles led to the corresponding hydrochloride salt.
Λ/-(DL-f/?reo-1 -Hvdroxy-3-oxo-1 -(pyhdin-4-yl)-3-(pyrrolidin-1 -yl)propan-2- vPhexanamide hydrochloride Compound 51 .
The compound was prepared according to method F with hexanoyl chloride (59 mg, 0.484 mmol) and DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-4- yl)-1-(pyrrolidin-1-yl)propan-1 -one dihydrochlohde Compound 22. /V-(DL- f/?reo-1 -Hydroxy-3-oxo-1 -(pyhdin-4-yl)-3-(pyrrolidin-1 -yl)propan-2- yl)hexanamide hydrochloride Compound 51 was obtained as an off white solid. (56 mg, 34 % yield).
Figure imgf000107_0001
Compound 51
MW: 369.89; Yield: 34 %; Off White Solid; Mp (0C): 182.0. 1H-NMR (CD3OD, δ): 0.84 (t, 3H, J = 6.7, CH3), 1 .10-1.32 (m, 4H, CH2), 1 .35- 1.50 (m, 2H, CH2), 1.80-2.00 (m, 4H, CH2), 2.05-2.30 (m, 2H, CH2), 3.35-3.45 (m, 2H, CH2), 3.50-3.65 (m, 2H, CH2), 5.09 (d, 1 H, J = 3.7 Hz, N-CH), 5.38 (d, 1 H, J = 3.7 Hz, 0-CH), 8.14 (d, 2H, J = 6.3 Hz, ArH), 8.80 (d, 2H, J = 6.3 Hz, ArH).
MS-ESI m/z (% rel. int.): 334.2 ([MH]+ , 10).
HPLC: Method A, detection UV 214 nm, Compound 51 RT = 3.90 min, peak area 99.0 %. Λ/-(DL-f/?reo-1 -Hvdroxy-3-oxo-1 -(pyridin-4-yl)-3-(pyrrolidin-1 -yl)propan-2- vDheptanamide hydrochloride Compound 52.
The compound was prepared according to method F with heptanoyl chloride (72 mg, 0.484 mmol) and DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-4- yl)-1-(pyrrolidin-1-yl)propan-1 -one dihydrochloride Compound 22. /V-(DL- f/?reo-1 -Hydroxy-3-oxo-1 -(pyhdin-4-yl)-3-(pyrrolidin-1 -yl)propan-2- yl)heptanamide hydrochloride Compound 52 was obtained as an off white solid. (192 mg, 66 % yield).
Figure imgf000108_0001
Compound 52
MW: 383.91 ; Yield: 66 %; Off White Solid; Mp (0C): 187.1.
1H-NMR (CD3OD, δ): 0.88 (t, 3H, J =3.7 Hz, CH3), 1.15-1.37 (m, 6H, CH2), 1.37 (m, 2H, CH2), 1.85-2.02 (m, 4H, CH2), 1.18-2.27 (m, 2H, CH2), 3.37-3.50
( m, 2H, N-CH2), 3.55-3.70 (m, 2H, NCH2), 5.14 (d, 1 H, N-CH), 5.42 (d, 1 H, O-
CH), 8.19 (d, 2H, J = 6.3 Hz, ArH), 8.83 (d, 2H, J = 6.3 Hz, ArH).
13C-NMR (CD3OD, δ): 14.4, 23.6, 25.0, 26.7, 27.0, 29.9, 32.6, 36.4, 47.5,
,56.7, 72.6, 126.6, 142.0, 164.5, 169.2, 175.9. MS-ESI m/z (% rel. Int.): 348.2 ([MH]+, 10).
HPLC: Method A, detection UV 254 nm, Compound 52 RT = 4.10 min, peak area 99.0 %.
Λ/-(DL-f/?reo-1 -Hydroxy-3-oxo-1 -(pyhdin-4-yl)-3-(pyrrolidin-1 -yl)propan-2- vDoctanamide hydrochloride Compound 53.
The compound was prepared according to method F with octanoyl chloride (78 mg, 0.484 mmol) and DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-4- yl)-1-(pyrrolidin-1-yl)propan-1 -one dihydrochloride Compound 22. /V-(DL- f/?reo-1 -Hydroxy-3-oxo-1 -(pyridin-4-yl)-3-(pyrrolidin-1 -yl)propan-2- yl)octanamide hydrochloride Compound 53 was obtained as an off white solid. (131 mg, 75 % yield).
Figure imgf000109_0001
Compound 53 MW: 397.94; Yield: 75 %; Off White Solid; Mp (0C): 185.9.
1H-NMR (CD3OD, δ): 0.91 (t, 3H, J = 6.4 Hz, CH3), 1.12-1 .37 (m, 8H, CH2), 1.40-1.52 (m, 2H, CH2), 1.81 (m, 4H, CH2), 2.12-2.25 (m, 2H, CH2), 3.40-3.52 (m, 2H, N-CH2), 3.55-3.65 (m, 2H, N-CH2), 5.14 (d, 1 H, J = 3.7 Hz, N-CH), 5.43 (d, 1 H, J = 3.7 Hz, OCH), 8.19 (d, 2H, J = 6.3 Hz, ArH), 8.84 (d, 2H, J = 6.3 Hz, ArH).
13C-NMR (CD3OD, δ): 14.4, 23.7, 24.9, 25.0, 25.5, 26.8, 27.0, 30.1 , 30.2, 32.8, 34.3, 36.5, 47.5, 56.8, 71.6, 72.6, 126.7, 126.9, 142.1 , 143.7, 164.5, 169.2, 176.0. MS-ESI m/z (% rel. Int.): 362.2 ([MH]+, 10). HPLC: Method A, detection UV 254 nm, Compound 53 RT = 4.37 min, peak area 99.9 %.
Λ/-(DL-f/?reo-1 -Hydroxy-3-oxo-1 -(pyhdin-4-yl)-3-(pyrrolidin-1 -yl)propan-2- vPpalmitamide hydrochloride Compound 54. The compound was prepared according to method F with palmitoyl chloride (133 mg, 0.484 mmol) and DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-4- yl)-1-(pyrrolidin-1-yl)propan-1 -one dihydrochlohde Compound 22. /V-(DL- f/?reo-1 -Hydroxy-3-oxo-1 -(pyhdin-4-yl)-3-(pyrrolidin-1 -yl)propan-2- yl)palmitamide hydrochloride Compound 54 was obtained as an off white solid. (105 mg, 47 % yield).
Figure imgf000110_0001
Compound 54
MW: 510.15; Yield: 47 %; White Solid; Mp (0C): 185.9. 1H-NMR (CD3OD, δ): 0.92 (t, 3H, CH3), 1.18 -1.42 (m, 24H, CH2), 1.42-1.58 (m, 2H, CH2), 1.85 (m, 4H, CH2), 2.15 (m, 2H, CH2), 3.41 -3.50 (m, 2H, CH2), 3.50-3.68 (m, 2H, CH2), 5.14 (d, 1 H, J = 3.5 Hz, N-CH), 5.42 (d, 1 H, J = 3.5 Hz, 0-CH), 8.18 (d, 2H, J = 6.0 Hz, ArH), 8.82 (d, 2H, J = 5.7 Hz, ArH). 13C-NMR (CD3OD, δ): 14.4, 23.7, 25.0, 26.8, 27.0, 30.3, 30.4, 30.5, 30.6, 30.7, 30.8, 33.1 , 36.4, 47.5, 56.8, 72.6, 126.6, 142.1 , 164.5, 169.2, 175.9. MS-ESI m/z (% rel. Int.): 474.2 ([MH]+, 40).
HPLC: Method A, detection UV 254 nm, Compound 54 RT = 6.36 min, peak area 97.0%.
Λ/-(DL-f/?reo-1 -Hydroxy-3-oxo-1 -(pyhdin-4-yl)-3-(pyrrolidin-1 -yl)propan-2- vPbenzamide hydrochloride Compound 55.
The compound was prepared according to method F with benzoyl chloride (141 mg, 0.484 mmol) and DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-4- yl)-1-(pyrrolidin-1-yl)propan-1 -one dihydrochlohde Compound 22. /V-(DL- f/?reo-1 -Hydroxy-3-oxo-1 -(pyhdin-4-yl)-3-(pyrrolidin-1 -yl)propan-2- yl)benzamide hydrochloride Compound 55 was obtained as an off white solid. (67 mg, 34 % yield).
Figure imgf000110_0002
Compound 55 MW: 375.85; Yield: 34 %; Off White Solid; Mp (0C): 212. 1H-NMR (CD3OD, δ): 1.69-1.91 (m, 4H, CH2), 3.25-3.40 (m, 2H, N-CH2), 3.40- 3.58 (m, 2H, N-CH2), 5.22 (d, 1 H, J = 3.7 Hz, N-CH), 5.43 (d, 1 H, J = 3.5 Hz, 0-CH), 7.32 (t, 2H, J = 7.8 Hz, ArH), 7.40 (t, 1 H, J = 6.9 Hz, ArH), 7.63 (d, 2H, J = 7.1 Hz, ArH), 8.08 (d, 2H, J = 6.6 Hz, ArH), 8.66 (d, 2H, J = 6.1 Hz, ArH). 13C-NMR (CD3OD, δ): 25.0, 27.1 , 47.6, 57.6, 72.7, 126.6, 128.4, 129.7, 133.3, 134.4, 142.1 , 164.5, 169.0, 169.7. MS-ESI m/z (% rel. Int.): 340.2 ([MH]+, 5).
HPLC: Method A, detection UV 254 nm, Compound 55 RT = 3.66 min, peak area 99.0 %.
Λ/-(DL-f/?reo-1 -Hvdroxy-3-oxo-1 -(pyhdin-4-yl)-3-(pyrrolidin-1 -yl)propan-2-yl)-4- methoxy-benzamide hydrochloride Compound 56.
The compound was prepared according to method F with 4-methoxybenzoyl chloride (82 mg, 0.484 mmol) and DL-f/?reo-2-amino-3-hydroxy-3-(pyridin-4- yl)-1-(pyrrolidin-1-yl)propan-1 -one dihydrochlohde Compound 22. /V-(DL- f/?reo-1 -Hydroxy-3-oxo-1 -(pyridin-4-yl)-3-(pyrrolidin-1 -yl)propan-2-yl)-4- methoxy-benzamide hydrochloride Compound 56 was obtained as an off white solid. (105 mg, 58 % yield).
Figure imgf000111_0001
Compound 56
MW: 405.87; Yield: 58 %; Off White Solid; Mp(0C): 205.3 (dec).
1H-NMR (CD3OD, δ): 1.82-2.08 (m, 4H, CH2), 3.45-3.55 (m, 2H, CH2-N), 3.60-
3.70 (m, 2H, NCH2), 3.86 (s, 3H, 0-CH3), 5.35 (d, 1 H, J = 3.7 Hz, N-CH), 5.56
(d, 1 H, J = 3.6 Hz, 0-CH), 6.99 (dd, 2H, J = 6.9 Hz, J = 1 .9 Hz), 7.76 (dd, 2H, J = 6.9 Hz, J = 1.9 Hz, ArH), 8.21 (d, 2H, J = 6.6 Hz, ArH), 8.79 (d, 2H, J = 6.6
Hz, ArH).
13C-NMR (CD3OD, δ): 25.0, 27.1 , 47.6, 56.0, 57.5, 72.7, 1 14.9, 1 15.2, 126.3,
126.6, 130.4, 133.7, 142.1 , 164.4, 164.5, 169.1. MS-ESI m/z (% rel. Int.): 370.2 ([MH]+, 10).
HPLC: Method A, detection UV 254 nm, Compound 56 RT = 3.76 min, peak area 99 %.
3,4-Dichloro-Λ/-(DL-f/?reo-1 -Hydroxy-3-oxo-1 -(pyhdin-4-yl)-3-(pyrrolidin-1 - yl)propan-2-yl)benzamide Compound 57.
The compound was prepared according to method F with 3,4- dichlorobenzoyl chloride (101 mg, 0.484 mmol) and DL-f/?reo-2-amino-3- hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochlohde Compound 22. 3,4-Dichloro-Λ/-(DL-f/?reo-1 -hydroxy-3-oxo-1-(pyhdin-4-yl)-3-
(pyrrolidin-1-yl)propan-2-yl)benzamide Compound 57 was obtained as an off white solid. (92 mg, 55 % yield).
Figure imgf000112_0001
Compound 57 MW: 444.74; Yield: 55 %; Off White Solid; Mp (0C): 319.5 (dec).
1H-NMR (CD3OD, δ): 1.82-2.05 (m, 4H, CH2), 3.40-3.70 (m, 4H, N-CH2), 5.33
(d, 1 H, J = 3.9 Hz, N-CH), 5.55 (d, 1 H, J = 4.0 Hz, 0-CH), 7.61-7.75 (m, 2H,
ArH), 7.96 (d, 1 H, J = 1 .5 Hz, ArH), 8.22 (d, 2H, J = 6.4 Hz, ArH), 8.81 (d, 2H,
J = 6.0 Hz, ArH). 13C-NMR (CD3OD, δ): 25.0, 27.0, 57.8, 72.6, 126.6, 128.3, 130.7, 131.9,
133.8, 134.7, 137.2, 142.2, 164.3, 167.2, 168.8.
MS-ESI m/z (% rel. Int.): 408.0, ([MH]+, 10)
HPLC: Method A, detection UV 254 nm, Compound 57 RT = 4.28 min, peak area 99.9 %.
Preparation of Compound 58, Compound 59, Compound 60, Compound 61 , Compound 62, Compound 63, Compound 64, Compound 65, Compound 66, Compound 67, Compound 68, Compound 69. General procedures:
Method G (in CH2CI2): To a stirred solution of DL-f/?reo-2-amino-3-hydroxy-3-(pyhdin-4-yl)-1-
(pyrrolidin-1-yl)propan-1-one dihydrochlohde Compound 22 (0.15 g, 0.49 mmol) in 10 mL of CH2CI2 at +4 0C were added thethylamine (200 μl, 1 .45 mmol) and very slowly acid chloride in 3 mL of CH2CI2. The mixture was stirred overnight at RT under nitrogen and then partitioned between CH2CI2 and 1 N aqueous sodium carbonate. The organic layer was evaporated and the obtained residue purified by column chromatography on silica (EtOAc:MeOH = 95:5). The hydrochloride salt was obtained in MeOH at 0 0C with 0.3 M HCI in diethylether to give after evaporation of solvents and drying the acylated compound.
Method H (in MeOH):
To a stirred solution of DL-f/?reo-2-amino-3-hydroxy-3-(pyhdin-4-yl)-1- (pyrrolidin-1-yl)propan-1-one dihydrochlohde Compound 22 (0.20 g, 0.65 mmol) in 3 mL of MeOH were added triethylamine (180 μl, 1.30 mmol) and aldehyde (or ketone). The mixture was stirred overnight at RT under nitrogen and then was added AcOH (200 μl_, 3.2 mmol) and NaBH3CN. After 5 h at 20 0C, MeOH was evaporated and the residue was partitioned between CH2CI2 and 1 N aqueous sodium carbonate. The organic layer was evaporated and the obtained residue was purified by column chromatography on silica (EtOAc:MeOH or CH2CI2:Me0H). The hydrochloride salt was obtained in MeOH at 0 0C with 0.3 M HCI in diethylether to give after evaporation of solvents and drying the alkylated compound.
Benzyl DL-f/?reo-3-hydroxy-1 -oxo-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-2- ylcarbamate hydrochloride Compound 58. The compound was prepared according to method G with benzyl chloroformate (91 mg, 0.53 mmol). After work-up benzyl DL-f/?reo-3-hydroxy- 1 -oxo-3-(pyrid i n-4-yl )- 1 -(pyrrolidin-1 -yl)propan-2-ylcarbamate hydrochloride Compound 58 was obtained as a white solid (90 mg, 46 % yield).
Figure imgf000114_0001
Compound 58
MW: 405.9; Yield: 46.0 %; White Solid; Mp (0C): 185.3. Rf. 0.38 (MeOH:EtOAc = 10:90) free base.
1H-NMR (CD3OD, δ): 1.87-2.03 (m, 4H, 2xCH2), 3.40-3.48 (m, 2H, CH2N), 3.56-3.62 (m, 2H, CH2N), 4.85-5.04 (m, 3H, CH2O, CHO), 5.39 (d, 1 H, J = 2.8 Hz, NH), 7.26-7.36 (m, 5H, ArH), 8.12 (d, 2H, J = 6.0 Hz, ArH), 8.69 (d, 2H, J = 6.0 Hz, ArH).
13C-NMR (CD3OD, δ): 25.0, 27.0, 47.5, 48.0, 58.8, 67.9, 72.7, 126.6 (2xC), 129.1 , 129.2, 129.5, 138.1 , 141 .9 (2xC), 158.1 , 164.4, 169.2. MS-ESI m/z (% rel. Int.): 370.1 ([MH]+, 15), 219.0 (100). HPLC: Method A, detection UV 254 nm, Compound 58 RT = 4.10 min, peak area 99.8 %.
Λ/-(DL-f/?reo-3-hvdroxy-1 -oxo-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-2- vDdecanamide hydrochloride Compound 59.
The compound was prepared according to method G with decanoyl chloride (1 1 1 μl_, 0.53 mmol). After work-up Λ/-(DL-f/?reo-3-hydroxy-1 -oxo-3- (pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-2-yl)decanamide hydrochloride Compound 59 was obtained as a white solid (1 15 mg, 55 % yield).
Figure imgf000114_0002
Compound 59 MW: 425.99; Yield: 55 %; White Solid; Mp (0C): 184.8. Rf. 0.22 (MeOH:EtOAc = 5:95) free base.
1H-NMR (CD3OD, δ): 0.90 (t, 3H, J = 7.0 Hz, CH3), 1.26-1.34 (m, 12H, 6xCH2), 1.42-1.50 (m, 2H, CH2), 1.86-1.98 (m, 4H, 2xCH2), 2.13-2.20 (m, 2H, CH2CO), 3.41-3.46 (m, 2H, CH2N), 3.52-3.61 (m, 2H, CH2N), 5.12 (d, 1 H, J = 3.8 Hz, CH), 5.40 (d, 1 H, J = 3.7 Hz, CH), 8.16 (d, 2H, J = 6.5 Hz, ArH), 8.97 (d, 2H, J = 6.7 Hz, ArH).
13C-NMR (CD3OD, δ): 14.4, 23.7, 25.0, 26.8, 27.0, 30.3, 30.4, 30.6, 33.0, 36.5, 47.5, 56.8, 72.6, 126.6 (2xC), 142.1 (2xC), 164.4, 169.2, 175.9. MS-ESI m/z (% rel. Int.): 390.1 ([MH]+, 20), 219.1 (100). HPLC: Method A, detection UV 254 nm, Compound 59 RT = 4.9 min, peak area 99.5 %.
DL-f/?reo-2-(Benzylamino)-3-hvdroxy-3-(pyridin-4-yl)-1-(pyrrolidin-1 -yl)propan- 1-one dihydrochloride Compound 60. The compound was prepared according to method H with benzaldehyde (78 mg, 0.72 mmol). After column chromatography (EtOAc:MeOH = 95:5) and HCI treatment DL-f/?reo-2-(benzylamino)-3- hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochloride Compound 60 was obtained as a white solid (1 14 mg, 46 % yield).
Figure imgf000115_0001
Compound 60
MW: 398.33; Yield: 46 %; White Solid; Mp (0C): 131 .5. Rf. 0.60 (MeOH:EtOAc = 10:90) free base.
1H-NMR (CD3OD, δ): 1.35-1.72 (m, 4H, 2xCH2), 2.10-2.18 (m, 1 H1 CH2N), 2.78-2.86 (m, 1 H, CH2N), 3.18-3.24 (m, 2H, CH2N), 4.22 (d, 1 H, J = 8.5 Hz, CH), 4.26-4.36 (m, 2H, CH2N), 5.18 (d, 1 H, J = 8.5 Hz, CH), 7.43-7.51 (m, 5H, BzH), 7.86 (d, 2H, J = 6.6 Hz, ArH), 8.69 (d, 2H, J = 6.6 Hz, ArH). 13C-NMR (CD3OD, δ): 24.7, 26.3, 47.3, 47.9, 51 .4, 63.9, 72.4, 126.2, 130.3, 131.0, 131.4, 131.5, 148.9, 156.1 , 163.9. MS-ESI m/z (% rel. Int.): 326.1 ([MH]+, 100), 227.0 (80). HPLC: Method A, detection UV 254 nm, Compound 60 RT = 4.30 min, peak area 98.2 %.
DL-f/?reo-3-Hvdroxy-2-(niethylamino)-3-(pyridin-4-yl)-1 -(pyrrolidin-1-yl)propan- 1-one dihydrochloride Compound 61.
The compound was prepared according to method H with paraformaldehyde (21 mg, 0.65 mmol). After colummn chromatography
(EtOAc: MeOH = 7:3) and HCI treatment DL-f/?reo-3-hydroxy-2-(methylamino)- 3-(pyhdin-4-yl)-1-(pyrrolidin-1-yl)propan-1-one dihydrochloride Compound 61 was obtained as a pale yellow solid (28 mg, 13 % yield).
Figure imgf000116_0001
Compound 61
MW: 322.23; Yield: 13 %; Pale Yellow Solid.
Rf : 0.20 (MeOH:EtOAc = 30:70) free base.
1H-NMR (CD3OD, δ): 1.60-1.80 (m, 4H, 2xCH2), 2.61 (s, 1 H, CH3), 2.68-2.76
(m, 1 H, CH2N), 3.24-3.57 (m, 3H, CH2N), 4.53 (d, 1 H, J = 6.8 Hz, CH), 5.26 (d, 1 H, J = 7.0 Hz, CH), 8.1 1 (d, 2H, J = 5.8 Hz, ArH), 8.85 (d, 2H, J = 5.6 Hz,
ArH).
13C-NMR (CD3OD, δ): 24.8, 26.7, 32.8, 47.7, 48.3, 65.4, 71.7, 126.7 (2xC),
143.4 (2xC), 161.1 , 163.9.
MS-ESI m/z (% rel. Int.): 251.1 ([MH]+, 10), 151 .0 (100). HPLC: Method A, detection UV 254 nm, Compound 61 RT = 0.70 min, peak area 97.5 %.
DL-f/?reo-3-Hvdroxy-2-(pentylamino)-3-(pyridin-4-yl)-1-(pyrrolidin-1-yl)propan- 1-one dihydrochloride Compound 62. The compound was prepared according to method H with valeraldehyde (60 mg, 0.68 mmol). After column chromatography (EtOAc: MeOH = 95:5) and HCI treatment DL-f/?reo-3-hydroxy-2- (pentylamino)-3-(pyridin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochlohde Compound 62 was obtained as a white solid (107 mg, 44 % yield).
Figure imgf000117_0001
Compound 62
MW: 378.34; Yield: 44 %; White Solid; Mp (0C): 86.6.
Rf. 0.30 (MeOH:EtOAc = 20:80) free base. 1H-NMR (CD3OD, δ): 0.95 (t, 3H, J = 6.4 Hz, CH3), 1.32-1.40 (m, 6H, 3xCH2),
1.70-1.87 (m, 4H, 2xCH2), 2.70-2.75 (m, 1 H, CH2N), 2.90-3.00 (m, 1 H, CH2N),
3.10-3.39 (m, 3H1 CH2N), 3.46-3.60 (m, 1 H, CH2N), 4.61 (d, 1 H, J = 7.5 Hz,
CH), 5.39 (d, 1 H, J = 7.5 Hz, CH), 8.22 (d, 2H, J = 6.2 Hz, ArH), 8.97 (d, 2H, J
= 6.2 Hz, ArH). 13C-NMR (CD3OD, δ): 9.3, 14.1 , 23.2, 24.8, 26.7, 26.8, 29.7, 47.6, 47.9, 64.5,
72.1 , 126.8 (2xC), 143.3 (2xC), 161 .1 , 164.1.
MS-ESI m/z (% rel. Int.): 306.3 ([MH]+, 15), 207.1 (100).
HPLC: Method A, detection UV 254 nm, Compound 62 RT = 3.60 min, peak area 98.5 %.
DL-f/?reo-3-Hvdroxy-2-(hexylamino)-3-(pyridin-4-yl)-1-(pyrrolidin-1-yl)propan- 1-one dihydrochloride Compound 63.
The compound was prepared according to method H with hexanal (71 mg, 0.68 mmol). After column chromatography (EtOAc:MeOH = 95:5) and HCI treatment DL-f/?reo-3-hydroxy-2-(hexylamino)-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 - yl)propan-1-one dihydrochloride Compound 63 was obtained as a beige solid (1 12 mg, 45 % yield).
Figure imgf000118_0001
Compound 63
MW: 392.36; Yield: 45 %; Beige Solid; Mp (0C): 108.2.
Rf. 0.35 (MeOH:EtOAc = 20:80) free base. 1H-NMR (CD3OD, δ): 0.94 (t, 3H, J = 7.2 Hz, CH3), 1.30-1.42 (m, 6H, 3xCH2),
1.66-1.90 (m, 6H, 3xCH2), 2.68-2.74 (m, 1 H, CH2N), 2.90-2.99 (m, 1 H, CH2N),
3.09-3.16 (m, 1 H, CH2N), 3.32-3.39 (m, 1 H, CH2N), 3.47-3.60 (m, 2H, CH2N),
4.60 (d, 1 H, J = 7.7 Hz, CH), 5.38 (d, 1 H, J = 7.7 Hz, CH), 8.24 (s, 2H, ArH),
8.97 (s, 2H, ArH). 13C-NMR (CD3OD, δ): 14.3, 23.4, 24.8, 26.7, 27.1 , 27.2, 32.4, 47.6, 64.4,
72.1 , 126.9 (2xC), 143.2 (2xC), 161 .3, 164.1.
MS-ESI m/z (% rel. Int.): 320.1 ([MH]+, 30), 221 .1 (100).
HPLC: Method A, detection UV 254 nm, Compound 63 RT = 3.80 min, peak area 97.8 %.
DL-f/?reo-3-Hvdroxy-2-(heptylamino)-3-(pyridin-4-yl)-1-(pyrrolidin-1-yl)propan- 1-one dihydrochloride Compound 64.
The compound was prepared according to method H with heptaldehyde (82 mg, 0.68 mmol). After column chromatography with
EtOAc: MeOH = 95:5 and HCI treatment DL-f/?reo-3-hydroxy-2-(heptylamino)- 3-(pyhdin-4-yl)-1-(pyrrolidin-1-yl)propan-1-one dihydrochloride Compound 64 was obtained as a white solid (121 mg, 47 % yield).
Figure imgf000118_0002
Compound 64 MW: 406.39; Yield: 47 %; White Solid; Mp (0C): 242.4.
Rf : 0.40 (MeOH:EtOAc = 20:80) free base.
1H-NMR (CD3OD, δ): 0.92 (t, 3H, J = 7.0 Hz, CH3), 1.32-1.40 (m, 8H, 4xCH2),
1.65-1.90 (m, 6H, 3xCH2), 2.71-2.76 (m, 1 H, CH2N), 2.90-2.99 (m, 1 H, CH2N), 3.09-3.38 (m, 1 H, CH2N), 3.47-3.62 (m, 2H, CH2N), 4.61 (d, 1 H, J = 7.5 Hz,
CH), 5.39 (d, 1 H, J = 7.5 Hz, CH), 8.24 (d, 2H, J = 6.0 Hz, ArH), 8.97 (d, 2H, J
= 5.9 Hz, ArH).
13C-NMR (CD3OD, δ): 14.4, 23.6, 24.8, 26.7, 27.1 , 27.5, 29.9, 32.7, 47.7,
64.4, 72.1 , 126.9 (2xC), 143.2 (2xC), 161.4, 164.1. MS-ESI m/z (% rel. Int.): 334.1 ([MH]+, 45), 235.1 (100).
HPLC: Method A, detection UV 254 nm, Compound 64 RT = 4.00 min, peak area 97.5 %.
DL-f/?reo-2-(4-Methylbenzylamino)-3-hvdroxy-3-(pyhdin-4-yl)-1-(pyrrolidin-1- yl)propan-1-one Compound 65.
The compound was prepared according to method H with 4- methylbenzaldehyde (86 mg, 0.70 mmol). After column chromatography (EtOAc:MeOH = 95:5) and HCI treatment DL-f/?reo-2-(4-methylbenzylamino)- 3-hydroxy-3-(pyhdin-4-yl)-1-(pyrrolidin-1-yl)propan-1-one Compound 65 was obtained as a white solid (137 mg, 51 % yield).
Figure imgf000119_0001
Compound 65
MW: 412.35; Yield: 51 %; White Solid; Mp (0C): 87.5. Rf : 0.20 (MeOH:EtOAc = 5:95) free base. 1H-NMR (CD3OD, δ): 1.49-2.12 (m, 4H, 2xCH2), 2.30-2.40 (m, 1 H, CH2N), 2.35. (s, 3H, CH3), 2.75-2.95 (m, 1 H, CH2N), 3.18-3.25 (m, 2H, CH2N), 4.12- 4.32 (m, 3H, CH2N, CH), 5.30 (d, 1 H, J = 7.9 Hz, CH), 7.24-7.39 (m, 4H, BzH), 8.1 1 (d, 2H, J = 6.7 Hz, ArH), 8.89 (d, 2H, J = 6.6 Hz, ArH). 13C-NMR (CD3OD, δ): 21 .2, 24.7, 26.3, 26.4, 47.4, 47.9, 51.2, 63.4, 72.4, 126.1 , 126.6 (2xC), 128.2, 130.9, 131.4, 131 .5, 141 .4, 143.5 (2xC), 148.9, 156.1 ; 160.5, 163.8.
MS-ESI m/z (% rel. Int.): 340.1 ([MH]+, 10), 104.9 (100).
HPLC: Method A, detection UV 254 nm, Compound 65 RT = 3.70 min, peak area 97.3 %.
DL-f/?reo-2-(4-Chlorobenzylamino)-3-hvdroxy-3-(pyhdin-4-yl)-1-(pyrrolidin-1 - yl)propan-1-one Compound 66.
The compound was prepared according to method H with 4- chlorobenzaldehyde (98 mg, 0.70 mmol). After column chromatography
(EtOAc:MeOH = 95:5) and HCI treatment DL-f/?reo-2-(4-chlorobenzylamino)- 3-hydroxy-3-(pyhdin-4-yl)-1-(pyrrolidin-1-yl)propan-1-one Compound 66 was obtained as a white solid (126 mg, 45 % yield).
Figure imgf000120_0001
Compound 66
MW: 432.77; Yield: 45 %; White Solid; Mp (0C): 122.7. Rf : 0.20 (MeOH: EtOAc = 5:95) free base.
1H-NMR (CD3OD, δ): 1 .54-1.75 (m, 4H, 2xCH2), 2.41-2.49 (m, 1 H, CH2N),
3.08-3.15 (m, 1 H, CH2N), 3.19-3.27 (m, 2H, CH2N), 4.25-4.47 (m, 3H, CH2N,
CH), 5.34 (d, 1 H, J = 7.9 Hz, CH), 7.46-7.55 (m, 4H, BzH), 8.15 (d, 2H, J =
6.0 Hz, ArH), 8.92 (d, 2H, J = 5.7 Hz, ArH). 13C-NMR (CD3OD, δ): 24.7, 26.5, 47.5, 47.8, 50.8, 63.8, 72.3, 126.7 (2xC),
130.3, 133.4, 137.1 , 143.5 (2xC), 149.0, 160.6, 163.8.
MS-ESI m/z (% rel. Int.): 360.1/362.1 ([MH]+, 20), 124.9 (100). HPLC: Method A, detection UV 254 nm, Compound 66 RT = 3.70 min, peak area 97.0 %.
DL-f/?reo-2-(4-Methoxybenzylamino)-3-hvdroxy-3-(pyridin-4-yl)-1-(pyrrolidin-1 - yl)propan-1-one one Compound 67.
The compound was prepared according to method H with 4- methoxybenzaldehyde (95 mg, 0.70 mmol). After column chromatography
(EtOAc:MeOH = 95:5) and HCI treatment DL-f/?reo-2-(4- methoxybenzylamino)-3-hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 - one one Compound 67 was obtained as a white solid (123 mg, 44 % yield).
Figure imgf000121_0001
Compound 67
MW: 428.35; Yield: 44 %; White Solid; Mp (0C): 193.2. Rf. 0.20 (MeOH:EtOAc = 5:95) free base.
1H-NMR (CD3OD, δ): 1.51-1.72 (m, 4H, 2xCH2), 2.38-2.41 (m, 1 H, CH2N), 2.94-3.01 (m, 1 H, CH2N), 3.18-3.28 (m, 2H, CH2N), 3.81 (s, 3H, CH3O), 4.18- 4.34 (m, 3H, CH2N, CH), 5.31 (d, 1 H, J = 7.3 Hz, CH), 6.97 (d, 2H, J = 8.5 Hz, ArH), 7.42 (d, 2H, J = 8.5 Hz, ArH), 8.13 (d, 2H, J = 6.5 Hz, ArH), 8.91 (d, 2H, J = 6.3 Hz, ArH).
13C-NMR (CD3OD, δ): 24.7, 26.5, 47.5, 47.9, 51 .1 , 55.9, 63.2, 72.3, 1 15.5, 122.9, 126.8 (2xC), 133.1 , 143.3 (2xC), 160.9, 162.4, 163.8. MS-ESI m/z (% rel. Int.): 356.1 ([MH]+, 10), 120.9 (100). HPLC: Method A, detection UV 254 nm, Compound 67 RT = 3.50 min, peak area 98.6 %. DL-f/7reo-2-(3,4-Dichlorobenzylamino)-3-hvdroxy-3-(pyridin-4-yl)-1-(pyrrolidin- 1-yl)propan-1-one dihvdrochloride Compound 68.
The compound was prepared according to method H with 3,4- dichlorobenzaldehyde (122 mg, 0.70 mmol). After column chromatography (EtOAc:MeOH = 95:5) and HCI treatment DL-f/?reo-2-(3,4- dichlorobenzylamino)-3-hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 - one dihydrochloride Compound 68 was obtained as a white solid (153 mg, 50 % yield).
Figure imgf000122_0001
Compound 68
MW: 467.22; Yield: 50 %; White Solid; Mp (0C): 190.3.
Rf. 0.20 (MeOH:EtOAc = 5:95) free base. 1H-NMR (CD3OD, δ): 1.58-1.76 (m, 4H, 2xCH2), 2.48-2.55 (m, 1 H, CH2N),
2.85-3.00 (m, 1 H, CH2N), 3.18-3.26 (m, 2H, CH2N), 4.23-4.41 (m, 2H, CH2N),
4.54 (d, 1 H, J = 7.7 Hz, CH), 5.34 (d, 1 H, J = 7.2 Hz, CH), 7.48 (d, 1 H, J = 8.3
Hz, ArH), 7.63 (dd, 1 H, J = 8.2 Hz, J =1.4 Hz, ArH), 7.75 (s, 1 H, ArH), 8.16 (d,
2H, J = 5.4 Hz, ArH), 8.92 (d, 2H, J = 5.3 Hz, ArH). 13C-NMR (CD3OD, δ): 23.2, 25.0, 46.0, 46.4, 48.9, 62.5, 70.7, 125.3, 130.2,
130.6, 130.7, 132.3, 133.6, 141.9, 159.2, 162.3.
MS-ESI m/z (% rel. Int.): 394.1/396.1 ([MH]+, 40), 1 10.0 (100).
HPLC: Method A, detection UV 254 nm, Compound 68 RT = 3.90 min, peak area 99.0 %.
DL-f/?reo-2-(4-Methoxybenzylamino)-3-hvdroxy-3-(pyridin-4-yl)-1-(pyrrolidin-1 - yl)propan-1-one Compound 69. The compound was prepared according to method H with cyclohexanone (75 μl_, 0.70 mmol). After column chromatography (EtOAc: MeOH = 95:5) and HCI treatment DL-f/?reo-2-(4- methoxybenzylamino)-3-hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 - one Compound 69 as a white solid (154 mg, 61 % yield).
Figure imgf000123_0001
Compound 69
MW: 390.35; Yield: 61 %; White Solid; Mp (0C): 144.1.
Rf. 0.25 (MeOH: EtOAc = 5:95) free base. 1H-NMR (CD3OD, δ): 1.16-2.15 (m, 15H, 7xCH2, CH), 2.65-2.72 (m, 1 H,
CH2N), 3.07-3.15 (m, 1 H, CH2N), 3.43-3.65 (m, 2H, CH2N), 4.61 (d, 1 H, J =
7.8 Hz, CH-N), 5.35 (d, 1 H, J = 7.8 Hz, CH-O), 8.21 (d, 2H, J = 6.3 Hz, ArH),
8.95 (d, 2H, J = 6.1 Hz1 ArH).
13C-NMR (CD3OD, δ): 23.2, 24.0, 24.2, 24.4, 25.2, 28.2, 29.3, 46.1 , 57.4, 60.5, 71.0, 125.4 (2xC), 141.8 (2xC), 159.5, 162.6.
MS-ESI m/z (% rel. Int.): 318.2 ([MH]+, 40; 219.1 , 100).
HPLC: Method A, detection UV 254 nm, Compound 69 RT = 3.40 min, peak area 99.7 %.
Preparation of (±)-f/?reo-2-amino-3-(furan-2-yl)-3-hvdroxy-1 -(pyrrolidin-1 - yl)propan-1 -one hydrochloride Compound 201.
frans-5-(Furan-2-yl)-4,5-dihvdrooxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 04136D. BLE 04136D was prepared in accordance with method D using 2- furaldehyde (0.449 mL, 5.42 mmol), KOH (0.276 mg, 4.92 mmol) in methanol (5 mL) and 2-isocyano-1 -(pyrrolidin-1 -yl)ethanone BLE 04098 (0.75 g, 5.42 mmol). After work-up the residue was purified by column chromatography (SiO2, cyclohexane: EtOAc = 100:0 to 0:100) to led, after evaporation, to trans- 5-(furan-2-yl)-4,5-dihydrooxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 04138D (0.742 g, 58.5 % yield) as a pale yellow oil.
Figure imgf000124_0001
(±)-trans
BLE 04136D
MW: 234.25; Yield: 58.5 %; Pale Yellow Oil.
1H-NMR (CDCI3,): 1.80-2.10 (m, 4H, CH2), 3.47-3.60 (m, 3H, CH2N), 3.93- 4.03 (m, 1 H, CH2N), 4.94 (dd, 1 H, J = 7.4 Hz, J = 2.2 Hz, CH-N), 6.14 (d, 1 H, J = 7.4 Hz, CH-O), 6.37 (dd, 1 H, J = 3.3 Hz, J = 1 .8 Hz, CH=C), 6.47 (d, 1 H, J = 3.3 Hz, CH=C), 6.92 (d, 1 H, J = 2.2 Hz, 0-CH=N), 7.44 (t, 1 H, J = 1.6 Hz, OCH=C).
(±)-f/?reo-2-Amino-3-(furan-2-yl)-3-hvdroxy-1 -(pyrrolidin-1 -yl)propan-1 -one hydrochloride Compound 201.
Compound 201 was prepared following method E with frans-(5-(furan- 2-yl)-4,5-dihydrooxazol-4-yl)(pyrrolidin-1-yl)methanone BLE 04136D (0.30 g, 1.28 mmol), hydrochloric acid 37 % (0.3 mL) and methanol (10 ml_). After overnight at RT and work-up (±)-f/?reo-2-amino-3-(furan-2-yl)-3-hydroxy-1 - (pyrrolidin-1-yl)propan-1-one hydrochloride (0.22 g, 66 % yield) was obtained as a pale brown solid.
Figure imgf000124_0002
Compound 201
MW: 260.72; Yield: 66 %; Pale Brown Solid; Mp (0C): 159.8 1H-NMR (CD3OD1): 1.62-1.95 (m, 4H, 2xCH2), 2.72-2.85 (m, 1 H, CH2N), 3.22-3.35 (m, 1 H, CH2N), 3.38-3.55 (m, 2H, CH2N), 4.35 (d, 1 H, J = 8.5 Hz, CH-N), 4.91 (d, 1 H, J = 8.5 Hz, CH-O), 6.45 (m, 2H, ArH), 7.52-7.57 (m, 1 H, ArH).
13C-NMR (CD3OD1): 24.9, 26.9, 47.4, 47.6, 57.4, 67.8, 109.9, 1 1 1.9, 144.4, 153.0, 166.0. MS-ESI m/z (% rel. Int.): 225.1 ([MH]+, 18), 207.1 (100).
HPLC: Method A, detection UV 254 nm, Compound 201 RT = 2.87 min, peak area 92.0 %.
Preparation of (-H2f?,3S)-2-amino-3-hvdroxy-3-(pyridin-4-yl)-1 -(pyrrolidin-1 - yl)propan-1-one dihvdrochloride Compound 203 and (+)-(2S,3f?)-2-amino-3- hvdroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihvdrochlohde Compound 204.
Extraction of the free base of Compound 22: (±)-f/?reo-2-Amino-3-hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 - one dihydrochloride Compound 22 (350 mg, 1 .14 mmol) was dissolved in 20 ml. of a K2CO3 (10 %) solution and the aqueous mixture was then saturated with NaCI. The aqueous phase was extracted by a mixture CH2CI2:2-Pr0H = 9:1 (6x15 ml_). The organic phase was dried over MgSO4 and evaporated to afford 226 mg (85 % yield) of the free base of Compound 22. Analytical chiral separation:
20 μl_ of a 1 mg/mL solution of Compound 22 were injected on Chiralpak AD: flow-rate = 1 mL/min, temperature = 25 0C, mobile phase: hexane:ethanol = 1 :1 , detection by UV at 220 nm and by polarimeter, Rt (-) = 8.20 min, Rt (+) = 10.61 min, k (-) = 1 .72, k (+) = 2.51 , α = 1.47 and resolution Rs = 3.08.
Semi-preparative chiral separation:
A solution of 100 mg/mL was prepared and 10 μL of this solution were injected every 4.5 min on Chiralpak AD, flow-rate = 1 mL/min, mobile phase hexane:ethanol = 4:6, detection by UV at 254 nm. 135 successive injections were done. The two main fractions were identified by UV and collected in two different flasks. The solvent was removed in vacuo at 30 0C. The resulting solid was dissolved in 50 mL of CH2CI2 and then filtered on a 0.45 Dm millipore membrane. After evaporation Of CH2CI2, the solid was dissolved in 50 mL of methanol and then filtered. The salts were regenerated according to the procedure reported above. Regeneration of the salt:
After the chiral separation, about 63 mg of each enantiomer of the free base were dissolved in 100 mL of ethanol and 6.7 mL of HCI (0.2 N, 5 eq) were added. The solvent was evaporated, then 50 mL of ethanol were added and then removed in vacuo and the products were dried over P2Os under vacuum overnight. The enantiomeric purity of the products was checked by analytical injection of the regenerated salts:
(-H2f?,3S)-2-Amino-3-hvdroxy-3-(pyridin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihvdrochloride Compound 203.
Figure imgf000126_0001
HCI HCI (-ythreo
Compound 203
MW: 308.20; 83 mg obtained; Yield: 23.5 %; White Solid; Mp (0C): 183.5 Enantiomeric excess = 99.3 % G 25 D = - 22.7 (MeOH, c = 0.51 ).
(+H2S,3f?)-2-Amino-3-hvdroxy-3-(pyridin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihvdrochlohde Compound 204.
Figure imgf000126_0002
(+)-threo
Compound 204 MW: 308.20; 82 mg obtained; Yield: 23.5 %; White Solid; Mp (0C): 176.9 Enantiomeric excess = 98.5 % G 25D = + 23.1 (MeOH, c = 1 ).
Preparation of (+)-(2S,3f?)-2-amino-3-hvdroxy-1 -(pyrrolidin-1 -yl)-3-(thiophen- 3-yl)propan-1-one hydrochloride Compound 205 and (-)-(2f?,3S)-2-amino-3- hydroxy-1 -(pyrrolidin-1 -yl)-3-(thiophen-3-yl)propan-1 -one hydrochloride Compound 206.
Extraction of the free base:
(±)-f/?reo-2-amino-3-hydroxy-1 -(pyrrolidin-1 -yl)-3-(thiophen-3-yl)propan- 1-one hydrochloride Compound 23 (243 mg, 0.88 mmol) was dissolved in 10 ml. of a Na2COs (10 %) solution and the aqueous mixture was then saturated with NaCI. The aqueous phase was extracted by 5 x 15 ml. of a mixture CH2Cl2:2-Pr0H = 9:1. The organic phase was dried over MgSO4 and evaporated to afford 190 mg (90 %) of the free base of Compound 23. Analytical chiral separation:
20 μl_ of a 1 mg/mL solution of Compound 23 were injected on an analytical Chiralpak AD: flow-rate = 1 mL/min, temperature = 25 0C, mobile phase: ethanol, detection by UV at 220 nm and by polahmeter, Rt (+) = 4.98 min, Rt (-) = 6.23 min, k (+) = 0.55, k (-) = 0.93, α = 1.17 and resolution Rs = 3.34.
Regeneration of the salt:
After the chiral separation, about 70 mg of each enantiomer of the free base were dissolved in 100 ml. of ethanol and 3.6 ml. of HCI (0.2 N, 2.5 eq) were added. The solvent was evaporated then 50 ml. of ethanol were added and then removed in vacuo. The product was dissolved in 2 ml. of methanol and 3 ml. of ethyl acetate were added. The solvents were removed to give a white solid and then, the solids were dried over P2Os under vacuum overnight. Semi-preparative chiral separation:
A 175 mg/mL solution of the free base was prepared and 6 μl_ of this solution were injected every 3 min on an analytical Chiralpak AD, flow-rate = 1 mL/min, mobile phase ethanol, detection by UV at 254 nm. 150 successive injections were done. The two main fractions were identified by UV and collected in two different flasks. The solvent was removed in vacuo at 30 0C. The resulting solid was dissolved in 50 mL of CH2CI2 and then filtered on a 0.45 Dm millipore membrane. After evaporation Of CH2CI2, the solid was dissolved in 50 mL of methanol and then filtered. The salt was regenerated according to the procedure reported above.
The enantiomeric purity of the products was checked by analytical injection of the regenerated salts:
(+H2S,3f?)-2-Amino-3-hvdroxy-1 -(pyrrolidin-1 -yl)-3-(thiophen-3-yl)propan-1 - one hydrochloride Compound 205.
Figure imgf000128_0001
Compound 205
MW: 276.78; 83 mg obtained; Yield: 34 %; White Solid; Mp (0C): too hygroscopic. Enantiomeric excess = 99.5 % G25 D = + 20.4 (MeOH, c = 0.5).
(-H2f?,3S)-2-Amino-3-hvdroxy-1 -(pyrrolidin-1 -yl)-3-(thiophen-3-yl)propan-1 - one hydrochloride Compound 206.
Figure imgf000128_0002
Compound 206
MW: 276.78; 77 mg obtained; Yield: 32 %; White Solid; Mp (0C): too hygroscopic.
Enantiomeric excess = 99.0 % G25 D = - 20.0 (MeOH, c = 0.52). Preparation of (-)-f/?reo-2-amino-3-hvdroxy-3-(pyridin-3-yl)-1 -(pyrrolidin-1 - yl)propan-1-one dihvdrochloride Compound 207 and (+)-f/?reo-2-amino-3- hvdroxy-3-(pyhdin-3-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihvdrochlohde Compound 208.
Semi-preparative separation was performed on Chiralpak AD (250x10 mm): The semi-preparative chiral separation needed three steps :
First step A: A 80 mg/mL solution of (±)-f/?reo-2-amino-3-hydroxy-3- (pyhdin-3-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one Compound 20 free base (220 mg) racemate was prepared and 200 μl_ of this solution were injected every 8 min on Chiralpak AD, flow-rate = 4 mL/min, mobile phase: ethanol, detection by UV at 290 nm. Two main fractions were collected after 13 successive injections:
• 1 A containing about 61 mg of (-) enantiomer Compound 207 free base with ee > 97 %.
• 2A containing about 135 mg of mixture +/- enantiomers in a 74/26 ratio.
Second step B: A 30 mg/mL solution of fraction 2A was prepared and 100 μl_ of this solution were injected every 6 min on Chiralpak AD, flow-rate = 4 mL/min, mobile phase: ethanol, detection by UV at 290 nm. Two main fractions were collected after 45 successive injections:
• 1 B containing about 27 mg of (-) enantiomer Compound 207 free base with ee > 97 %.
• 2B containing about 105 mg of mixture (+)/(-) in a 93/7 ratio. Third step C: A 15 mg/mL solution of fraction 2B was prepared and 250 μL of this solution were injected every 6 min on Chiralpak AD, flow-rate = 4 mL/min, mobile phase: ethanol, detection by UV at 254 nm. Two main fractions were collected after 28 successive injections:
• 1 C containing about 7 mg of (-) enantiomer Compound 207 free base with ee > 97 %.
• 2C containing about 89 mg of (+) enantiomer with ee > 97 %. Fractions 1 A, 1 B and 1 C of (-) enantiomer Compound 207 free base were mixed together. Fraction 2C of (+) enantiomer Compound 208 free base was taken alone. For the both enantiomers, the solvent was removed in vacuo at 30 0C. The resulting solid was dissolved in 50 ml. of CH2CI2 and then filtered on a 0.45 Dm millipore membrane. After evaporation of CH2CI2, the solid was dissolved in 50 ml. of methanol and then filtered. The salt was regenerated according to the procedure reported below. The intermediate fraction collected contains 25 mg of a mixture of the both enantiomers in 50/50 (+)/(-) ratio and some impurities. Regeneration of the salt (dihydrochlohde):
After the chiral separation, about 90-95 mg of each enantiomers of the free base were dissolved in 100 ml. of ethanol and 10 ml. of HCI (0.2 N, 5 eq) were added. The solvent was evaporated and 50 ml. of ethanol were added and then removed in vacuo. The product was dissolved in 1 ml. of methanol and 5 ml. of ethyl acetate were added to precipitate the salt. The solvents were removed to give a white solid and then, the solids were dried over P2Os under vacuum overnight.
The enantiomeric purity of the products was checked by analytical HPLC injection of the regenerated dihydrochloride salts: (-H/7reo-2-Amino-3-hvdroxy-3-(pyridin-3-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihvdrochloride Compound 207.
Figure imgf000130_0001
Compound 207
MW: 308.20; 124 mg obtained; Yield: 43 %; White Solid; Mp (0C): 120.4 Enantiomeric excess = 97.8 % measured by HPLC at 220 nm (Chiralpak AD) RT = 6.24 min, eluent ethanol, flow 1 mL/min. Q25 D = - 15.9 (MeOH, c = 1 ). (+)-f/?reo-2-Amino-3-hvdroxy-3-(pyridin-3-yl)-1-(pyrrolidin-1-yl)propan-1-one dihydrochloride Compound 208.
Figure imgf000131_0001
Compound 208 MW: 308.20; 1 17 mg obtained; Yield: 40.5 %; White Solid; Mp (0C): 120.1 Enantiomeric excess = 98.0 % measured by HPLC at 220 nm (Chiralpak AD) RT = 7.39 min, eluent ethanol, flow 1 mL/min. G25 D = + 15.8 (MeOH, c = 1 ).
Preparation of (±)-f/?reo-2-amino-3-hvdroxy-3-(2-iodophenyl)-1 -(pyrrolidin-1 - yl)propan-1 -one hydrochloride Compound 209.
frans-(4,5-Dihvdro-5-(2-iodophenyl)oxazol-4-yl)(pyrrolidin-1-yl)methanone VIB
01090A. VIB 01090A was prepared in accordance with method D using 2- isocyano-1 -(pyrrolidin-1 -yl)ethanone SLA 09100 (327.9 mg, 2.155 mmol), potassium hydroxide (121 mg, 2.155 mmol) in methanol (2.2 mL) and 2-iodo- benzaldehyde (500 mg, 2.155 mmol). The solution was stirred for 3 h at 0 0C.
After work-up the crude product was purified by column chromatography (florisil, EtOAc:MeOH = 9:1 ) to obtain after evaporation frans-(4,5-dihydro-5-
(2-iodophenyl)oxazol-4-yl)(pyrrolidin-1-yl)methanone VIB 01090A as a yellow oil (364 mg, 44 % yield).
Figure imgf000131_0002
(±)-trans VIB 01090A
MW: 384.22; Yield: 44 %; Yellow Oil. Rf : 0.51 (EtOAc:MeOH = 9:1 ).
1H-NMR (CDCI3, δ): 1 .85-2.07 (m, 4H, 2xCH2), 3.50-3.62 (m, 3H, CH2), 3.78- 3.90 (m, 1 H, CH2), 4.57 (dd, 1 H, J = 5.6 Hz, J = 1.9 Hz, CH-N), 6.19 (d, 1 H, J = 5.6 Hz, CH-O), 7.05 (dt, 1 H, J = 7.7 Hz, J = 1.6 Hz, ArH), 7.15 (d, 1 H, J = 1.9 Hz, HC=N), 7.27 (dd, 1 H, J = 7.9 Hz, J =1 .6 Hz, ArH), 7.39 (t, 1 H, J = 7.3 Hz, ArH), 7.87 (d, 1 H, J = 7.8 Hz, ArH).
13C-NMR (CDCI3, δ): 24.3, 26.0, 46.3, 46.6, 74.8, 84.4, 95.0, 126.5, 128.4, 129.9, 139.8, 142.2, 155.7, 167.1.
(±)-f/?reo-2-Amino-3-hvdroxy-3-(2-iodophenyl)-1 -(pyrrolidin-1 -yl)propan-1 -one hydrochloride Compound 209.
Compound 209 was prepared following method E with trans-{4,5- dihydro-5-(2-iodophenyl)oxazol-4-yl)(pyrrolidin-1-yl)methanone VIB 01090A (0.345 g, 0.89 mmol), HCI 37 % (0.22 ml.) and methanol (4 ml_). After heating at 50 0C for 3 h and work-up, a trituration with EtOAc followed by filtration and drying afforded to (±)-f/?reo-2-amino-3-hydroxy-3-(2-iodophenyl)-1 -(pyrrolidin- 1-yl)propan-1-one hydrochloride Compound 209 as a white solid (287 mg, 74 % yield).
Figure imgf000132_0001
HCI
(±) Compound 209
MW: 396.65; Yield: 74 %; White Solid; Mp (0C): 164.0
1H-NMR (CD3OD, δ): 1.47-1.90 (m, 4H, 2xCH2), 1.95-2.10 (m, 1 H, CH2), 3.25-
3.55 (m, 3H, CH2), 4.23 (d, 1 H, J = 9.0 Hz, CH"N), 5.20 (d, 1 H, J = 9.0 Hz,
CH-O), 7.1 1 (t, 1 H, J = 7.4 Hz, ArH), 7.50 (t, 1 H, J = 7.5 Hz, ArH), 7.78 (d, 2H, J = 7.9 Hz, ArH), 7.88 (d, 2H, J = 7.7 Hz, ArH).
13C-NMR (CD3OD, δ): 24.8, 26.8, 47.8, 48.1 , 58.6, 75.8, 98.3, 130.1 , 130.8,
131.8, 141.1 , 143.8, 165.6.
MS-ESI m/z (% rel. Int.): 360.9 ([MH]+, 100), 342.9 (40). HPLC: Method A, detection UV 254 nm, Compound 209 RT = 3.88 min, peak area 97.8 %.
Preparation of (±)-f/?reo-2-amino-3-hvdroxy-3-hvdroxy-3-(4-iodophenyl)-1 - (pyrrolidin-1-yl)propan-1-one dihydrochloride Compound 210.
frans-(4,5-Dihvdro-5-(4-iodophenyl)oxazol-4-yl)(pyrrolidin-1-yl)methanone VIB 01090B.
VIB 01090B was prepared in accordance with method D using 2- isocyano-1-(pyrrolidin-1-yl)ethanone SLA 09100 (327.9 mg, 2.155 mmol), potassium hydroxide (121 mg, 2.155 mmol) in methanol (2.2 mL) and 4- iodobenzaldehyde (500 mg, 2.155 mmol). The solution was stirred for 3 h at 0 0C. After work-up, the crude product was washed in a minimum amount of MeOH and filtered to obtain after drying frans-(4,5-dihydro-5-(4- iodophenyl)oxazol-4-yl)(pyrrolidin-1-yl)methanone VIB 01090B as a white solid (0.377 g, 52 % yield).
Figure imgf000133_0001
(±ytrans VIB 01090B
MW: 384.22; Yield: 52 %; White Solid; Mp (0C): 1 15.1 1H-NMR (CDCI3, δ): 1 .75-2.10 (m, 4H, 2xCH2), 3.35-3.60 (m, 3H, CH2), 3.88- 4.02 (m, 1 H, CH2), 4.53 (dd, 1 H, J = 7.7 Hz, J = 1.9 Hz, CH-N), 6.09 (d, 1 H, J = 7.7 Hz, CH-O), 6.92-7.1 1 (m, 3H, 2xArH & CH=N), 7.69-7.70 (dd, 2H, J = 8.4 Hz, J = 1.7 Hz1 ArH).
13C-NMR (CDCI3,δ): 24.1 , 26.0, 46.2, 46.4, 75.7, 80.8, 94.0, 127.6 (2xC), 137.9 (2xC), 139.4, 155.2, 166.4.
(±)-f/?reo-2-Amino-3-hvdroxy-3-hvdroxy-3-(4-iodophenyl)-1-(pyrrolidin-1- yl)propan-1 -one dihvdrochloride Compound 210. Compound 210 was prepared following method E with trans-{4,5- dihydro-5-(4-iodophenyl)oxazol-4-yl)(pyrrolidin-1-yl)methanone VIB 01090B (0.345 g, 0.89 mmol), hydrochloric acid 37 % (0.24 ml.) and methanol (4.4 ml_). After heating at 50 0C for 3 h and work-up, a trituration with EtOAc followed by filtration and drying afforded to (±)-f/?reo-2-amino-3-hydroxy-3-(4- iodophenyl)-1 -(pyrrolidin-1 -yl)propan-1-one hydrochloride Compound 210 as a white solid (247.5 mg, 64 % yield).
Figure imgf000134_0001
(±)
Compound 210 MW: 396.65; Yield: 64 %; White Solid; Mp (0C): 184.4
1H-NMR (CD3OD, δ): 1 .35-1 .9 (m, 4H, 2xCH2), 2.20-2.33 (m, 1 H, CH2), 3.18-
3.40 (m, 3H, CH2), 4.1 1 (d, 1 H, J = 8.9 Hz, CH"N), 4.82 (d, 1 H, J = 8.9 Hz,
CH-O), 7.22 (d, 2H, J = 8.2 Hz, ArH), 7.76 (d, 2H, J = 8.2 Hz, ArH).
13C-NMR (CD3OD, δ): 24.8, 26.6, 47.3, 47.7, 59.2, 73.6, 95.1 , 129.8 (2xC), 138.9 (2xC), 140.6, 166.1 .
MS-ESI m/z (% rel. Int.): 360.9 ([MH]+, 100), 342.9 (85).
HPLC: Method A, detection UV 254 nm, Compound 210 RT = 4.08 min, peak area 96.8%.
Preparation of (±)-f/?reo-2-amino-3-hvdroxy-3-(3-iodophenyl)-1 -(pyrrolidin-1 - yl)propan-1 -one hydrochloride Compound 21 1 .
frans-(4,5-dihvdro-5-(3-iodophenyl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone SLA 09104. To a stirred and cooled (0 0C) solution of KOH (0.144 g, 2.57 mmol) in methanol (10 mL) was added 3-iodobenzaldehyde (0.500 mg, 2.15 mmol) and 1 -isocyano-3-(pyrrolidin-1 -yl)propan-2-one (0.295 g, 2.13 mmol). The solution was stirred 24 h with continued cooling and then concentrated. Water (50 mL) was added and the solution was extracted with EtOAc (3 x 50 mL). The organic layer was washed with brine (50 ml_), dried over MgSO4, filtered and evaporated. frans-(4,5-Dihydro-5-(3-iodophenyl)oxazol-4-yl)(pyrrolidin-1 - yl)methanone SLA 09104 was obtained (0.63 g, 80 % yield) as a pale yellow solid.
Figure imgf000135_0001
c (±)-trans
SLA 09104
MW: 370.19; Yield: 80 %; Pale Yellow Solid.
1H-NMR (CDCI3, δ): 1 .80-2.08 (m, 4H, 2xCH2), 3.42-3.58 (m, 3H, 1 .5xCH2), 3.90-3.96 (m, 1 H, 0.5xCH2), 4.56 (dd, 1 H, J = 7.7 Hz, J = 2.2 Hz, CH-N), 6.100 (d, 1 H, J = 7.7 Hz, CH-O), 7.01 (d, 1 H, J = 2.2 Hz, CH=N), 7.1 1 (t, 1 H, J = 6.8 Hz, ArH), 7.29 (m, 1 H, ArH), 7.66 (m, 2H, ArH).
13C-NMR (CDCI3, δ): 24.2, 26.0, 46.5, 46.6, 75.8, 80.4, 94.7, 125.0, 130.6, 134.6, 137.5, 142.0, 155.2, 166.3. 5 (±)-f/?reo-2-Amino-3-hvdroxy-3-(3-iodophenyl)-1 -(pyrrolidin-1 -yl)propan-1 -one hydrochloride Compound 21 1. frans-(4,5-Dihydro-5-(3-iodophenyl)oxazol-4-yl)(pyrrolidin-1- yl)methanone SLA 09104 (0.620 g, 1.67 mmol) was dissolved in methanol (5 mL). The solution was stirred at room temperature and a solution of HCI (370 %, 1 mL) was added via syringe and the mixture was stirred at 50 0C for 6 h. The mixture was concentrated and triturated with EtOAc. After filtration and drying (±)-f/?reo-2-amino-3-hydroxy-3-(3-iodophenyl)-1 -(pyrrolidin-1 -yl)propan- 1-one hydrochloride Compound 211 was obtained (586 mg, 88 % yield) as a white solid.
Figure imgf000135_0002
5 (±)
Compound 21 1 MW: 396.65; Yield: 88 %; White Solid; Mp (0C): 183.7 1H-NMR (CD3OD, δ): 1.35-1.50 (m, 1 H, 0.5xCH2), 1.58-1.82 (m, 3H, 1.5xCH2), 2.08-2.18 (m, 1 H, 0.5xCH2), 3.21 -3.45 (m, 4H, 2xCH2), 4.09 (d, 1 H, J = 9.1 Hz, CH-N), 4.80 (d, 1 H, J = 9.2 Hz, CH-O), 7.19 (t, 1 H, J = 7.1 Hz, ArH), 7.49 (d, 1 H, J = 7.6 Hz, ArH), 7.74 (m, 2H, ArH). MS-ESI m/z (% rel. Int.): 360.9 ([MH]+, 100), 342.9 (40). HPLC: Method A, detection UV 254 nm, Compound 21 1 RT = 4.07 min, peak area 93.4 %.
Preparation of (±)-f/?reo-2-amino-3-hydroxy-1 -(4-methylpiperazin-1 -yl)-3- (pyhdin-4-yl)propan-1-one thhvdrochlohde Compound 212.
2-lsocvano-1-(4-methyl-piperazin-1 -yl)-ethanone VIB 01 128.
Prepared in accordance with Method B with methyl isocyanoacetate
(1 .0 g, 10.09 mmol) and Λ/-methylpiperazine (1.37 ml_, 15.14 mmol). The reaction mixture was stirred overnight at RT and then concentrated. The residue was dissolved in dichloromethane (50 mL) and evaporated. The residue was coevaporated three times with a mixture of CH2Cl2:cyclohexane = 50:50 (3x10 mL). After drying 2-isocyano-1-(4-methyl-piperazin-1-yl)- ethanone VIB 01 128 was obtained as yellow solid (1.67 g, 99 % yield).
Figure imgf000136_0001
VIB 01 128 MW: 167.21 ; Yield: 99 %; Yellow Solid; Mp (0C) = 106.0
1H NMR (CDCI3, δ): 2.32 (m, 3H, Me) 2.38-2.50 (m, 4H, 2xCH2), 3.42 (t, 2H, J = 4.7 Hz, CH2), 3.66 (t, 2H, J = 4.7 Hz, CH2), 4.30 (s, 2H, CH2). 13C-NMR (CDCI3, δ): 42.4, 44.4, 45.5, 46.0, 54.3, 54.5, 160.8, 161.2. frans-(4,5-Dihvdro-5-(pyridin-4-yl)oxazol-4-yl)(4-methylpiperazin-1- vDmethanone VIB 01 130.
To a stirred and cooled (0 0C) solution of KOH (0.335 g, 5.98 mmol) in 7 ml. MeOH were added successively pyridine-4-carbaldehyde (0.705 g, 6.58 mmol) and 2-isocyano-1-(4-methyl-piperazin-1-yl)-ethanone VIB 01 128 (1.00 g, 6.58 mmol). The mixture was stirred at 0 0C until precipitation and concentrated. The mixture was partitioned between EtOAc (20 mL) and H2O (10 mL). The aqueous layer was extracted twice with EtOAc (60 mL). The EtOAc fractions were combined, washed twice with brine (2x10 mL), dried over MgSO4 and filtered. After evaporation and drying frans-(4,5-dihydro-5- (pyhdin-4-yl)oxazol-4-yl)(4-methylpiperazin-1-yl)methanone VIB 01 130 was obtained (1.282 g, 78 % yield) as a yellow oil.
Figure imgf000137_0001
(±)-trans VIB 01 130 MW: 274.32; Yield: 78 %; Yellow Oil.
1H NMR (CDCI3, δ): 2.32 (m, 3H, Me) 2.35-2.58 (m, 4H, 2xCH2), 3.50-3.70 (m, 2H, CH2-N), 3.80-4.00 (m, 2H, CH2-N), 4.59 (dd, 1 H, J = 7.8 Hz, J = 2.2 Hz, CH-N), 6.27 (d, 1 H, J = 7.8 Hz, 0-CH), 7.02 (d, 1 H, J = 2.2 Hz, OCH=N), 7.23 (d, 2H, J = 4.7 Hz, ArH) , 8.62 (dd, 2H, J = 4.5 Hz, J = 1 .6 Hz, ArH). 13C-NMR (CDCI3, δ): 42.6, 45.7, 46.0, 54.6, 55.1 , 74.7, 79.6, 120.0 (2xC), 148.6, 150.3 (2xC), 154.8, 166.0.
MS-ESI m/z (% rel. Int.): 275.2 ([MH]+, 40), 190.1 (35), 147.0 (40), 127.0 (100). HPLC: Method A, detection UV 254 nm, VIB 01 130 RT = 0.70 min, peak area 99.9 %.
(±)-f/?reo-2-Amino-3-hvdroxy-1-(4-methylpiperazin-1-yl)-3-(pyhdin-4-yl)propan- 1-one trihvdrochloride Compound 212. To a solution of frans-(4,5-dihydro-5-(pyridin-4-yl)oxazol-4-yl)(4- methylpiperazin-1-yl)methanone VIB 01 130 (1 .235 g, 4.50 mmol) in MeOH (15 ml.) was added HCI 37 % (1.6 ml_). After heating (50 0C) the mixture for 3.5 h a white solid precipitated and the reaction mixture was concentrated and the crude product was coevaporated twice with ethyl acetate. After trituration with ethyl acetate, filtration and drying, (±)-f/?reo-2-amino-3-hydroxy-1 -(4- methylpiperazin-1 -yl)-3-(pyhdin-4-yl)propan-1 -one trihydrochloride Compound 212 (1.62 g, 96 % yield) was obtained as a white solid.
Figure imgf000138_0001
Compound 212
MW: 373.71 ; Yield: 96 %; White Solid; Mp (0C): 203.1 1H-NMR (CD3OD, δ): 2.95 (s, 3H, Me), 3.10-3.40 (m, 4H, 2xCH2), 3.48-3.80 (m, 2H, CH2), 4.19-4.49 (m, 1 H, 0.5xCH2), 4.51-4.79 (m, 1 H, 0.5xCH2), 5.06 (d, 1 H, J = 4.3 Hz, CH-NH2), 5.35-5.63 (m, 1 H, CH-O), 8.30 (s broad, 2H, ArH), 8.95 (d, 2H, J = 6.7 Hz, ArH).
MS-ESI m/z (% rel. Int.): 265.1 ([MH]+, 5), 248.1 (20), 156.1 (20), 148.0 (100). HPLC: Method A, detection UV 254 nm, Compound 212 RT = 0.68 min, peak area 99.9 %.
Preparation of (±)-f/?reo-2-amino-1 -(3,3-difluoropyrrolidin-1 -yl)-3-hydroxy-3- (pyhdin-4-yl)propan-1-one dihvdrochlohde Compound 213.
1-(3,3-Difluoropyrrolidin-1 -yl)-2-isocvanoethanone VIB 01 158. To stirred and cooled (0 0C) methyl isocyanoacetate (96 % technical grade, 345 mg, 3.48 mmol) was slowly added 3,3-difluoropyrrolidine hydrochloride (500 mg, 3.48 mmol), thethylamine (487 μL, 3.48 mmol) and MeOH (1 mL). The mixture was stirred for 15 h at RT and then concentrated. The resulting oil was coevaporated twice from EtOAc. 1-(3,3- Difluoropyrrolidin-1 -yl)-2-isocyanoethanone VIB 01 158 was obtained as a yellow oil (305 mg, 60 % yield) and used in the next step without purification.
Figure imgf000139_0001
MW: 174.17; Yield: 60 %; Yellow Oil.
1H NMR (CDCI3, δ): 2.30-2.62 (m, 2H, CH2) 3.65-3.90 (m, 4H, CH2), 4.25 (d, 2H, J = 17.5 Hz1 CH2).
frans-(3,3-Difluoropyrrolidin-1-yl)(4,5-dihvdro-5-(pyhdin-4-yl)oxazol-4- vDmethanone VIB 01 160.
To a stirred and cooled (0 0C) solution of KOH (0.098 g, 1 .75 mmol) in MeOH (2 mL) were added successively pyhdine-4-carbaldehyde (0.206 g 1.92 mmol) and 1 -(3,3-difluoropyrrolidin-1 -yl)-2-isocyanoethanone VIB 01 158 (0.305 g, 1.75 mmol). The mixture was stirred at 0 0C for 3 h. After evaporation of MeOH, the mixture was partitioned between EtOAc (50 mL) and H2O (40 mL). The aqueous layer was extracted with EtOAc (4x50 mL). The fractions were combined, washed twice with brine (2x20 mL), dried over MgSO4 and filtered. After evaporation and drying frans-(3,3-difluoropyrrolidin- 1-yl)(4,5-dihydro-5-(pyhdin-4-yl)oxazol-4-yl)methanone VIB 01 160 (0.33 g, 67 % yield) was obtained as a yellow oil.
Figure imgf000139_0002
VIB 01 160
MW: 281.26; Yield: 67 %; Yellow Oil.
1H NMR (CDCI3, δ): 2.30-2.60 (m, 2H, CH2) 3.67-3.99 (m, 3H, 1 .5xCH2), 4.22- 4.59 (m, 2H, CH-N & 0.5xCH2), 6.17 (d, 1 H, J = 7.8 Hz, CH-O), 7.04 (dd, 1 H, J = 3.2 Hz, J = 2.5 Hz, HC=N), 7.20-7.38 (m, 2H, ArH), 8.55-8.70 (m, 2H, ArH). (±)-f/?reo-2-Amino-1-(3,3-difluoropyrrolidin-1-yl)-3-hvdroxy-3-(pyridin-4- yl)propan-1 -one dihydrochloride Compound 213.
To a solution of frans-(3,3-difluoropyrrolidin-1 -yl)(4,5-dihydro-5-(pyridin- 4-yl)oxazol-4-yl)methanone VIB 01 160 (0.305 g, 1 .08 mmol) in methanol (4 ml.) was added hydrochloric acid 37 % (395 μl_). After heating (500C) the mixture for 3.5 h the reaction mixture was concentrated and the crude product was coevaporated twice with EtOAc. After trituration with EtOAc, filtration and drying, (±)-f/?reo-2-amino-1 -(3,3-difluoropyrrolidin-1 -yl)-3-hydroxy-3-(pyridin-4- yl)propan-1-one dihydrochloride Compound 213 (269 mg, 44 % yield) was obtained as a beige solid.
Figure imgf000140_0001
Compound 213
MW: 344.19; Yield: 44 %; Beige Solid; Mp (0C): 182.2 1H-NMR (CD3OD, δ): 2.25-2.62 (m, 2H, CH2 ), 3.30-4.20 (m, 4H, CH2), 4.55
(d, 1 H, J = 5.5 Hz , 0.35xCH-N), 4.66 (d, 1 H, J = 5.5 Hz , 0.65xCH"N), 5.42
(m, 1 H, CH-O), 8.23 (d, 2H, J = 4.8 Hz, ArH), 8.94 (d, 2H, J = 5.9 Hz, ArH).
MS-ESI m/z (% rel. Int.): 272.2 ([MH]+, 15), 254.1 (15), 178.1 (20), 148.1
(100), 137.1 (95). HPLC: gradient of solvent B:solvent A = 5:95 to 100% solvent B in 7 min.
Solvent A was H2O with 0.1 % Et3N and solvent B was CH3CN with 0.1 %
Et3N; detection UV 254 nm, Compound 213 RT = 4.70 min, peak area 98.1 %.
Preparation of (2S.3f?V & (2f?.3S)-2-amino-1-((S)-3-fluoropyrrolidin-1-yl)-3- hvdroxy-3-(pyridin-4-yl)propan-1 -one dihvdrochlorides Compounds 214.
1-((S)-3-Fluoropyrrolidin-1-yl)-2-isocvanoethanone VIB 01 166.
To stirred and cooled (0 0C) methyl isocyanoacetate (96 % technical grade, 1 ,18 g, 1 1.9 mmol) was slowly added (S)-(+)-3-fluoropyrrolidine hydrochloride (97%, 1.5 g, 1 1.9 mmol), triethylamine (1.67 ml_, 1 1.9 mmol) and MeOH (3 ml_). The mixture was stirred for 15 h at RT and concentrated. The mixture was stirred for 15 h at RT and concentrated. Water was added (50 ml.) and the mixture was extracted with EtOAc (3x50 ml_), dried over MgSO4, filtered and evaporated to obtained crude 1 -((S)-3-fluoropyrrolidin-1 - yl)-2-isocyanoethanone VIB 01 166 (1.47 g, 79 % yield) as a brown oil which was used in the next step without purification.
Figure imgf000141_0001
VIB 01 166
MW: 156.152; Yield: 79 %; Brown Oil.
1H NMR (CDCI3, δ): 1.85-2.50 (m, 2H, CH2), 3.40-4.35 (m, 6H, 3xCH2), 5.17- 5.47 (m, 1 H1 CHF).
MS-ESI m/z (% rel. Int.): 171.1 ([MH++Na], 100), 157.1 ([MH]+, 82), 130.1 (95).
frans-((S)-3-Fluoropyrrolidin-1 -yl)((4S,5f?)- & 4R5S)-4,5-dihvdro-5-(pyridin-4- yl)oxazol-4-yl)methanones VIB 01 168.
To a stirred and cooled (0 0C) solution of KOH (0.526 g, 9.39 mmol) in MeOH (10 ml.) were added successively pyhdine-4-carbaldehyde (1.10 g, 10.33 mmol) and 1-((S)-3-fluoropyrrolidin-1 -yl)-2-isocyanoethanone VIB 01 166 (1 .47 g, 9.39 mmol). The mixture was stirred at 0 0C to RT for 24 h. After evaporation of MeOH, the mixture was partitioned between EtOAc (40 ml.) and H2O (20 ml_). The aqueous layer was extracted with EtOAc (6x40 ml_). The EtOAc fractions were combined, washed twice with brine (2x10 ml_), dried over MgSO4, filtered and evaporated. The crude product was purified by column chromatography (florisil, EtOAc:MeOH = 9:1 ). After evaporation and drying frans-((S)-3-fluoropyrrolidin-1 -yl)((4S,5R)- & 4R,5S)-4,5-dihydro-5- (pyhdin-4-yl)oxazol-4-yl)methanones VIB 01 168 (590 mg, diastereoisomehc mixture in ratio 1 :1 , 24 % yield) were obtained as a yellow oil.
Figure imgf000142_0001
VIB 01 168
MW: 263.27; Yield: 24 %; Yellow Oil.
1H NMR (CDCI3, δ): 1.89-2.46 (m, 2H, CH2) 3.5-4.45 (m, 4H, CH2), 4.46-4.60 (m, 1 H, CH-N), 5.16-5.45 (m, 1 H, CH-F), 6.12-6.25 (m, 1 H, CH-O), 6.95-7.18 (m, 1 H, CH=N), 7.20-7.40 (m, 2H, ArH), 8.50-8.70 (m, 2H, ArH).
(2S.3f?)- & (2R3S)-2-Amino-1 -((S)-3-fluoropyrrolidin-1 -yl)-3-hvdroxy-3- (pyridin-4-yl)propan-1-one dihvdrochlorides Compounds 214. To a solution of ((S)-3-fluoropyrrolidin-1 -yl)((4S,5R)- & 4R,5S)-4,5- dihydro-5-(pyridin-4-yl)oxazol-4-yl)methanones VIB 01 168 (0.590 g, 2.24 mmol) in methanol (8 mL) was added hydrochloric acid 37 % (686 μl_). After heating (50 0C) the mixture for 3.5 h the reaction mixture was concentrated and the crude product was coevaporated twice with EtOAc. After trituration with EtOAc, filtration and drying (2S,3R)- & (2R,3S)-2-amino-1 -((S)-3- fluoropyrrolidin-1 -yl)-3-hydroxy-3-(pyhdin-4-yl)propan-1 -one dihydrochlohdes Compounds 214 (474 mg, diastereoisomehc mixture in ratio 1 :1 , 68 % yield) were obtained as a pale yellow solid.
Figure imgf000142_0002
Compounds 214
MW: 326.2; Yield: 68 %; Pale Yellow Solid; Mp (0C): 173.1. 1H-NMR (CD3OD, δ): 1.82-2.38 (m, 2H, CH2), 2.80-4.15 (m, 4H, 2xCH2), 4.35- 4.68 (m, 1 H, CH"N), 5.00-5.50 (m, 2H, CH-O & CH-F), 8.1 1 -8.32 (m, 2H, ArH), 8.82-9.00 (m, 2H, ArH). MS-ESI m/z (% rel. Int.): 254.2 ([MH]+, 15), 237.1 (20), 148.1 (100), 137.1 (70). HPLC: gradient of solvent B:solvent A = 5:95 to 100% solvent B in 7 min. Solvent A was H2O with 0.1 % Et3N and solvent B was CH3CN with 0.1 % Et3N; detection UV 254 nm, Compounds 214 RT = 4.35 min, peak area 99.0 %.
Preparation of (2S,3f?)- & (2R3S)-2-amino-1-((f?)-3-fluoropyrrolidin-1-yl)-3- hvdroxy-3-(pyridin-4-yl)propan-1 -one dihvdrochlorides Compounds 215.
1-((f?)-3-Fluoropyrrolidin-1-yl)-2-isocvanoethanone BLE 04170.
To stirred and cooled (0 0C) methyl isocyanoacetate (96 % technical grade, 0.79 g, 7.96 mmol) was slowly added (f?)-(-)-3-fluoropyrrolidine hydrochloride (1 .0 g, 7.96 mmol), thethylamine (1 .1 1 mL, 7.96 mmol) and MeOH (2.5 mL). The mixture was stirred for 15 h at RT and concentrated. Water was added (50 mL) and the mixture was extracted with EtOAc (3x50 mL), dried over MgSO4, filtered and evaporated to obtained crude 1 -((f?)-3- fluoropyrrolidin-1-yl)-2-isocyanoethanone BLE 04170 as a brown oil (0.96 g, 77 % yield) which was used in the next step without further purification.
Figure imgf000143_0001
BLE 04170
MW: 156.152; Yield: 77 %; Brown Oil.
1H NMR (CDCI3, δ): 1.85-2.50 (m, 2H, CH2), 3.40-4.35 (m, 6H, 3xCH2), 5.17-
5.47 (m, 1 H1 CHF).
frans-((f?)-3-Fluoropyrrolidin-1 -yl)((4S.5f?)- & 4R5S)-4.5-dihvdro-5-(pyridin-4- yl)oxazol-4-yl)methanones BLE 04172.
To a stirred and cooled (0 0C) solution of KOH (0.34 g, 6.06 mmol) in MeOH (4.5 mL) were added successively pyridine-4-carbaldehyde (0.71 g, 6.66 mmol) and a solution of BLE 04170 (0.95 g, 6.06 mmol) in MeOH (2.5 ml_). The mixture was stirred at 0 0C to RT for 15 h. After evaporation of MeOH, the mixture was partitioned between EtOAc (40 mL) and H2O (20 ml_). The aqueous layer was extracted with EtOAc (3x40 mL). The EtOAc fractions were combined, washed with brine (50 mL), dried over MgSO4, filtered and evaporated. The crude product was purified by column chromatography (florisil, EtOAc:MeOH = 9:1 ). After evaporation and drying trans-{{R)-3- fluoropyrrolidin-1 -yl)((4S,5R)- & 4R,5S)-4,5-dihydro-5-(pyridin-4-yl)oxazol-4- yl)methanones BLE 04172 (653 mg, diastereoisomeric mixture in ratio 1 :1 , 41 % yield) were obtained as a colorless oil.
Figure imgf000144_0001
trans
Figure imgf000144_0002
BLE 04172
MW: 263.27; Yield: 41 %; Colorless Oil.
1H NMR (CDCI3, δ): 1.89-2.46 (m, 2H, CH2) 3.5-4.45 (m, 4H, 2xCH2), 4.46- 4.60 (m, 1 H, CH-N), 5.16-5.45 (m, 1 H, CH-F), 6.12-6.25 (m, 1 H, CH-O), 6.95- 7.18 (m, 1 H, CH=N), 7.20-7.40 (m, 2H, ArH), 8.50-8.70 (m, 2H, ArH).
(2S.3f?)- & (2R3S)-2-Amino-1 -((f?)-3-fluoropyrrolidin-1 -yl)-3-hvdroxy-3- (pyridin-4-yl)propan-1-one dihvdrochlorides Compounds 215.
To a solution of frans-((R)-3-fluoropyrrolidin-1 -yl)((4S,5R)- & 4R,5S)- 4,5-dihydro-5-(pyridin-4-yl)oxazol-4-yl)methanones BLE 04172 (0.65 g, 2.47 mmol) in methanol (8 mL) was added hydrochloric acid 37 % (757 μL). After heating (50 0C) the mixture for 3.5 h the reaction mixture was concentrated and the crude product was coevaporated twice with EtOAc. After trituration with EtOAc, filtration and drying (2S,3R)- & (2R,3S)-2-amino-1 -((R)-3- fluoropyrrolidin-1 -yl)-3-hydroxy-3-(pyhdin-4-yl)propan-1 -one dihydrochlohdes Compounds 215 (544 mg, diastereoisomeric mixture in ratio 1 :1 , 68 % yield) were obtained as a pale yellow solid.
Figure imgf000145_0001
Compounds 215
MW: 326.2; Yield: 68 %; Pale Yellow Solid; Mp (0C): 134.5 1H-NMR (CD3OD, δ): 1.82-2.38 (m, 2H, CH2), 2.80-4.15 (m, 4H, CH2), 4.35- 4.68 (m, 1 H, CH"N), 5.00-5.50 (m, 2H, CH-O & CHF), 8.1 1 -8.32 (m, 2H, ArH), 8.82-9.00 (m, 2H1 ArH).
Preparation of Λ/-((±H/?reo-1 -hvdroxy-3-oxo-1 -(pyridin-4-yl)-3-(pyrrolidin-1 - yl)propan-2-yl)formamide hydrochloride Compound 216.
frans-(4,5-Dihydro-5-(pyhdin-4-yl)oxazol-4-yl)(pyrrolidin-1 - yl)methanone Compound 19 (0.200 g, 0.81 mmol) was dissolved in methanol (5 ml_). Dowex 50WX8-200 (0.5 ml_, washed beforehand by a 0.5 M solution of HCI then water) was added at RT. The mixture was stirred for 1 h at 50 0C and after cooling was filtered off. MeOH was evaporated and the residue was dried to give 150 mg of a pasty product (150 mg, 61 .0 % yield). The free base form was dissolved in a minimum of a mixture of EtOAc:MeOH = 95:5 and a 0.4 M solution of HCI in ether (1.3 ml_, 0.52 mmol) was added at + 4 0C. After evaporation of solvents, the product was precipitated with diethyl ether, filtered and dried to obtain Λ/-((±)-f/?reo-1-hydroxy-3-oxo-1 -(pyhdin-4-yl)-3- (pyrrolidin-1-yl)propan-2-yl)formamide hydrochloride Compound 216 as a beige solid (100 mg, 41.0 % yield).
Figure imgf000145_0002
Compound 216 MW: 299.75; Yield: 41.0 %; Beige Solid; Mp (0C): 203.1 Rf. 0.35 (CH2CI2:Me0H = 90:10) free base.
1H-NMR (CD3OD, δ): 1.87-2.00 (m, 4H, 2xCH2), 3.41-3.46 (m, 2H, CH2-N),
3.62-3.66 (m, 2H, CH2-N), 5.18 (d, 1 H, J = 3.6 Hz, CH-N), 5.40 (d, 1 H, J = 3.8
Hz, CH-O), 7.99 (s, 1 H, CH=O), 8.16 (d, 2H, J = 5.8 Hz, ArH), 8.81 (d, 2H, J = 5.5 Hz, ArH).
13C-NMR (CD3OD, δ): 25.0, 27.0, 47.5, 48.3, 55.6, 72.4, 126.6 (2xC), 142.1
(2xC), 163.4, 164.1 , 168.5.
MS-ESI m/z (% rel. Int.): 264.1 ([MH]+, 10), 148.0 (100).
HPLC: Method A, detection UV 254 nm, Compound 216 RT = 1.30 min, peak area 98.0 %.
Preparation of Λ/-((±)-f/?reo-2-acetoxy-2-pyridin-4-yl-1 -(pyrrolidine- 1 -carbonyl)- ethvD-formamide hydrochloride Compound 217.
Λ/-((±)-f/?reo-1 -hydroxy-3-oxo-1 -(pyhdin-4-yl)-3-(pyrrolidin-1 -yl)propan- 2-yl)formamide hydrochloride Compound 216 (0.220 g, 0.80 mmol) was dissolved in CH2CI2 (10 ml.) with triethylamine (280 μl_, 2 mmol) at 0 0C. Then acetic anhydride (160 μl_, 1.6 mmol) was added slowly and the mixture was stirred for 72 h at RT. The solvent was evaporated and the residue was dried under vacuum. After column chromatography (SiO2, EtOAc:MeOH = 90:10) to give a pasty product (100 mg, 41 % yield). The product obtained was dissolved in MeOH (10 mL) and a solution of HCI (0.4 M, 1 ml.) in Et2O was added at 0 0C. Evaporation of the volatiles led to acetic acid N-((±)-threo-2- acetoxy-2-pyhdin-4-yl-1-(pyrrolidine-1-carbonyl)-ethyl)-formamide hydrochloride Compound 217 was obtained as a white solid (1 10 mg, 40 % yield).
Figure imgf000146_0001
(±) HCI
Compound 217 MW: 341.79; Yield: 40.0 %; White Solid; Mp (0C): 173.9. Rf. 0.25 (EtOAc:MeOH = 90:10, free base).
1H-NMR (CD3OD1): 1.80-2.00 (m, 4H, 2xCH2), 2.19 (s, 3H, CH3) 3.29-3.64 (m, 4H, 2xCH2-N), 5.34 (s, 1 H, N-CH), 6.44 (s, 1 H, 0-CH), 8.01 (s, 1 H, CHO), 8.16 (s, 2H, ArH), 8.85 (s, 2H, ArH). 13C-NMR (CD3OD, δ): 20.5, 25.0, 27.0, 47.6, 47.7, 53.9, 73.4, 126.7 (2xC), 142.8 (2xC), 144.3, 163.3, 166.9, 170.9. MS-ESI m/z (% rel. Int.): 306.1 ([MH]+, 10), 261 .1 (100). HPLC: Method A, detection UV 254 nm, Compound 217 RT = 0.90 min, peak area 97.0 %.
Preparation of ferf-butyl (±)-f/?reo-1-hvdroxy-3-oxo-1 -(pyridin-4-yl)-3- (pyrrolidin-1-yl)propan-2-ylcarbamate hydrochloride Compound 218.
(±)-f/?reo-2-Amino-3-hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 - one dihydrochloride Compound 22 (2.90 g, 9.3 mmol) was dissolved in CH2CI2 (250 ml.) with Et3N (4.3 mL, 30.7 mmol) at 0 0C. Di-terf-butyl dicarbonate (2.45 g, 1 1 .2 mmol) in CH2CI2 (50 mL) was added slowly and the mixture was stirred for 15 h at RT. Brine (50 mL) was added and the product was extracted with CH2CI2. After drying over MgSO4 and filtration, CH2CI2 was evaporated and the residue was dried in vacuum. After column chromatography (SiO2, EtOAc:MeOH = 90:10), terf-butyl (±)-f/?reo-1-hydroxy- 3-OXO-1 -(pyhdin-4-yl)-3-(pyrrolidin-1-yl)propan-2-ylcarbamate TTA 08100 was obtained as a beige solid (2.00 g, 64 % yield). A Sample of TTA 08100 (55 mg) was dissolved in CH2CI2 (1 mL) and Et2O (30 mL) and a solution of HCI (0.1 M, 2 mL) in Et2O was added at 0 0C. Evaporation of the volatiles led to ferf-butyl (±)-f/?reo-1 -hydroxy-3-oxo-1 -(pyrid i n-4-yl )-3-(py rro lid i n- 1 -yl)propan-2- ylcarbamate hydrochloride Compound 218 (50 mg, 52% yield) as a white solid.
Figure imgf000147_0001
Compound 218
MW: 371.86; Yield: 52.0 %; White Solid; Mp (0C): 141.2.
Rf. 0.30 (EtOAc:MeOH = 90:10, free base).
1H-NMR (CD3OD, δ): 1.33 (s, 9H, 3xCH3) 1 .91-2.00 (m, 4H, 2xCH2), 3.44-3.49 (m, 2H, CH2), 3.60-3.64 (m, 2H, CH2-N), 4.78 (s, 1 H, N-CH), 5.38 (s, 1 H, O-
CH), 8.16 (s, 2H, ArH), 8.83 (s, 2H, ArH).
13C-NMR (CD3OD, δ): 25.0, 27.1 , 28.5 (3xC), 47.5, 48.0, 58.3, 72.8, 81 .0,
126.7 (2xC), 142.0 (2xC), 157.3, 164.6, 169.5.
MS-ESI m/z (% rel. Int.): 336.1 ([MH]+, 20), 219.1 (100). HPLC: Method A, detection UV 254 nm, Compound 218 RT = 3.8 min, peak area 98.0 %.
Preparation of (±)-erv^/?ro-2-amino-3-hvdroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 - yl)propan-1 -one dihvdrochlohde Compound 219.
ferf-Butyl (±H/?reo-1 -hydroxy-3-oxo-1 -(pyridin-4-yl)-3-(pyrrolidin-1 -vDpropan- 2-ylcarbamate VIB 01080.
To a solution of (±)-f/?reo-2-amino-3-hydroxy-3-(pyhdin-4-yl)-1- (pyrrolidin-1-yl)propan-1-one dihydrochlohde Compound 22 (3.25 g, 10.54 mmol) in CH2CI2 (250 mL) in a 500 ml. round bottom flask equipped with a magnetic stirrer and under nitrogen atmosphere was added via syringe at 0 0C triethylamine (4.69 mL, 33.73 mmol). A solution of di-ferf-butyldicarbonate (2.76 g, 12.65 mmol) in CH2CI2 (75 mL) was added at 0 0C dropwise via a dropping funnel. The reaction mixture was abandoned at 0° C for 2 h then at RT overnight. A solution of brine (130 mL) was added and the solution was extracted with CH2CI2 (3 x 75 mL), dried over MgSO4, filtered and evaporated. The crude product was purified by column chromatography (SiO2, EtOAc: MeOH = 90:10). After evaporation to dryness of the combined fractions, ferf-butyl (±)-f/?reo-1 -hydroxy-3-oxo-1 -(pyridin-4-yl)-3-(pyrrolidin-1 - yl)propan-2-ylcarbamate VIB 01080 (2.79 g, 79 % yield) was obtained as a yellow solid.
Figure imgf000149_0001
01080
MW: 335.4; Yield: 79 %; Yellow Solid; Mp (0C): 160.5 Rf. 0.31 (EtOAc:MeOH = 90:10).
1H-NMR (CDCI3,.: 1.26 (s, 9H, 3xCH3), 1.71 -2.00 (m, 4H, 2xCH2), 3.25-3.60
(m, 4H, 2xCH2N), 4.61 (dd, 1 H, J = 9.8 Hz, J = 2.6 Hz, N-CH), 4.96 (s, 1 H,
OH), 5.08 (d, 1 H, J = 2.6 Hz, NH), 5.47 (d, 1 H, J = 9.8 Hz, 0-CH), 7.35 (d, 2H,
J = 5.7 Hz, ArH), 8.58 (d, 2H, J = 5.4 Hz, ArH). 13C-NMR (CDCI3, δ): 24.0 (2xC), 25.9 (2xC), 28.1 (3xC), 46.1 , 46.7, 56.1 ,
60.4, 72.8, 80.2, 121.4 (2xC), 148.7, 149.5 (2xC), 155.6, 169.4.
MS-ESI m/z (% rel. Int.): 336.1 ([MH]+, 45), 280 (18), 219 (100); 148 (38).
ferf-Butyl (±)-1 ,3-dioxo-3-(pyridin-4-yl)-1 -(pyrrolidin-1 -yl)propan-2-ylcarbamate TTA 08120.
To a solution of terf-butyl (±)-f/?reo-1-hydroxy-3-oxo-1-(pyridin-4-yl)-3- (pyrrolidin-1-yl)propan-2-ylcarbamate VIB 01080 (3.92 g, 1 1.70 mmol) in CH2CI2 (320 mL) in a 500 mL round bottom flask equipped with a magnetic stirrer and under nitrogen atmosphere was added slowly Dess-Martin pehodinane (4.96 g, 1 1 .70 mmol) at RT. The reaction mixture was stirred at RT for 0.5 h and CH2CI2, washed with a mixture of saturated sodium bicarbonate (100 mL), 1 M sodium thiosulfate (50 mL), brine (50 mL) and dried over MgSO4, filtered and evaporated. Diethyl ether (250 mL) was added and the precipitate was filtered off. After evaporation of the filtrate, the crude product was purified by column chromatography (SiO2, CH2CI2:Et0Ac = 4:6). After evaporation to dryness of the combined fractions ferf-butyl (±)-1 ,3-dioxo- 3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-2-ylcarbamate TTA 08120 (3.0 g, 77 % yield) was obtained as a white solid.
Figure imgf000150_0001
TTA 08120
MW: 333.38; Yield: 77 %; White Solid; Mp (0C): 125.4 Rf. 0.25 (CH2CI2:Et0Ac = 4:6). 1H-NMR (CDCI3, δ): 1 .36 (s, 9H, 3xCH3), 1.86-2.00 (m, 4H, 2xCH2), 3.47-3.71 (m, 4H, 2xCH2N), 5.62 (d, 1 H, J = 7.4 Hz, N-CH), 6.07 (d, 1 H, J = 7.4 Hz, NH), 7.80 (dd, 2H, J = 4.6 Hz, J = 1.3 Hz, ArH), 8.80 (dd, 2H, J = 4.5 Hz, J = 1.5 Hz1 ArH).
13C-NMR (CDCI3, δ): 24.0, 26.0, 28.1 (3xC), 46.8, 47.0, 61.0, 80.9, 121.5 (2xC), 141.4, 150.8 (2xC), 155.1 , 163.8, 194.3.
MS-ESI m/z (% rel. Int.): 334.1 ([MH]+, 10), 173.1 (30), 129.1 (100).
ferf-Butyl (±)-e/yf/7/O-3-hvdroxy-1 -oxo-3-(pyridin-4-yl)-1 -(pyrrolidin-1 -vDpropan- 2-ylcarbamate TTA 08124P. To a solution of terf-butyl (±)-1 ,3-dioxo-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 - yl)propan-2-ylcarbamate TTA 08120 (2.28 g, 6.80 mmol) in MeOH (50 ml.) in a 250 mL round bottom flask equipped with a magnetic stirrer and under nitrogen atmosphere was added slowly sodium borohydhde (285 mg, 7.50 mmol) at RT. The reaction mixture was stirred at RT for 0.5 h and was cooled at 4 0C. A solution of 2 M NaOH (10 mL) was added and MeOH was evaporated at 30 0C. EtOAc (300 mL) was added and the organic phase was washed with brine (20 mL), dried over MgSO4, filtered and evaporated to give a crude compound (ratio erythro:threo = 80:20 estimated by 1H NMR 300 MHz) TTA 08124 (2.1 g, 92 % yield). The crude product was recrystallized in EtOAc and after filtration and drying ferf-butyl (±)-eryf/?ro-3-hydroxy-1-oxo-3- (pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-2-ylcarbamate TTA 08124P (1.30 g, ratio erythro-.threo = 96:4 estimated by 1H NMR 300 MHz analysis, 57 % yield) was obtained as a white solid.
Figure imgf000151_0001
(±)-erythro TTA 08124P
MW: 335.40; Yield: 57 %; White Solid; Mp (0C): 170.3 Rf. 0.45 (EtOAc : MeOH = 85:15).
1H-NMR (CDCI3, δ): 1 .43 (s, 9H, 3xCH3), 1.64-1.83 (m, 4H, 2xCH2), 2.88-2.96 (m, 1 H, CH2N), 3.23-3.29 (m, 1 H, CH2N), 3.34-3.43 (m, 1 H, CH2N), 3.56-3.63 (m, 1 H, CH2N), 4.66 (dd, 1 H, J = 8.9 Hz, J = 3.7 Hz, N-CH), 4.91 (dd, 1 H, J = 8.4 Hz, J = 3.4 Hz, 0-CH), 5.42 (d, 1 H, J = 8.8 Hz, OH), 5.75 (d, 1 H, J = 8.8 Hz, NH), 7.30 (d, 2H, J = 5.9 Hz, ArH), 8.57 (d, 2H, J = 5.8 Hz, ArH). Ratio erythro:threo = 96:4.
13C-NMR (CDCI3, δ): 24.0, 25.7, 28.1 (3xC), 45.7, 46.8, 55.2, 74.5, 80.5, 120.9 (2xC), 149.6, 149.8 (2xC), 155.4, 168.8. MS-ESI m/z (% rel. Int.): 336.1 ([MH]+, 10), 280.1 (20), 1 10.0 (100).
(±)-e/yf/?ro-2-Amino-3-hvdroxy-3-(pyridin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihvdrochlohde Compound 219.
To a solution of terf-butyl (±)-eryf/?ro-3-hydroxy-1 -oxo-3-(pyhdin-4-yl)-1- (pyrrolidin-1-yl)propan-2-ylcarbamate TTA 08124P (1 .15 g, 3.40 mmol) in MeOH (30 mL) in a 250 mL round bottom flask equipped with a magnetic stirrer was added HCI 37 % (3 mL, 35 mmol) at RT. The reaction mixture was stirred at RT for 0.4 h at 45 0C and MeOH was evaporated at 45 0C to give after drying, a white solid TTA 08136 (ratio erythro:threo = 96:4 estimated by 1H NMR 300 MHz analysis). Amberlite IRA-400 (Cl") (10 g) was washed successively with water (2x10 mL), 0.5 N NaOH (3x20 mL), water (2x10 mL) and MeOH (3x10 mL). A solution of TTA 08136 in MeOH (30 mL) was stirred with washed Amberlite IRA-400 for 5 min at RT. After filtration, the MeOH was evaporated and the free base form was purified by column chromatography (SiO2, CHCI3:EtOH 95° = 86:14) to give TTA 08136A (445 mg, 55 % yield) as a beige solid (no threo isomer detected by 1H NMR 300 MHz and HPLC). TTA 08136A (193 mg) was stirred at RT in ethyl acetate (5 mL) with a solution of 0.1 N HCI in isopropanol (17 mL). Solvents were evaporated at 33 0C, MeOH (0.5 mL) was added and the salt was precipitated by addition of EtOAc (20 mL) to give nearly quantitatively after filtration and drying (±)-eryf/7ro-2-amino- 3-hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochloride Compound 219 as a white solid (no threo isomer detected by 1H NMR 300 MHz and HPLC).
Figure imgf000152_0001
2.HCI (±)-erythro
Compound 219
MW: 308.20; Yield: 55 %; White Solid; Mp (0C): 154.1.
Rf: 0.18 (CHCI3:EtOH 95° = 86:14, free base). 1H-NMR (CD3OD, δ): 1.94-2.04 (m, 4H, 2xCH2), 3.45-3.56 (m, 2H, CH2N),
3.66-3.78 (m, 2H, CH2N), 4.71 (d, 1 H, J = 5.2 Hz N-CH), 5.50 (d, 1 H, J = 5.1
Hz, 0-CH), 8.12 (d, 2H, J = 5.6 Hz, ArH), 8.92 (d, 2H, J = 5.4 Hz, ArH).
13C-NMR (CD3OD, δ): 25.0, 27.0, 47.8, 58.4, 70.0, 127.2 (2xC), 143.1 (2xC),
159.8, 164.8. MS-ESI m/z (% rel. Int.): 236.1 ([MH]+, 10), 219.1 (55), 1 10.0 (100).
Preparation of (-)-(2f?,3f?)-2-amino-3-hvdroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 - yl)propan-1-one dihvdrochloride Compound 220 and (+H2S,3S)-2-amino-3- hvdroxy-3-(pyridin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihvdrochlohde Compound 221.
Analytical chiral separation:
20 μL of a 1 mg/mL solution of Compound 219 were injected on Chiralpak AD: flow-rate = 1 mL/min, temperature = 25 0C, mobile phase: hexane:ethanol = 8:2, detection UV at 220 nm and by polahmeter, Rt (-) = 16.26 min, Rt (+) = 19.02 min, k(-) = 4.38, k(+) = 5.30, α = 1.21 and resolution Rs = 1.90. Semi-preparative separation was performed on Chiralpak AS (250x10 mm) : A 40 mg/mL solution of (±)-e/yf/?ro-2-amino-3-hydroxy-3-(pyridin-4-yl)-
1-(pyrrolidin-1-yl)propan-1-one Compound 219 racemate was prepared and 100 μl_ of this solution were injected every 4 min on Chiralpak AS, flow-rate = 5 mL/min, mobile phase hexane:ethanol = 1 :1 , UV detection at 254 nm. 56 successive injections were done. The two main fractions were identified on UV and collected in two different flasks. The solvent was removed in vacuo at 30 0C. The resulting solid was dissolved in 50 ml. of CH2CI2 and then filtered on a 0.45 Dm millipore membrane. After evaporation Of CH2CI2, the solid was dissolved in 50 ml. of methanol and then filtered. The salt was regenerated according to the procedure reported for Compound 203 and Compound 204.
The enantiomeric purity of the compounds was checked by analytical injection on Chiralpak AD of the regenerated salts:
(-H2f?,3f?)-2-amino-3-hvdroxy-3-(pyridin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihvdrochloride Compound 220.
Figure imgf000153_0001
2.HCI
{-)-erythro
Compound 220
MW: 308.20; 133 mg obtained; White Solid; Mp (0C): too hygroscopic. Rf. 0.18 (CHCI3:EtOH 95° = 86:14, free base).
Enantiomeric excess = 99.1 % measured by HPLC at 220 nm (Chiralpak AD) G25 D = - 6.4 (MeOH, c = 1 ) 1H-NMR (CD3OD, δ): 1.94-2.03 (m, 4H, 2xCH2), 3.45-3.55 (m, 2H, CH2-N), 3.63-3.76 (m, 2H, CH2-N), 4.68 (d, 1 H, J = 5.1 Hz, N-CH), 5.48 (d, 1 H, J = 5.5 Hz, 0-CH), 8.08 (d, 2H, J = 6.4 Hz, ArH), 8.90 (d, 2H, J = 6.6 Hz, ArH). 13C-NMR (CD3OD, δ): 23.4, 25.4, 46.2, 56.7, 68.4, 125.7 (2xC), 141.3 (2xC), 158.5, 163.2.
(+)-(2S,3S)-2-amino-3-hvdroxy-3-(pyridin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochloride Compound 221 .
Figure imgf000154_0001
Compound 221
MW: 308.20; 140 mg obtained; White Solid; Mp (0C): too hygroscopic.
Rf. 0.18 (CHCI3:EtOH 95° = 86:14, free base). Enantiomeric excess = 98.6 % measured by HPLC at 220 nm (Chiralpak AD)
G25 D = + 6.3 (MeOH, c = 1 ).
1H-NMR (CD3OD, δ): 1.94-2.02 (m, 4H, 2xCH2), 3.44-3.52 (m, 2H, CH2-N),
3.64-3.74 (m, 2H, CH2-N), 4.68 (d, 1 H, J = 5.2 Hz, N-CH), 5.48 (d, 1 H, J = 4.3
Hz, 0-CH), 8.09 (d, 2H, J = 6.4 Hz, ArH), 8.90 (d, 2H, J = 6.1 Hz, ArH). 13C-NMR (CD3OD, δ): 23.4, 25.4, 46.2, 56.8, 68.4, 125.7 (2xC), 141.3 (2xC),
158.5, 163.2.
Preparation of (±)-f/?reo-3-hvdroxy-2-(isopropylamino)-3-(pyhdin-4-yl)-1 - (pyrrolidin-1-yl)propan-1-one dihvdrochloride Compound 223. To a stirred solution of (±)-f/?reo-2-amino-3-hydroxy-3-(pyhdin-4-yl)-1-
(pyrrolidin-1-yl)propan-1-one dihydrochloride Compound 22 (0.20 g, 0.65 mmol) in 3 ml. of MeOH were added triethylamine (180 μl, 1.30 mmol) and acetone (75 μl_, 1.00 mmol). The mixture was stirred overnight at RT under nitrogen and then was added AcOH (200 μl_, 3.2 mmol) and NaBH3CN (85 mg, 1 .3 mmol). After 5 h at 20 0C, MeOH was evaporated and the residue was partitioned between CH2CI2 and 1 N aqueous sodium carbonate. The organic layer was evaporated and the obtained residue was purified by column chromatography (SiO2, EtOAc:MeOH = 9:1 ). HCI treatment in MeOH gave (±)-f/?reo-3-hydroxy-2-(isopropylamino)-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 - yl)propan-1 -one dihydrochlohde Compound 223 (56 mg, 25 % yield) as a white solid.
Figure imgf000155_0001
2. HCl (±ythreo
Compound 223 MW: 350.28; Yield: 25 %; White Solid; Mp (0C): 189.8.
Rf : 0.20 (EtOAc:MeOH= 9:1 , free base).
1H-NMR (CD3OD, δ): 1.32 (d, 3H, J = 6.5 Hz, CH3), 1.40 (d, 3H, J = 6.5 Hz,
CH3), 1 .55-1.84 (m, 4H, 2xCH2), 2.66-2.74 (m, 1 H, CH), 3.27-3.54 (m, 4H,
CH2-N), 4.60 (d, 1 H, J = 7.9 Hz, N-CH), 5.38 (d, 1 H, J = 7.8 Hz, 0-CH), 8.24 (d, 2H, J = 6.3 Hz, ArH), 8.98 (d, 2H, J = 6.3 Hz, ArH).
13C-NMR (CD3OD, δ): 18.7, 20.1 , 24.7, 26.7, 47.7, 48.5, 52.2, 62.4, 72.5,
127.0 (2xC), 143.2 (2xC), 161.1 , 164.1.
MS-ESI m/z (% rel. Int.): 278.1 ([MH]+, 25), 179.1 (100).
HPLC: Method A, detection UV 254 nm, Compound 223 RT = 1.30 min, peak area 99.0 %.
Preparation of (±)-2-amino-3-(hvdroxyimino)-3-(pyridin-4-yl)-1 -(pyrrolidin-1 - yl)propan-1-one dihvdrochloride Compound 224.
(±)-2-ferf-Butyloxycarbonylamino-3-(hvdroxyimino)-3-(pyridin-4-yl)-1 - (pyrrolidin-1-yl)propan-1-one TTA 08160. To a stirred solution of ferf-butyl (±)-1 ,3-dioxo-3-(pyridin-4-yl)-1 - (pyrrolidin-1-yl)propan-2-ylcarbamate TTA 08120 (0.20 g, 0.60 mmol) in 10 mL of dioxane were added Et3N (125 μl, 0.90 mmol) and hydroxylamine hydrochloride (65 mg, 0.90 mmol). The mixture was stirred 2 h at 1 10 0C in a sealed tube then dioxane was evaporated. The obtained residue was purified by column chromatography (SiO2, CH2CI2:Me0H = 9:1 to 97:3) to give (±)-2- te/t-butyloxycarbonylamino-3-(hydroxyimino)-3-(pyridin-4-yl)-1 -(pyrrolidin-1 - yl)propan-1 -one TTA 08160 (100 mg, 48 % yield) as an oil.
Figure imgf000156_0001
TTA 08160
MW: 348.40; Yield: 48 %; Oil.
Rf: 0.30 (CH2CI2:Me0H = 97:3).
1H-NMR (CDCI3, δ): 1 .39 (s, 9H, 3xCH3), 1.81-1.98 (m, 4H, 2xCH2), 3.37-3.62
(m, 4H, 2xCH2-N), 5.44 (d, 1 H, J = 8.1 Hz, N-CH), 5.97 (d, 1 H, J = 8.1 Hz, NH), 7.37 (d, 2H, J = 5.6 Hz, ArH), 8.65 (d, 2H, J = 6.0 Hz, ArH).
13C-NMR (CDCI3, δ): 24.0, 26.0, 28.2 (3xC), 46.5, 46.7, 55.8, 80.3, 123.1
(2xC), 139.7, 149.4 (2xC), 155.1 , 166.4.
MS-ESI m/z (% rel. Int.): 349.2 ([MH]+, 85), 293.2 (100).
HPLC: Method A, detection UV 254 nm, TTA 08160 RT = 3.90 min, peak area 97.0 %.
(±)-2-Amino-3-(hvdroxyimino)-3-(pyridin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochlohde Compound 224.
(±)-2-ferf-Butyloxycarbonylamino-3-(hydroxyimino)-3-(pyridin-4-yl)-1- (pyrrolidin-1 -yl)propan-1 -one TTA 08160 (100 mg, 0.29 mmol) was dissolved in MeOH (2 mL) and a solution of 1 M HCI in MeOH (1 mL, 3.00 mmol) was added and the mixture was heated for 10 min at 45 0C. MeOH was evaporated and the residue was dried to give crude (±)-2-amino-3- (hydroxyimino)-3-(pyridin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one TTA 08164. Amberlite IRA-400 (Cl") (1 mL, 1.4 mmol) was washed successively with water (2x10 mL), NaOH 0.5 N (3x20 mL), water (2x10 mL) and MeOH (3x10 mL). A solution of TTA 08164 in MeOH (30 mL) was stirred with washed Amberlite IRA-400 for 5 min at RT. After filtration, the MeOH was evaporated and the free base form was purified by column chromatography (SiO2, CH2C^MeOH = 9:1 ). HCI Treatment in MeOH gave (±)-2-amino-3-(hydroxyimino)-3-(pyridin- 4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochloride Compound 224 (29 mg, 31 % yield) as a beige solid.
Figure imgf000157_0001
(±)
Compound 224
MW: 321.20; Yield: 31 %; Beige Solid; Mp (0C): 225.2
Rf: 0.30 (EtOAc:MeOH= 9:1 , free base).
1H-NMR (CD3OD, δ): 1 .85-2.04 (m, 4H, 2xCH2), 3.23-3.73 (m, 4H, CH2-N), 5.58 (d, 1 H, J = 4.4 Hz, CH), 8.23 (d, 2H, J = 5.0 Hz, ArH), 9.03 (d, 2H, J =
5.0 Hz, ArH).
13C-NMR (CD3OD, δ): 24.9,26.9, 47.9, 48.0, 55.6, 128.7 (2xC), 143.8 (2xC),
145.5, 148.8, 163.8.
MS-ESI m/z (% rel. Int.): 249.2 ([MH]+, 10), 1 15.0 (100). HPLC: Method A, detection UV 254 nm, Compound 224 RT = 0.60 min, peak area 99.0 %.
Preparation of (±)-f/?reo-2-(4-iodobenzylamino)-3-hvdroxy-3-(pyhdin-4-yl)-1 - (pyrrolidin-1-yl)propan-1-one hydrochloride Compound 225. To a stirred solution of (±)-f/?reo-2-amino-3-hydroxy-3-(pyhdin-4-yl)-1-
(pyrrolidin-1-yl)propan-1-one dihydrochloride Compound 22 (500 mg, 1 .62 mmol) in 7.5 mL of MeOH at RT under nitrogen was added dropwise thethylamine (495 μL, 3.56 mmol) and 4-iodobenzaldehyde (413 mg, 1.73 mmol). The mixture was stirred for 5 h at RT under nitrogen. Acetic acid (463 μl_, 8.1 mmol) and NaBHsCN (356 mg, 5.67 mmol) were added. The mixture was stirred for another 15 h at RT. The mixture was partitioned between EtOAc (750 ml.) and an 10 % aqueous solution of potassium carbonate. The organic layer was washed with brine (2 x 20 ml_), dried over MgSO4 and filtered. After evaporation, the crude product was purified by column chromatography (SiO2, EtOAc: MeOH = 85:15) to give (±)-threo-2-(4- iodobenzylamino)-3-hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one VIB 01096 as a yellow solid (380 mg, 52 % yield). To a solution of {±)-threo-2- (4-iodobenzylamino)-3-hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one VIB 01096 (380 mg, 8.43 mmol) in methanol (10 ml.) was added a solution of 0.5 M aqueous hydrochloric acid (7 ml_). After stirring the mixture at RT for 0.5 h the reaction mixture was concentrated and the crude product was coevaporated twice with EtOAc. After trituration with EtOAc, filtration and drying, (±)-f/?reo-2-(4-iodobenzylamino)-3-hydroxy-3-(pyhdin-4-yl)-1- (pyrrolidin-1 -yl)propan-1 -one hydrochloride Compound 225 (373 mg, 44 % yield) was obtained as a pale yellow solid.
Figure imgf000158_0001
Compound 225 MW: 524.37; Yield 44 %; Pale Yellow Solid; Mp (0C): 194.7
1H-NMR (CD3OD, δ): 1.42-1.80 (m, 4H, 2xCH2), 2.32-2.50 (m, 1 H, CH2), 2.96- 3.12 (m, 1 H, CH2), 3.12-3.25 (m, 2H, CH2), 4.27 (q, 2H, J = 13.3 Hz, CH2), 4.42 (d, 1 H, J = 7.8 Hz, HC-N), 5.31 (d, 2H, J = 7.7 Hz, HC-O), 7.29 (d, 2H, J = 8.1 Hz, ArH), 7.80 (dd, 2H, J = 2.9 Hz, J = 8.1 Hz, ArH), 8.15 (d, 2H, J = 5.5 Hz, ArH), 8.91 (d, 2H, J = 5.5 Hz, ArH).
13C-NMR (CD3OD, δ): 24.7, 26.5, 47.5, 48.4, 51 .1 , 63.7, 72.3, 96.9, 126.9 (2xC), 131.0, 133.6 (2xC), 139.5 (2xC), 143.2 (2xC), 161.1 , 163.8. MS-ESI m/z (% rel. Int.): 451.9 ([MH]+, 100), 363.8 (45), 342.9 (70), 216.9 (75), 148.0 (30).
HPLC: Method A, detection UV 254 nm, Compound 225 RT= 3.83 min, peak area 98.7 %.
Preparation of (±)-f/?reo-2-(2-iodobenzylamino)-3-hvdroxy-3-(pyhdin-4-yl)-1 - (pyrrolidin-1-yl)-propan-1-one dihydrochlohde Compound 226.
To a stirred solution of (±)-f/?reo-2-amino-3-hydroxy-3-pyhdin-4-yl-1- pyrrolidin-1-yl-propan-1 -one Compound 22 (500 mg, 1.62 mmol) in 10 ml methanol was added successively Et3N (496 μl_, 3.57 mmol) and 2-iodo- benzaldehyde (414 mg, 1.78 mmol) in 1 ml methanol. The mixture was stirred 6 h at RT under nitrogen and acetic acid (464 μl_, 8.1 1 mmol) and sodium cyanoborohydhde (356 mg, 5.57 mmol) were added. The mixture was stirred overnight at 200C and evaporated to give a residue which was partitioned between CH2CI2 and 1 N aqueous potassium hydroxyde. The organic layer was washed with brine and dried over MgSO4, filtered and evaporated. The crude product was purified by column chromatography (SiO2, EtOAc:MeOH = 9:1 ). After evaporation and drying a white solid (±)-threo-2-(2- iodobenzylamino)-3-hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)-propan-1 -one was obtained (419.6 mg, 57 % yield). The product was dissolved in methanol (10 ml_). The solution was stirred at room temperature and a solution of HCI (1 M, 7.4 ml.) was added via syringe at RT for 10 min. The mixture was concentrated and triturated with EtOAc. After filtration and drying, (±)-threo-2- (2-iodobenzylamino)-3-hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)-propan-1 - one dihydrochloride Compound 226 was obtained (195 mg, 25 % yield) as a white solid.
Figure imgf000159_0001
Compound 226 MW: 524.22; Yield: 25 %; White Solid; Mp (0C): 148.0
Rf : 0.50 (MeOH:EtOAc = 20:80, free base).
1H-NMR (CD3OD, δ): 1.33-1.80 (m, 4H, 2xCH2), 2.12-2.25 (m, 1 H, CH2), 2.75-
2.88 (m, 1 H, CH2), 3.14-3.25 (m, 1 H, CH2), 3.25-3.31 (m, 1 H, CH2), 4.14 (d, 1 H, J = 8.4 Hz, NH-CH), 4.37-4.52 (m, 2H, NH-CH2) 5.10 (d, 1 H, J = 8.3 Hz,
CH-O), 7.18 (t, 1 H, J = 7.6 Hz, ArH), 7.49 (d, 1 H, J = 7.5 Hz, ArH), 7.61 (d,
1 H, J = 7.7 Hz, ArH), 7.68 (d, 2H, J = 5.2 Hz, ArH), 7.97 (d, J = 7.9 Hz, 1 H,
ArH), 8.66 (d, J = 4.9 Hz, 2H, ArH).
13C-NMR (CD3OD, δ): 24.7, 26.5, 47.6, 47.8, 55.3, 64.3, 72.7, 101.8, 124.4 (2xC), 130.3, 132.0, 132.5, 135.1 , 141.7 (2xC), 148.2, 154.0, 164.2.
MS-ESI m/z (% rel. Int.): 451.9 ([MH]+, 100), 352.9 (55), 342.9 (30).
HPLC: Method A, detection UV 254 nm, Compound 226 RT = 3.72 min, peak area 98.95 %.
Preparation of (±)-f/?reo-2-(3-iodobenzylamino)-3-hvdroxy-3-(pyhdin-4-yl)-1 - (pyrrolidin-1-yl)-propan-1-one dihydrochlohde Compound 227.
To a stirred solution of (±)-f/?reo-2-amino-3-hydroxy-3-pyhdin-4-yl-1- pyrrolidin-1-yl-propan-1-one Compound 22 (500 mg, 1.62 mmol) in 10 ml methanol was added successively Et3N (496 μl_, 3.55 mmol) and 3-iodo- benzaldehyde (414 mg, 1.78 mmol) in 1 ml methanol. The mixture was stirred 6 h at RT under nitrogen and acetic acid (464 μl_, 8.1 1 mmol) and sodium cyanoborohydhde (356 mg, 5.57 mmol) were added. The mixture was stirred overnight at 20 0C then evaporated and the residue was partitioned between CH2CI2 and 1 N aqueous potassium hydroxyde. The organic layer was washed with brine and dried. The organic layer was evaporated. The crude product was purified by column chromatography (SiO2, CH2CI2:Me0H = 95:5). After evaporation and drying a white solid (±)-f/?reo-2-(3-iodobenzylamino)-3- hydroxy-3-(pyhdin-4-yl)-1-(pyrrolidin-1-yl)-propan-1-one (437 mg, 60 % yield). This product was dissolved in methanol (10 ml_). The solution was stirred at RT and a solution of HCI (1 M, 7.7 ml.) was added at room temperature for 10 min. The mixture was concentrated and triturated with diethyl ether. After filtration and drying, (±)-f/?reo-2-(3-iodobenzylamino)-3-hydroxy-3-(pyhdin-4- yl)-1-(pyrrolidin-1-yl)-propan-1-one dihydrochloride Compound 227 was obtained (205 mg, 26 % yield) as a white solid.
Figure imgf000161_0001
Compound 227 MW: 524.22; Yield: 26 %; White Solid; Mp (0C): 142.8 Rf: 0.30 (MeOH:CH2CI2 = 5:95, free base).
1H-NMR (CD3OD, δ): 1.25-1.80 (m, 4H, 2xCH2), 2.03-2.12 (m, 1 H, CH2), 2.80- 2.92 (m, 1 H, CH2), 3.1 1-3.25 (m, 2H, CH2), 4.06-4.30 (m, 3H, CH-N & N-CH2), 5.04 (d, 1 H, J = 8.8 Hz, CH-OH), 7.23 (t, 1 H, J = 7.8 Hz, ArH), 7.51 (d, 1 H, J = 7.5 Hz, ArH), 7.67 (d, 2H, J = 5.8 Hz, ArH), 7.80 (d, 1 H, J = 5.2 Hz, ArH), 7.87 (s, 1 H, ArH), 8.66 (d, J = 5.2 Hz, 2H, ArH).
13C-NMR (CD3OD, δ): 24.7, 26.5, 47.3, 47.7, 50.5, 64.3, 72.8, 95.3, 124.5 (2xC), 130.9, 132.0, 133.9, 140.1 , 140.5, 148.3, 153.7, 164.2. MS-ESI m/z (% rel. Int.): 451.9 ([MH]+, 100), 352.9 (40), 342.9 (50). HPLC: Method A, detection UV 254 nm, Compound 227 RT = 3.80 min, peak area 98.7 %.
Preparation of (+)-(2S.3f?)-2-(3-iodobenzylamino)-3-hvdroxy-3-(pyhdin-4-yl)-1 - (pyrrolidin-1-yl)-propan-1-one dihvdrochlohde Compound 228. To a stirred solution of (+)-(2S,3f?)-2-amino-3-hydroxy-3-(pyridin-4-yl)-
1-(pyrrolidin-1-yl)propan-1-one dihydrochloride Compound 204 (204 mg, 0.67 mmol) in MeOH (6 mL) was added Et3N (202 μl_, 1.46 mmol) and, via syringe, a solution of 3-iodobenzaldehyde (169 mg, 0.73 mmol) in methanol (1 mL). The mixture was stirred 5 h at RT under nitrogen. CH3COOH (190 μL, 3.30 mmol) and sodium cyanoborohydride (146 mg, 2.32 mmol) were added and the reaction mixture was stirred overnight at RT. MeOH was evaporated and the residue was partitioned between CH2CI2 and a solution of 1 N aqueous potassium carbonate. The organic layer was washed with brine, dried over MgSO4, filtered and evaporated. The crude product was purified by column chromatography (SiO2, CH2CI2:Me0H = 95:5). After evaporation and drying (2S,3R)-2-(3-iodobenzylamino)-3-hydroxy-3-(pyridin-4-yl)-1-(pyrrolidin-1-yl)- propan-1-one (246 mg) was obtained as a white solid. The product was dissolved in methanol (5 mL) and a solution of HCI in MeOH (1 M, 2.5 mL) was added via syringe and the solution was stirred at RT for 0.6 h. The mixture was concentrated and the resulting solid triturated with Et2O. After filtration and drying (+)-(2S,3f?)-2-(3-iodobenzylamino)-3-hydroxy-3-(pyridin-4- yl)-1-(pyrrolidin-1-yl)-propan-1-one dihydrochloride Compound 228 (280 mg, 80 % yield) was obtained as a white solid.
Figure imgf000162_0001
Compound 228
MW: 524.22; Yield: 80 %; White Solid; Mp (0C): 107.4
G22 D = + 101.9 (MeOH, c = 1.02). Rf. 0.30 (MeOH:CH2CI2 = 5:95, free base).
1H-NMR (CD3OD, δ): 1.52-1.82 (m, 4H, 2xCH2), 2.38-2.45 (m, 1 H, CH2), 3.01 -
3.09 (m, 1 H1 CH2), 3.18-3.32 (m, 2H, CH2), 4.20-4.29 (dd, 2H, J = 28.1 Hz1 J
= 13.4 Hz, NH-CH2), 4.40 (d, 1 H, J = 7.9 Hz, N-CH), 5.30 (d, J = 7.9 Hz, 1 H,
CH-O), 7.25 (t, 1 H, J = 5.4 Hz, ArH), 7.54 (d, 1 H, J = 7.1 Hz, ArH), 7.81 (d, 1 H, J = 7.2 Hz, ArH), 7.89 (d, 1 H, J = 1.3 Hz, ArH), 8.13 (d, J = 5.5 Hz, 2H,
ArH), 8.90 (d, J = 5.3 Hz, 2H, ArH).
MS-ESI m/z (% rel. Int.): 452.1 ([MH]+, 100), 353.0 (65), 343.1 (80).
HPLC: Method A, detection UV 254 nm, Compound 228 RT = 3.87 min, peak area 97.0 %.
Preparation of (2R3S)-2-(3-iodobenzylamino)-3-hvdroxy-3-(pyridin-4-yl)-1 - (pyrrolidin-1-yl)-propan-1-one dihvdrochloride Compound 229. Similar to Compound 228 with (-)-(2R,3S)-2-amino-3-hydroxy-3- (pyridin-4-yl)-1-(pyrrolidin-1-yl)propan-1-one dihydrochloride Compound 203 (77 mg, 0.25 mmol) in 2.3 ml of MeOH, Et3N (76 μl_, 0.79 mmol), 3- iodobenzaldehyde (64 mg, 0.275 mmol), CH3COOH (78.6 μl_, 1.37 mmol) and NaBH3CN (60.5 mg, 0.96 mmol).
(2f?,3S)-2-(3-iodobenzylamino)-3-hydroxy-3-(pyhdin-4-yl)-1-(pyrrolidin- 1-yl)-propan-1-one dihydrochloride Compound 229 (70 mg, 53.5 % yield) was obtained as a pale yellow solid.
Figure imgf000163_0001
Compound 229
MW: 524.22; Yield: 53.5 %; Pale Yellow Solid; Mp (0C): 179.7
Rf. 0.30 (MeOH:CH2CI2 = 5:95, free base).
1H-NMR (CD3OD, δ): idem to Compound 228.
MS-ESI m/z (% rel. Int.): 452.1 ([MH]+, 100), 353.0 (70), 343.1 (60). HPLC: Method A, detection UV 254 nm, Compound 228 RT = 3.78 min, peak area 99.0 %.
Preparation of (±)-f/?reo-2-amino-3-hvdroxy-1 -morpholino-3-(thiophen-3- yl)propan-1 -one hydrochloride Compound 230. frans-(4,5-Dihvdro-5-(thiophen-3-yl)oxazol-4-yl)(morpholino)methanone SLA
09052A.
SLA 09052A was prepared in accordance with method D using thiophene-3-carbaldehyde (0.768 ml_, 5.35 mmol), KOH (0.273 mg, 4.86 mmol) in methanol (5 mL) and 2-isocyano-1-morpholinoethanone SLA 071 18 (0.75 g, 4.86 mmol). The solution was stirred for 2 h at 0 0C. After work-up the residue was purified by column chromatography (florisil, EtOAc). After evaporation and drying, frans-(4,5-dihydro-5-(thiophen-3-yl)oxazol-4- yl)(morpholino)methanone SLA 09052A (0.327 g, 25 % yield) was obtained as a yellow oil.
Figure imgf000164_0001
SLA 09052A MW: 266.32; Yield: 25 %; Yellow Oil.
1H-NMR (CDCI3, δ): 3.40-4.00 (m, 8H, 4xCH2), 4.67 (dd, 1 H, J = 7.3 Hz, J = 2.2 Hz, CH-N), 6.29 (d, 1 H, J = 7.3 Hz, CH-O), 6.97 (d, 1 H, J = 2.2 Hz, CH=N), 7.01 (dd, 1 H, J = 5.0 Hz, J = 1.3 Hz, CH=C), 7.28-7.40 (m, 2H, CH=C).
(±)-f/?reo-2-Amino-3-hvdroxy-1-niorpholino-3-(thiophen-3-yl)propan-1-one hydrochloride Compound 230.
To a solution of frans-(4,5-dihydro-5-(thiophen-3-yl)oxazol-4- yl)(morpholino)methanone SLA 09052A (0.327 g, 1 .12 mmol) in MeOH (5 mL) was added HCI 37 % (2 mL). After heating (50 0C) the mixture for 24 h the reaction mixture was concentrated and the crude product was coevaporated twice with EtOAc. After trituration with EtOAc, filtration and drying (±)-threo-2- amino-3-hydroxy-1 -morpholino-3-(thiophen-3-yl)propan-1 -one hydrochloride Compound 230 (276 mg, 77 % yield) was obtained as a pale yellow solid.
Figure imgf000164_0002
HCI (±)
Compound 230
MW: 292.78; Yield: 77 %; Pale Yellow Solid; Mp (0C): 209.1 1H-NMR (CD3OD, δ): 2.77-2.92 (m, 2H, CH2), 3.20-3.60 (m, 6H, 3xCH2), 4.49 (d, 1 H, J = 8.7 Hz, CH-N), 4.95 (d, 1 H, J = 8.8 Hz, CHO), 7.18 (d, 1 H, J = 4.5 Hz, CH=C), 7.44 (d, 1 H, J = 1.7 Hz CH=C), 7.44 (dd, 1 H, J = 4.5 Hz, J = 1.7 Hz CH=C). Preparation of (±)-f/?reo-2-amino-3-hvdroxy-1 -(piperidin-1 -yl)-3-(thiophen-3- yl)propan-1 -one hydrochloride Compound 231 .
frans-(4,5-Dihydro-5-(thiophen-3-yl)oxazol-4-yl)(piperidin-1-yl)methanone SLA 09052B.
SLA 09052B was prepared in accordance with method D using thiophene-3-carbaldehyde (0.778 ml_, 5.43 mmol), KOH (0.273 mg, 4.94 mmol) in methanol (5 mL) and 2-isocyano-1-(pipehdin-1-yl)ethanone SLA 071 16B (0.75 g, 4.94 mmol). The solution was stirred for 2 h at 0 0C. After work-up (without any further purification) and drying frans-(4,5-dihydro-5- (thiophen-3-yl)oxazol-4-yl)(pipehdin-1-yl)methanone SLA 09052B was obtained as a yellow oil (1.29 g, 99 % yield).
Figure imgf000165_0001
SLA 09052B
MW: 264.34; Yield: 99 %; Yellow Oil.
1H-NMR (CDCI3, δ): 1 .47-1.75 (m, 6H, 3xCH2), 3.42-3.82 (m, 4H, 2xCH2), 4.72 (dd, 1 H, J = 7.2 Hz, J = 2.2 Hz, CH-N), 6.28 (d, 1 H, J = 7.2 Hz, CH-O), 6.96 (d, 1 H, J = 2.2 Hz, CH=N), 7.01 (dd, 1 H, J = 5.0 Hz, J = 1.2 Hz, CH=C), 7.30-7.35 (m, 2H, CH=C).
13C-NMR (CDCI3,δ): 24.5, 25.5, 26.5, 43.7, 46.8, 73.4, 78.0, 122.6, 125.1 , 127.2, 140.5, 155.0, 166.3.
(±H/7reo-2-Amino-3-hydroxy-1 -(piperidin-1 -yl)-3-(thiophen-3-yl)propan-1 -one hydrochloride Compound 231.
To a solution of frans-(4,5-dihydro-5-(thiophen-3-yl)oxazol-4- yl)(piperidin-1-yl)methanone SLA 09052B (1.29 g, 4.88 mmol) in methanol (5 mL) was added hydrochloric acid 37 % (5 mL). After heating (50 0C) the mixture for 24 h the reaction mixture was concentrated and the crude product was coevaporated twice with EtOAc. After trituration with EtOAc, filtration and drying (±)-f/?reo-2-amino-3-hydroxy-1 -morpholino-3-(thiophen-3-yl)propan-1 - one hydrochloride Compound 231 was obtained as a pale yellow solid (1.07 g, 75 % yield).
Figure imgf000166_0001
HCI
(±)
Compound 231
MW: 290.81 ; Yield: 75 %; Pale Yellow Solid; Mp (0C): 210.6 1H-NMR (CD3OD, δ): 0.82-0.97 (m, 1 H, 0.5xCH2), 1.30-1.70 (m, 5H, 2.5xCH2), 2.75-2.91 (m, 1 H, CH2), 3.12-3.55 (m, 3H, 1 .5xCH2), 4.51 (d, 1 H, J = 8.4 Hz, CH-N), 4.94 (d, 1 H, J = 8.5 Hz, CHO), 7.16 (d, 1 H, J = 4.9 Hz, CH=C), 7.44 (d, 1 H, J = 2.5, Hz CH=C), 7.44 (dd, 1 H, J = 4.5 Hz, J = 3.2 Hz CH=C).
Preparation of (±)-f/?reo-2-amino-3-hydroxy-1 -morpholino-3-(pyhdin-3- yl)propan-1-one dihvdrochloride Compound 232.
frans-(4,5-Dihvdro-5-(pyridin-3-yl)oxazol-4-yl)(morpholino)methanone SLA 09050A. SLA 09050A was prepared in accordance with method D using pyridine-3-carbaldehyde (0.65 mL, 4.86 mmol), KOH (0.273 mg, 4.86 mmol) in methanol (10 mL) and 2-isocyano-1 -morpholinoethanone SLA 071 18 (0.75 g, 4.86 mmol). The solution was stirred for 20 h at 0 0C. After work-up (without any further purification), evaporation and drying frans-(4,5-dihydro-5-(pyridin- 3-yl)oxazol-4-yl)(morpholino)methanone SLA 09050A was obtained as a yellow solid (0.92 g, 72.5 % yield).
Figure imgf000167_0001
SLA 09050A
MW: 261.28; Yield: 72.5 %; Yellow Solid.
1H-NMR (CDCI3,δ): 3.42-4.00 (m, 8H, 4xCH2), 4.63 (dd, 1 H, J = 7.7 Hz, J = 2.3 Hz, CH-N), 6.29 (d, 1 H, J = 7.7 Hz, CH-O), 7.02 (d, 1 H, J = 2.2 Hz, CH=N), 7.33 (m, 1 H, ArH), 7.60-7.66 (m, 1 H, ArH), 8.57-8.62 (m, 2H, ArH). 13C-NMR (CDCI3,δ): 43.0, 46.3, 66.7, 66.8, 74.6, 79.2, 123.5, 133.4, 135.1 , 147.5, 150.0, 155.0, 166.3. MS-ESI m/z (% rel. Int.): 262.1 ([MH]+, 55), 108.0 (100).
(±)-f/?reo-2-Amino-3-hvdroxy-1-morpholino-3-(pyridin-3-yl)propan-1-one dihydrochloride Compound 232.
To a solution of frans-(4,5-dihydro-5-(pyhdin-3-yl)oxazol-4- yl)(morpholino)methanone SLA 09050A (0.91 1 g, 3.48 mmol) in methanol (10 mL) was added hydrochloric acid 37 % (5 mL). After heating (500C) the mixture for 2.25 h the reaction mixture was concentrated and the crude product was coevaporated twice with EtOAc and the crude product was dissolved in a 1 N solution of K2CO3 which was extracted with a mixture CH2C^iPrOH = 9:1 (6 x 100 mL). The combined organic layer was dried over MgSO4, filtered and evaporated to led to a crude product which was purified by column chromatography (SiO2, EtOAC:MeOH = 70:30). After evaporation the product was dissolved in a solution of HCI in MeOH (0.5M, 29 mL) and stirred at RT for 1 .5 h. The product was co-evaporated twice with EtOAc. After trituration with EtOAc, filtration and drying, (±)-f/?reo-2-amino-3-hydroxy-1- morpholino-3-(pyhdin-3-yl)propan-1-one dihydrochloride Compound 232 (270 mg, 24 % yield) was obtained as a yellow solid.
Figure imgf000168_0001
(±)
Compound 232
MW: 324.2; Yield: 24 %; Yellow Solid.
1H-NMR (CD3OD, δ): 3.20-3.78 (m, 8H, 4xCH2), 4.87 (d, 1 H, J = 5.5 Hz, CH" 5 N), 5.40 (d, 1 H, J = 5.4 Hz, CHO), 8.20 (m, 1 H, ArH), 8.77 (d, 1 H, J = 8.3 Hz, ArH), 8.93 (d, 1 H, J = 5.5 Hz, ArH), 9.03 (s, 1 H, ArH).
Preparation of (±H/7reo-2-amino-3-hvdroxy-1 -piperidin-3-(pyridin-3-yl)propan- 1-one dihydrochloride Compound 233. 0 frans-(4,5-Dihvdro-5-(pyridin-3-yl)oxazol-4-yl)(piperidin)methanone SLA 09050B.
SLA 09050B was prepared in accordance with method D using pyhdine-3-carbaldehyde (0.512 mL, 5.42 mmol), KOH (0.277 mg, 4.93 mmol)5 in methanol (10 mL) and 2-isocyano-1 -pipehdinethanone SLA 071 16B (0.75 g, 4.93 mmol). The solution was stirred for 20 h at 0 0C. After work-up (without any further purification), evaporation and drying, frans-(4,5-dihydro-5-(pyridin- 3-yl)oxazol-4-yl)(piperidin)methanone SLA 09050B (0.917 g, 72 % yield) was obtained as a yellow solid.
Figure imgf000168_0002
SLA 09050B
MW: 259.30; Yield: 72 %; Yellow Solid.
1H-NMR (CDCI3,δ): 1 .59-1.73 (m, 6H, 3xCH2), 3.44-3.83 (m, 4H, 2xCH2), 4.68
(dd, 1 H, J = 7.6 Hz, J = 2.3 Hz, CH-N), 6.30 (d, 1 H, J = 7.6 Hz, CH-O), 7.025 (d, 1 H, J = 2.2 Hz, CH=N), 7.32 (m, 1 H, ArH), 7.60-7.66 (m, 1 H, ArH), 8.57-
8.62 (m, 2H1 ArH). 13C-NMR (CDCI3,δ): 24.4, 25.5, 26.4, 43.8, 46.9, 74.6, 79.5, 123.6, 133.5, 135.4, 147.5, 149.9, 154.8, 165.9.
(±)-f/?reo-2-Amino-3-hvdroxy-1-piperidin-3-(pyridin-3-yl)propan-1-one dihydrochloride Compound 233.
To a solution of frans-(4,5-dihydro-5-(pyridin-3-yl)oxazol-4- yl)(piperidin)methanone SLA 09050B (0.917 g, 3.54 mmol) in methanol (10 ml.) was added hydrochloric acid 37 % (5 mL). After heating at 50 0C the mixture for 2.25 h the reaction mixture was concentrated and the crude product was coevaporated twice with EtOAc. After trituration with EtOAc, filtration and drying, the crude product was dissolved in a 1 N solution of K2CO3 and the product was extracted CH2CI2:iPr0H = 9:1 (6 x 100 mL). The crude product was purified by column chromatography (EtOAc:MeOH = 7:3) (±)-f/?reo-2-amino-3-hydroxy-1 -piperidin-3-(pyhdin-3-yl)propan-1 -one was obtained pale yellow solid (223.5 mg). The product was dissolved in a solution of HCI in MeOH (0.5 M, 18 mL) and stirred at RT for 1.5 h. After trituration with EtOAc, filtration and drying, (±)-f/?reo-2-amino-3-hydroxy-1-pipehdin-3- (pyhdin-3-yl)propan-1-one dihydrochloride Compound 233 (208 mg, 18 % yield) was obtained as a yellow solid.
Figure imgf000169_0001
Compound 233
MW: 322.2; Yield: 18 %; Yellow Solid.
1H-NMR (CD3OD, δ): 1.15-1.80 (m, 6H, 4xCH2), 3.10-3.80 (m, 4H, 2xCH2), 4.88 (d, 1 H, J = 5.6 Hz, CH-N), 5.33 (d, 1 H, J = 5.6 Hz, CHO), 8.19 (t, 1 H, J = 7.1 Hz, ArH), 8.74 (d, 1 H, J = 7.9 Hz, ArH), 8.93 (d, 1 H, J = 5.6 Hz, ArH), 9.01 (s, 1 H1 ArH). Preparation of (2f?.3S)-2-((f?)-2-hvdroxy-1 -phenylethylamino)-3-hvdroxy-3- (pyridin-4-yl)-1-(pyrrolidin-1-yl)propan-1-one dihvdrochloride Compound 234.
(f?)-5-Phenylmorpholin-2-one EBE 06134.
To a solution of phenylbromoacetate (18.58 g, 86 mmol) in CH3CN under nitrogen was added a solution of (R)-phenylglycinol (10.17 g, 74 mmol) and di-isopropylethylamine (34 ml_, 195 mmol) in CH3CN. The volatiles were removed under reduced pressure keeping the bath temperature below 25 0C to obtain an oil that was treated with EtOAc (120 mL) and stirred for 15 min. The resulting white precipitate was removed by filtration. The filtrate was concentrated under reduced pressure and the desired product was isolated using column chromatography (SiO2) with a step gradient of 10 % to 50 % [v/v] EtOAc in cyclohexane to give after evaporation (f?)-5-phenylmorpholin-2- one EBE 06134 (3.17 g, 24 % yield) as a white solid.
Figure imgf000170_0001
MW: 177.2; Yield: 24 %; White Solid; Mp (0C): 50.3
Rf\ 0.30 (EtOAc:cyclohexane = 50:50). 1H-NMR (CDCI3, δ): 1.99 (s, 1 H, NH), 3.89 (q, 2H, J = 17.8 Hz, N-CH2), 4.18
(dd, 1 H, J = 3.7 Hz, J = 10.3 Hz, 0-CH), 4.29 (t, 1 H, J = 10.5 Hz, N-CH), 4.40
(dd, 1 H, J = 3.7 Hz, J = 10.5 Hz, 0-CH), 7.30-7.45 (m, 5H, ArH).
13C-NMR (CDCI3, δ): 46.8, 54.8, 72.8, 125.3 (2xC), 127.0, 127.3 (2xC), 135.9,
166.0. [G]22D = + 30.3 ° (c = 1.00, MeOH).
(1 S,3f?,5f?,8af?)-Tetrahvdro-5-phenyl-1 ,3-di(pyhdin-4-yl)oxazolor4,3- ciπ ,41oxazin-8(3H)-one EBE 06136.
A solution of (R)-5-phenylmorpholinon-2-one EBE 06134 (3.0 g, 16.9 mmol) and pyridine-4-carboxaldehyde (5.43 g, 50.7 mmol) in toluene (75 mL) was refluxed in a soxhlet extractor filled with molecular sieves 4A (25 g) for 16 hours. All the volatiles were evaporated and the desired product was purified by column chromatography (SiO2) using a gradient of 80 % to 100 % [v/v] EtOAc in cyclohexane to give after evaporation (1 S,3f?,5f?,8af?)-tetrahydro-5- phenyl-1 ,3-di(pyridin-4-yl)oxazolo[4,3-c][1 ,4]oxazin-8(3H)-one EBE 06136 (1.7 g, 46 % yield) as a pale yellow solid.
Figure imgf000171_0001
EBE 06136
MW: 373.4; Yield: 46 %; Pale Yellow Solid; Mp (0C): 155.6 Rf. 0.20 (EtOAc). 1H-NMR (CDCI3, δ): 4.10-4.17 (m, 1 H, N-CH), 4.25 (dd, 1 H, J = 3.3 Hz, N- CH), 4.36-4.54 (m, 2H, 0-CH), 5.38 (d, 1 H, O-CH, J = 8.2 Hz), 5.53 (s, 1 H, N- CH), 7.20-7.35 (m, 8H, ArH), 7.40-7.50 (m, 1 H, ArH), 8.51 (dd, 2H, J = 1.4 Hz, J = 4.5 Hz), 8.57 (dd, 2H, J = 1.4 Hz, J = 4.5 Hz, ArH).
((2f?,4S,5f?)-3-((f?)-2-Hvdroxy-1-phenylethyl)-2,5-di(pyridin-4-yl)oxazolidin-4- yl)(pyrrolidin-1 -yl)methanone EBE 06138.
To a solution of (1 S,3f?,5f?,8af?)-tetrahydro-5-phenyl-1 ,3-di(pyridin-4- yl)oxazolo[4,3-c][1 ,4]oxazin-8(3H)-one EBE 06136(1.7 g, 4.55 mmol) in CH2CI2 was added pyrrolidine (1.90 ml_, 22.8 mmol) and the solution was stirred under nitrogen at 25 0C for 16 h. All the volatiles were evaporated and the resulting product was isolated using column chromatography (SiO2) with a gradient of 0-20 % [v/v] MeOH in EtOAc to give after evaporation ((2R,4S,5R)-3-((R)-2-hydroxy-1-phenylethyl)-2,5-di(pyhdin-4-yl)oxazolidin-4- yl)(pyrrolidin-1 -yl)methanone EBE 06138 (0.665 g, 33 % yield) as a white solid.
Figure imgf000172_0001
EBE 06138
MW: 444.5; Yield: 33 %; White Solid; Mp (0C): 63.6.
Rf. 0.25 (MeOH:EtOAc = 20:80). 1H-NMR (CDCI3, δ); 1 .70-1.90 (m, 4H, 2xCH2), 2.25 (bs, 1 H, OH), 2.75-2.85
(m, 1 H, CH-N), 2.95-3.05 (m, 1 H, N-CH), 3.50-3.60 (m, 2H, N-CH2), 3.80-4.15
(m, 4H, 2xCH +CH2-O), 5.10 (d, 1 H, J = 4.7 Hz, CH), 6.32 (s, 1 H, CH), 7.18-
7.32 (m, 7H, ArH), 7.45 (d, 2H, J = 5.9 Hz, ArH), 8.54 (d, 2H, J = 6.0 Hz, ArH),
8.64 (d, 2H, J = 6.0 Hz). 13C-NMR (CDCI3, δ): 23.9, 26.0, 46.3, 46.6, 60.3, 63.8, 65.2, 94.1 , 121.1
(2xC), 123.0 (2xC), 128.0, 128.3 (2xC), 128.5 (2xC), 137.8, 147.8. MS-ESI m/z (% rel. Int.): 445.1 ([MH]+, 20).
HPLC: Method A, detection at 254 nm, EBE 06138 RT = 3.50 min, peak area
99 %. [G]22 D = -16.0 ° (c = 1.00, CHCI3).
(2f?,3S)-2-((f?)-2-Hvdroxy-1-phenylethylamino)-3-hvdroxy-3-(pyhdin-4-yl)-1- (pyrrolidin-1-yl)propan-1-one dihydrochloride Compound 234.
To a solution of ((2R,4S,5R)-3-((R)-2-hydroxy-1 -phenylethyl)-2,5- di(pyridin-4-yl)oxazolidin-4-yl)(pyrrolidin-1-yl)methanone EBE 06138 (600 mg, 1.34 mmol) in MeOH (6 ml.) was added a solution of 1 N HCI (6 ml.) and the reaction mixture was stirred at 60 0C for 2 h, evaporated to dryness to give a white solid (745 mg). This crude product was dissolved in CH2CI2 (10 mL) and Na2CO3 (sat. sol.). The organic layer was separated was separated and the aqueous layer was washed with CH2CI2 (5><2 mL). The combined organic layer were dried over Na2SO4, filtered over cotton mixed with silica gel (600 mg), evaporated and loaded on a silica gel column of 25 g. The desired product was eluted using a gradient of MeOH 0 % to 20 % in EtOAc to give after evaporation (2f?,3S)-2-((f?)-2-hydroxy-1 -phenylethylamino)-3-hydroxy-3- (pyridin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one (320 mg, 67 % yield) as a white solid.
Figure imgf000173_0001
Compound 234
MW = 355.43; Yield: 67 %; White Solid; Mp (0C) = 76.4
Rf: 0.3 (MeOH:EtOAc = 20:80).
1H NMR (CDCI3, δ): 1.18-1.30 (m, 2H, CH2), 1.30-1 .48 (m, 2H, CH2), 1 .80-
1.90 (m, 1 H, CH2), 2.23-2.33 (m, 1 H, N-CH2), 2.85-2.95 (m, 2H, N-CH2), 3.00-
3.12 (m, 2H, N-CH + N-CH), 3.72 -3.85 (m, 3H, 0-CH2 + NH), 4.58 (d, 1 H, J =
8.4 Hz, 0-CH), 7.18-7.35 (m, 7H, ArH), 8.51 (d, 2H, J = 6.0 Hz, ArH).
[α]22D = - 68.8 ° (c = 1 .00, CHCI3).
To a solution of (2R,3S)-2-((R)-2-hydroxy-1-phenylethylamino)-3- hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one in MeOH (100 mg), in MeOH (1 ml.) was added a solution HCI (1 N, 1.1 ml.) at 0 0C and the solution was stirred for 10 min to give after evaporation (2f?,3S)-2-((f?)-2-hydroxy-1- phenylethylamino)-3-hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochlohde Compound 234 (120 mg, 99 % yield) as a white solid.
Figure imgf000173_0002
2 HCl
Compound 234
MW: 430.4; Global Yield: 66 %; White Solid; Mp (0C): 38.6. 1H-NMR (CD3OD, δ): 1.38-1.58 (m, 4H, CH2), 2.12-2.22 (m, 1 H, N-CH2), 2.80- 2.90 (m, 1 H1 N-CH2), 3.00-3.10 (m, 2H1 N-CH2), 3.91 (dd, 1 H, J = 4.3 Hz, J = 1 1.3 Hz, CH2-O), 4.02-4.12 (m, 1 H, CH2-O), 4.55-4.65 (m, 2H, N-CH), 5.27 (d, 1 H, J = 8.9 Hz, 7.35-7.45 (m, 3H, ArH), 7.45-7.58 (m, 2H, ArH). 13C-NMR (CD3OD, δ): 24.5, 26.3, 47.2, 48.0, 63.5, 64.4, 67.3, 72.6, 126.7, 130.2, 131.5, 132.3, 143.3, 160.7, 163.8.
Preparation of Λ/-((±H/?reo-1 -hvdroxy-3-oxo-1 -(pyridin-4-yl)-3-(pyrrolidin-1 - yl)propan-2-yl)acetamide hydrochloride Compound 235. To a suspension of (±)-f/?reo-2-amino-3-hydroxy-3-(pyridin-4-yl)-1-
(pyrrolidin-1-yl)propan-1-one dihydrochloride Compound 22 (300 mg, 0.89 mmol) in CH2CI2 (6 ml.) was added Et3N (370 μl_, 2.67 mmol) and the mixture was stirred for 10 min, cooled at 4 0C and a solution of acetic anhydride (65 ml_, 0.89 mmol) in CH2CI2 was added dropwise for 10 min. The reaction mixture was brought to room temperature, stirred for 16 h and washed with water (3x4 ml_), NaOH (0.5 N) (3><4 ml.) evaporated to give an oily residue that was purified using column chromatography (SiO2) with a gradient of 0 % to 20 % MeOH in EtOAc. The product in MeOH at 4 0C was treated with a solution of 1 N HCI (4 ml.) and all the volatiles were evaporated. Product was precipitated using a mixture of methanol in EtOAc to obtain after evaporation Λ/-((±)-f/?reo-1 -hydroxy-3-oxo-1 -(pyridin-4-yl)-3-(pyrrolidin-1 -yl)propan-2- yl)acetamide hydrochloride Compound 235 (209 mg, 52 % yield).
Figure imgf000174_0001
(±) Compound 235 MW: 313.78; Yield: 52 %; White Solid; Mp (0C): 181.3 Rf. 0.20 (MeOH:EtOAc = 20:80, free base). 1H-NMR (CD3OD, δ): 1.82-2.05 (m, 7H, 2χCH2 & CH3), 3.35-3.45 (m, 2H, CH2), 3.50-3.65 (m, 2H, CH2), 5.10 (d, 1 H, J = 3.8 Hz, N-CH), 5.39 (d, 1 H, J = 3.9 Hz, 0-CH), 8.16 (d, 2H, J = 6.2 Hz, ArH), 8.81 (d, 2H, J = 6.7 Hz, ArH). 13C-NMR (CD3OD, δ): 22.1 , 25.0, 27.0, 47.5, 48.2, 57.0, 72.0, 126.6 (2xC), 142.0 (2C), 142.0 (2xC), 164.4, 169.0, 173.0.
Preparation of (f?)-2-amino-3-(pyridin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihvdrochlohde Compound 236.
ferf-Butyl (f?)-1 -oxo-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-2-ylcarbamate EBE 06172.
Λ/-Boc-(S)-3-(pyhdin-4-yl)alanine (0.500 g, 1.88 mmol) was dissolved in CH2CI2 (15 ml.) and DIEA (361 μl_, 2.07 mmol) was added. The mixture was cooled to 0 0C and isobutyl chloroformate (270 μl_, 2.07 mmol) was added. The mixture was stirred for 10 min and pyrrolidine (267 mg, 3.76 mmol) in CH2CI2 (5 ml.) was added. This mixture was stirred for 15 min at 4 0C, 12 h at 25 0C, washed successively with NaH2PO4, saturated sodium hydrogen carbonate, water and brine. The organic layer was dried over magnesium sulfate and evaporated to dryness. The residue obtained was purify by column chromatography (SiO2) using a gradient of MeOH 0-10% [v/v] in
EtOAc to give ferf-butyl (f?)-1 -oxo-3-(pyhdin-4-yl)-1 -(pyrrolidin-1-yl)propan-2- ylcarbamate EBE 06172 (146 mg, 24 % yield) as a white solid.
Figure imgf000175_0001
EBE 06172 MW: 319.4; Yield: 24 %; White Solid; Mp (0C): 52.5. Rf\ 0.30 (MeOH:EtOAc = 20:80).
1H-NMR (CDCI3, δ): 1 .40 (s, 9H, (CH3)3), 1.65-1.90 (m, 4H, CH2), 2.80-2.90 (m, 1 H, CH), 2.90-3.00 (m, 2H, CH2), 3.28-3.38 (m, 1 H, N-CH), 3.40-3.50 (m, 2H, N-CH2), 4.60-4.70 (m, 1 H, N-CH), 5.38-5.48 (m, 1 H, NH), 7.16 (d, 2H, J = 4.5 Hz, ArH), 8.50 (d, 2H, J = 4.5 Hz, ArH). MS-ESI m/z (% rel. Int.): 320.2 ([MH] +, 20).
HPLC: Method A, detection at 254 nm, EBE 06172 RT = 3.82 min, peak area 85 %.
(f?)-2-Amino-3-(pyridin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochloride Compound 236.
To a solution of TFA (2 ml.) in CH2CI2 (8 ml.) was added (R)-I -oxo-3- (pyhdin-4-yl)-1 -(pyrrolidin-1-yl)propan-2-ylcarbamate EBE 06172 (146 mg, 0.654 mmol) and the mixture was stirred for 2 h at 25 0C. The volatiles were evaporated and the product was treated with a suspension on amberlite-400 (OH" form, 2 g) in MeOH. The suspension was filtered and washed with MeOH (3 x 5 ml_). The combined methanol fractions were evaporated under reduced pressure and the desired product was isolated using column chromatography (SiO2) with a mixture of EtOAc: MeOH: N H4OH = 70:30:4 to give an oily residue that was treated with a solution of 0.1 N HCI in iPrOH for 10 min. Evaporation of the volatile afforded (f?)-2-amino-3-(pyridin-4-yl)-1- (pyrrolidin-1-yl)propan-1-one dihydrochloride Compound 236 (78 mg, 47 % yield) as a pale yellow solid.
Figure imgf000176_0001
2 HCl
Compound 236
MW: 255.75; Yield: 47 %; Pale Yellow Solid; Mp (0C): 127.5 Rf\ 0.30 (EtOAc:MeOH:NH4OH = 70:30:4, free base). 1H-NMR (CD3OD, δ): 1.80-2.05 (m, 4H, CH2), 3.28-3.80 (m, 6H, CH2, N-CH2), 3.70 (t, 1 H, J = 6.7 Hz, CH-N), 8.10 (d, 2H, J = 5.9 Hz, ArH), 8.88 (d, 2H, J = 5.6 Hz, ArH).
13C-NMR (CD3OD, δ): 23.4, 25.4, 35.6, 46.1 , 46.5, 51.1 , 128.5 (2χC), 146.3 (2χC), 156.1 , 165.0. Preparation of ferf-butyl 5-((±)-f/?reo-1 -hvdroxy-3-oxo-1-(pyridin-4-yl)-3- (pyrrolidin-1 -vDpropan^-ylcarbamovDpentylcarbamate Compound 237.
To a solution of Λ/-Boc-aminohexanoic acid (342 mg, 1.48 mmol) in THF (10 ml.) was added Λ/-methylmorpholine (163 μl_, 1.48 mmol). The solution was stirred for 5 min, cooled at -15 0C and treated dropwise with isobutyl chloroformate (21 1 μl_, 1.48 mmol). This solution was added via a stainless steal cannula to a solution of (±)-f/?reo-2-amino-3-hydroxy-3-(pyhdin- 4-yl)-1-(pyrrolidin-1-yl)propan-1-one dihydrochlohde Compound 22 (500 mg, 1.48 mmol) and Λ/-methyl-morpholine (489 mg, 1 .47 mmol) in THF (10 mL) at -15°C. The reaction mixture was kept for 0.5 h at -15 0C followed by 2 h at 25 0C with continuous stirring. After evaporation of the solvent, the residue was partitioned between EtOAc and H2O, washed with NaH2PO4, saturated aqueous NaHCO3, dried over sodium sulfate and purified by column chromatography (SiO2) with a gradient of 0 % to 10 % [v/v] MeOH in EtOAc to give ferf-butyl 5-((±)-f/?reo-1 -hydroxy-3-oxo-1 -(pyhdin-4-yl)-3-(pyrrolidin-1- yl)propan-2-ylcarbamoyl)pentylcarbamate Compound 237 (455 mg, 69 % yield) as a white solid.
Figure imgf000177_0001
Compound 237
MW: 448.6; Yield: 69 %; White Solid. Rf: 0.20 (EtOAc:MeOH = 90: 10).
1H-NMR (CD3OD, δ): 1.05-1.15 (m, 2H, CH2), 1.35-1.55 (m, 13H, 2χCH2 + C(CHs)3), 1 .75-1.95 (m, 4H, 2χCH2), 2.00-2.20 (m, 2H, O=CCH2), 3.05 (q, 2H, J = 6.7 Hz, N-CH2), 3.20-3.35 (m, 1 H, N-CH), 3.38-3.50 (m, 2H, N-CH2), 3.65- 3.75 (m, 1 H, N-CH), 4.72 (bs, 1 H, NH), 4.98 (dd, 1 H, J = 8.8 Hz, J = 3.6 Hz), 5.08 (d, 1 H, J = 3.3 Hz, OCH), 5.23 (bs, 1 H, OH), 6.50 (d, 1 H, J = 8.7 Hz, NH), 7.35 (d, 2H, J = 6.0 Hz, ArH), 8.58 (d, 2H, J = 4.6 Hz, J = 1.4 Hz, ArH). MS-ESI m/z (% rel. Int.): 449.2 ([MH] +, 30), 349.2 (100). HPLC: Method A, detection at 254 nm, Compound 237 RT = 4.03 min, peak area 99.9 %.
Preparation of 6-amino-Λ/-((±)-f/?reo-1 -hydroxy-3-oxo-1 -(pyhdin-4-yl)-3- (pyrrolidin-1-yl)propan-2-yl)hexanamide Compound 238.
To a solution of terf-butyl 5-((±)-f/?reo-1-hydroxy-3-oxo-1-(pyhdin-4-yl)- 3-(pyrrolidin-1-yl)propan-2-ylcarbamoyl)pentylcarbamate Compound 237 (81 mg, 0.181 mmol) in CH2CI2 (8 ml.) was added TFA (2 ml.) at 0 0C and stirred for 2 h at 0 0C. All the volatiles were evaporated to give a residue that was treated with a suspension of Amberlite-400 (OH") in MeOH. After filtration, the filtrate was evaporated and the product was isolated by column chromatography (SiO2) with CH2CI2:MeOH:NH4OH = 10:5:0.4 to afford 6- amino-Λ/-((±)-f/?reo-1 -hydroxy-3-oxo-1 -(pyridin-4-yl)-3-(pyrrolidin-1 -yl)propan- 2-yl)hexanamide Compound 238 (40 mg, 64 % yield) as a white solid.
Figure imgf000178_0001
Compound 238
MW: 448.6; Yield: 64 %; White Solid; Mp (0C): 134.4 Rf. 0.30 (CH2CI2:MeOH:NH4OH = 10:5:0.4).
1H NMR (CDCI3, δ): 1.12-1 .30 (m, 2H, CH2), 1.30-1.50(m, 2H, CH2), 1.50-1.65 (m, 2H, CH2), 1 .65-1.95 (m, 4H, CH2), 2.10-2.30 (m, 2H, CH2), 2.55-2.70 (t, 2H, J = 6.9 Hz, CH2), 3.10 -3.20 (m, 2H, CH2), 3.28-3.50 (m, 2H, CH2), 3.60- 3.70 (m, 1 H, CH), 4.95 (dd, 1 H, J = 5.1 Hz, J = 8.4Hz, 0-CH), 5.02 (d, 1 H, J = 5.0 Hz, OH), 7.1 1 Hz (d, J = 8.48 Hz, 1 H, ArH), 7.35 (dd, 2H, J = 4.4 Hz, J = 1.5 Hz, ArH), 8.55 (dd, J = 1.5 Hz, J = 4.6 Hz, 2H, ArH). 13C NMR (CDCI3, δ): 24.0, 25.1 , 25.8, 25.9, 32.5, 35.8, 41 .7, 46.0, 46.9, 55.6, 72.6, 121.3 (2χC), 149.2, 149.5 (2χC), 168.9, 173.7.
Preparation of (±)-erv^/?ro-2-amino-3-hvdroxy-3-(pyridin-4-yl)-1 -(pyrrolidin-1 - yl)butan-1-one dihydrochloride Compound 239.
(±)-c/s-(5-Methyl-5-pyridin-4-yl-4,5-dihvdro-oxazol-4-yl)-pyrrolidin-1-yl- methanone EBE 06180.
To a stirred solution of KOH (223 mg, 39.7 mmol) in MeOH was added 4-acetylpyhdine (478 mg, 39.7 mmol) and 2-isocyano-1-(pyrrolidin-1- yl)ethanone BLE 04098 (500 mg, 3.2 mmol). The reaction mixture was stirred for 3 h at 0 0C and then concentrated. The residue was partitioned between EtOAc and H2O. The organic layer was washed with brine, dried over MgSO4, filtered and evaporated to give a yellow oil that was purified by column chromatography (SiO2) with 20 % [v/v] MeOH in EtOAc to give cis- and trans- (±)-(5-methyl-5-pyridin-4-yl-4,5-dihydro-oxazol-4-yl)-pyrrolidin-1-yl-methanone. The mixture was further purified by column chromatography (SiO2) using a gradient of 2 % to 5 % [v/v] MeOH [v/v] in CH2CI2 to obtain the pure c/s-(±)-(5- methyl-5-pyhdin-4-yl-4,5-dihydro-oxazol-4-yl)-pyrrolidin-1-yl-methanone EBE 06180 (122 mg, 51 % yield) as white solid.
Figure imgf000179_0001
EBE 06180
MW: 259.3; Yield: 51 %; White Solid; Mp (0C): 140.9 Rf\ 0.30 (EtOAc:MeOH = 80:20).
1H-NMR (CDCI3, δ): 1 .45-1.75 (m, 4H, 2xCH2), 1.81 (s, 3H, CH3), 2.75-2.90 (m, 1 H, N-CH2), 3.10-3.22 (m, 1 H, N-CH2), 3.30-3.40 (t, 2H, J = 6.7 Hz, N- CH2), 4.83 (d, 1 H, J = 1.7 Hz, NCH), 7.22 (d, 1 H, J = 1 .4 Hz, N=CH), 7.27 (d, 2H, J = 6.0 Hz, Ar), 8.57 (d, 2H, J = 6.1 Hz, ArH).
13C-NMR (CDCI3, δ): 23.6, 26.0, 27.9, 46.0, 46.5, 77.8, 87.2, 120.2, 148.9, 149.6 (2xC), 155.6 (2xC), 165.4. (±)-e/yf/?ro-2-Amino-3-hvdroxy-3-(pyridin-4-yl)-1 -(pyrrolidin-1 -yl)butan-1 -one dihydrochloride Compound 239.
To a solution c/s-(±)-(5-methyl-5-pyridin-4-yl-4,5-dihydro-oxazol-4-yl)- pyrrolidin-1-yl-methanone EBE 06180 (50 mg, 0.19 mmol) in MeOH (1 ml.) was added a solution of 1 N HCI (1 mL) and the reaction mixture was heated at 60 0C for 2 h. All the volatiles were evaporated to give (±)-eryf/7ro-2-amino- 3-hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)butan-1 -one dihydrochloride Compound 239 (54 mg, 87 % yield) as a white solid.
Figure imgf000180_0001
Compound 239
MW: 322.23; Yield: 87 %; White Solid; Mp (0C): 140.9 Rf. 0.1 (EtOAc:MeOH = 80:20, free base).
1H-NMR (CDCI3, δ): 1 .85-2.05 (m, 7H, CH3 + 2χCH2), 3.35-3.65 (m, 4H, 2χN- CH2), 4.61 (s, 1 H, 0-CH), 8.23 (d, 2H, J = 4.5 Hz, ArH), 8.89 (d, 2H, J = 4.3 Hz, ArH).
Preparation of (S)-2-amino-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihvdrochlohde Compound 240.
ferf-Butyl (S)-1 -oxo-3-(pyridin-4-yl)-1 -(pyrrolidin-1 -yl)propan-2-ylcarbamate EBE 06190.
To a solution of Λ/-Boc-(2S)-3-(pyhdin-4-yl)alanine (500 mg, 1.9 mmol) in THF (12 mL) was added Λ/-methylmorpholine (200 μl_, 1.9 mmol) and the solution was stirred for 5 min, cooled at - 15 0C and treated dropwise with isobutyl chloroformate (249 μl_, 1.9 mmol). The mixture was stirred for 10 min and pyrrolidine (1.08 g, 15.2 mmol) was added and allowed to warm to 25 0C with stirring for 3 h. The solvent were removed under reduced pressure and the residue was partitioned between EtOAc and NaH2PO4 pH = 7.2. The aqueous layer was discarded and the organic layer was washed with aqueous saturated NaHCθ3, dried over Na2SO4, filtered and evaporated. The resulting solid was purified by column chromatography (SiO2) with a gradient of O % to 10 % [v/v] MeOH in EtOAc to give terf-butyl (S)-1-oxo-3-(pyridin-4-yl)-1- (pyrrolidin-1-yl)propan-2-ylcarbamate EBE 06190 (167 mg, 28 % yield) as a white solid.
Figure imgf000181_0001
EBE 06190 MW: 319.4; Yield: 28 %; White Solid; Mp (0C): 130.0 Rf. 0.30 (EtOAc:MeOH = 90:10).
1H-NMR (CDCI3, δ): 1 .40 (s, 9H, C(CH3)3, 1.65-1.90 (m, 4H, CH2), 2.80-2.95 (m, 1 H, CH), 2.95-3.05 (m, 2H, CH2), 3.30-3.45 (m, 1 H, NCH), 3.45-3.55 (m, 2H, N-CH2), 4.60-4.75 (m, 1 H, N-CH), 5.42 (d, 1 H, J = 8.8 Hz, NH), 7.16 (dd, 2H, J = 4.5 Hz, J = 1.5 Hz, ArH), 8.51 (dd, 2H, J = 1.5 Hz, J = 4.5 Hz, ArH). 13C-NMR (CDCI3, δ): 24.1 , 25.8, 28.3 (3xC), 39.1 , 45.8, 46.5, 52.6, 79.9, 124.8 (2χC), 145.7, 149.8 (2χC), 155.0, 169.2.
(S)-2-Amino-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihvdrochloride Compound 240.
To a solution of ferf-butyl (S)-1-oxo-3-(pyridin-4-yl)-1 -(pyrrolidin-1 - yl)propan-2-ylcarbamate EBE 06190 (100 mg, 0.313 mmol) in CH2CI2 (3 ml.) at 4 0C was added TFA (479 μl_, 6.26 mmol) and MeOH (0.3 ml.) and the reaction was stirred for 2 h. All the volatiles were evaporated to give a product that was treated with a suspension of amberlite-400 (OH" form, 5 g) in MeOH. The suspension was filtered and washed with MeOH (5x5 ml_). The combined methanol fractions were evaporated under reduced pressure and the desired product was isolated by column chromatography (SiO2) with a gradient of 0 % to 30 % [v/v] MeOH in EtOAc to give (S)-2-amino-3-(pyridin-4-yl)-1-(pyrrolidin- 1-yl)propan-1-one. The product was dissolved in MeOH, cooled at 5 0C and a solution of HCI (0.1 N) (9 mL) was added dropwise. All the volatiles were evaporated to give (S)-2-amino-3-(pyhdin-4-yl)-1-(pyrrolidin-1-yl)propan-1 -one dihydrochloride Compound 240 (91 mg, 99 % yield) as a white solid.
Figure imgf000182_0001
2 HCI
Compound 240
MW: 292.21 ; Yield: 99 %; White Solid; Mp (0C): 195.9
Rf. 0.10 (EtOAc:MeOH = 90: 10, free base). 1H-NMR (CD3OD, δ): 1.80-2.05 (m, 4H, CH2), 3.30-3.40 (m, 5H, N-CH2 + N-
CH), 3.60-3.75 (m, 1 H, CH), 4.72 (t, 1 H, J = 7.3 Hz, CH), 8.08 (d, 2H, J = 5.4
Hz, ArH), 8.87 (d, 2H, J = 5.3 Hz, ArH).
13C-NMR (CD3OD, δ): 25.0, 26.9, 37.2, 47.6, 48.0, 57.2, 130.0 (2χC), 143.1
(2χC), 157.3, 166.6. MS-ESI m/z (% rel. Int.): 220.1 ([MH]+, 10), 203.1 (50).
Preparation of (±)-f/?reo-2-amino-3-(6-(trifluoromethyl)pyhdin-3-yl)-3-hvdroxy- 1-(pyrrolidin-1-yl)propan-1 -one dihvdrochloride Compound 241 .
(±)-frans-(5-(6-(Trifluoromethyl)pyridin-3-yl)-4,5-dihvdrooxazol-4-yl)(pyrrolidin- 1-yl)methanone EBE 06196.
To a solution of KOH (184 mg, 3.28 mmol) in MeOH (10 mL) at 4 0C was added 6-(trifluoromethyl)pyhdine-3-carbaldehyde (575 mg, 3.28 mmol) and and 2-isocyano-1-(pyrrolidin-1 -yl)ethanone BLE 04098 (500 mg, 3.28 mmol). The mixture was stirred for 2 h at 4 0C and allowed to warm to 25 0C. All the volatiles were evaporated and the resulting product was partitioned between brine and EtOAc. The organic layer was separated, dried over Na2SO4, filtered to give after evaporation of the solvent (±)-trans-{5-{6- (trifluoromethyl)pyπdin-3-yl)-4,5-dihydrooxazol-4-yl)(pyrrolidin-1-yl)methanone EBE 06196 (801 mg, 78 % yield) as a pale brown solid.
Figure imgf000183_0001
MW: 310.13; Yield: 78 %; Pale brown solid.
1H-NMR (CDCI3, δ): 1 .75-2.10 (m, 4H, 2xCH2), 3.40-3.60 (m, 3H, 2χN-CH2), 3.90-4.00 (m, 1 H, N-CH), 4.57 (dd, 1 H, J = 8.0 Hz, J = 2.2 Hz, N-CH), 6.31 (d, 1 H, J = 8.0 Hz, 0-CH), 7.06 (d, 1 H, J = 2.2 Hz, N=CH), 7.71 (d, 1 H, J = 8.1 Hz, ArH), 7.84 (dd, 1 H, J = 1.8 Hz, J = 8.1 Hz, ArH), 8.70 (d, 1 H, J = 1.5 Hz, ArH).
(±)-f/?reo-2-Amino-3-(6-(trifluoromethyl)pyhdin-3-yl)-3-hvdroxy-1-(pyrrolidin-1 - yl)propan-1 -one dihvdrochloride Compound 241.
To a solution of (±)-frans-(5-(6-(thfluoromethyl)pyhdin-3-yl)-4,5- dihydrooxazol-4-yl)(pyrrolidin-1 -yl)methanone EBE 06196 (400 mg, 1 .28 mmol) in MeOH (2 ml.) was added HCI (37 %) (10 ml.) and the mixture was heated at 60 0C for 2 h with continuous stirring. After evaporation the resulting white solid was treated with a suspension of amberlite-400 (OH" form) in MeOH. The suspension was filtered and washed with MeOH (5x5 ml_). The combined methanol fraction were evaporated under reduced pressure and the desired product was isolated by column chromatography (SiO2) with a gradient of 0 % to 8 % [v/v] MeOH in EtOAc to obtain (±)-f/?reo- 2-amino-3-(6-(trifluoromethyl)pyhdin-3-yl)-3-hydroxy-1-(pyrrolidin-1-yl)propan- 1-one (287 mg). To a solution (±)-f/?reo-2-amino-3-(6-(thfluoromethyl)pyridin- 3-yl)-3-hydroxy-1-(pyrrolidin-1-yl)propan-1-one (209 mg, 0.69 mmol) in MeOH (2 ml.) at 4 0C and treated HCI 37 % (10 ml_). All the volatiles were evaporated to give (±)-f/?reo-2-amino-3-(6-(trifluoromethyl)pyhdin-3-yl)-3- hydroxy-1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochloride Compound 241 as a white solid (269 mg, 74 % yield).
Figure imgf000184_0001
(±)
Compound 241 MW: 376.2; Yield: 74 %; White Solid; Mp (0C): 190.0. Rf : 0.3 (EtOAc:MeOH = 92:8, free base).
1H-NMR (CD3OD, δ): 1.40-1.70 (m, 2H, CH2), 1.70-1 .90 (m, 2H, CH2), 2.35- 2.50 (m, 1 H, N-CH), 3.15-3.35 (m, 1 H, N-CH2), 3.35-3.45 (m, 2H, N-CH2), 4.32 (d, 1 H, J = 8.3 Hz, N-CH), 5.1 1 (d, 1 H, J = 8.2 Hz, 0-CH), 7.88 (d, 1 H, J = 8.3 Hz, ArH), 8.16 (d, 1 H1 J = 8.0 Hz, ArH), 8.73 (s, 1 H1 ArH).
13C-NMR (CD3OD, δ): 24.8, 26.6, 47.4, 47.9, 58.7, 71.3, 121.7, 122.9 (d, 1 C, J = 273.2 Hz, CF3), 137.9, 140.3, 149.0, 149.7, 165.8.
Preparation of (±)-f/?reo-2-amino-3-hydroxy-1 -(thiazolidin-3-yl)-3-(pyhdin-4- yl)propan-1-one dihydrochloride Compound 242.
(±)-f/?reo-2-Amino-3-hydroxy-3-(pyridin-4-yl)propanoic acid dihydrochlohde
EBE 10038B.
To a stirred solution of KOH (2.57 g, 35.4 mmol) in MeOH (35 mL) at 0
0C was added 4-pyridinecarboxaldehyde (3.80 g, 35.4 mmol) and tert- butylisocyano acetate (5 g, 35.4 mmol). The solution was stirred for 3 h at 0
0C and concentrated to obtain intermediate frans-4,5-dihydro-5-(pyridin-4- yl)oxazole-4-carboxylate as a pale yellow solid EBE 10038A.
Figure imgf000184_0002
EBE 10038A 1H NMR (CD3OD, δ): 4.29 (dd, 1 H, J = 1.7 Hz, J = 7.6 Hz, N-CH), 5.67 (d, 1 H, J = 7.6 Hz, 0-CH), 7.29 (d, 1 H, J = 1.9 Hz, N=CH), 7.46 (d, 2H, J = 4.7 Hz, ArH), 8.55 (dd, 2H, J = 1.7 Hz, J = 4.4 Hz, ArH).
The solid EBE 10038A was dissolved in MeOH (100 ml_), stirred 5 min at 0 0C and treated with HCI (12 N) (10.5 ml_). The reaction was heated at 60 0C for 2 h, cooled down to 4 0C to form a precipitate that was filtered. The filtrate was evaporated and dried to obtain (±)-f/?reo-2-amino-3-hydroxy-3- (pyhdin-4-yl)propanoic acid dihydrochlohde EBE 10038B with 20 % of (±)- erythro isomer (9 g, 99 % crude yield) as a pale beige solid.
Figure imgf000185_0001
(±)
EBE 10038B
MW: 255.10; Yield: 99 %; Pale Beige Solid.
1H-NMR (CD3OD, δ): 3.94 (d, 1 H, J = 3.9 Hz, N-CH), 4.61 (t, 1 H, J = 3.0 Hz, 0-CH), 8.31 (d, 2H, ArH), 8.95 (m, 2H, ArH). Only the formula and the 1H- NMR description of major threo isomer is shown. MS-ESI m/z (% rel. Int.): 183.1 ([MH]+, 5).
(±)-f/?reo-Λ/-Boc-2-amino-3-(pyridin-4-yl)-3-hvdroxy-propionic acid EBE 10040. A solution of di-ferf-butyldicarbonate (9.28 g, 42.5 mmol) in dioxane (40 ml.) was added to a pre-mixed ice cold solution of (±)-f/?reo-2-amino-3- hydroxy-3-(pyhdin-4-yl)propanoic acid dihydrochlohde EBE 10038B (9.01 g, 35.4 mmol) in a solution of 1 N NaOH (145 ml_). The biphasic mixture was stirred at 5 0C for 30 min and allowed to warm to room temperature for 3.5 h, concentrated, cooled in an ice bath, acidified to pH 4-5 and extracted with n- butanol. The combined extracts were dried over Na2SO4 and filtered to give (±)-f/?reo-Λ/-Boc-2-amino-3-(pyhdin-4-yl)-3-hydroxy-propionic acid EBE 10040 (3.34g, 33 % yield) as a pale yellow solid.
Figure imgf000186_0001
EBE 10040
MW: 282.2; Yield: 33 %; Pale Yellow Solid.
1H-NMR (DMSO-d6, δ): 1.24 (s, 9H, C(CH3)3), 4.32 (d, 1 H, J = 9.4 Hz, N-CH), 5.13 (s, 1 H, 0-CH), 6.35 (d, 1 H, J = 9.4 Hz, NH), 7.37 (d, 2H, J = 3.8 Hz, ArH), 8.48 (d, 2H, J = 3.6 Hz), 2 OH not seen.
13C-NMR (DMSO-d6, δ): 27.9 (3xC), 59.0, 71.3, 78.2, 121 .4, 122.8, 148.9, 150.3, 151.0, 155.2, 171 .6. MS-ESI m/z (% rel. Int.): 283.2 ([MH] +, 10). HPLC: Method A, detection at 254 nm, EBE 10040 RT = 3.17 min, peak area 95.9 %.
ferf-butyl (±)-f/?reo-1 -hvdroxy-3-oxo-1 -(pyhdin-4-yl)-3-(thiazolidin-3-yl)propan- 2-ylcarbamate EBE 10042. To a solution of (±)-f/?reo-Λ/-Boc-2-amino-3-(pyhdin-4-yl)-3-hydroxy- propionic acid EBE 10040 (500 mg, 1 .77 mmol) in CH2CI2 (10 ml.) at 0 0C was added thethylamine (253 μl_, 3.54 mmol), hydroxybenzothazole (239 mg, 1 .77 mmol), 1 -(3-dimethylaminopropyl)-3-ethylcarbo-diimide hydrochloride (EDCI) (340 mg, 1.77 mmol) and thiazolidine (140 μl_, 1.77 mmol). The reaction mixture was stirred for 2 h at 0 0C, allowed to warm to room temperature, stirred for 16 h and diluted in CH2CI2 (90 ml_). The mixture was washed with brine (3*10 ml_), 1 N NaOH (3^10 ml_), dried over Na2SO4, filtered to give a crude oil that was purified using column chromatography (SiO2) with a gradient of 3 % to 4 % MeOH in CH2CI2. After evaporation terf-butyl (±ythreo- 1 -hydroxy-3-oxo-1 -(pyhdin-4-yl)-3-(thiazolidin-3-yl)propan-2-ylcarbamate EBE 10042 (200 mg, 32 % yield) was obtained as a yellow oil.
Figure imgf000187_0001
EBE 10042
MW: 353.4; Yield: 32 %; Yellow Oil. Rf. 0.2 (CH2CI2:Me0H = 97:3). 1H-NMR (CDCI3, δ): 1 .29 (s, 9H, (CH3)3), 2.90-3.05 (m, 2H, S-CH2), 3.60-3.95 (m, 2H, N-CH2), 4.40-4.72 (m, 3H, N-CH + S-CH2-N), 5.10-5.15 (m, 1 H, OCH), 5.68 (m, 1 H, NH), 7.34 (d, 2H, J = 6.0 Hz, ArH), 8.53 (d, 2H, J = 6.0 Hz, ArH), OH not seen.
13C-NMR (CDCI3, δ), minor rotamer in parenthesis: 28.1 (29.2) [3xC], 31.1 , (48.5) 48.7, 49.0 (49.1 ), (56.4) 56.7, 72.3, 80.5, 121.4 [2xC], 148.7, 149.4 [2xC], 155.5, (169.0) 169.2. MS-ESI m/z (% rel. Int.): 354.2 ([MH]+, 20).
HPLC: Method A, detection at 254 nm, EBE 10042 RT = 3.87 min, peak area 98.1 %.
(±)-f/?reo-2-Amino-3-hvdroxy-1-(thiazolidin-3-yl)-3-(pyridin-4-yl)propan-1-one dihydrochlohde Compound 242.
To a solution of terf-butyl (±)-f/?reo-1-hydroxy-3-oxo-1-(pyridin-4-yl)-3-
(thiazolidin-3-yl)propan-2-ylcarbamate EBE 10042 (150 mg, 424 mg) in MeOH (10 ml.) at 4 0C was added a solution of 1 N HCI in MeOH (12 mL). The reaction mixture was allowed to warm at room temperature and stirred for 1 h.
All the volatiles were evaporated to give an oily residue that was dissolved in
MeOH and treated with EtOAc to form a precipitate. The volatiles were evaporated to give (±)-f/?reo-2-amino-3-hydroxy-1 -(thiazolidin-3-yl)-3-(pyridin- 4-yl)propan-1-one dihydrochlohde Compound 242 (135 mg, 98 % yield) as a pale yellow solid.
Figure imgf000188_0001
(±)
Compound 242
MW: 326.24; Yield: 98 %; Pale Yellow solid; Mp (0C): 210.6 1H-NMR (CDCI3, δ): 2.90-3.15 (m, 2H, S-CH2), 3.55-3.90 (m, 1 H, N-CH), 4.00- 4.15 (m, 1 H, N-CH), 4.18-4.53 (m, 1 H, N-CH), 4.62-4.78 (m, 2H, N-CH2-S), 5.38-5.49 (m, 1 H, 0-CH), 8.25 (d, 2H, J = 5.7 Hz, 2H, ArH), 8.94 (d, 2H, J = 5.7 Hz, ArH).
13C-NMR (CDCI3, δ), minor rotamer in parenthesis: (29.9) 31.8, 34.8, (51.1 ) 52.1 , (57.8) 58.2, 71.0 (71 .3), 126.8 [2xC], 143.2 [2xC], 161 .4, (164.8) 165.2. MS-ESI m/z (% rel. Int.): 254.2 ([MH]+, 15).
Preparation of (±)-f/?reo-2-amino-3-hydroxy-1 -(indolin-1 -yl)-3-(pyhdin-4- yl)propan-1-one dihydrochloride Compound 243.
To a solution of potassium frans-4,5-dihydro-5-(pyridin-4-yl)oxazole-4- carboxylate EBE 10038A (500 mg, 2.60 mmol) in CH2CI2 (13 ml.) were added HOBT (352 mg, 2.60 mmol), EDCI (499 mg, 2.60 mmol) and indoline (292 ml_, 2.60 mmol). The reaction mixture was stirred 2 h at 0 0C, allowed to warm to RT and stirred for 16 h. The reaction mixture was diluted in CH2CI2 (100 ml_), wash with brine (3x25 ml_), NaOH 1 N (3^25 ml_), dried over MgSO4, filtered to give after evaporation frans-(4,5-dihydro-5-(pyridin-4-yl)oxazol-4- yl)(1 /-/-indol-1-yl)methanone SLA 09182 (533 mg, 70 % yield) as a pale brown oil. To a solution of SLA 09182 (533 mg, 2.6 mmol) in MeOH (12 mL) was added a solution of HCI 37 % (880 μL). The reaction was stirred for 3 h at 50 0C and concentrated under reduced pressure. The resulting product was dissolved in MeOH and treated with amberlite (OH" form), filtered to give after evaporation a residue that was purified using silica gel chromatography with a gradient of MeOH 0 % - 10 % in CH2CI2 to yield (±)-f/?reo-2-amino-3-hydroxy- 1-(indolin-1 -yl)-3-(pyridin-4-yl)propan-1-one. The hydrochloride salt was formed by treatment with a solution of 1 M HCI in MeOH (3.2 mL) to give after evaporation (±)-f/?reo-2-amino-3-hydroxy-1 -(indolin-1 -yl)-3-(pyridin-4- yl)propan-1 -one dihydrochloride Compound 243 (149 mg, 16 % yield) as a white solid.
Figure imgf000189_0001
(±) Compound 243
MW: 356.25; Yield: 16 %; White Solid; Mp (0C): 202.5
Rf. 0.30 (CH2CI2: MeOH = 90:10, free base)
1H-NMR (CD3OD,δ): 3.65-3.80 (m, 1 H, N-CH2), 4.15-4.28 (m, 1 H, CH2), 3.65-
3.80 (m, 1 H1 CH2), 4.15-4.30 (m, 1 H, N-CH2), 4.64 (d, 1 H, J = 5.4 Hz, N-CH), 5.45 (d, 1 H, J = 5.3 Hz, 0-CH), 6.70 (t, 1 H, J = 7.4 Hz, ArH), 7.12 (dd, 2H, J =
7.7 Hz, ArH), 8.07 (d, 1 H1 J = 7.9 Hz, ArH), 8.15 (d, 2H, J = 6.2 Hz, ArH), 8.79
(d, 2H, J = 6.2 Hz, ArH).
13C-NMR (CD3OD,δ): 28.9, 30.8, 58.4, 71.1 , 1 18.5, 126.1 , 126.4, 126.5 (2xC),
128.4, 133.6, 142.9, 143.6 (2xC), 161.0, 164.7. MS-ESI m/z (% rel. Int.): 284.2 ([MH]+, 10).
Preparation of (±)-f/?reo-2-amino-3-(3,5-dichloropyridin-4-yl)-3-hydroxy-1 -(2H- pyrrol-1 (5/-/)-yl)propan-1-one dihvdrochloride Compound 245.
frans-(5-(3.5-Dichloropyhdin-4-yl)-4.5-dihvdrooxazol-4-yl)(2/-/-pyrrol-1 (5/-/)- vDmethanone SLA 09022.
To a stirred and cooled (0 0C) solution of KOH (0.315 g, 5.62 mmol) in methanol (70 mL) was added a mixture of 3,5-dichloropyhdine-4- carbaldehyde (0.989 mg, 5.62 mmol) and 2-isocyano-1-(2H-pyrrol-1 (5H)- yl)ethanone SLA 07178 (0.696 g, 5.1 1 mmol). The solution was stirred 2 h with continued cooling and then concentrated. The residue was partitioned between EtOAc (50 mL) and water (50 mL). The organic layer was combined with additional EtOAc extracts (3x50 mL), washed with brine (70 mL) and dried with MgSO4, filtered and evaporated to obtain a crude product which was purified by column chromatography (florisil, EtOAc:cyclohexane = 80:20) to obtain after evaporation frans-(5-(3,5-dichloropyridin-4-yl)-4,5- dihydrooxazol-4-yl)(2H-pyrrol-1 (5H)-yl)methanone SLA 09022 (1.267 g, 80 % yield) as a pale yellow solid.
Figure imgf000190_0001
(±ytrans
SLA 09022
MW: 312.15; Yield: 80 %; Pale Yellow Solid. Rf\ 0.15 (EtOAc: cyclohexane = 80:20).
1H-NMR (CDCI3, δ): 4.30-4.32 (m, 3H, 1.5xCH2), 4.77-4.83 (m, 1 H, 0.5xCH2), 4.86 (dd, 1 H, J = 2.2 Hz, J = 8.8 Hz, CH-N), 5.84-5.88 (m, 2H, CH=CH), 6.86 (d, 1 H, J = 8.7 Hz, CH-O), 6.96 (d, 1 H, J = 2.2 Hz, 0-CH=N), 8.52 (s, 2H, ArH).
13C-NMR (CDCI3, δ): 52.1 , 52.6, 59.2, 75.1 , 123.9, 124.4, 131.5, 139.5, 147.6, 148.3, 151.0, 153.9, 164.3.
(±)-f/?reo-2-Amino-3-(3.5-dichloropyridin-4-yl)-3-hvdroxy-1-(2/-/-pyrrol-1 (5/-/)- yl)propan-1-one dihvdrochloride Compound 245.
To a stirred solution of frans-(5-(3,5-dichloropyridin-4-yl)-4,5- dihydrooxazol-4-yl)(2H-pyrrol-1 (5H)-yl)methanone SLA 09022 (1.26 g, 4.04 mmol) in methanol (20 mL) was added HCI 37 % (1 .5 mL). The reaction mixture was stirred for 3 h at RT, concentrated and the resulting yellow oil was co evaporated twice with EtOAc and triturated with EtOAc to obtain after filtration and drying under vacuum (±)-f/?reo-2-amino-3-(3,5-dichloropyridin-4- yl)-3-hydroxy-1 -(2/-/-pyrrol-1 (5/-/)-yl)propan-1-one dihydrochloride (1.13 g, 75 % yield) as a pale yellow solid.
Figure imgf000191_0001
Compound 245
MW: 375.16; Yield: 75 %; Pale yellow solid; Mp (0C): 176.5 Rf. 0.15 (CH2CI2: MeOH = 90:10, free base).
1H-NMR (CD3OD, δ): 3.30-3.32 (m, 1 H, 0.5xCH2), 3.90-3.97 (m, 1 H, 0.5xCH2), 4.25-4.37 (m, 2H, CH2), 4.76 (d, 1 H, J = 10.3 Hz, N-CH), 5.60-5.65 (m, 1 H, =CH), 5.70 (d, 1 H, J = 10.3 Hz, 0-CH) 5.78-5.82 (m, 1 H, =CH), 8.57 (s, 2H, ArH).
13C-NMR (CD3OD, δ): 56.6, 56.7, 57.3, 72.4, 128.0, 128.9, 136.3(2C), 145.8, 152.5(2C), 168.1 .
Preparation of Λ/-(1 -oxo-3-(pyridin-4-yl)-1 -(pyrrolidin-1 -yl)prop-2-en-2- vDformamide hydrochloride (cis:trans isomers mixture) Compound 246.
To a stirred solution of triphenylphosphine (200 mg, 0.76 mmol) in 5 ml. of CH3CN were added at 20 0C diethylazodicarbonate (120 μl, 0.76 mmol), Et3N (55 μl, 0.38 mmol) and Λ/-((±)-f/?reo-1-hydroxy-3-oxo-1 -(pyridin-4- yl)-3-(pyrrolidin-1-yl)propan-2-yl)formamide hydrochloride Compound 216 (1 15 mg, 0.38 mmol). The mixture was stirred 2 h at 70 0C then solvent was evaporated. The obtained residue was purified by column chromatography (SiO2, EtOAc:MeOH = 9:1 ) to give Λ/-(1-oxo-3-(pyridin-4-yl)-1 -(pyrrolidin-1 - yl)prop-2-en-2-yl)formamide TTA 08074A (80 mg, 43 % yield). HCI treatment in EtOAc with HCI 0.4 N in diethyl ether (1 ml_, 0.4 mmol) gave after evaporation and drying Λ/-(1-oxo-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)prop-2-en-2- yl)formamide hydrochloride (transxis isomers mixture) Compound 246 (60 mg, 28 % yield) as a pale yellow pasty product.
Figure imgf000192_0001
cisArans mixture Compound 246
MW: 281.74; Yield: 28 %; Pale Yellow Pasty Product. Rf : 0.24 (EtOAc:MeOH = 9:1 ). 1H-NMR (CD3OD, δ): 1.80-2.05 (bs, 4H, 2xCH2), 3.25-3.45 (bs, 2H, CH2-N), 3.55-3.70 (bs, 2H, CH2-N), 6.87 (s, 1 H, CH), 7.82 (d, 2H, J = 5.0 Hz, ArH), 8.31 (s, 1 H, HC=O), 8.65 (d, 2H, J = 5.0 Hz, ArH).
13C-NMR (CD3OD, δ): 25.1 , 26.5, 47.1 , 48.9, 109.0, 125.6 (2xC), 127.7, 142.2 (2xC), 142.6, 154.9, 161 .6. MS-ESI m/z (% rel. Int.): 246.1 ([MH]+, 5), 175.1 (100).
HPLC: Method A, detection UV 254 nm, Compound 246 RT = 1.90 min, peak area 95.0 %.
Preparation of (2S.3R)- & (2f?.3S)-2-amino-1-((f?)-3-hvdroxypyrrolidin-1-yl)-3- hvdroxy-3-(pyhdin-4-yl)propan-1 -one dihvdrochlorides Compound 247.
1-((f?)-3-hvdroxypyrrolidin-1-yl)-2-isocvanoethanone VIB 01 172.
To stirred and cooled (0 0C) methyl isocyanoacetate (96 % technical grade, 1.7 g, 17.21 mmol) was slowly added (f?)-(+)-3-pyrrolidinol (1.5 g, 17.21 mmol) and MeOH (5 ml_). The mixture was stirred for 3 h at RT and concentrated. Brine was added (30 mL) and the mixture was extracted with EtOAc (3x50 mL), dried over MgSO4, filtered and evaporated to obtained crude 1-((f?)-3-hydroxypyrrolidin-1-yl)-2-isocyanoethanone VIB 01 172 (1.3 g, 49 % yield) as a yellow solid.
Figure imgf000192_0002
VIB 01 172 MW: 154.17; Yield: 49 %; Yellow Solid; Mp (0C) = 55.9 1H-NMR (CDCI3, δ): 1 .95-2.20 (m, 2H, CH2 ), 2.60-2.82 (m, 1 H, OH), 3.30- 3.68 (m, 4H, 2xCH2), 4.20-4.36 (m, 2H, CH2), 4.49-4.65 (m, 1 H, CH-O). MS-ESI m/z (% rel. Int.): 155.1 ([MH]+, 90).
frans-((f?)-3-hvdroχypyrrolidin-1 -yl)((4S.5f?)- & (4f?.5S)-4.5-dihvdro-5-(pyridin- 4-yl)oxazol-4-yl)methanones VIB 01 174.
To a stirred and cooled (0 0C) solution of KOH (0.40 g, 7.13 mmol) in MeOH (8 mL) were added successively pyhdine-4-carbaldehyde (0.84 g, 7.84 mmol) and 1-((f?)-3-hydroxypyrrolidin-1 -yl)-2-isocyanoethanone VIB 01 172 (1 .10 g, 7.13 mmol). The mixture was stirred at 0 0C to RT for 24 h. After evaporation of MeOH, the mixture was partitioned between EtOAc (50 mL) and H2O (10 mL). The aqueous layer was further extracted with EtOAc (2x50 mL). The EtOAc fractions were combined, washed twice with brine (2x10 mL), dried over MgSO4, filtered and evaporated. After evaporation and drying fra/7s-((R)-3-hydroxypyrrolidin-1 -yl)((4S,5R)- & (4R,5S)-4,5-dihydro-5-(pyridin- 4-yl)oxazol-4-yl)methanones VIB 01 174 (490 mg, diastereoisomehc mixture in ratio 1 :1 , 26 % yield) were obtained as a crude pale yellow solid.
Figure imgf000193_0001
trans
VIB 01 174
MW: 261.28; Yield: 26 %; Pale Yellow Solid.
1H NMR (CDCI3,δ): 1.88-2.22 (m, 2H, CH2), 3.50-3.80 (m, 3H, 1.5xCH2), 3.95- 4.20 (m, 1 H, 0.5xCH2), 4.40-4.65 (m, 2H, CH-N & CH-O), 4.74 (s, 1 H, OH), 6.18-6.22 (m, 1 H, CH-O), 7.00-7.12 (m, 1 H, HC=N), 7.20-7.30 (m, 2H, ArH), 8.52-8.68 (m, 2H, ArH). MS-ESI m/z (% rel. Int.): 262.2 ([MH]+, 45), 235.2 (75), 148 (100). (2S.3f?)- & (2R3S)-2-Amino-1 -((f?)-3-hvdroxypyrrolidin-1 -yl)-3-hvdroxy-3- (pyridin-4-yl)propan-1-one dihydrochlorides Compound 247.
To a solution of frans-((R)-3-hydroxypyrrolidin-1-yl)((4S,5R)- & (4R,5S)- 4,5-dihydro-5-(pyridin-4-yl)oxazol-4-yl)methanones VIB 01 174 (0.49 g, 1.87 mmol) in methanol (6.5 ml.) was added hydrochloric acid 37 % (575 μl_). After heating (50 0C) the mixture for 2 h the reaction mixture was concentrated and the crude product was coevaporated twice with EtOAc. After trituration with EtOAc, filtration and drying (2S,3R)- & (2R,3S)-2-amino-1-((R)-3- hydroxypyrrolidin-1-yl)-3-hydroxy-3-(pyhdin-4-yl)propan-1-one dihydrochlorides (420 mg, diastereoisomeric mixture in ratio 1 :1 , 69 % yield) were obtained as a pale pink solid.
Figure imgf000194_0001
2HCI 2HCI threo threo
Compound 247 MW: 324.2; Yield: 69 %; Pale Pink Solid; Mp (0C): 177.0
1H-NMR (CD3OD, δ): 1 .88-2.22 (m, 2H, CH2), 2.70-3.80 (m, 4H, 2xCH2),
4.20-4.65 (m, 2H, CH-O & CH-N), 5.20-5.45 (m, 1 H, CH-O), 8.10-8.25 (m, 2H,
ArH), 8.80-9.00 (m, 2H, ArH), 2xOH & NH2 not seen.
MS-ESI m/z (% rel. Int.): 252.2 ([MH]+, 37), 235.1 (63), 148.0 (100).
Preparation of (-)-f/?reo-2-amino-1 -((S)-3-fluoropyrrolidin-1 -yl)-3-hvdroxy-3- (pyhdin-4-yl)propan-1-one dihydrochloride Compound 248 and (+)-threo-2- amino-1 -((S)-3-fluoropyrrolidin-1 -yl)-3-hvdroxy-3-(pyridin-4-yl)propan-1 -one dihvdrochlohde Compound 249.
Extraction of the free base:
(2S,3R)- & (2R,3S)-2-amino-1 -((S)-3-fluoropyrrolidin-1 -yl)-3-hydroxy-3- (pyhdin-4-yl)propan-1-one dihydrochlorides Compound 214 (300 mg, 0.92 mmol) were dissolved in 10 ml. of a Na2COs (10 %) solution and the aqueous mixture was then saturated with NaCI. The aqueous phase was extracted by 5 x 15 mL of a mixture CH2Cb^-PrOH (9:1 ). The organic phase was dried over MgSO4 and evaporated to afford 163 mg (70 %) of the corresponding free base of Compound 214.
Analytical chiral separation:
20 μl_ of a 1 mg/mL solution of Compound 214 were injected on Chiralpak AD: flow-rate = 1 mL/min, temperature = 25 0C, mobile phase hexane:ethanol = 7:3, detection on UV 220 nm and on polahmeter, first eluted diastereoisomer Compound 248 Rt1 (-) = 20.94 min, second eluted diastereoisomer Rt2(+) = 24.77 min, k1 (-) = 5.93, k2(+) = 7.20, α = 1.21 and resolution Rs = 1 .21. The integration of the UV signal gives 42 % for the first diastereoisomer compound 248 and 58 % for the second Compound 249 (the UV response is different for the two diastereoisomers).
Semi-preparative chiral separation:
170 mg of the free base of Compound 214 were dissolved in 6 mL of ethanol, and 30 μl_ of this solution were injected every 9 min on Chiralpak AD- H, flow-rate = 2 mL/min, mobile phase hexane:ethanol = 7:3, detection on UV 220 nm. 195 successive injections were done. The two main fractions were identified on UV and collected in two different flasks. The solvent was removed in vacuo at 30 0C. The resulting solid was dissolved in 50 mL of CH2CI2 and then filtered on a 0.45 Dm millipore membrane. After evaporation of CH2CI2, the solid was dissolved in 50 mL of methanol and then filtered. For the free base of the first diastereoisomer, a new series of injections was needed to remove two UV-visible impurities collected in the same flask: in the same chromatographic conditions, 30 injections of 100 μL of a 25 mg/mL solution were made every 20 min. The salts Compound 248 and Compound 249 were regenerated according to the same procedure reported for Compound 203 and Compound 204. The regenerated salts Compound 248 and Compound 249 were injected in analytical conditions, the diastereoisomehc excesses for Compound 248 and Compound 249 were determined to be higher than 96 %.
(-)-f/?reo-2-Amino-1-((S)-3-fluoropyrrolidin-1-yl)-3-hvdroxy-3-(pyridin-4- yl)propan-1-one dihydrochloride Compound 248.
Figure imgf000196_0001
Compound 248
MW: 326.19; 70 mg obtained; Pale Yellow Solid; Mp (0C): too hygroscopic. Diastereoisomehc excess > 96 % measured by HPLC at 220 nm (Chiralpak
AD). α25 D = - 2.0 (methanol, c = 1 ).
1H-NMR (CD3OD, δ): 1.85-2.38 (m, 2H, CH2), 2.72-4.05 (m, 4H, 2xCH2), 4.49-
4.62 (m, 1 H, CH-N), 5.10-5.48 (m, 2H, CH-O & CH-F), 8.1 1 -8.25 (m, 2H, ArH), 8.82-8.98 (m, 2H, ArH).
(+)-f/?reo-2-Amino-1-((S)-3-fluoropyrrolidin-1-yl)-3-hydroxy-3-(pyhdin-4- yl)propan-1-one dihvdrochloride Compound 249.
Figure imgf000196_0002
(+)-threo Compound 249
MW: 326.19; 85 mg obtained; Pale Yellow Solid; Mp (0C): too hygroscopic.
Diastereoisomehc excess > 96 % measured by HPLC at 220 nm (Chiralpak
AD).
Q25D = + 31 .7 (methanol, c = 1 ). 1H-NMR (CD3OD, δ): 1.82-2.38 (m, 2H, CH2), 2.90-4.00 (m, 4H, 2xCH2), 4.35- 4.60 (m, 1 H, CH-N), 5.00-5.48 (m, 2H, CH-O & CH-F), 8.1 1 -8.25 (m, 2H, ArH), 8.82-9.00 (m, 2H1 ArH).
Preparation of (±)-f/?reo-2-amino-Λ/-ethyl-3-hvdroxy-Λ/-methyl-3-pyridin-4-yl- propanamide dihydrochloride Compound 250.
frans-Λ/-Ethyl-4,5-dihvdro-Λ/-methyl-5-(pyridin-4-yl)oxazole-4-carboxamide SLA 09190. To a solution of potassium frans-4,5-dihydro-5-(pyhdin-4-yl)oxazole-4- carboxylate EBE 10038A (501 mg, 2.60 mmol) in CH2CI2 (12 mL) were added HOBT (352 mg, 2.60 mmol), EDCI (500 mg, 2.60 mmol) and N- methylethanamine (223 mL, 2.60 mmol). The reaction mixture was stirred 2 h at 0 0C, allowed to warm to room temperature and stirred for 16 h. The reaction mixture was diluted in CH2CI2 (100 mL), washed with brine (2x25 mL), 1 N NaOH (2x25 mL), dried over MgSO4, filtered to give after evaporation frans-Λ/-ethyl-4,5-dihydro-Λ/-methyl-5-(pyridin-4-yl)oxazole-4- carboxamide SLA 09190 (144 mg, 24 % yield) as a pale brown oil.
Figure imgf000197_0001
(±)-trans SLA 09190
MW: 233.27; Yield: 24 %; Pale Brown Oil.
1H NMR (CDCI3, δ): 1.15-1 .30 (m, 3H, CH3), 3.22 (s, 3H, CH3-N), 3.40-3.80 (m, 2H, N-CH2), 4.59 (dd, 1 H, J = 2.2 Hz, J = 7.8 Hz, N-CH), 6.24 (d, J = 7.7 Hz, OCH), 7.02 (d, 1 H, J = 1.0 Hz, N=CH), 7.23 (d, 2H, J = 4.8 Hz, ArH), 8.61 (d, 2H, J = 4.6 Hz, ArH).
MS-ESI m/z (% rel. Int.): 234.2 ([MH]+, 30).
(±)-f/?reo-2-Amino-Λ/-ethyl-3-hvdroxy-Λ/-methyl-3-pyridin-4-yl-propanamide dihvdrochloride Compound 250. To a solution of Λ/-ethyl-4,5-dihydro-Λ/-methyl-5-(pyridin-4-yl)oxazole-4- carboxamide SLA 09190 (144 mg, 0.6 mmol) in MeOH (5 ml.) was added a solution of HCI 37 % (240 μl_). The reaction was stirred for 3 h at 50 0C and concentrated under reduced pressure. The resulting product was dissolved in MeOH and treated with amberlite (OH" form), filtered to give after evaporation a residue that was purified by column chromatography (SiO2, with a gradient of MeOH 10 % in CH2CI2) to yield to 2-amino-Λ/-ethyl-3-hydroxy-Λ/-methyl-3- pyridin-4-yl-propionamide. The hydrochloride salt was formed by treatment of this free base with a solution of HCI 1 M in MeOH (1 ml.) to give after evaporation 2-amino-Λ/-ethyl-3-hydroxy-Λ/-methyl-3-pyhdin-4-yl-propanamide dihydrochloride Compound 250 as a pale yellow solid (80 mg, 44 % yield).
Figure imgf000198_0001
2 HCI (±)
Compound 250 MW: 296.19; Yield: 44 %; Pale Yellow Solid; Mp (0C): 1 14.7 Rf. 0.30 (CH2CI2:Me0H = 90:10, freebase).
1H-NMR (CD3OD,δ): 0.96-1.10 (m, 3H, CH3), 2.81 (s, 1.8H major rotamer, 0.6xCH3), 2.88 (s, 1.2H minor rotamer, 0.4CH3), 3.18-3.55 (m, 2H, CH2), 4.66 (d, 0.4H minor rotamer, J = 6.6 Hz, 0.4xN-CH), 4.69 (d, 0.6H major rotamer, J = 6.3 Hz, 0.6xN-CH), 5.24 (d, 0.4H minor rotamer, J = 6.9 Hz, 0.4xO-CH), 5.27 (d, 0.6H major rotamer, J = 6.3 Hz, 0.6xO-CH), 8.08 (t, 2H, J = 6.5 Hz, ArH), 8.86 (d, 2H, J = 5.0 Hz, ArH).
13C-NMR (CD3OD,δ): 12.0, (13.6), (33.4), 35.3, 44.5, (45.6), 56.1 , (56.2), 71.6, (72.1 ), 126.2 (2χC), 144.5, 144.6, 159.6, 166.5. ( ) Minor rotamer in parenthesis.
Preparation of (2R3S)-2-(3,4-dichlorobenzylamino)-3-hvdroxy-3-(pyridin-4-yl)- 1-(pyrrolidin-1-yl)propan-1 -one dihvdrochloride Compound 251.
To a solution of (-)-(2f?,3S)-2-amino-3-hydroxy-3-(pyridin-4-yl)-1- (pyrrolidin-1-yl)propan-1-one dihydrochloride Compound 203 (175 mg, 0.57 mmol) and Et3N (175 μl_, 1 .25 mmol) in MeOH (5 mL) in a 50 ml. round bottom flask equipped with a magnetic stirrer and under nitrogen atmosphere was added slowly at RT 3,4-dichlorobenzaldehyde (1 12 mg, 0.63 mmol). The reaction mixture was stirred at RT for 20 h. Then AcOH (65 μl_, 1.15 mmol) and NaBH3CN (50 mg, 0.74 mmol) were added. The reaction mixture was stirred at RT for another 15 h. MeOH was evaporated and EtOAc (100 mL) was added. The organic phase was washed with a mixture of saturated sodium carbonate (5 mL) and brine (20 mL), then with brine (10 mL) and dried over MgSO4, filtered and evaporated. The crude product was purified by column chromatography (SiO2, eluent EtOAc:MeOH = 95:5) to give an oil (-)- (2f?,3S)-2-amino-3-hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one (182 mg, 81 % yield). This free base (182 mg, 0.46 mmol) was dissolved in MeOH (2 mL) at 4 0C and a solution of HCI 0.1 N in isopropanol (10.2 mL, 1.01 mmol) was added. After evaporation at 30 0C, a mixture of EtOAc:MeOH = 95:5 was added to yield, after evaporation and drying, to (2f?,3S)-2-(3,4- dichlorobenzylamino)-3-hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 - one dihydrochloride Compound 251 as a white solid (208 mg, 78 % yield).
Figure imgf000199_0001
Compound 251 MW: 467.22; Yield: 78 %; White Solid; Mp (0C): 195.1 Rf: 0.22 (EtOAc:MeOH = 95:5, free base).
1H-NMR (CD3OD, δ): 1.55-1.77 (m, 4H, 2xCH2), 2.46-2.53 (m, 1 H, 0.5xN- CH2), 3.20-3.30 (m, 3H, 1.5xN-CH2), 4.25 (d, 1 H, J = 13.3 Hz, 0.5xN-CH2), 4.38 (d, 1 H, J = 13.3 Hz, 0.5xN-CH2), 4.52 (d, 1 H, J = 7.7 Hz, N-CH), 5.33 (d, 1 H, J = 7.7 Hz, 0-CH), 7.48 (dd, 1 H, J = 8.3 Hz, J = 1.7 Hz, ArH), 7.62 (dd, 1 H, J = 8.3 Hz, J = 1.2 Hz, ArH), 7.74 (s, 1 H, ArH), 8.15 (d, 2H, J = 5.6 Hz, ArH), 8.91 (d, 2H, J = 5.6 Hz, ArH). 13C-NMR (CD3OD,δ): 24.7, 26.6, 47.6, 48.2, 50.4, 64.0, 72.2, 126.8 (2xC), 131.7, 132.2, 132.3, 133.8, 133.9, 135.1 , 143.5 (2xC), 160.6, 163.8. MS-ESI m/z (% rel. Int.): 394.1/396.1/398.1 ([MH]+, 60/45/10), 219.2 (100). HPLC: Method A, detection UV 254 nm, Compound 251 RT = 3.83 min, peak area 99.5 %.
Preparation of (2S,3f?)-2-(3,4-dichlorobenzylamino)-3-hvdroxy-3-(pyhdin-4-yl)- 1-(pyrrolidin-1-yl)propan-1 -one dihvdrochlohde Compound 252.
Same experimental as forCompound 251 preparation starting from (+)- (2S,3f?)-2-amino-3-hydroxy-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochlohde Compound 204 (175 mg, 0.57 mmol). After purification by column chromatography (2S,3f?)-2-(3,4-dichlorobenzylamino)-3-hydroxy-3- (pyhdin-4-yl)-1-(pyrrolidin-1-yl)propan-1-one was obtained (187 mg, 83 % yield). This free base (187 mg, 0.47 mmol) was dissolved in MeOH (2 ml.) at 4 0C and a solution of HCI 0.1 N in isopropanol (10.4 ml_, 1 .04 mmol) was added. After evaporation at 30 0C, a mixture of EtOAc:MeOH = 95:5 was added to yield, after evaporation and drying, to Compound 252 as a white solid (212 mg, 80 % yield).
Figure imgf000200_0001
Compound 252
MW: 467.22; Yield: 80 %; White Solid; Mp (0C): 187.5 Rf. 0.22 (EtOAc:MeOH = 95:5, free base).
1H-NMR (CDCI3,δ): 1 .55-1.77 (m, 4H, 2xCH2), 2.46-2.53 (m, 1 H, 0.5xN-CH2), 3.20-3.30 (m, 3H, 1 .5xN-CH2), 4.25 (dd, 1 H, J = 13.3 Hz, 0.5xN-CH2), 4.40 (d, 1 H, J = 13.3 Hz, 0.5xN-CH2), 4.52 (d, 1 H, J = 7.7 Hz, N-CH), 5.33 (d, 1 H, J = 7.4 Hz, 0-CH), 7.48 (d, 1 H, J = 8.3 Hz, ArH), 7.62 (dd, 1 H, J = 8.3 Hz, J = 0.8 Hz, ArH), 7.74 (s, 1 H, ArH), 8.15 (d, 2H, J = 5.7 Hz, ArH) 8.91 (d, 2H, J = 5.7 Hz, ArH).
13C-NMR (CD3OD, δ): 24.7, 26.6, 47.6, 48.2, 50.4, 64.0, 72.3, 126.8 (2xC), 131.7, 132.2, 132.3, 133.8, 133.9, 135.1 , 143.5 (2xC), 160.6, 163.8. MS-ESI m/z (% rel. Int.): 394.1/396.1/398.1 ([MH]+, 60/45/10), 219.2 (100). HPLC: Method A, detection UV 254 nm, Compound 252 RT = 3.83 min, peak area 99.5 %.
Preparation of (£)-3-(pyridin-4-yl)-1 -(pyrrolidin-1 -yl)prop-2-en-1 -one hydrochloride Compound 253.
To a solution of 3-(4-pyhdinyl)acrylic acid ( 1 .01 g, 6.77 mmol) in CHCI3 (20 ml.) in a 100 ml. round bottom flask equipped with a magnetic stirrer and under nitrogen atmosphere was added 1-hydroxybenzothazole (1.1 1 g, 8.21 mmol). The reaction mixture was stirred at RT for 10 min. Then 1-(3- dimethylaminopropyl)-3-ethylcarbodiimide (1.56 g, 8.15 mmol) was added. The reaction mixture was stirred at 4 0C for 10 min. Then pyrrolidine (1.1 1 ml_, 18.3 mmol) was added slowly and the reaction mixture was stirred for 15 h at +4 0C to RT. Dichloromethane (200 mL) was added and organic phase was washed with brine (100 mL), a solution of NaOH 0.5 N (100 mL) and brine (50 mL). The organic phase was dried over MgSO4, filtered, and evaporated to obtain (£)-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 -yl)prop-2-en-1 -one (1.30 g, 94 % yield). This free base (1.3 g, 6.40 mmol) was dissolved in MeOH (10 mL) at 4 0C and a solution of HCI 0.1 N in isopropanol (79 mL, 7.9 mmol) was added. After evaporation at 30 0C and drying, (£)-3-(pyhdin-4-yl)-1 -(pyrrolidin-1 - yl)prop-2-en-1-one hydrochloride Compound 253 was obtained as a beige solid (1 .40 g, 87 % yield).
Figure imgf000201_0001
Compound 253
MW: 238.71 ; Yield: 87 %; Beige Solid; Mp (0C): 229.4 Rf. 0.35 (EtOAc : MeOH = 95:5, free base).
1H-NMR (CD3OD, δ): 1.92-2.10 (m, 4H, 2xCH2), 3.56 (t, 2H, J = 6.7 Hz, N-
CH2), 3.80 (t, 2H, J = 6.7 Hz, N-CH2), 7.59 (d, 1 H, J = 15.6 Hz, CH=C), 7.68
(d, 1 H, J = 15.6 Hz, CH=C), 8.35 (d, 2H, J = 5.7 Hz, ArH) 8.86 (d, 2H, J = 5.6 Hz, ArH).
13C-NMR (CD3OD, δ): 25.2, 27.0, 47.6, 48.2, 126.7 (2xC), 131 .9, 136.5, 143.1
(2xC), 154.5, 164.6.
MS-ESI m/z (% rel. Int.): 203.2 ([MH]+, 100).
HPLC: Method A, detection UV 254 nm, Compound 253 RT = 3.18 min, peak area 99.5 %.
Preparation of (±)-f/?reo-2-amino-3-hydroxy-3-(1 -methyl-1 /-/-imidazol-2-yl)-1 - (pyrrolidin-1-yl)propan-1-one dihydrochloride Compound 254.
frans-(4,5-Dihydro-5-(1 -methyl-1 /-/-imidazol-2-yl)oxazol-4-yl)(pyrrolidin-1 - vDmethanone LPO 01 190B.
To a stirred and cooled (0 0C) solution of potassium hydroxide (0.33 g, 5.0 mmol) in MeOH (6 mL) were added a mixture of 1 -methyl-2-imidazole carboxaldehyde (0.56 g, 5.0 mmol) and 2-isocyano-1-(pyrrolidin-1- yl)ethanone BLE 04098 (0.70 g, 5.0 mmol). The solution was stirred 3 h at 4 0C and then concentrated. The residue was partitioned between EtOAc (100 mL) and water (20 mL). The organic layer was washed with brine (10 mL) and dried over MgSO4, filtered and evaporated. Concentration afforded a crude product which was purified by column chromatography (florisil, EtOAc:MeOH = 95:5 to 90:10) to yield, after evaporation and drying, to frans-(4,5-dihydro-5- (1 -methyl-1 /-/-imidazol-2-yl)oxazol-4-yl)(pyrrolidin-1 -yl)methanone LPO 01 19OB (0.32 g, 25 % yield) as a brown oil.
Figure imgf000202_0001
(±)-trans LPO 01 190B MW: 248.28; Yield: 25 %; Brown Oil.
Rf\ 0.30 (EtOAc:MeOH = 9:1 , free base).
1H-NMR (CDCI3, δ): 1 .94-2.12 (m, 4H, 2xCH2), 3.50 (t, 2H, J = 6.5 Hz, N-
CH2), 3.69-4.13 (m, 5H, N- CH2 ,N-CH3), 5.68 (dd, 1 H, J = 7.7 Hz, J = 2.3 Hz, CH-N), 6.19 (d, 1 H, J = 7.7 Hz, CH-O), 6.82 (d, 1 H, J = 2.2 Hz, CH=N), 6.94
(d, 1 H, J = 1.1 Hz, ArH), 7.00 (d, 1 H, J = 1 .1 Hz, ArH).
13C-NMR (CDCI3, δ): 24.2, 26.0, 32.9, 46.4, 46.7, 71.2, 72.7, 123.0, 127.9,
143.7, 153.7, 166.6.
MS-ESI m/z (% rel. Int.): 267.3 ([MH+18]+, 10), 196.2 (100). HPLC: Method A, detection UV 254 nm, LPO 01 19OB RT = 3.92 min, peak area 99.5 %.
(±)-f/?reo-2-Amino-3-hydroxy-3-(1 -methyl-1 /-/-imidazol-2-yl)-1 -(pyrrolidin-1 - yl)propan-1-one dihydrochloride Compound 254. A solution of frans-(4,5-dihydro-5-(1 -methyl-1 /-/-imidazol-2-yl)oxazol-4- yl)(pyrrolidin-1 -yl)methanone LPO 01 19OB (320 mg, 1.29 mmol) and HCI 37 % (0.4 mL, 13 mmol) in MeOH (6 mL) was stirred at 50 0C for 3 h in a 50 mL round bottom flask. The solvent was evaporated and the product was precipitated by a mixture of MeOH: EtOAc: Et2O = 3:12:5 (20 mL). Solvents were evaporated at 30 0C to give, after evaporation and drying, (±)-threo-2- amino-3-hydroxy-3-(1 -methyl-1 /-/-imidazol-2-yl)-1 -(pyrrolidin-1 -yl)propan-1 -one dihydrochloride Compound 254 as a pale yellow solid (305 mg, 76 % yield).
Figure imgf000203_0001
2.HCI (±ythreo
Compound 254 MW: 31 1.21 ; Yield: 76 %; Pale Yellow Solid; Mp (0C): 183.4 Rf\ 0.30 (CH2CI2:Me0H = 95:5, free base).
1H-NMR (CD3OD, δ): 1.75-1.99 (m, 4H, 2xCH2), 2.80-2.88 (m, 1 H, 0.5xN- CH2), 3.30-3.70 (m, 3H, 1.5xN-CH2), 3.95 (s, 3H, N-CH3), 4.62 (d, 1 H, J = 8.3 Hz, N-CH), 5.51 (d, 1 H, J = 8.3 Hz, 0-CH), 7.66 (s, 2H, ArH). 13C-NMR (CD3OD, δ): 24.9, 26.9, 36.0, 47.9 (2xC), 56.2, 65.1 , 121.1 , 126.5,
145.1 , 164.4.
MS-ESI m/z (% rel. Int.): 239.3 ([MH]+, 10), 134.1 (100).
HPLC: Method A, detection UV 254 nm, RT = 0.8 min, peak area 99.5 %.
Preparation of (2S,3f?)- & (2R3S)-2-amino-1-((S)-3-fluoropyrrolidin-1-yl)-3- hvdroxy-3-(pyhdin-3-yl)propan-1 -one dihydrochlorides Compound 255.
frans-((S)-3-fluoropyrrolidin-1 -yl)((4S.5f?)- & (4f?.5S)-4.5-dihvdro-5-(pyridin-4- yl)oxazol-4-yl)methanones SLA 1 1014.
To a stirred and cooled 0 0C solution of KOH (0.216 mg, 4.22 mmol) in methanol (5 mL) was added 1-((S)-3-fluoropyrrolidin-1-yl)-2- isocyanoethanone VIB 01 166 (0.600 g, 4.22 mmol) and pyridine-3- carbaldehyde (0.40 mL, 3.84 mmol). The solution was stirred for 20 h at 0 0C. After evaporation under reduced pressure, the residue obtained was partitioned between EtOAc and H2O. The product was extracted with EtOAc (4x50 mL) and washed with brine (25 mL), dried over MgSO4, filtered and evaporated to yield to a product that was purified using chromatography (florisil, EtOAc:MeOH = 95:5), frans-((S)-3-fluoropyrrolidin-1-yl)((4S,5R)- & (4f?,5S)-4,5-dihydro-5-(pyridin-4-yl)oxazol-4-yl)methanones SLA 1 1014 were obtained as a yellow solid (0.464 g, diastereoisomehc mixture in ratio about 1 :1 , 46 % yield).
Figure imgf000204_0001
SLA 1 1014 MW: 263.27; Yield: 46 %; Yellow Solid; Mp (0C) = 171 .7 Rf. 0.25 (EtOAc:MeOH = 95:5).
1H-NMR (CDCI3, δ): 1 .85-2.45 (m, 2H, CH2), 3.50-4.10 (m, 3H, CH2 & N-CH), 4.25-4.65 (m, 2H, N-CH2), 5.15-5.25 (m, 0.5H, O.δxCHF), 5.35-5.45 (m, 0.5H, 0.5xCHF), 6.21 (d, 1 H, J = 7.6 Hz, O-CH), 7.04 (d, 1 H, J = 2.1 Hz, N=CH), 7.30-7.38 (m, 1 H, ArH), 7.60-7.68 (m, 1 H, ArH), 8.55-8.65 (m, 2H, ArH). MS-ESI m/z (% rel. Int.): 264.1 ([MH]+, 18).
(2S,3f?)- & (2R3S)-2-Amino-1 -((S)-3-fluoropyrrolidin-1 -yl)-3-hvdroxy-3- (pyridin-3-yl)propan-1-one dihydrochlorides Compound 255.
To a solution of frans-((S)-3-fluoropyrrolidin-1-yl)((4S,5R)- & (4R,5S)- 4,5-dihydro-5-(pyridin-4-yl)oxazol-4-yl)methanones SLA 1 1014 (0.450 g, 1.71 mmol) in methanol (40 mL) was added HCI 37 % (5 ml_). After heating at 50 0C for 3 h, the mixture was concentrated and the crude product was co- evaporated twice with EtOAc. Trituration with EtOAc and filtration yielded, after drying, to (2S,3R)- & (2R,3S)-2-amino-1-((S)-3-fluoropyrrolidin-1-yl)-3- hydroxy-3-(pyhdin-3-yl)propan-1 -one dihydrochlorides Compound 255 (540 mg, diastereoisomeric mixture in ratio about 1 :1 , 97 % yield) as a yellow solid.
Figure imgf000205_0001
Compound 255
MW: 326.19; Yield: 97 %; Yellow Solid; Mp (0C): 168.9
1H-NMR (CD3OD, δ): 1.85-2.40 (m, 2H, CH2), 3.45-4.20 (m, 4H, 2xCH2), 4.40- 4.75 (m, 1 H, N-CH), 5.30-5.60 (m, 2H, O-CH & CHF), 8.15-8.25 (m, 1 H, ArH), 8.70-8.80 (m, 1 H, ArH), 8.90-9.10 (m, 2H, ArH). MS-ESI m/z (% rel. Int.): 254.1 ([MH]+, 81 .38), 236.2 (25).
((S)-3-fluoropyrrolidin-1 -yl)((4S.5f?)- & (4R5S)-4.5-dihvdro-5-(thiophen-3- yl)oxazol-4-yl)methanones SLA 1 1016. To a stirred and cooled 0 0C solution of KOH (0.216 mg, 3.85 mmol) in methanol (8 mL) was added 1-((S)-3-fluoropyrrolidin-1-yl)-2- isocyanoethanone VIB 01 166 (0.600 g, 4.22 mmol) and thiophene-3- carbaldehyde (0.37 mL, 3.85 mmol). The solution was stirred for 20 h at 0 0C. After evaporation under reduced pressure, the residue obtained was partitioned between EtOAc and H2O. The product was extracted with EtOAc (4x50 mL) and washed brine (25 ml_), dried over MgSO4, filtered and evaporated to yield to a product that was purified using chromatography (florisil, gradient EtOAc:MeOH = 95:5 to 85:15), ((S)-3-fluoropyrrolidin-1 - yl)((4S,5R)- and (4R,5S)-4,5-dihydro-5-(thiophen-3-yl)oxazol-4-yl)methanones SLA 1 1016 were obtained as a yellow solid (0.41 1 g, diastereoisomeric mixture in ratio about 1 :1 , 32 % yield) .
Figure imgf000206_0001
Figure imgf000206_0002
trans
SLA 1 1016 MW: 268.31 ; Yield: 32 %; Yellow Solid; Mp (0C) = 132.9 Rf. 0.35 (EtOAc:MeOH = 80:20).
1H-NMR (CDCI3, δ): 1 .80-2.45 (m, 2H, CH2), 3.50-4.10 (m, 3H, CH-N & CH2), 4.20-4.70 (m, 2H, CH2), 5.15-5.45 (m, 1 H, CHF), 6.18-6.25 (m, 1 H, 0-CH), 6.99 (d, 1 H, J = 2.2 Hz, N=CH), 7.00-7.15 (m, 1 H, ArH), 7.28-7.35 (m, 1 H, ArH), 7.32-7.40 (m, 1 H1 ArH).
MS-ESI m/z (% rel. Int.): 269.0 ([MH]+, 10).
(2S,3f?)- & (2R3S)-2-Amino-1 -((S)-3-fluoropyrrolidin-1 -yl)-3-hvdroxy-3- (thiophen-3-yl)propan-1-one dihvdrochlorides Compound 256. To a solution of ((S)-3-fluoropyrrolidin-1-yl)((4S,5R)- & (4R,5S)-4,5- dihydro-5-(thiophen-3-yl)oxazol-4-yl)methanones SLA 11016 (0.400 g, 1.49 mmol) in methanol (50 mL) was added hydrochloric acid 37 % (4 mL). After heating at 50 0C for 3 h, the mixture was concentrated and the crude product was co-evaporated twice with EtOAc. Trituration with EtOAc and filtration and drying afforded (2S,3R)- & (2R,3S)-2-amino-1-((S)-3-fluoropyrrolidin-1-yl)-3- hydroxy-3-(thiophen-3-yl)propan-1-one dihydrochlohdes Compound 256 (451 mg, diastereoisomeric mixture about 1 :1 , 91 % yield) as a yellow solid.
Figure imgf000207_0001
Compound 256
MW: 331.23; Yield: 91 %; Yellow Solid; Mp (0C): 221.6 1H-NMR (CD3OD, δ): 1.25-2.05 (m, 2H, CH2), 2.10-2.50 (m, 1 H, 0.5xCH2), 3.20-3.65 (m, 3H, 1 .5xCH2), 3.90-4.10 (m, 1 H, CH-N), 4.70-5.10 (m, 2H, O- CH & CHF), 6.92-6.99 (m, 1 H, ArH), 7.21-7.32 (m, 2H, ArH). MS-ESI m/z (% rel. Int.): 259.1 ([MH]+, 25).

Claims

What is claimed is:
1. The use of a compound in the manufacture of a medicament for treating a condition selected from the group consisting of neuropathic pain, inflammatory pain, headache pain, somatic pain, visceral pain, and referred pain, wherein the compound has the following structure:
Figure imgf000208_0001
Formula 1 where Ri is H or alkyl of 1 to 6 carbons,
R2 is H, alkyl of 1 to 6 carbons or the Ri and R2 groups together with the nitrogen form a saturated or unsaturated 4, 5, 6 or 7 membered ring that optionally includes one or two heteroatoms independently selected from N, O and S, said 4, 5, 6 or 7 membered ring optionally being substituted with one or two COOH, CH2OH, OH, B(OH)2, halogen groups, cyano groups, or with one or two alkyl groups having 1 to 6 carbons, or one or two carbons of said rings being attached to an oxygen to form keto groups and said 4, 5, 6 or 7 membered ring optionally being condensed with an aromatic or non-aromatic 5 or 6 membered ring that optionally includes 1 or heteroatoms selected from N, O and S; R3 is independently selected from H, alkyl of 1 to 20 carbons, cycloalkyl of 3 to 6 carbons, aryl or heteroaryl, aryl-alkyl where the alkyl moiety has 1 to 4 carbons, aryl-(hydroxy)alkyl where the alkyl moiety has 1 to 4 carbons,, heteroaryl-alkyl where the alkyl moiety has 1 to 4 carbons, or hetero- (hydroxy)alkyl where the alkyl moiety has 1 to 4 carbons, said aryl or heteroaryl groups being optionally substituted with 1 to 3 groups independently selected from the group consisting of halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 3 carbons, or R3 is CO-
R7, SO2R7 or CO-O-R7 where R7 is H, alkyl of 1 to 1 to 20 carbons, alkyl of 1 to 20 carbons substituted with and NH2 group or with an NH-COalkyl group where the alkyl group has one to 6 carbons, aryl or heteroaryl, aryl-alkyl where the alkyl moiety has 1 to 4 carbons or heteroaryl-alkyl where the alkyl moiety has 1 to 4 carbons, said aryl or heteroaryl groups being optionally substituted with 1 to 3 groups independently selected from the group consisting of halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 3 carbons; R4 is H, alkyl of 1 to 6 carbons or CO-Rs where Rs is alkyl of 1 to 6 carbons; the wavy lines represent bonds connected to carbons having R or S configuration; the dashed lines represent a bond or absence of a bond with the proviso that the ring containing the dashed lines is aromatic; m, n and q are integers independently selected from O, 1 , 2 or 3 with the proviso that the sum of m, n and q is 2 or 3; s is zero (O) or when X is N then s is zero (O) or 1 ;
W, X and Y independently represent a CH, CR5, CRε or a heteroatom selected independently of N, O and S, and R5 and Re are independently selected from H, halogen, alkyl of 1 to 6 carbons, halogen substituted alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 3 carbons, phenyl, or
R5 and Re together with the atoms to which they are attached jointly form a carbocyclic or a heterocyclic ring, the carbocyclic ring having 5 or 6 atoms in the ring, the heterocyclic ring having 5 or 6 atoms in the ring and 1 to 3 heteroatoms independently selected from N, O and S; said carbocyclic or heterocyclic ring jointly formed by R5 and Re being optionally substituted with 1 to 6 Rg groups where Rg is independently selected from halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 3 carbons or a pharmaceutically acceptable salt of said compound with the proviso that Formula 1 does not cover compounds where R4 is H, Ri and R2 jointly with the nitrogen form a pyrrolidino or morpholino ring, the sum of m, n and q is 3, and where none of W, X and Y represent a heteroatom with the further proviso that the formula does not cover the compounds of the formula below
Figure imgf000210_0001
(±)-erythro
2. The use of claim 1 , wherein the sum of the integers m, n and q is 3.
3. The use of claim 2, wherein one of the W, X and Y groups is N.
4. The use of Claim 3, wherein R5 and R& together with the atoms to which they are attached jointly form a carbocyclic or a heterocyclic ring.
5. The use of Claim 2, wherein none of the W, X and Y groups is a heteroatom.
6. The use of Claim 5 wherein R5 and R& together with the atoms to which they are attached jointly form a carbocyclic or a heterocyclic ring.
7. The use of Claim 1 , wherein the sum of the integers m, n and q is 2 and at least one of W, X and Y is a heteroatom.
8. The use of Claim 7, wherein only one of W, X and Y represents a heteroatom.
9. The use of Claim 7, wherein two of the W, X and Y groups each represent an independently selected heteroatom.
10. The use of Claim 1 , wherein R4 is H.
11. The use of Claim 10, wherein both R3 groups are H.
12. The use of Claim 10, wherein one R3 group is H and the other R3 group is CO-R7 or alkyl of 1 to 20 carbons.
13. The use of Claim 10, wherein R3 is independently selected from H and alkyl of 1 to 10 carbons.
14. The use of Claim 1 , wherein R4 is CO-Rs.
15. The use of Claim 1 , wherein Ri and R2 together with the nitrogen form a 4, 5, 6 or 7 membered ring.
16. The use of Claim 15, wherein Ri and R2 together with the nitrogen form a
5 membered ring.
17. The use of Claim 15, wherein Ri and R2 together with the nitrogen form a
6 membered ring.
18. The use of Claim 1 , wherein both R5 and R& are hydrogen.
19. The use of a compound in the manufacture of a medicament for treating a condition selected from the group consisting of neuropathic pain, inflammatory pain, headache pain, somatic pain, visceral pain, and referred pain, wherein the compound has the following structure:
Figure imgf000211_0001
where Ri is H or alkyl of 1 to 6 carbons,
R2 is H, alkyl of 1 to 6 carbons or the Ri and R2 groups together with the nitrogen form a saturated or unsaturated 4, 5, 6 or 7 membered ring that optionally includes one or two heteroatoms independently selected from N, O and S, said 4, 5, 6 or 7 membered ring optionally being substituted with one or two COOH, CH2OH, OH, B(OH)2, halogen groups, cyano groups or with one or two alkyl groups having 1 to 6 carbons; R3 is H, CO-R7 or CO-O-R7 where R7 is H, alkyl of 1 to 1 to 20 carbons, cycloalkyl of 3 to 6 carbons, aryl or heteroaryl, aryl-alkyl, aryl(hydroxy)alkyl, heteroaryl-alkyl or heteroalkyl(hydroxy)alkyl where the alkyl moiety has 1 to 4 carbons, said aryl or heteroaryl groups being optionally substituted with 1 to 3 groups independently selected from the group consisting of halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 3 carbons;
R4 is H, alkyl of 1 to 6 carbons, or CO-Rs where Rs is alkyl of 1 to 6 carbons, R5 and Rε are independently selected from H, halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 3 carbons, or Rs and Re together with the atoms to which they are attached jointly form a carbocyclic or a heterocyclic ring, the carbocyclic ring having 5 or 6 atoms in the ring, the heterocyclic ring having 5 or 6 atoms in the ring and 1 to 3 heteroatoms independently selected from N, O and S; said carbocyclic or heterocyclic ring jointly formed by Rs and R& being optionally substituted with 1 to 6 Rg groups where Rg is independently selected from halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons, and the wavy lines represent bonds of the alpha or beta configuration, or any pharmaceutically acceptable salt of said compound.
20. The use of Claim 19, wherein R4 is H.
21. The use of Claim 19, wherein both R3 groups are H.
22. The use of Claim 19, wherein both R5 and R& are hydrogen.
23. The use of Claim 22, wherein the compound has the formula
Figure imgf000212_0001
or a pharmaceutically acceptable salt of said compound.
24. The use of Claim 23, wherein R4 is H.
25. The use of Claim 23, wherein both R3 groups are H.
26. The use of Claim 23, wherein the compound has the formula
Figure imgf000212_0002
or a pharmaceutically acceptable salt of said compound.
27. The use of Claim 23, wherein the compound has the formula
Figure imgf000212_0003
or a pharmaceutically acceptable salt of said compound.
28. The use of Claim 23, wherein the compound has the formula
Figure imgf000213_0001
2.HCI (±)-threo
or any other pharmaceutically acceptable salt of said compound.
29. The use of Claim 23, wherein the compound has the formula
Figure imgf000213_0002
2. HCI
or any other pharmaceutically acceptable salt of said compound.
30. The use of Claim 23, wherein the compound has the formula
Figure imgf000213_0003
or any other pharmaceutically acceptable salt of said compound.
31. The use of Claim 23, wherein the compound has the formula
Figure imgf000213_0004
2.HCI threo or any other pharmaceutically acceptable salt of said compound.
32. The use of Claim 23, wherein the compound has the formula
Figure imgf000213_0005
2.HCI or any other pharmaceutically acceptable salt of said compound.
33. The use of Claim 23, wherein the compound has the formula
Figure imgf000214_0001
or any other pharmaceutically acceptable salt of said compound.
34. The use of Claim 23, wherein the compound has the formula
Figure imgf000214_0002
2.HCI (±ythreo or any other pharmaceutically acceptable salt of said compound.
35. The use of Claim 23, wherein the compound has the formula
Figure imgf000214_0003
or any other pharmaceutically acceptable salt of said compound.
36. The use of Claim 23, wherein the compound has the formula
Figure imgf000214_0004
(-ythreo or any other pharmaceutically acceptable salt of said compound.
37. The use of Claim 23, wherein the compound has the formula
Figure imgf000214_0005
(±)-erythro or a pharmaceutically acceptable salt of said compound.
38. The use of Claim 19, wherein the compound has the formula
Figure imgf000215_0001
2. HCI (±)-erythro or any other pharmaceutically acceptable salt of said compound.
39. The use of Claim 19, wherein the compound has the formula
Figure imgf000215_0002
2.HCI (±)-threo or any other pharmaceutically acceptable salt of said compound.
40. The use of Claim 19, wherein the compound has the formula
Figure imgf000215_0003
or any other pharmaceutically acceptable salt of said compound.
41. The use of Claim 19, wherein the compound has the formula
Figure imgf000215_0004
or any other pharmaceutically acceptable salt of said compound.
42. The use of Claim 19, wherein the compound has the formula
Figure imgf000215_0005
2. HCI (±)-threo or any other pharmaceutically acceptable salt of said compound.
43. The use of Claim 19, wherein the compound has the formula
Figure imgf000216_0001
or any other pharmaceutically acceptable salt of said compound.
44. The use of Claim 19, wherein the compound has the formula
Figure imgf000216_0002
(±ythreo or any other pharmaceutically acceptable salt of said compound.
45. The use of Claim 19, wherein the compound has the formula
Figure imgf000216_0003
or any other pharmaceutically acceptable salt of said compound.
46. The use of Claim 45, wherein the compound has the formula
Figure imgf000216_0004
or any other pharmaceutically acceptable salt of said compound.
47. The use of Claim 45, wherein the compound has the formula
Figure imgf000217_0001
or any other pharmaceutically acceptable salt of said compound.
48. The use of a compound in the manufacture of a medicament for treating a condition selected from the group consisting of neuropathic pain, inflammatory pain, headache pain, somatic pain, visceral pain, and referred pain, wherein the compound has the following structure:
Figure imgf000217_0002
where Ri is H or alkyl of 1 to 6 carbons,
R2 is alkyl of 1 to 6 carbons or the Ri and R2 groups together with the nitrogen form a saturated or unsaturated 4, 5, 6 or 7 membered ring that optionally includes one or two heteroatoms independently selected from N, O and S, said 4, 5, 6 or 7 membered ring optionally being substituted with one or two COOH, CH2OH, OH, B(OH)2, halogen groups, cyano groups, or with one or two alkyl groups having 1 to 6 carbons, R3 is H, CO-R7 or CO-O-R7 where R7 is H, alkyl of 1 to 1 to 20 carbons, aryl or heteroaryl, cycloalkyl of 3 to 6 carbons, aryl-alkyl aryl(hydroxy)alkyl, heteroaryl-alkyl or heteroaryl(hydroxy)alkyl where the alkyl moiety has 1 to 4 carbons, said aryl or heteroaryl groups being optionally substituted with 1 to 3 groups independently selected from the group consisting of halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 3 carbons;
R4 is H, alkyl of 1 to 6 carbons, or CO-Rs where Rs is alkyl of 1 to 6 carbons; R5 and Rε are independently selected from H, halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons, thioxy of 1 to 3 carbons and phenyl, or Rs and Re together with the atoms to which they are attached jointly form a carbocyclic or a heterocyclic ring, the carbocyclic ring having 5 or 6 atoms in the ring, the heterocyclic ring having 5 or 6 atoms in the ring and 1 to 3 heteroatoms independently selected from N, O and S; said carbocyclic or heterocyclic ring jointly formed by Rs and R& being optionally substituted with 1 to 6 Rg groups where Rg is independently selected from halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 3 carbons, and the wavy lines represent bonds connected to carbons having R or S of configuration, with the proviso that the claim does not cover compounds where R4 is H, Ri and R2 jointly with the nitrogen form a pyrrolidino or morpholino ring, or any pharmaceutically acceptable salt of said compound.
49. The use of Claim 48, wherein the compound has the formula
Figure imgf000218_0001
(±)-threo or any other pharmaceutically acceptable salt of said compound.
50. The use of a compound in the manufacture of a medicament for treating a condition selected from the group consisting of neuropathic pain, inflammatory pain, headache pain, somatic pain, visceral pain, and referred pain, wherein the compound has the following structure:
Figure imgf000218_0002
where X is O or S; Ri is H or alkyl of 1 to 6 carbons,
R2 is H, alkyl of 1 to 6 carbons or the Ri and R2 groups together with the nitrogen form a saturated or unsaturated 5 or 6 membered ring that optionally includes one or two heteroatoms independently selected from N, O and S, said 4, 5, 6 or 7 membered ring optionally being substituted with one or two COOH, CH2OH, OH, B(OH)2, halogen groups or cyano groups with one or two alkyl groups having 1 to 6 carbons, R3 is H, CO-R7 or CO-O-R7 where R7 is H, alkyl of 1 to 1 to 20 carbons, aryl or heteroaryl, aryl-alkyl, aryl(hydroxy)alkyl, heteroaryl-alkyl or heteroaryl(hydroxy)alkyl where the alkyl moiety has 1 to 4 carbons, said aryl or heteroaryl groups being optionally substituted with 1 to 3 groups independently selected from the group consisting of halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 3 carbons; R4 is H, alkyl of 1 to 6 carbons, or CO-Rs where Rs is alkyl of 1 to 6 carbons; R5 and Rε are independently selected from H, halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to3 carbons, or R5 and Rε together with the atoms to which they are attached jointly form a carbocyclic or a heterocyclic ring, the carbocyclic ring having 5 or 6 atoms in the ring, the heterocyclic ring having 5 or 6 atoms in the ring and 1 to 3 heteroatoms independently selected from N, O and S; said carbocyclic or heterocyclic ring jointly formed by R5 and R& being optionally substituted with 1 to 6 Rg groups where Rg is independently selected from halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons, and the wavy lines represent bonds connected to carbons having R or S configuration, or any pharmaceutically acceptable salt of said compound with the further proviso that the claim does not cover the compound shown below
Figure imgf000219_0001
51. The use of Claim 50, wherein the compound has the formula
Figure imgf000219_0002
or a pharmaceutically acceptable salt of said compound.
52. The use of Claim 51 , wherein the compound has the formula
Figure imgf000220_0001
(±)-erythro or a pharmaceutically acceptable salt of said compound.
53. The use of Claim 51 , wherein the compound has the formula
Figure imgf000220_0002
HCl (±)-threo or any other pharmaceutically acceptable salt of said compound.
54. The use of Claim 53, wherein the compound has the formula
Figure imgf000220_0003
(+ ythreo or any other pharmaceutically acceptable salt of said compound.
55. The use of Claim 53, wherein the compound has the formula
Figure imgf000220_0004
56. The use of a compound in the manufacture of a medicament for treating a condition selected from the group consisting of neuropathic pain, inflammatory pain, headache pain, somatic pain, visceral pain, and referred pain, wherein the compound has the following structure:
Figure imgf000221_0001
Formula 1 where Ri is H or alkyl of 1 to 6 carbons,
R2 is H, alkyl of 1 to 6 carbons or the Ri and R2 groups together with the nitrogen form a saturated or unsaturated 4, 5, 6 or 7 membered ring that optionally includes one or two heteroatoms independently selected from N, O and S, said 4, 5, 6 or 7 membered ring optionally being substituted with one or two COOH, CH2OH, OH, B(0H)2, halogen or cyano groups groups or with one or two alkyl groups having 1 to 6 carbons, or one or two carbons of said rings being attached to an oxygen to form keto groups and said 4, 5, 6 or 7 membered ring optionally being condensed with an aromatic or non-aromatic
5 or 6 membered ring that optionally includes 1 or heteroatoms selected from N, O and S;
R3 is independently selected from H, alkyl of 1 to 20 carbons, cycloalkyl of 3 to
6 carbons, aryl or heteroaryl, aryl-alkyl where the alkyl moiety has 1 to 4 carbons, aryl-(hydroxy)alkyl where the alkyl moiety has 1 to 4 carbons, heteroaryl-alkyl where the alkyl moiety has 1 to 4 carbons or hetero- (hydroxy)alkyl where the alkyl moiety has 1 to 4 carbons, said aryl or heteroaryl groups being optionally substituted with 1 to 3 groups independently selected from the group consisting of halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 3 carbons, or R3 is CO- R7, SO2R7 or CO-O-R7 where R7 is H, alkyl of 1 to 1 to 20 carbons, alkyl of 1 to 20 carbons substituted with and NH2 group or with an NH-COalkyl group where the alkyl group has one to 6 carbons aryl or heteroaryl, aryl-alkyl where the alkyl moiety has 1 to 4 carbons or heteroaryl-alkyl where the alkyl moiety has 1 to 4 carbons, said aryl or heteroaryl groups being optionally substituted with 1 to 3 groups independently selected from the group consisting of halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 3 carbons; the wavy lines represent bonds connected to carbons having R or S configuration; the dashed lines represent a bond or absence of a bond with the proviso that the ring containing the dashed lines is aromatic;
Rg and R-io are one of the combinations selected from the group consisting of
(1 ) Rg is NOR11 and R10 does not exist, (2) Rg is ORn and R10 is alkyl of 1 to 6 carbons, (3) when the dashed lines between carbons 2 and 3 of the propionic acid moiety represent a bond then Rg is H or alkyl of 1 to 6 carbons and R-io does not exist;
R11 is H, alkyl of 1 to 6 carbons or CO-R-12 where R12 is alkyl of 1 to 6 carbons; m, n and q are integers independently selected from O, 1 , 2 or 3 with the proviso that the sum of m, n and q is 2 or 3; s is zero (0) or when X is N then s is zero (0) or 1 ;
W, X and Y independently represent a CH, CR5, CRε or a heteroatom selected independently of N, O and S, and
R5 and Rε are independently selected from H, halogen, alkyl of 1 to 6 carbons, halogen substituted alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 3 carbons, phenyl, or
R5 and Rε together with the atoms to which they are attached jointly form a carbocyclic or a heterocyclic ring, the carbocyclic ring having 5 or 6 atoms in the ring, the heterocyclic ring having 5 or 6 atoms in the ring and 1 to 3 heteroatoms independently selected from N, O and S; said carbocyclic or heterocyclic ring jointly formed by R5 and Re being optionally substituted with 1 to 6 R12 groups where R12 is independently selected from halogen, alkyl of 1 to 6 carbons, alkoxy of 1 to 6 carbons and thioxy of 1 to 3 carbons or a pharmaceutically acceptable salt of said compound.
57. The use of Claim 56, wherein the sum of the integers m, n and q is 3.
58. The use of Claim 56, wherein one of the W, X and Y groups is N.
59. The use of Claim 56, wherein the Ri and R2 groups together with the nitrogen form a 5-membered ring.
60. The use of Claim 56, wherein Rg is NORn and R-io does not exist.
61. The use of Claim 56, wherein Rg is methyl and R-io is ORn.
62. The use of Claim 56, wherein the dashed line between carbons 2 and 3 represents a bond.
63. The use according to any of the preceding claims, wherein the pain is selected from the group consisting of cancer pain, pain associated with HIV neuropathy, pain associated with drug-induced neuropathy, pain associated with diabetic neuropathy, complex regional pain syndrome, pain associated with post-herpetic neuralgia, pain associated with trigeminal neuralgia, postoperative pain, pain associated with irritable bowel syndrome, pain associated with functional dyspepsia, and pain associated with interstitial cystitis.
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