EP4719382A1 - Inhibitors of type 1 methionyl-trna synthetase and methods of using them - Google Patents

Inhibitors of type 1 methionyl-trna synthetase and methods of using them

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Publication number
EP4719382A1
EP4719382A1 EP24811541.2A EP24811541A EP4719382A1 EP 4719382 A1 EP4719382 A1 EP 4719382A1 EP 24811541 A EP24811541 A EP 24811541A EP 4719382 A1 EP4719382 A1 EP 4719382A1
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Prior art keywords
alkyl
compound
compound according
alkoxy
halogen
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German (de)
French (fr)
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Erkang FAN
Frederick S. Buckner
Zhongsheng ZHANG
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University of Washington
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University of Washington
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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
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/4985—Pyrazines or piperazines ortho- or peri-condensed with heterocyclic ring systems
    • 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
    • A61K31/4353—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 ortho- or peri-condensed with heterocyclic ring systems
    • A61K31/437—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 ortho- or peri-condensed with heterocyclic ring systems the heterocyclic ring system containing a five-membered ring having nitrogen as a ring hetero atom, e.g. indolizine, beta-carboline
    • 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
    • A61K31/56—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids
    • A61K31/575—Compounds containing cyclopenta[a]hydrophenanthrene ring systems; Derivatives thereof, e.g. steroids substituted in position 17 beta by a chain of three or more carbon atoms, e.g. cholane, cholestane, ergosterol, sitosterol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca

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  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Epidemiology (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)

Abstract

The present disclosure is generally directed to compositions useful in the inhibition of MetRS and methods for treating diseases that are ameliorated by the inhibition of MetRS.

Description

INHIBITORS OF TYPE 1 METHIONYL-tRNA SYNTHETASE AND
METHODS OF USING THEM
CROSS REFERENCE TO RELATED APPLICATIONS
[0001] This application claims priority to U.S. Provisional Patent Application No. 63/504,112, filed May 24, 2023 and to U.S. Provisional Patent Application No. 63/504,110, filed May 24, 2023, the contents of each of which are hereby incorporated by reference.
STATEMENT OF GOVERNMENT SUPPORT
[0002] This invention was made with government support under Grant Nos. R01AI152358 and R44AI134190, awarded by the National Institutes of Health. The government has certain rights in the invention.
BACKGROUND OF THE DISCLOSURE
Field of Disclosure
[0003] The present disclosure is generally directed to compositions and methods for treating diseases that are ameliorated by the inhibition of methionyl-tRNA synthetase (MetRS).
Technical Background
[0004] Gram-positive bacteria such as Staphylococcus, Streptococcus, and Enterococcus are major human pathogens responsible for a myriad of clinical syndromes. Antibiotic resistant strains such as methicillin resistant Staphylococcus aureus (MRSA) and vancomycin resistant Enterococcus (VRE) are widespread and limit the effectiveness of available antibiotics. CDC statistics show that there are 80,461 severe infections and 11,285 deaths due to MRSA per year in the USA. Similarly, there are 20,000 infections and 1,300 deaths per year due to VRE. Concern about the diminishing availability of effective antibiotics has led to urgent calls for the development of new antibiotics, such as the “10 x ’20 Initiative” by the Infectious Diseases Society of America. Launched in 2010, this is a plea to the global community to produce 10 new systemic antibiotics by 2020. As of 2016, six new systemic antibiotics have been approved (ceftaroline, dalbavancin, tedizolid, oritavancin, ceftolozane/tazobactam, and ceftazidime/avibactam) representing progress. However, none of these new antibiotics works by a novel mechanism of action (with the exception of the new beta-lactamase inhibitor avibactam) and thus may be at risk for rapid evolution of resistant bacterial strains. Moreover, many patients with existing drug allergies (e.g. to beta lactam drugs) or other contraindications to these drag classes may not benefit from these antibiotics. As a result, antibiotics acting by novel mechanisms of action are urgently needed to strengthen the treatment options.
[0005] Protozoan pathogens are also responsible for important diseases in humans and animals. These include Cryptosporidia, Cyclospora, and Giardiathat cause diarrheal disease mainly in low resource settings. Chronic or recurring infections with these pathogens contribute to stunting of growth and cognitive development in children. Cryptosporidium and Giardia are important pathogens in livestock and companion animals. Trichomonas is the most prevelant non-viral sexually transmitted infection in the world. It causes vaginitis in women and urethritis in men. Leishmania and Trypanosoma species are vector-bome protozoa that are responsible for leishmaniasis, human African trypanosomiasis (T. brace!) and Chagas disease (T. cruzi). These diseases are potentially fetal and contribute significantly to lost disease adjusted life years (DALY s) in large swaths of the tropics. For all of these protozoa, treatments options are problematic due to factors such as resistance, toxicity, high cost, or difficult treatment regimes. Improved, modem therapeutics acting by new mechanisms are needed.
[0006] Targeting the prokaryotic protein synthesis machinery has been a highly successfill strategy for developing antibiotics. Aminoglycosides, tetracyclines, macrolides, ketolides, and oxazolidinones are major classes of antibiotics that all interfere with bacterial protein translation.
[0007] With respect to the target, bacteria and all living organisms contain a complement of tRNA synthetases that are responsible for charging tRNAs with their corresponding amino acids for subsequent delivery to the ribosome. tRNA synthetases, including methionyl-tRNA synthetase (MetRS), catalyze a two-step reaction as follows:
In the first step, a highly reactive aminoacyl adenylate (aa~AMP) is formed through condensing ATP with the carboxylate of the amino acid. The second step uses this activated species to transfer the amino acid to the 3’-end of the tRNA (aa-tRNA). The bacterial MetRS enzymes are categorized in two forms (MetRS 1 and MetRS2) based on sequence similarity and sensitivity to inhibitors. Bacteria generally have a single MetRS enzyme with most Gram positive genera containing the MetRS 1 form (Staphylococcus, Streptococcus, Enterococcus, Bacillus, Clostridium, and others) and most Gram negative bacteria containing the MetRS2 form (Escherichia. Klebsiella. Pseudomonas, Haemophilus, Bacteroides, and others). Exceptions include Bacillus anthracis and a subset of Streptococcus pneumoniae which contain both MetRSl and MetRS2 isofomis. In mammals, distinct tRNA synthetases typically operate in the cytoplasm and the mitochondria. The human mitochondrial MetRS encoded in the mitochondrial genome has close sequence homology to bacterial enzymes of the MetRSl variety, whereas the human cytoplasmic MetRS is nuclear encoded with close homology to the MetRS2 variety.
[0008] Inhibition of tRNA synthetases represents another possible approach to target prokaryotic protein translation. The widely used antibiotic, mupirocin, works by inhibiting the bacterial isoleucyl-tRNA synthetase. Mupirocin is used as an ointment to treat or decolonize patients with cutaneous infections due to Staphylococcus or Streptococcus, however, its use is limited to the topical route of administration. Another bacterial tRNA synthetase inhibitor, a boron-containing compound targeting the bacterial leucyl-tRNA synthetase (GSK2251052) made it to phase 2 trials for Gram negative infections. Unfortunately, its development was discontinued due to high rates of resistance occurring during treatment, which may be related to the targeting of the editing domain of the enzyme rather than the catalytic domain. Investigators at GlaxoSmithKline reported on inhibitors to the S. aureus MetRS as having excellent antibiotic potency, but poor oral bioavailability that restricted its development (pre-New Drag Application) to topical use for skin infections and to oral use for Clostridium difficile infections where oral absorption is not needed.
[0009] Therefore, there remains a need for broad spectrum antibiotics that act by inhibiting MetRS but also have good bioavailability, pharmacokinetic properties, and efficacy in various in vivo models.
SUMMARY
[0010] In a first aspect, the present disclosure provides compounds of formula (I): or a pharmaceutically acceptable salt thereof, wherein Ri is halogen; X is Cl or F; and R2 is hydrogen, halogen, -NO2, -CN, C1-C6 alkyl, C1-C6 haloalkyl, -NH2, -(C1-C6 alkyl)NH2, -NH( C1-C6 alkyl), -NH(C2-C6 alkyl)OH, -NH(C2-C6 alkyl)NH2, -N(C1-C6 alkyl)2, -OH, hydroxy(C1-C6 alkyl), C1-C5 alkoxy, C1-C6 haloalkoxy, -O(C2-C6 alkyl)OH, and -O(C2- C6 alkyl)NH2, R4C(0)(Co-C6 alkyl)-, R4C(0)0(Co-C6 alkyl)-, R.C(0)NH(Co-C6 alkyl)-, R4S(0)(Co-C6 alkyl)-, R4S(0)0(Co-C6 alkyl)-, R4S(0)NH(Co-C6 alkyl)-, R4S(0)2(Co-C6 alkyl)-, R4S(0)20(Co-C6 alkyl)-, R4S(0)2NH(Co-C6 alkyl)-, aryl(Co-C6 alkyl)-, heteroaryl(Co- C6 alkyl)-, or heterocyclyl(Co-C6 alkyl)-, and wherein each aryl, heteroaryl, or heterocyclyl moiety is optionally substituted with one or more R3; wherein each R3 is independently selected from halogen, -NO2, -CN, oxo, thioxo, C1-C6 alkyl, C1-C6 haloalkyl, -NH2, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, amino(C1-C6 alkyl), -NH(C2-C6 alkyl)OH, -NH(C2-C6 alkyl)NH2, -OH, hydroxy(C1-C6 alkyl), C1-C6 alkoxy, C1-C6 haloalkoxy', -O(C2-C6 alkyl)OH, -O(C2-C6 alkyl)NH2, R4C(0)(Co-C6 alkyl)-, R4C(0)0(Co-C6 alkyl)-, R4C(0)NH(Co-C6 alkyl)-, R4S(0)(Co-C6 alkyl)-, R4S(0)0(Co-C6 alkyl)-, R4S(0)NH(Co-C6 alkyl)-, R4S(O)2(Co-C6 alkyl)-, R4S(0)20(Co- C6 alkyl)-, and R4S(0)2NH(Co-C6 alkyl)-; each R4 is independently selected from C1-C6 alkyl, C1-C6 haloalkyl, -NH2, -NH(C1-C6 alkyl), -NH(C2-C6 alkyl)OH, -NH(C2-C6 alkyl)NH2, -N(C1-C6 alkyl)2, -OH, -(C1-C6 alkyl)OH, C1-C6 alkoxy, C1-C6 haloalkoxy, -O(C2-C alkyl)OH, and -O(C2-C6 alkyl)NH2.
[0011] In a second aspect, the present disclosure provides compounds of formula (II): or a pharmaceutically acceptable salt thereof, wherein m is an integer 0, 1, or 2; n is an integer 2, 3, 4, or 5;
A is a heteroaryl group optionally substituted with one or two R2; each Ri is independently selected from halogen, -CN, C1-C6 alkyl, C1-C6 haloalkjd, -NH2, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, -OH, C1-C6 alkoxy, C1-C6 haloalkoxy, -SH, -S(C1-C6 alkyl), hydroxy(C1-C6 alkyl), C1-C6 alkoxy(C1-Cfi alkyl), and amino(C1-C6 alkyl); and
Rs is halogen, -NO2, -CN, C1-C6 alkyl, C1-C6 aloalkyl, -NH2, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, -OH, C1-C6 alkoxy, C1-C6 haloalkoxy, hydroxy(C1-C6 alkyl), C1-C6 alkoxy(C1-C6 alkyl), amino(C1-G6 alkyl), -C1-G6 alkyl-NH(C1-C6 alkyl), -C1-C6 alkyl-N(C1-C6 alkyl)2, -CONH2, -CONH(C1-C6 alkyl), -CON(C1-C6 alkyl)2, -CO(C1-C6 alkyl), -NHCO(C1- C6 alkoxy), -NHCO(C1-C6 alkyl), -NHCONH2, -NHCONH(C1-C6 alkyl), -S(0)o-z-(C1-C6 alkyl), -S(0)1-2NH(Co-C6 alkyl), -S(0)1-2N(Co-C6 alkyl)2, -NH-S(O)1-2-(C1-C6 alkyl), -NH- S(O)1-2NH(C1-C6 alkyl), -NH-S(O)1-2-aryl, -NH-S(O)1-2-heteroaryl, aryl(Co-C6 alkyl), heteroaryl(Co-C6 alkyl), or heterocyclyl(Co-C6 alkyl), and wherein each alkyl, aryl, heteroaryl, or heterocyclyl moiety is optionally substituted with one or more R4; wherein each R2 is independently selected from halogen, -NO2, -CN, C1-C6 alkyl, C1-C6 haloalkyl, -NH2, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, -OH, C1-C6 alkoxy, and C1-C6 haloalkoxy. each R4 is independently selected from independentiy selected from halogen, -NO2, -CN, C1- C6 alkyl, C1-C6 haloalkyl, -NH2, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, -OH, oxo, C1-C6 alkoxy, C1-C6 haloalkoxy, -SH, thioxo, -S(C1-C6 alkyl), hydroxy(C1-C6 alkyl), C1-C6 alkoxy(C1-C6 alkyl), amino(C1-C6 alkyl), -CH2-NH(C1-C6 alkyl), -CH2-N(C1-C6 alkyl)2, -CONH2, -CONH(C1-C6 alkyl), -CON(C1-C6 alkyl)2, -CONH-OH, -CONH-NH2, -COH, -COzH, -COz(C1-C6 alkyl), -CO(C1-C6 alkyl), -OCO(C1-C6 alkyl), -OCONH(C1-C6 alkyl), -NHCO(C1-C6 alkoxy), -NHCO(C1-C6 alkyl), -NHCONH2, -NHCONH(C1-C6 alkyl), -NH-S(O)1-2-(C1-C6 alkyl), -NH-S(O)1-2-aryl, -NH-S(O)1-2-heteroaryl, -CH2- NHCONH2, -CH2-NHCONH(C1-C6 alkyl), and -CH2-OCO(C1-C6 alkyl).
[0012] In a third aspect, the present disclosure provides methods for inhitibing MetRS in a pathogen, comprising exposing the pathogen to an effective amount of a compound according to the first or second aspects of the disclosure, or a pharmaceutical composition comprising one ore more compounds according to the first or second aspects of the disclosure.
[0013] In a fourth aspect, the present disclosure provides methods for treating diseases that are ameliorated by the inhibition of MetRS comprising administering to a patient in need of such treatment a therapeutically effective amount of a compound according to the first or second aspects of the disclosure, or a pharmaceutical composition comprising one ore more compounds according to the first or second aspects of the disclosure.
[0014] In a lst embodiment of the first aspect of the disclosure, the disclosure provides a compound of the first aspect of the disclosure, wherein Ri is chloro or fluoro. [0015] In a 2nd embodiment of the first aspect of the disclosure, the disclosure provides a compound of the first aspect of the disclosure, wherein Ri is chloro.
[0016] In a 3rd embodiment of the first aspect of the disclosure, the disclosure provides a compound of the first aspect of the disclosure, wherein Ri is fluoro.
[0017] In a 4th embodiment of the first aspect of the disclosure, the disclosure provides a compound of the first aspect or of any of the 1st through 3rd embodiments of the first aspect of the disclosure, wherein R2 is hydrogen.
[0018] In a 5th embodiment of the first aspect of the disclosure, the disclosure provides a compound of the first aspect or of any of the 1st through 3rd embodiments of the first aspect of the disclosure, wherein R2 is hydrogen, halogen, -NOa, -CN, C1-C6 alkyl, C1-C6 haloalkyl, -NH2, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, -OH, C1-C6 alkoxy, C1-C6 haloalkoxy, aryl(Co-C6 alkyl), heteroaryl(Co-C6 alkyl), or heterocyclyl(Co-C6 alkyl), and wherein each aryl, heteroaryl, or heterocyclyl moiety is optionally substituted with one or more R3; wherein each each R2 is independently selected from halogen, -NOa, -CN, C1-C6 alkyl, C1-C6 haloalkyl, -NHa, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, -OH, C1-C6 alkoxy, and C1-C6 haloalkoxy.
[0019] In a 6th embodiment of the first aspect of the disclosure, the disclosure provides a compound of the first aspect or of any of the 1st through 3rd embodiments of the first aspect of the disclosure, wherein R2 is hydrogen, halogen, -CN, C1-C6 alkyl, C1-C6 haloalkyl, aryl(Co-C6 alkyl), heteroaryl(Co-C6 alkyl), or heterocyclyl(Co-Q alkyl), and wherein each aryl, heteroaryl, or heterocyclyl moiety is optionally substituted with one or more R3.
[0020] In a 7th embodiment of the first aspect of the disclosure, the disclosure provides a compound of the 6th embodiment of the first aspect of the disclosure, wherein R2 is hydrogen, halogen, -CN, C1-C6 alkyl, or C1-C6 haloalkyl.
[0021] In an 8th embodiment of the first aspect of the disclosure, the disclosure provides a compound of the first aspect or of any of the 1st through 3rd embodiments of the first aspect of the disclosure, wherein R2 is hydrogen, aryl(Co-C6 alkyl), heteroaryl(Co-C6 alkyl), or heterocyclyl(Co-C6 alkyl), and wherein each ary l, heteroaryl, or heterocyclyl moiety is optionally substituted with one or more R3. [0022] In a 9th embodiment of the first aspect of the disclosure, the disclosure provides a compound of the first aspect or of any of the 1th through 3rd, 5th through 6th, and 8th embodiments of the first aspect of the disclosure, wherein R2 is hydrogen or heteroaryl(Co-C6 alkyl).
[0023] In a 10 th embodiment of the first aspect of the disclosure, the disclosure provides a compound of the first aspect or of any of the 1st through 9th embodiments of the first aspect of the disclosure, wherein X is F.
[0024] In an 11th embodiment of the first aspect of the disclosure, the disclosure provides a compound of the first aspect or of any of the 1st through 9th embodiments of the first aspect of the disclosure, wherein X is Cl.
[0025] In a 12th embodiment of the first aspect of the disclosure, the disclosure provides a compound of the first aspect of the disclosure which is any of the compounds of Formula (I) provided in Tables 1-3.
[0026] In a 13th embodiment of the first aspect of the disclosure, the disclosure provides a compound of the first aspect of the disclosure, which is: or a pharmaceutically acceptable salt thereof.
[0027] In a 14th embodiment of the first aspect of the disclosure, the disclosure provides a compound of the first aspect of the disclosure, which is: or a pharmaceutically acceptable salt thereof. [0028] In a 15th embodiment of the first aspect of the disclosure, the disclosure provides a pharmaceutical composition comprising one or more compounds of the first aspect or of any of the 1st through 14th embodiments of the first aspect of the disclosure, and a pharmaceutically acceptable carrier, diluent, or excipient.
[0029] In a 16th embodiment of the first aspect of the disclosure, the disclosure provides a pharmaceutical composition of the 15 th embodiment of the first aspect disclosure, further comprising one or more additional antibacterial compounds.
[0030] In a 1st emodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect of the disclosure, wherein each Ri is independently selected from halogen, C1-C6 alkyl, C1-C6 haloalkyl, -NH2, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, -OH, C1-C6 alkoxy, and C1-C6 haloalkoxy.
[0031] In a 2nd emodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect of the disclosure, wherein wherein each Ri is independently selected from halogen, C1-C6 alkyl, C1-C6 haloalkyl, -OH C1-C6 alkoxy, and C1-C6 haloalkoxy.
[0032] In a 3rd emodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect of the disclosure, wherein each Ri is independently selected from halogen, C1-C6 haloalkyl, -OH and C1-C6 alkoxy.
[0033] In a 4th emodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect of the disclosure, wherein each Ri is independently selected from halogen and C1-C6 alkoxy.
[0034] In a 5th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the 1st through 4th embodiments of the second aspect of the disclosure, wherein n is 2 or 3.
[0035] In a 6th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the 1st through 4th embodiments of the second aspect of the disclosure, wherein n is 2.
[0036] In a 7th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the 1st through 4th embodiments of the second aspect of the disclosure, wherein n is 3. [0037] In an 8th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the lat through 4th embodiments of the second aspect of the disclosure, which is of formula:
[0038] In a 9th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the 1st through 8th embodiments of the second aspect of the disclosure, wherein A is a 9-member heteroaryl group optionally substituted with one or two R2.
[0039] In a 10th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the 1st through 8th embodiments of the second aspect of the disclosure, wherein A is indole, azaindole, benzimidazole, or imidazopyridine, each optionally substituted with one or two R2.
[0040] In an 11th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the 1st through 8th embodiments of the second aspect of the disclosure, wherein A is indole, azaindole, benzimidazole, or imidazopyridine, each optionally substituted with one or two R2.
[0041] In a 12th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the 1st through 8th embodiments of the second aspect of the disclosure, wherein A is imidazopyridine, optionally substituted with one or two R2.
[0042] In a 13th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the 1st through 4th embodiments of the second aspect of the disclosure, which is of formula: [0043] In a 14th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the 1st through 4th embodiments of the second aspect of the disclosure, which is of formula:
[0044] In a 15th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the 1 st through 14th embodiments of the second aspect of the disclosure, wherein each R?. is independently selected from halogen, Ct-C-6 alkyl, C1-C6 haloalkyl, -OH, C1-C6 alkoxy, and C1-C6 haloalkoxy.
[0045] In a 16“ embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of tire 1st through 14th embodiments of tire second aspect of the disclosure, wherein each R2 is independently selected from halogen, C1-C6 haloalkyl, -OH, and C1-C6 alkoxy.
[0046] In a 17th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the 1st through 14th embodiments of the second aspect of the disclosure, wherein each R2 is independently selected from halogen and C1-C6 alkoxy.
[0047] In a 18th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the 1st through 14th embodiments of the second aspect of the disclosure, wherein each R? is independently halogen.
[0048] In a 19“ embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of tire 1st through 14th embodiments of tire second aspect of the disclosure, wherein A is substituted with one R2, which is chloro.
[0049] In a 20th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the 1st through 4th embodiments of the second aspect of the disclosure, which is of formula:
[0050] In a 21st embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the 1” through 20th embodiments of the second aspect of the disclosure, wherein m is 0.
[0051] In a 22nd embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the 1st through 20th embodiments of the second aspect of the disclosure, wherein m is 0 or 1.
[0052] In a 23rd embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the 1st through 20th embodiments of the second aspect of the disclosure, wherein m is 1.
[0053] In a 24th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the 23rd embodiment of the second aspect of the disclosure, which is of formula:
[0054] In a 25th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the 1st through 20th embodiments of the second aspect of the disclosure, wherein m is 2.
[0055] In a 26th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the 25rd embodiment of the second aspect of the disclosure, which is of formula:
[0056] In a 27th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect or of any of the 1st through 26th embodiments of the second aspect of the disclosure, wherein Rs is halogen, -NH2, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, -OH, C1-C6 alkoxy, C1-C6 haloalkoxy, hydroxy(C1-C6 alkyl), C1-C6 alkoxy(C1-C6 alkyl), amino(C1-C6 alkyl), -C1-C6 alkyl-NH(C1-C6 alkyl), -C1-C6 alkyl-N(C1-C6 alkyl)2, -S(O)1-2-(C1-C6 alkyl), aryl(Co-C6 alkyl), heteroaryl(Co-C6 alkyl), or heterocyclyl(Co-C6 alkyl), and wherein each alkyl, aryl, heteroaryl, or heterocyclyl moiety is optionally substituted with one or more R*.
[0057] In a 28th embodiment of the second aspect of the disclosure, the disclosure provides a compound of the second aspect of the disclosure which is any of the compounds of Formula (II) provided in Table 4.
[0058] In a 29th embodiment of the second aspect of the disclosure, the disclosure provides a pharmaceutical composition comprising one or more compounds of the second aspect or of any of the 1st through 28th embodiments of the second aspect of the disclosure, and a pharmaceutically acceptable carrier, diluent, or excipient.
[0059] In a 30th embodiment of the second aspect of the disclosure, the disclosure provides a pharmaceutical composition of the 29th embodiment of the second aspect disclosure, further comprising one or more additional antibacterial compounds.
[0060] In a 1st embodiment of the third aspect of the disclosure, the disclosure provides a method for inhibiting MetRS in a pathogen, the method comprising exposing the pathogen to an effective amount of a compound of the first aspect or of any of the 1st through 14th embodiments of the first aspect of the disclosure, or of a compound of the second aspect or of any of the 1st through 28th embodiments of the second aspect of the disclosure.
[0061] In a 2nd embodiment of the third aspect of the disclosure, the disclosure provides a method of the 1st embodiment of the third aspect of the disclosure, wherein the pathogen is a bacteria or a protozoa. [0062] In a 1th embodiment of the fourth aspect of the disclosure, the disclosure provides a method for treating diseases that are ameliorated by the inhibition of MetRS, the method comprising administering to a patient in need thereof a therapeutically effective amount of a compound of the first aspect or of any of the 1st through 14th embodiments of the first aspect of the disclosure, or of a compound of the second aspect or of any of the 1st through 28th embodiments of the second aspect of the disclosure, or of a pharmaceutical composition of the 15th or 16th embodiments of the first aspect of the disclosure, or of a pharmaceutical composition of the 29th or 30th embodiments of the second aspect of the disclosure.
[0063] In a 2nd embodiment of the fourth aspect of the disclosure, the disclosure provides a method of the 1st embodiment of the fourth aspect of the disclosure, further comprising administering one or more additional antibacterial compounds in combination with the compound or the pharmaceutical composition.
[0064] In a 3rd embodiment of the fourth aspect of the disclosure, the disclosure provides a method of the 1st embodiment of the fourth aspect of the disclosure, further comprising administering a second antibacterial compound in combination with the compound or the pharmaceutical composition.
[0065] In a 4th embodiment of the fourth aspect of the disclosure, the disclosure provides a method of the 3rd embodiment of the fourth aspect of the disclosure, wherein the compound is the second antibacterial compound is fusidic acid.
[0066] In a 5th embodiment of the fourth aspect of the disclosure, the disclosure provides a method of the 1st embodiment of the fourth aspect of the disclosure, wherein the disease is a protozoan infection.
[0067] In a 6th embodiment of the fourth aspect of the disclosure, the disclosure provides a method of the 5th embodiment of the fourth aspect of the disclosure, wherein the protozoan infection is selected from the group consisting of Cryptosporidia, Cyclospora, Giardia, Leishmania, Trichomonas, and Trypanosoma. [0068] In a 7th embodiment of the fourth aspect of the disclosure, the disclosure provides a method of the 1st embodiment of the fourth aspect of the disclosure, wherein the disease is a bacterial infection.
[0069] In an 8th embodiment of the fourth aspect of the disclosure, the disclosure provides a method of the 7th embodiment of the fourth aspect of the disclosure, wherein the bacteria is a Gram positive bacteria.
[0070] In a 9th embodiment of the fourth aspect of the disclosure, the disclosure provides a method of the 8th embodiment of the fourth aspect of the disclosure, wherein the Gram positive bacteria is selected from one of Staphylococcus, Streptococcus, Enterococcus, Clostridia, Bacillus. Listeria, Corynebacteria, Arcanobacteria, Rothia, and Rhodococcus.
[0071] In a 10th embodiment of the fourth aspect of the disclosure, the disclosure provides a method of the 7th embodiment of the fourth aspect of the disclosure, wherein the bacteria is a Gram negative bacteria.
[0072] In an 11th embodiment of the fourth aspect of the disclosure, the disclosure provides a method of the 10th embodiment of the fourth aspect of the disclosure, wherein the Gram negative bacteria is Brucella, Campylobacter, and Helicobacter.
[0073] In a 12th embodiment of the fourth aspect of the disclosure, the disclosure provides a method of the 7th embodiment of the fourth aspect of the disclosure, wherein the bacteria is Mycobacteria.
[0074] In a 13th embodiment of the fourth aspect of the disclosure, the disclosure provides a method of the 12th embodiment of the fourth aspect of the disclosure, wherein the Mycobacteria is selected from M. tuberculosis, M. avium, M. abscessus, M. kansasii, M. chelonae, M. marinum, M. ulcerans, and M. haemophilum.
[0075] In a 14th embodiment of the fourth aspect of the disclosure, the disclosure provides a method of the 7th embodiment of the fourth aspect of the disclosure, wherein the bacteria is Mycoplasma.
[0076] In a 15th embodiment of the fourth aspect of the disclosure, the disclosure provides a method of the 1st embodiment of the fourth aspect of the disclosure, wherein the disease is a Trypanosomatid protozoa infection. [0077] In a 16th embodiment of the fourth aspect of the disclosure, the disclosure provides a method of the 15 th embodiment of the fourth aspect of the disclosure, wherein the Trypanosomatid protozoa is one of Trypanosoma brucei, Trypanosoma cruzi, and Leishmania species.
[0078] In a 17th embodiment of the fourth aspect of the disclosure, the disclosure provides a method of the 7th embodiment of the fourth aspect of the disclosure, wherein the bacteria is selected from Staphylococcus aureus, methicillin resistant Staphylococcus aureus (MRSA), Enterococcus faecalis, Enterococcus faecium, and vancomycin resistant Enterococcus (VRE).
[0079] In a fifth aspect, the disclosure provides a compound selected from the group consisting of or a pharmaceutically acceptable salt thereof.
[0080] In a 1st embodiment of the fifth aspect of the disclosure, the disclosure provides a pharmaceutical composition comprising one or more compounds of the fifth aspect of the disclosure, and a pharmaceutically acceptable carrier, diluent, or excipient.
[0081] In 2nd embodiment of the fifth aspect of the disclosure, the disclosure provides a pharmaceutical composition of the 1st embodiment of the fifth aspect disclosure, further comprising one or more additional antibacterial compounds. [0082] In a 3rd embodiment of the fifth aspect of the disclosure, the disclosure provides methods of using the compounds of the fifth aspect of the disclosure, and/or the pharmaceutical compositions of the 1st and 2nd embodiments of the fifth aspect of the disclsoure, in accordance with the methods of the third and fourth aspects of the disclosure, and the embodiments associated therewith.
BRIEF DESCRIPTION OF THE DRAWINGS
[0083] FIG. 1 shows the results of the Time kill assays of 2541, fosidic acid, and the combination at 0.25X MIC (FIG. 1A) and 0.125X MIC (FIG. IB). A decrease of CFU counts >2-logs at 24 hr of the combination compared to the drug alone is indicative of synergism.
DETAILED DESCRIPTION
[0084] Before the disclosed processes and materials are described, it is to be understood that the aspects described herein are not limited to specific embodiments, apparatus, or configurations, and as such can, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular aspects only and, unless specifically defined herein, is not intended to be limiting.
[0085] In view of the present disclosure, the methods and compositions described herein can be configured by the person of ordinary skill in the art to meet the desired need. In general, the disclosed materials, methods, and apparatus provide improvements in treatment of bacterial and/or protozoan infections. The disclosure provides novel MetRS inhibitor compounds that show promising antibiotic and antiprotozoal activity. At the same time, in certain embodiments, the compounds of the disclosure show good oral bioavailability and pharmacokinetic properties.
[0086] Thus, one aspect of the disclosure provides compounds of formula (I): or a pharmaceutically acceptable salt thereof, wherein Ri is halogen;
X is Cl or F; and
R2 is hydrogen, halogen, -NO2, -CN, C1-C6 alkyl, Ci-Cc, haloalkyl, -NTH 2, -(C1-C6 alkyl)NH2, -NH(CI-C6 alkyl), -NH(C 2-C6 alkyl)OH, -NH(C2-C6 alkyl)NH2, -N(C 1i -C6 alkyl)2, -OH, hydroxy(Ci-C6 alkyl), C1-C6 alkoxy, C1-C6 haloalkoxy, -O('C2-C6 alkyl)OH, and -O(C?- C6 alkyl)NH2, R4C(0)(Co-C« alkyl)-. R4C(0)0(Co-C6 alkyl)-, R4C(O)NH(Cu-C6 alkyl)-, R4S(O)(C0-C6 alkyl)-, R4S(0)0(Co-C6 alkyl)-, R4S(0)NH(Co-C6 alkyl)-, R4S(0)2(Co~Cb alkyl)-, R4S(O)2O(C.3-C6 alkyl)-, R4S(0)2NH(Co-C6 alkyl)-, aryl(Co-C6 alkyl)-, heteroaryl(Co- Cs alkyl)-, or heterocyclylCo-C6 alkyl)-, and wherein each aryl, heteroaryl, or heterocyclyl moiety is optionally substituted with one or more R3; wherein each R3 is independently selected from halogen, -NO2, -CN, oxo, thioxo, C1-C alkyl, C1-C6 haloalkyl, -NH?., -NH(C1-C6 alkyl), -N(C1-C6 alkyl)?., arnino(C1-C6 alkyl), ~NH(C2-C6 alky 1)OH, -NH(C2-C6 alkyl)NH2, -OH, hydroxy(Ci-C6 alkyl), C1-C6 alkoxy, C1-C6 haloalkoxy, -O(C2~C6 alkyl)OH, -O(C2-C6 alkyl)NH2, R4C(0)(Co-C6 alkyl)-, R4C(0)0(CO-C.6 alkyl)-, R4C(O)NH(Cc-C6 alkyl)-, R4S(0)(Co-C6 alkyl)-, R4S(0)0(Co-C6 alkyl)-, R4S(0)NH(Co-C6 alkyl)-, R4S(0)2(Co-C6 alkyl)-, R4S(0)?.0(Co- C6 alkyl)-, and R4S(0)2NH(Co-C6 alkyl)-; each R4 is independently selected from C1-C6 alkyl, C1-C6 haloalkyl, -NH2, -NH(C1-C6 alkyl), -NH(C2-C6 alkyl)()H, -NH(C2.-C6 alkyl)NH2, -N(C1-C6 alkyl)2, -OH, -(C1-C6 alkyl)OH, C1-C6 alkoxy, Ci-Cs haloalkoxy, -O(C2-C6 alkyl)OH, and -O(C2~C6 alkyl)NH2.
[0087] A second aspect of the disclosure provides compounds of formula (II): or a pharmaceutically acceptable salt thereof, wherein m is an integer 0, 1, or 2; n is an integer 2, 3, 4, or 5;
A is a heteroaryl group optionally substituted with one or two R?.; each Ri is independently selected from halogen, -CN, C1-C6 alkyl, C1-C6 haloalkyl, -NH2,
-NH(C1-C6 alkyl), -N(C1-C 6 alky 1): 2, -OH, C1-C6 alkoxy, C1-C6 haloalkoxy, -SH, -S(C1-C6 alkyl), hydroxy (C1-C5 alkyl), C1-C6 alkoxy(C1-C6 alkyl), and amino(C1-C'6 alkyl); and R3 is halogen, -NO2, -CN, C1-C6 alkyl, C1-C6 haloalkyl, -NH2, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, -OH, C1-C6 alkoxy, C1-C6 haloalkoxy, hydroxy(C1-C6 alkyl), C1-C6 alkoxy(C1-C6 alkyl), amino(C1-C6 alkyl), -C1-C6 alkyl-NH(C1-C6 alkyl), -C1-C6 alkyl-N(C1-C6 alkyl)2, -CONH?, -CONH(C1-C6 alkyl), -CON(C1-C6 alkyl)2, -CO(C1-C6 alkyl), -NHCO(C1- C6 alkoxy), -NHCO(C1-C6 alkyl), -NHCONH2, -NHCONH(C1-C6 alkyl), -S(0)o-2-(C1-C6 alkyl), -S(0)i-2NH(Co-C6 alkyl), -NH-S(O)1-2-(C1-C6 alkyl), -NH- S(O)i-2NH(C1-C6 alkyl), -NH-S(O)1-2-aryl, -NH-S(O)1-2-heteroaryl, aryl(Co-C6 alkyl), heteroaryl(Co-C6 alkyl), or heterocyclyl(Co-C6 alkyl), and wherein each alkyl, aryl , heteroaryl, or heterocyclyl moiety is optionally substituted with one or more R4; wherein each R2 is independently selected from halogen, -NCh, -CN, C1-C6 alkyl, C1-C6 haloalkyl, -NH2, -NH(C1-G6 alkyl), -N(C1-C6 alkyl)2, -OH, C1-C6 alkoxy, and C1-C6 haloalkoxy. each R4 is independently selected from independently selected from halogen, -NO2, -CN, C1- C6 alkyl, C1-C6 haloalkyl, -NH2, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, -OH, oxo, C1-C6 alkoxy, C1-C6 haloalkoxy, -SH, thioxo, -S(C1-C6 alkyl), hydroxy(C1-C6 alkyl), C1-C6 alkoxy(C1-C6 alkyl), amino(C1-C6 alkyl), -CH2-NH(C1-C6 alkyl), -CH2-N(C1-C6 alkyl)2, -CONH2, -CONH(C1-C6 alkyl), -CON(C1-C6 alkyl)2, -CONH-OH, -CONH-NH2, -COH, -CO2H, -CO2(C1-C6 alkyl), -CO(C1-C6 alkyl), -OCO(C1-C6 alkyl), -OCONH(C1-C6 alkyl), -NHCO(C1-C6 alkoxy), -NHCO(C1-C6 alkyl), -NHCONH2, -NHCONH(C1-C6 alkyl), -NH-S(O)I-2-(C1-C6 alkyl), -NH-S(O)1-2-aryl, -NH-S(O)1-2-heteroaryl, -CH2- NHCONH2, -CH2-NHCONH(C1-C6 alkyl), and -
Applications
[0088] The compounds of the disclosure are capable of inhibiting the activity of MetRS.
Inhibition of MetRS may be either in vivo and/or in vitro. Accordingly, the disclosure provides methods of inhibiting MetRS in pathogens comprising exposing the pathogen to an effective amount of either a compound of the disclosure (e.g., compounds formula (I) or of formula (II)), or a pharmaceutical composition comprising one or more of compounds of the disclosure. The disclosure also provides methods for treating diseases that are ameliorated by the inhibition of MetRS by administering to a patient in need of such treatment a therapeutically effective amount of either a compound of the disclosure (e.g., compounds formula (I) or of formula (II)), or a pharmaceutical composition comprising one or more of compounds of the disclosure. [0089] In certain embodiments, the pathogens include bacteria or protozoa.
[0090] In certain embodiments, the diseases that are ameliorated by the inhibition of MetRS by the compounds of the present disclosure include bacterial and protozoan infections. In certain embodiments, the diseases that are ameliorated by the inhibition of MetRS by the compounds of the present disclosure include bacterial diseases including, but not limited to, skin and soft tissue infections, bacteremia, sepsis, pneumonia, bone and joint infections, prosthetic joint infections, pharyngitis, dental abscess, cardiovascular infections, intraabdominal infections, sexually transmitted infections, urinary tract infections, infections of the eye, infections of the central nervous system, brucellosis, tuberculosis, Clostridium difficile infection, and infections caused by non-tuberculous mycobacteria, and protozoan diseases that including, but not limited to, Chagas disease, leishmaniasis, cryptosporidiosis, giardiasis, and trichopmoniasis.
[0091] Examples of protozoa, or examples of protozoan infection include those caused by, but not limited to, Cryptosporidia, Cyclospora, Giardia, Leishmania, Trichomonas, and Trypanosoma. In certain embodiments, protozoan infection is caused by' one of Trypanosoma brucei, Trypanosoma cruzi, or Leishmania species.
[0092] Examples of bacteria, or examples of bacterial diseases include those caused by, but not limited to, Gram positive bacteria, such as Staphylococcus, Streptococcus, Enterococcus, Clostridia, Bacillus, Listeria, Corynebacteria, Arcanobacteria, Rothia, and Rhodococcus, Gram negative bacteria, such as Brucella, Campylobacter, and Helicobacter, Mycobacteria, such as M. tuberculosis, M. avium, M. abscessus,M. kansasii,M. chelonae,M. marinum,M. ulcerans, andL M. haemophilum, and Mycoplasma. Some particular embodiments include infections caused by Staphylococcus aureus, including methicillin resistant Staphylococcus aureus (MRSA), Enterococcus faecalis or Enterococcus faecium, including vancomycin resistant enterococcus (VRE).
[0093] In certain embodiments, the methods of the disclosure further comprise administering one or more additional antibacterial compounds in combination with a compound of the disclosure. In certain embodiments, the methods further comrprise administering a second antibacterial compound in combination with a compound of the disclosure. In certain embodiments, one or more of the compounds of the disclosure may be used in combination with one or more additional antibacterial compounds in the methods disclosed herein. As used herein, “in combination” may be concurrent administration, i.e. the compounds are administered at the same time, or it may be sequential administration, i.e. the compounds may be administered at different times. The one or more additional antibacterial compounds may be, but are not limited to, a quinolone, an acridine, a phenothiazine, an aminoglycoside, a macrolide, an amphenicol, a steroid, an ansamycin, an antifolate, a polymyxin, a glycopeptide, a cephalosporin, a lactam, a fusidane-type, or any combination thereof. In certain embodiments, the antibacterial compound is fusidic acid. In certain other embodiments, the methods of the disclosure comprise treating diseases that are ameliorated by the inhibition of MetRS by providing to a patient in need of such treatment a therapeutically effective amount of Compound 2541 in combination with fusidic acid. As disclosed herein, the combination of Compound 2541 and fusidic acid has a synergistic effect when used in accordance with the methods disclosed herein. In certain embodiments, the one or more additional antibacterial compounds may be administered in an amount below its minimum inhibitory concentration (MIC) established in the absence of the one or more compounds. For example, the second antibacterial compound may be administered in an amount less than 90% of, e.g., less than 75%, or less than 50%, or less than 25%, or less than 10%, or even less than 1% of the minimum inhibitory concentration (MIC).
[0094] The development of the compositions of the present application is highly significant as it solves the problem of poor oral bioavailability of MetRS inhibitors. Thus, in certain embodiments, the compounds of the disclosure are orally administered. In other embodiments, the compositions of the disclosure may be administered systemically (e.g., transdermal, intranasal or by suppository), or parenterallly (e.g., intramuscular, intravenous or subcutaneous).
Pharmaceutical Compositions
[0095] In another aspect, the present disclosure provides compositions comprising one or more of the compounds as described above with respect to formula (I) and formula (II) and an appropriate carrier, excipient or diluent. The exact nature of the carrier, excipient or diluent will depend upon the desired use for the composition, and may range from being suitable or acceptable for veterinary uses to being suitable or acceptable for human use. The composition may optionally include one or more additional compounds. In certain embodiments, the composition may include one or more additional antibacterial compounds in addition to the one or more compounds as described above with respect to formula (I) and formula (II).
[0096] When used to treat or prevent such diseases, the compounds described herein may be administered singly, as mixtures of one or more compounds or in mixture or combination with other agents usefill for treating such diseases and/or the symptoms associated with such diseases. The compounds may also be administered in mixture or in combination with agents useful to treat other disorders or maladies. The compounds may be administered in the form of compounds per se, or as pharmaceutical compositions comprising a compound.
[0097] Pharmaceutical compositions comprising the compound(s) may be manufactured by means of mixing, dissolving, granulating, dragee-making levigating, emulsifying, encapsulating, entrapping or lyophilization processes. The compositions may be formulated in a manner using one or more physiologically acceptable carriers, diluents, excipients or auxiliaries which facilitate processing of the compounds into preparations which can be used pharmaceutically. [0098] The compounds may be formulated in the pharmaceutical composition per se, or in the form of a hydrate, solvate, N-oxide or pharmaceutically acceptable salt. Typically, such salts are more soluble in aqueous solutions than the corresponding free acids and bases, but salts having lower solubility than the corresponding free acids and bases may also be formed.
[0099] Pharmaceutical compositions may take a form suitable for virtually any mode of administration, including, for example, topical, ocular, oral, buccal, systemic, nasal, injection, transdermal, rectal, vaginal, etc., or a form suitable for administration by inhalation or insufflation.
[0100] For topical administration, the compound(s) may be formulated as solutions, gels, ointments, creams, suspensions, etc. Systemic formulations include those designed for administration by injection, e.g., subcutaneous, intravenous, intramuscular, intrathecal or intraperitoneal injection, as well as those designed for transdermal, transmucosal oral or pulmonary administration.
[0101] Usefol injectable preparations include sterile suspensions, solutions or emulsions of the active compound(s) in aqueous or oily vehicles. The compositions may also contain formulating agents, such as suspending, stabilizing and/or dispersing agent. The formulations for injection may be presented in unit dosage form, e.g., in ampules or in multidose containers, and may contain added preservatives. Alternatively, the injectable formulation may be provided in powder form for reconstitution with a suitable vehicle, including but not limited to sterile pyrogen free water, buffer, dextrose solution, etc., before use. To this end, the active compound(s) may be dried by any technique, such as lyophilization, and reconstituted prior to use.
[0102] For transmucosal administration, penetrants appropriate to the barrier to be permeated are used in the formulation.
[0103] For oral administration, the pharmaceutical compositions may take the form of, for example, lozenges, tablets or capsules prepared by means with pharmaceutically acceptable excipients such as binding agents (e.g., pregelatinised maize starch, polyvinylpyrrolidone or hydroxypropyl methylcellulose); fillers (e.g., lactose, microcrystalline cellulose or calcium hydrogen phosphate); lubricants (e.g., magnesium stearate, talc or silica); disintegrants (e.g., potato starch or sodium starch glycolate); or wetting agents (e.g., sodium lauryl sulfete). The tablets may be coated by several methods, for example, sugars, films or enteric coatings. [0104] Liquid preparations for oral administration may take the form of, for example, elixirs, solutions, syrups or suspensions, or they may be presented as a dry product for constitution with water or other suitable vehicle before use. Such liquid preparations may be prepared by means with pharmaceutically acceptable additives such as suspending agents (e.g., sorbitol syrup, cellulose derivatives or hydrogenated edible fats); emulsifying agents (e.g., lecithin or acacia); non-aqueous vehicles (e.g., almond oil, oily esters, ethyl alcohol, cremophore™ or fractionated vegetable oils); and preservatives (e.g., methyl or propyl-p-hydroxybenzoates or sorbic acid). The preparations may also contain buffer salts, preservatives, flavoring, coloring and sweetening agents as appropriate.
[0105] Preparations for oral administration may be suitably formulated to give controlled release of the compound. For buccal administration, the compositions may take the form of tablets or lozenges formulated in a manner. For rectal and vaginal routes of administration, the compound(s) may be formulated as solutions (for retention enemas) suppositories or ointments containing suppository bases such as cocoa butter or other glycerides.
[0106] For nasal administration or administration by inhalation or insufflation, the compound(s) can be conveniently delivered in the form of an aerosol spray from pressurized packs or a nebulizer with the use of a suitable propellant, e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, fluorocarbons, carbon dioxide or other suitable gas. In the case of a pressurized aerosol, the dosage unit may be determined by providing a valve to deliver a metered amount. Capsules and cartridges for use in an inhaler or insufflator (for example capsules and cartridges comprised of gelatin) may be formulated containing a powder mix of the compound and a suitable powder base such as lactose or starch. [0107] For ocular administration, the compound(s) may be formulated as a solution, emulsion, suspension, etc. suitable for administration to the eye. A variety of vehicles are suitable for administering compounds to the eye.
[0108] For prolonged delivery, the compound(s) can be formulated as a depot preparation for administration by implantation or intramuscular injection. The compound(s) may be formulated with suitable polymeric or hydrophobic materials (e.g., as an emulsion in an acceptable oil) or ion exchange resins, or as sparingly soluble derivatives, e.g., as a sparingly soluble salt. Alternatively, transdermal delivery systems manufactured as an adhesive disc or patch which slowly releases the compound(s) for percutaneous absorption may be used. To this end, permeation enhancers may be used to facilitate transdermal penetration of the compound(s). [0109] Alternatively, other pharmaceutical delivery systems may be employed. Liposomes and emulsions are examples of delivery vehicles that may be used to deliver compound(s). C6rtain organic solvents such as dimethyl sulfoxide (DMSO) may also be employed, although usually at the cost of greater toxicity.
[0110] The pharmaceutical compositions may, if desired, be presented in a pack or dispenser device which may contain one or more unit dosage forms containing the compound(s). The pack may, for example, comprise metal or plastic foil, such as a blister pack. The pack or dispenser device may be accompanied by instructions for administration.
[0111] The compound(s) described herein, or compositions thereof, will generally be used in an amount effective to achieve the intended result, for example in an amount effective to treat or prevent the particular disease being treated. By therapeutic benefit is meant eradication or amelioration of the underlying disorder being treated and/or eradication or amelioration of one or more of the symptoms associated with the underlying disorder such that the patient reports an improvement in feeling or condition, notwithstanding that the patient may still be afflicted with the underlying disorder. Therapeutic benefit also generally includes halting or slowing the progression of the disease, regardless of whether improvement is realized.
[0112] The amount of compound(s) administered will depend upon a variety of factors, including, for example, the particular indication being treated, the mode of administration, whether the desired benefit is prophylactic or therapeutic, the severity of the indication being treated and the age and weight of the patient, the bioavailability of the particular compound(s) the conversation rate and efficiency into active drug compound under the selected route of administration, etc.
[0113] Determination of an effective dosage of compound(s) for a particular use and mode of administration is well within the capabilities of those skilled in the art. Effective dosages may be estimated initially from in vitro activity and metabolism assays. For example, an initial dosage of compound for use in animals may be formulated to achieve a circulating blood or serum concentration of the metabolite active compound that is at or above an IC6o of the particular compound as measured in as in vitro assay. Calculating dosages to achieve such circulating blood or serum concentrations taking into account the bioavailability of the particular compound via the desired route of administration is well within the capabilities of skilled artisans. Initial dosages of compound can also be estimated from in vivo data, such as animal models. Animal models may be used fortesting the efficacy of the active metabolites to treat or prevent the various diseases described above. Animal models suitable for testing the bioavailability and/or metabolism of compounds into active metabolites can be used. Such information can be adapted to determine dosages of particular compounds suitable for human administration.
[0114] Dosage amounts will typically be in the range of from about 0.0001 mg/kg/day, 0.001 mg/kg/day or 0.01 mg/kg/day to about 1000 mg/kg/day, but may be higher or lower, depending upon, among other factors, the activity of the active compound, the bioavailability of the compound, its metabolism kinetics and other pharmacokinetic properties, the mode of administration and various other factors, discussed above. Dosage amount and interval may be adjusted individually to provide plasma levels of the compound(s) and/or active metabolite compound(s) which are sufficient to maintain therapeutic or prophylactic effect. For example, the compounds may be administered once per week, several times per week (e.g., every other day), once per day or multiple times per day, depending upon, among other things, the mode of administration, the specific indication being treated and the judgment of the prescribing physician. In cases of local administration or selective uptake, such as local topical administration, the effective local concentration of compound(s) and/or active metabolite compound(s) may not be related to plasma concentration. Skilled artisans will be able to optimize effective dosages without undue experimentation.
Definitions
[0115] The following terms and expressions used herein have the indicated meanings.
[0116] Throughout this specification, unless the context requires otherwise, the word “comprise” and “include” and variations (e.g., “comprises,” “comprising,” “includes,” “including”) will be understood to imply the inclusion of a stated component, feature, element, or step or group of components, features, elements or steps but not the exclusion of any other integer or step or group of integers or steps.
[0117] As used in the specification and the appended claims, the singular forms “a,” “an” and “the” include plural referents unless the context clearly dictates otherwise.
[0118] Terms used herein may be preceded and/or followed by a single dash, or a double dash, “=“, to indicate the bond order of the bond between the named substituent and its parent moiety; a single dash indicates a single bond and a double dash indicates a double bond. In the absence of a single or double dash it is understood that a single bond is formed between the substituent and its parent moiety; further, substituents are intended to be read “left to right’' unless a dash indicates otherwise. For example, C1-C6alkoxycarbonyloxy and -C6alkyl indicate the same functionality; similarly arylalkyl and -alkylaryl indicate the same functionality. [0119] The term “alkenyl” as used herein, means a straight or branched chain hydrocarbon containing from 2 to 10 carbons, unless otherwise specified, and containing at least one carbon- carbon double bond. Representative examples of alkenyl include, but are not limited to, ethenyl, 2-propenyl, 2-methyl-2-propenyl, 3-butenyl, 4-pentenyl, 5-hexenyl, 2-heptenyl, 2-methyl-l- heptenyl, 3-decenyl, and 3, 7-dimethylocta-2, 6-dienyl.
[0120] The term “alkoxy” as used herein, means an alkyl group, as defined herein, appended to the parent molecular moiety through an oxygen atom. Representative examples of alkoxy include, but are not limited to, methoxy, ethoxy, propoxy, 2-propoxy, butoxy, tert-butoxy, pentyloxy, and hexyloxy.
[0121] The term “alkyl” as used herein, means a straight or branched chain hydrocarbon containing from 1 to 10 carbon atoms unless otherwise specified. Representative examples of alkyl include, but are not limited to, methyl, ethyl, n-propyl, iso-propyl, n-butyl, sec-butyl, iso- butyl, tert-butyl, n-pentyl, isopentyl, neopentyl, n-hexyl, 3-methylhexyl, 2,2-dimethylpentyl, 2,3- dimethylpentyl, n-heptyl, n-octyl, n-nonyl, and n-decyl. When an “alkyl” group is a linking group between two other moieties, then it may also be a straight or branched chain; examples include, but are not limited to , and
[0122] The term "alkylene" refers to a bivalent alkyl group. An "alkylene chain" is a polymethylene group, i.e., -(CH2)n-, wherein n is a positive integer, preferably from one to six, from one to four, from one to three, from one to two, or from two to three. A substituted alkylene chain is a polymethylene group in which one or more methylene hydrogen atoms is replaced with a substituent. Suitable substituents include those described below for a substituted aliphatic group. An alkylene chain also may be substituted at one or more positions with an aliphatic group or a substituted aliphatic group.
[0123] The term “alkynyl” as used herein, means a straight or branched chain hydrocarbon group containing from 2 to 10 carbon atoms and containing at least one carbon-carbon triple bond. Representative examples of alkynyl include, but are not limited, to acetylenyl, 1-propynyl, 2- propynyl, 3-butynyl, 2-pentynyl, and 1-butynyl. [0124] The term “aryl,” as used herein, means a phenyl (i.e., monocyclic aryl), or a bicyclic ring system containing at least one phenyl ring or an aromatic bicyclic ring containing only carbon atoms in the aromatic bicyclic ring system. The bicyclic aryl can be azulenyl, naphthyl, or a phenyl fused to a monocyclic cycloalkyl, a monocyclic cycloalkenyl, or a monocyclic heterocyclyl. The bicyclic aryl is attached to the parent molecular moiety through any carbon atom contained within the phenyl portion of the bicyclic system, or any carbon atom with the napthyl or azulenyl ring. The fused monocyclic cycloalkyl or monocyclic heterocyclyl portions of the bicyclic aryl are optionally substituted with one or two oxo and/or thioxo groups. Representative examples of the bicyclic aryls include, but are not limited to, azulenyl, naphthyl, dihydroinden-l-yl, dihydroinden-2-yl, dihydroinden-3-yl, dihydroinden-4-yl, 2,3-dihydroindol- 4-yl, 2,3-dihydroindol-5-yl, 2,3-dihydroindol-6-yl, 2,3-dihydroindol-7-yl, inden-l-yl, inden-2-yl, inden-3-yl, inden-4-yl, dihydronaphthalen-2-yl, dihydronaphthalen-3-yl, dihydronaphthalen-4-yl, dihydronaphthalen-l-yl, 5,6,7,8-tetrahydronaphthalen-l-yl, 5,6,7,8-tetrahydronaphthalen-2-yl,
2.3-dihydrobenzofuran-4-yl, 2,3-dihydrobenzofuran-5-yl, 2,3-dihydrobenzofuran-6-yl,
2.3-dihydrobenzofuran-7-yl, benzo[d][l,3]dioxol-4-yl, benzo[d][l,3]dioxol-5-yl, 2H-chromen-2- on-5-yl, 2H-chromen-2-on-6-yl, 2H-chromen-2-on-7-yl, 2H-chromen-2-on-8-yl, isoindoline-1, 3- dion-4-yl, isoindoline- 1, 3-dion-5-yl, inden- l-on-4-yl, inden-l-on-5-yl, inden- l-on-6-yl, inden- 1- on-7-yl, 2,3-dihydrobenzo[b][l,4]dioxan-5-yl, 2,3-dihydrobenzo[b][l,4]dioxan-6-yl, 2H- benzo[b][l,4]oxazin3(4H)-on-5-yl, 2H-benzo[b][l,4]oxazin3(4H)-on-6-yl, 2H- benzo[b][l,4]oxazin3(4H)-on-7-yl, 2H-benzo[b][l,4]oxazin3(4H)-on-8-yl, benzo[d]oxazin- 2(3H)-on-5-yl, benzo[d]oxazin-2(3H)-on-6-yl, benzo[d]oxazin-2(3H)-on-7-yl, benzo[d]oxazin- 2(3H)-on-8-yl, quinazolin-4(3H)-on-5-yl, quinazolin-4(3H)-on-6-yl, quinazolin-4(3H)-on-7-yl, quinazolin-4(3H)-on-8-yl, quinoxalin-2(lH)-on-5-yl, quinoxalin-2(lH)-on-6-yl, quinoxalin- 2(lH)-on-7-yl, quinoxalin-2(lH)-on-8-yl, benzo[d]thiazol-2(3H)-on-4-yl, benzo[d]thiazol- 2(3H)-on-5-yl, benzo[d]thiazol-2(3H)-on-6-yl, and, benzo[d]thiazol-2(3H)-on-7-yl. In certain embodiments, the bicyclic aryl is (i) naphthyl or (ii) a phenyl ring fused to either a 5 or 6 membered monocyclic cycloalkyl, a 5 or 6 membered monocyclic cycloalkenyl, or a 5 or 6 membered monocyclic heterocyclyl.
[0125] The terms “cyano” and “nitrile” as used herein, mean a -CN group.
[0126] The term “cycloalkyl” as used herein, means a monocyclic or a bicyclic cycloalkyl ring system. Monocyclic ring systems are cyclic hydrocarbon groups containing from 3 to 8 carbon atoms, where such groups can be saturated or unsaturated, but not aromatic. In certain embodiments, cycloalkyl groups are fully saturated. Examples of monocyclic cycloalkyls include cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cycloheptyl, and cyclooctyl. Bicyclic cycloalkyl ring systems are bridged monocyclic rings or fused bicyclic rings. Bridged monocyclic rings contain a monocyclic cycloalkyl ring where two non-adjacent carbon atoms of the monocyclic ring are linked by an alkylene bridge of between one and three additional carbon atoms (i.e., a bridging group of the form -(CH2)w-, where w is 1, 2, or 3). Representative examples of bicyclic ring systems include, but are not limited to, bicyclo[3.1.1]heptane, bicyclo[2.2.1]heptane, bicyclo[2.2.2]octane, bicyclo[3.2.2]nonane, bicyclo[3.3.1]nonane, and bicyclo[4.2.1]nonane. Fused bicyclic cycloalkyl ring systems contain a monocyclic cycloalkyl ring fused to either a phenyl, a monocyclic cycloalkyl, a monocyclic cycloalkenyl, a monocyclic heterocyclyl, or a monocyclic heteroaryl. The bridged or fused bicyclic cycloalkyl is attached to the parent molecular moiety through any carbon atom contained within the monocyclic cycloalkyl ring. In certain embodiments, the fused bicyclic cycloalkyl is a 5 or 6 membered monocyclic cycloalkyl ring fused to either a phenyl ring, a 5 or 6 membered monocyclic cycloalkyl, a 5 or 6 membered monocyclic cycloalkenyl, a 5 or 6 membered monocyclic heterocyclyl, or a 5 or 6 membered monocyclic heteroaryl. [0127] The term “halo” or “halogen” as used herein, means -Cl, -Br, -I or -F.
[0128] The terms "haloalkyl" and "haloalkoxy" refer to an alkyl or alkoxy group, as the case may be, which is substituted with one or more halogen atoms.
[0129] The term “heteroaryl,” as used herein, means a monocyclic heteroaryl or a bicyclic ring system containing at least one heteroaromatic ring. The monocyclic heteroaryl can be a 5 or 6 membered ring. The 5 membered ring consists of two double bonds and one, two, three or four nitrogen atoms and optionally one oxygen or sulfur atom. The 6 membered ring consists of three double bonds and one, two, three or four nitrogen atoms. The 5 or 6 membered heteroaryl is connected to the parent molecular moiety through any carbon atom or any nitrogen atom contained within the heteroaryl. Representative examples of monocyclic heteroaryl include, but are not limited to, furyl, imidazolyl, isoxazolyl, isothiazolyl, oxadiazolyl, oxazolyl, pyridinyl, pyridazinyl, pyrimidinyl, pyrazinyl, pyrazolyl, pyrrolyl, tetrazolyl, thiadiazolyl, thiazolyl, thienyl, triazolyl, and triazinyl. The bicyclic heteroaryl consists of a monocyclic heteroaryl fused to a phenyl, a monocyclic cycloalkyl, a monocyclic cycloalkenyl, a monocyclic heterocyclyl, or a monocyclic heteroaryl. When the bicyclic heteroaryl contains a fused cycloalkyl, cycloalkenyl, or heterocyclyl ring, then the bicyclic heteroaryl group is connected to the parent molecular moiety through any carbon or nitrogen atom contained within the monocyclic heteroaryl portion of the bicyclic ring system. When the bicyclic heteroaryl is a monocyclic heteroaryl fused to a benzo ring, then the bicyclic heteroaryl group is connected to the parent molecular moiety through any carbon atom or nitrogen atom within the bicyclic ring system. Representative examples of bicyclic heteroaryl include, but are not limited to, benzimidazolyl, benzofuranyl, benzothienyl, benzoxadiazolyl, benzoxathiadiazolyl, benzothiazolyl, cinnolinyl, 5,6- dihydroquinolin-2-yl, 5,6-dihydroisoquinolin-l-yl, furopyridinyl, indazolyl, indolyl, isoquinolinyl, naphthyridinyl, quinolinyl, purinyl, 5,6,7,8-tetrahydroquinolin-2-yl, 5,6, 7,8- tetrahydroquinolin-3-yl, 5,6,7,8-tetrahydroquinolin-4-yl, 5,6,7,8-tetrahydroisoquinolin- 1-yl, thienopyridinyl, 4,5,6,7-tetrahydrobenzo[c][l,2,5]oxadiazolyl, and 6,7- dihydrobenzo[c][l,2,5]oxadiazol-4(5H)-onyl. In certain embodiments, the fused bicyclic heteroaryl is a 5 or 6 membered monocyclic heteroaryl ring fused to either a phenyl ring, a 5 or 6 membered monocyclic cycloalkyl, a 5 or 6 membered monocyclic cycloalkenyl, a 5 or 6 membered monocyclic heterocyclyl, or a 5 or 6 membered monocyclic heteroaryl.
[0130] The terms “heterocyclyl’' and “heterocycloalkyl” as used herein, mean a monocyclic heterocycle or a bicyclic heterocycle. The monocyclic heterocycle is a 3, 4, 5, 6 or 7 membered ring containing at least one heteroatom independently selected from the group consisting of O, N, and S where the ring is saturated or unsaturated, but not aromatic. The 3 or 4 membered ring contains 1 heteroatom selected from the group consisting of O, N and S. The 5 membered ring can contain zero or one double bond and one, two or three heteroatoms selected from the group consisting of O, N and S. The 6 or 7 membered ring contains zero, one or two double bonds and one, two or three heteroatoms selected from the group consisting of O, N and S. The monocyclic heterocycle is connected to the parent molecular moiety through any carbon atom or any nitrogen atom contained within the monocyclic heterocycle. Representative examples of monocyclic heterocycle include, but are not limited to, azetidinyl, azepanyl, aziridinyl, diazepanyl, 1,3-dioxanyl, 1,3-dioxolanyl, 1,3-dithiolanyl, 1,3-dithianyl, imidazolinyl, imidazolidinyl, isothiazolinyl, isothiazolidinyl, isoxazolinyl, isoxazolidinyl, morpholinyl, oxadiazolinyl, oxadiazolidinyl, oxazolinyl, oxazolidinyl, piperazinyl, piperidinyl, pyranyl, pyrazolinyl, pyrazolidinyl, pyrrolinyl, pyrrolidinyl, tetrahydrofuranyl, tetrahydrothienyl, thiadiazolinyl, thiadiazolidinyl, thiazolinyl, thiazolidinyl, thiomorpholinyl, 1,1-dioxidothiomorpholinyl (thiomorpholine sulfone), thiopyranyl, and trithianyl. The bicyclic heterocycle is a monocyclic heterocycle fused to either a phenyl, a monocyclic cycloalkyl, a monocyclic cycloalkenyl, a monocyclic heterocycle, or a monocyclic heteroaryl. The bicyclic heterocycle is connected to the parent molecular moiety through any carbon atom or any nitrogen atom contained within the monocyclic heterocycle portion of the bicyclic ring system. Representative examples of bicyclic heterocyclyls include, but are not limited to, 2,3-dihydrobenzofuran-2-yl, 2,3- dihydrobenzofuran-3-yl, indolin-l-yl, indolin-2-yl, indolin-3-yl, 2,3-dihydrobenzothien-2-yl, decahydroquinolinyl, decahydroisoquinolinyl, octahydro- IH-indolyl, and octahydrobenzofuranyl. In certain embodiments, the bicyclic heterocyclyl is a 5 or 6 membered monocyclic heterocyclyl ring fused to phenyl ring, a 5 or 6 membered monocyclic cycloalkyl, a 5 or 6 membered monocyclic cycloalkenyl, a 5 or 6 membered monocyclic heterocyclyl, or a 5 or 6 membered monocyclic heteroaryl
[0131] The term “oxo” as used herein means a =0 group.
[0132] The term “saturated” as used herein means the referenced chemical structure does not contain any multiple carbon-carbon bonds. For example, a saturated cycloalkyl group as defined herein includes cyclohexyl, cyclopropyl, and the like.
[0133] The term "substituted", as used herein, means that a hydrogen radical of the designated moiety is replaced with the radical of a specified substituent, provided that the substitution results in a stable or chemically feasible compound. The term "substitutable", when used in reference to a designated atom, means that attached to the atom is a hydrogen radical, which can be replaced with the radical of a suitable substituent.
[0134] The phrase "one or more” substituents, as used herein, refers to a number of substituents that equals from one to the maximum number of substituents possible based on the number of available bonding sites, provided that the above conditions of stability and chemical feasibility are met. Unless otherwise indicated, an optionally substituted group may have a substituent at each substitutable position of the group, and the substituents may be either the same or different. As used herein, the term "independently selected" means that the same or different values may be selected for multiple instances of a given variable in a single compound.
[0135] The term “thioxo” as used herein means a =S group.
[0136] The term “unsaturated” as used herein means the referenced chemical structure contains at least one multiple carbon-carbon bond, but is not aromatic. For example, a unsaturated cycloalkyl group as defined herein includes cyclohexenyl, cyclopentenyl, cyclohexadienyl, and the like.
[0137] It will be apparent to one skilled in the art that certain compounds of this disclosure may exist in tautomeric forms, all such tautomeric farms of the compounds being within the scope of the disclosure. Unless otherwise stated, structures depicted herein are also meant to include all stereochemical forms of the structure; i.e., the R and S configurations for each asymmetric center. Therefore, single stereochemical isomers as well as enantiomeric and diastereomeric mixtures of the present compounds are within the scope of the disclosure. Both the R and the S stereochemical isomers, as well as all mixtures thereof, are included within the scope of the disclosure.
[0138] “Pharmaceutically acceptable” refers to those compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problems or complications commensurate with a reasonable benefit/risk ratio or which have otherwise been approved by the United States Food and Drug Administration as being acceptable for use in humans or domestic animals.
[0139] “Pharmaceutically acceptable salt” refers to both acid and base addition salts.
[0140] “Therapeutically effective amount” refers to that amount of a compound which, when administered to a subject, is sufficient to effect treatment for a disease or disorder described herein. The amount of a compound which constitutes a “therapeutically effective amount” will vary' depending on the compound, the disorder and its severity, and the age of the subject to be treated.
[0141] ‘Treating” or “treatment” as used herein covers the treatment of a disease or disorder described herein, in a subject, preferably a human, and includes: i. inhibiting a disease or disorder, i.e., arresting its development; ii. relieving a disease or disorder, i.e., causing regression of the disorder; iii. slowing progression of the disorder; and/or iv. inhibiting, relieving, ameliorating, or slowing progression of one or more symptoms of the disease or disorder
[0142] “Subject” refers to a warm blooded animal such as a mammal, preferably a human, or a human child, which is afflicted with, or has the potential to be afflicted with one or more diseases and disorders described herein.
Methods of Preparation
[0143] Many general references providing chemical synthetic schemes and conditions usefol for synthesizing the disclosed compounds are available (see, e.g., Smith and March, March's Advanced Organic Chemistry: Reactions, Mechanisms, and Structure, Fifth Edition, Wiley- Interscience, 2001; or Vogel, A Textbook of Practical Organic Chemistry, Including Qualitative Organic Analysis, Fourth Edition, New York: Longman, 1978). [0144] Compounds as described herein can be purified by any of the means, including chromatographic means, such as HPLC, preparative thin layer chromatography, flash column chromatography and ion exchange chromatography. Any suitable stationary phase can be used, including normal and reversed phases as well as ionic resins. Most typically the disclosed compounds are purified via silica gel and/or alumina chromatography. See, e.g., Introduction to Modem Liquid Chromatography, 2nd Edition, ed. L. R. Snyder and J. J. Kirkland, John Wiley and Sons, 1979; and Thin Layer Chromatography, ed E. Stahl, Springer-Verlag, New York, 1969.
[0145] During any of the processes for preparation of the subject compounds, it may be desirable to protect sensitive or reactive groups on any of the molecules concerned. This may be achieved by means of protecting groups as described in works, such as J. F. W. McOmie, "Protective Groups in Organic Chemistry,” Plenum Press, London and New York 1973, in T. W. Greene and P. G. M. Wuts, "Protective Groups in Organic Synthesis,” Third edition, Wiley, New York 1999, in "The Peptides"; Volume 3 (editors: E. Gross and J. Meienhofer), Academic Press, London and New York 1981, in "Methoden der organischen Chemie,” Houben-Weyl, 4.sup.th edition, Vol. 15/1, Georg Thieme Verlag, Stuttgart 1974, in H.-D. Jakubke and H. Jescheit, "Aminosauren, Peptide, Proteine,” Verlag Chemie, Weinheim, Deerfield Beach, and Basel 1982, and/or in Jochen Lehmann, "Chemie der Kohlenhydrate: Monosaccharide and Derivate,” Georg Thieme Veriag, Stuttgart 1974. The protecting groups may be removed at a convenient subsequent stage.
[0146] The compounds disclosed herein can be made using procedures familiar to the person of ordinary' skill in the art and as described herein. For example, compounds of structural formula (I) and/or (II) can be prepared according to the Schemes below, or analogous synthetic schemes. One of skill in the art can adapt the reaction sequences to fit the desired target molecule. Of course, in certain situations one of skill in the art will use different reagents to affect one or more of the individual steps or to use protected versions of certain of the substituents. Additionally, one skilled in the art would recognize that compounds of the disclosure can be synthesized using different routes altogether.
EXAMPLES
[0147] The compounds and the methods of the disclosure are illustrated further by the hallowing examples, which are not to be construed as limiting the disclosure in scope or spirit to the specific procedures and compounds described in them. [0148] Unless otherwise stated, all chemicals were purchased from commercial suppliers and used without further purification. The microwave irradiation was performed in a CEM Discover System. The final purity of all compounds was determined by analytical LCMS with Poroshell 120 EC-C18 column (4.6 mm x 100 mm). The products were detected by UV at the detection frequency of 230 nm. All compounds were determined to be >95% pure by this method. The mass spectra were recorded with the Agilent LC/MSD iQ or with the Agilent Liquid Chromatograph - Ion Trap Mass Spectrometer. NMR spectra were recorded with Broker 500 MHz spectrometer at ambient temperature. The compounds of the disclosure were synthesized through several different routes, as represented by the Schemes below. Syntheses and compound characterization data are presented below.
Example 1: Preparation of Compounds of Formula fl)
Preparation of 2541 (5-chloro-2-((5-(2-chloro-4,5-dimethoxyphenyl)-6-fluoroimidazo[l,2- a]pyrazin-2-yl)methyl)-lH-imidazo[4,5-b]pyridine)
Scheme 1
[0149] Reagents and conditions (a) , rt, overnight, 21% yield; (b) 2-chloro- 4,5-dimethoxyphenylboronic acid, K2CO3, Pd(dppf)Ch DCM, dioxane, H20, 110°C 4h; (c) ethyl 4-chloroacetoacetate, MeCN, microwave irradiation; (d) LiOH, ethanol/water; (e) EDC, 6- chloropyridine-2,3-diamine, pyridine; (f) HOAc, microwave irradiation. GeneralProcedure 1 (2541,2659) [0150] AgzCo3 (217mg, 0.79mmol)wasaddedtoasolutionof2-amino-6-chloropyrazine (1.700g, 13.13mmol)andSelectfluor(4.673g, 13.13mmol)in 80ml MeCNand 8mlH2O. The mixturewasstirredatroomtemperatureovernight. Themixturewasfiltered,andthefiltratewas concentrated. TheresiduewasdissolvedwithEtOAcandwashedsuccessivelywithwaterand brine. TheorganiclayerwasdriedoverNa2SO4andconcentratedinvacuo.Theresiduewas purifiedbyflashcolumn chromatography(EtOAc/hexane)togive 2-amino-5-fluoro-6- chloropyrazinein21%yield.1HNMR(500MH2, CDCh) 57.41 (s, IH),4.61 (s,2H). LC/MS: (ESI)(M+H)+= 148.2. [0151] 2-amino-5-fluoro-6-chloropyrazine (369mg,2.51mmol),2-chloro-4,5- dimethoxyphenylboronicacid(759mg, 3.51mmol),K2CO3(1039mg, 7.53mmol)and Pd(dppf)Ck lCM(103mg, 0.125mmol)inwater(1.2ml)anddioxane (12ml)washeatedat 110 °Cfor4hunderN2. Themixturewaspurifiedbyflashchromatographyonsilicageltoobtain IntermediateA (50%). NMR(500MHz, CDCh) 57.57(d,J=2.1 H2, 1H), 6.96(s, IH), 6.93 (s, IH),4.57 (s,2H), 3.92(s, 3H), 3.89(s,3H).LC/MS: (ESI) (M+H)+=284.2. [0152] Ethyl4-chloroacetoacetate (240μl, 1.78mmol)wasaddedtoIntermediateA(207mg, 0.73mmol)solutioninanhydrousMeCN(Nml).Themixturewasmicrowaveirradiatedat 100°C for 1.5h. Afterthesolutionwasconcentrated,theresiduewasstirredwithhexane(10ml).Hexane wasdecantedandtheresiduewaspurifiedbyflashchromatographyon silicageltoobtain5-(2- chloro-4,5-dimethoxy-phenyl)-6-fluoro-imidazo[l,2-<z]pyrazin-2-carboxylicacidethylester (70%).1HNMR(500MHz, CDCh)58.72(s, 1H), 7.45 (s, 1H), 7.06 (s, IH), 6.91 (s, 1H),4.17 (q,J=7.1 H2, 2H), 3.96(s, 3H), 3.89(s,2H), 3.87 (s, 3H), 1.25 (t,J=7.1 Hz, 3H). LC/MS: (ESI)(M+H)+=380.2. [0153] 5-(2-chloro-4,5-dimethoxy-phenyl)-6-fluoro-imidazo[1,2-a]pyrazin-2-carboxylieacid ethyl ester(50.3mg,0.134mmol)wasaddedto2mlofethanoland6ml ofwater,mixedwith LiOH(12.8mg, 0.54mmol)andstirredfor Ihatroomtemperature. Thesolutionwasacidified with 1 Nhydrochloricacidandthe solventwascompletelyremovedinvacuo.Theresiduewas dissolvedin 10mlofpyridine,then6-chloropyridine-2,3-diamine (38.6mg, 0.268mmol)and EDChydrochloride (38.6mg, 0.20mmol)wereadded. Themixturewasstirredatroom temperatureovernight. The solventwasremovedonarotary evaporator. Theresiduewas dissolvedindichloromethane (20ml)andwashedwithwater(20ml).Theorganiclayerwas collected. Theaqueouslayerwasextractedwithdichloromethane (20ml)twice. Theorganic layer was combined. After the solvent was removed in vacuo, the residue was recrystallized in MeOH. After the solid was collected and washed with small amount of cold methanol, it was dissolved in 5ml of acetic acid. The solution was microwave irradiated at 125 °C for 1 hour.
After the solvent was removed on the rotary evaporator, the residue was purified by flash column chromatography to give 2541 in 66% yield in two steps. *HNMR (500 MH2, CDCh) 5 8.75 (s, 1H), 7.88 (s, 1H), 7.47 (s, 1H), 7.18 (d, J= 8.3 H2, 1H), 7.06 (s, 1H), 6.89 (s, 1H), 4.55 (s, 2H), 3.97 (s, 3H), 3.86 (s, 3H). LC/MS: (ESI) (M +H) = 473.2.
Alternative Synthetic Method of Intermediate A
[0154] (Yawei Tian et al.. Journal of Fluorine Chemistry, 218, 111-115; 2019.) Reagents and conditions (a) 2-chloro-4,5-dimethoxyphenylboronic acid Pd(dppf)Ch DCM, dioxane, H2O, 110°C, 2h (b) MeCN, A Selectfluor, 80°C, overnight.
[0155] 2-amino-6-chloropyrazine (129.6mg, l.Ommol), 2-chloro-4,5-dimethoxyphenylboronic acid (281mg, 1.3mmol), K2CO3 (414mg, 7.53mmol) and Pd(dppf)Ch DCM (40.8mg, O.OSmmol) in water (1 ,2ml) and dioxane (12ml) was heated at 110 °C for 2h under . The mixture was purified by flash chromatography on silica gel to obtain 2-amino-6-(2-chloro-4,5-dimethoxy- phenyl)pyrazine (70%). LC/MS: (ESI) (M +H)*= 266.1.
[0156] s (11.6mg, 0.04mmol) was added to a solution of 2-amino-6-chloropyrazine (185 mg, 0.70mmol) and Selectfluor (62.3mg, 0.175mmol) in 10ml of MeCN. The mixture was stirred at 80°C overnight. The mixture was filtered, and the filtrate was concentrated. The residue was purified by flash column chromatography to give Intermediate A in 8% yield.
[0157] 2659 was synthesized using 2-fluoro-4,5-dimethoxyphenylboronic acid following General Procedure 1
Scheme 2
[0158] Reagents and conditions (a) NBS, chloroform, 0°C 2h (b); tetramethyl-13»2-dio.xaborolan-2-yl)-lH-pyrazole Pd(dppf)Ch DCM, dioxane, H2O, microwave irradiation, 110°C Ih; (c) ethyl 4-chloroacetoacetate, MeCN, 80°C, two days; (d) LiOH, dioxaneAvater; (e) EDC, 6-chloropyridine-2,3-diamine, pyridine; (f) HOAc, microwave irradiation.
General Procedure 2 (2624, 2664-2674, 2684, 2685, 2686, and 2688)
[0159] NBS (166mg, 0.932mmol) was added to Intermediate A (220mg, 0.78mmol) in 25ml of anhydrous chloroform at 0°C for 2h. After the solution was removed, the residue was purified by flash chromatography on silica gel eluted with EtOAc/hexane to obtain 275mg of 3-bromo-5- fluoro-6-(2-chloro-4,5-dimethoxy-phenyl)-pyrazin-2-ylamine in yield 98%.
[0160] Ethyl 4-chloroacetoacetate (2ml) was added to 3-bromo-5-fluoro-6-(2-chloro-4,5- dimethoxy-phenyl)-pyrazin-2-ylamine (220mg, 0.61mmol) solution in anhydrous MeCN (8ml). The mixture was heated at 80 °C two days. After the solution was concentrated, the residue was stirred with hexane (20ml) and hexane was decanted. The procedure was repeated twice, and the oil residue was purified by flash chromatography on silica gel eluted with EtOAc/hexane to obtain Intermediate B (70 %).
[0161] Intermediate B (23.3 mg, 0.055mmol), 1-methyl- dioxaborolan-2-yl)-l H-pyrazole (17.0 mg, 0.082 mmol), K3PO4 H2O (37.7mg, 0.164 mmol) and Pd(dppf)Ch-DCM (2.23mg, 0.0027 mmol) in water (0.2 ml) and dioxane (2.0ml) was microwave irradiated at 100 °C for Ih . After the solvents were removed, the residue was purified by flash chromatography on silica gel to obtain 5-(2-chloro-4,5-dimethoxy-phenyl)-6- fluoro-8-(l-methyI-4-lH-pyrazole)imidazo pyrazin-2-carboxylic acid ethyl ester.
[0162] 5-(2-chloro-4,5-dimethoxy-phenyl)-6-fluoro-8-(l-methyl-4-lH-pyrazole)imidazo[l,2- a]pyrazin-2-carboxylic acid ethyl ester (23.4mg, 0.05 mmol) was added in 1 ml of dioxane and 1ml of water, mixed with LiOH (4.4mg, 0.18 mmol) and stirred for Ih at room temperature. The solution was acidified with 1 N HC1 and the solvent was completely removed in vacuo. The residue was dissolved in 2 ml of pyridine, then 6-chloropyridine-2,3-diamine (lO.Omg, 0.069 mmol) and EDC hydrochloride (13.3 mg, 0.069 mmol) were added. The mixture was stirred at room temperature overnight. The solvent was removed on a rotary evaporator. The residue was added in dichloromethane (20 ml) and washed with water (20ml). The organic layer was collected. The aqueous layer was extracted with dichloromethane (20ml) twice. The organic layer was combined and washed with brine. The organic layer was dried over and concentrated in vacuo. The residue was purified by flash chromatography on silica gel. The purified compound was dissolved in 5 ml of acetic acid and microwave irradiated at 125 °C for 45min. The solvent was removed in vacuo, the residue was purified by flash chromatography on silica gel to obtain to get 2624 in 70% yield. ’H NMR (500 MHz, CDCh) 5 11.25 (s, IH), 8.70 (s, IH), 8.53 (s, IH), 7.92 (d, J= 8.0 Hz, IH), 7.39 (s, IH), 7.20 (d, J = 8.5 H2, IH), 7.05 (s, IH), 6.93 (s, IH), 4.55 (s, 2H), 4.04 (s, 3H), 3.97 (s, 3H) ), 3.86 (s, 3H). LC/MS: (ESI) (M +H)+= 553.2.
[0163] 2664 was synthesized using l-ethyl-4-(4,4,5,5-tetraniethyl-l,3,2-dioxaborolan-2-yi)-
IH-pyr azole following General Procedure 2 LC/MS: (ESI) (M +H)+= 567.1.
[0164] 2665 was synthesized using pyrimtdin-5-ylboronic acid following General Procedure
2. LC/MS: (ESI) (M +H)+= 551.1.
[0165] 2666 was synthesized using l,3-dimethyI-5-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2- yl)-lH-pyrazole following General Procedure 2. LC/MS: (ESI) (M +H)+= 567.1.
[0166] 2667 was synthesized using l-cyclopropyl-4-(4.4,5,-tetramethyl-1.3,2-dioxaborolan- 2-yI)-lH-pyrazole fallowing General Procedure 2. LC/MS: (ESI) (M +H)+= 579.1.
[0167] 2668 was synthesized using pyridin-3-ylboronic acid following General Procedure 2.
LC/MS: (ESI) (M +H)+= 550.1.
[0168] 2669 was synthesized using 13-dimethyl-4-(4,4,5,5-tetrarnethyl-l,3,2-dioxaborolan-2- yl)-lH-pyrazole following General Procedure 2. LC/MS: (ESI) (M +H)+= 567.1. 2670
[0169] 2670 was synthesized using 1-methyl-5-(4,4.5,5-tetramethyl-l,3,2-dioxaboroIan-2-yl)-
IH-pyr azole following General Procedure 2. LC/MS: (ESI) (M +H)+= 553.1. 2671
[0170] 2671 was synthesized using pyridin-4-ylboronic acid following General Procedure 2.
LC/MS: (ESI) (M +H)+= 550.1. 2672
[0171] 2672 was synthesized using l-methyl-3-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2-yl)- IH-pyrazole following General Procedure 2. LC/MS: (ESI) (M +H)+= 553.1. 2673
[0172] 2673 was synthesized using 1^5-dimethyl-4-(4,4^i,5-tetramethyi-13,2-dioxaborolan-2- yl)-lH-pyrazole following General Procedure 2. LC/MS: (ESI) (M +H)+= 567.1. 2674
[0173] 2674 was synthesized using l,4-dimethyl-5-(4,4.5^-tetramethyi-13^-dioxaborolan-2- yl)-lH-pyrazole following General Procedure 2. LC/MS: (ESI) (M +H)+= 567.2. 2684
[0174] 2684 was synthesized using l,3,5-trimethyl-4-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2- yl)-lH-pyrazole following General Procedure 2. LC/MS: (ESI) (M +H)*= 581 .5. 2685
[0175] 2685 was synthesized using l-ethyl-4-methyl-5-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan- 2-yl)-lH-pyrazole following General Procedure 2. LC/MS: (ESI) (M +H)+= 581.5. 2686
[0176] 2686 was synthesized using l-methyl-5-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2-yl)- lH-pyrazole-4-caibonitrile following General Procedure 2. LC/MS: (ESI) (M +H) = 578.4. 2688 [0177] 2688 was synthesized using 3,5-dimethyl-l-(oxetan-3-yl)-4-(4,4,5,5-tetramethyl-l,3,2- dioxaborolan-2-yl)-lH-pyrazole following General Procedure 2. LC/MS: (ESI) (M +H)+= 623.5.
Compound Z 2675
[0178] Compound Z was synthesized using 2-(4-(4,4,i,5-tetramethyi-13,2-dioxaborolan-2- yl)-1H-pyrazol-l-yl)ethanol following General Procedure 2 and further hydrolyzed in LiOH/H2O to form 2675. LC/MS: (ESI) (M +H)+= 583.1.
Procedures for 2676, 2677, 2679, 2681, 2682, and 2687:
Intermediate AA
2676
[0179] Intermediate AA was prepared from 2-fluoro-4,5-dimethoxyphenylboronic acid following the earlier steps of General Procedure 2. Then 2676 was synthesized using Intermediate AA and pyrimidin-5-ylboronic acid following the later steps in General Procedure 2. LC/MS: (ESI) (M +H)+= 535.1.
2677
[0180] 2677 was synthesized using l-methylpyrazole-4-boronic acid, pinacol ester following the procedure for 2676. LC7MS: (ESI) (M +H)+= 537.1.
2679
[0181] 2679 was synthesized using l,5-Dimethyl-4-(4,4,5,5-tetrainethyl-l,3,2-dioxaborolan-2- yl)4H-pyrazole following the procedure for 2676. LC/MS: (ESI) (M +H)+= 551.5.
2681
[0182] 2681 was synthesized using 2-(4-(4,4,5,5-Tetramethyl-l,3,2-dioxaborolan-2-yl)-lH- pyrazol-l-yl)ethanol following the procedure for 2676 and 2675. LC/MS: (ELSI) (M +H)+= 567.1.
2682
[0183] 2682 was synthesized using l,4-dimethyl-5-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan-2- yl)-lH-pyrazole following the procedure for 2676. LC/MS: (ESI) (M +H)+= 551.1.
2687
[0184] 2687 was synthesized using l,3,5-trimethyl-4-(4,4,5,5-tetramethyl-l,3,2-dioxaborolan- 2-yl)-lH-pyrazole following the procedure for 2676. LC/MS: (ESI) (M +H)+= 565.5.
Scheme 3
Intermediate B Intermediate C
2634
[0185] Reagents and conditions (a) LiOH, dioxane/water; (b) EDC, 6-chloropyridine-2,3- diamine, pyridine; (c) ethylamine, DMF, microwave irradiation; (d) HOAc, microwave irradiation. [0186] Intermediate B ( 19.6 mg, 0.046 mmol) was added in 1 ml of dioxane and 1 ml of water, mixed with LiOH (4.4mg, 0.184mmol) and stirred fbr Ih at room temperature. The solution was acidified with 1 N HC1 and the solvent was completely removed in vacuo. The residue was dissolved in 2 ml of pyridine, then 6-chloropyridine-2,3-diamine (10 mg, 0.069 mmol) and EDC hydrochloride (13.3 mg, 0.069 mmol) were added. The mixture was stirred at room temperature overnight. The solvent was removed on a rotary' evaporator. The residue was added in dichloromethane (10 ml) and washed with water (10ml). The organic layer was collected. The aqueous layer was extracted with dichloromethane (10ml) twice. The organic layer was combined and washed with brine. The organic layer was dried over NaaSO* and concentrated in vacuo. The solvent was removed in vacuo, the residue was purified by flash chromatography on silica gel to get 19.3 mg of Intermediate C.
[0187] Ethylamine HC1 salt (60 mg, 0.74 mmol) and DIPEA (0.19 ml, 1.11 mmol) were added to Intermediate C (19.3 mg, 0.0368mmol) in 2ml of anhydrous DMF. The solution was microwave irradiated at 110 °C for Ih. After the solvent was removed on the rotary’ evaporator, the residue was dissolved in 30 ml of EtOAc and washed with 10 ml of water, 30ml of brine and dried over NazSO*. The solvent was removed in vacuo, the residue was purified by flash chromatography on silica gel. The oily compound was dissolved in 5 ml of acetic acid and microwave irradiated at 125 °C for 45min. The solvent was removed in vacuo, the residue was purified by flash chromatography on silica gel to obtain 1.9 mg of 2634. LC/MS: (ESI) (M +H)+= 516.1.
Scheme 4
Reagents and conditions (a) NaOEt, EtOH, room temperature; (b) HOAc, microwave irradiation. General Procedure 3 (2630, 2627)
[0188] Sodium ethoxide (0.304 mmol) in anhydrous EtOH (1ml) was added to Intermediate C (20. Img, 0.038mmol) in 2ml of anhydrous EtOH. The solution was stirred at room temperature for 2h or microwave irradiation at 60 °C for 30 min. The solvent was removed in vacuo, the residue was purified by flash chromatography on silica gel. The oily compound was dissolved in 2 ml of acetic acid and microwave irradiated at 125 °C for 45min. The solvent was removed in vacuo, the residue was purified by flash chromatography on silica gel to obtain 2630 in 65% yield in two steps. *H NMR (500 MH2, CDCb) 5 11.95 (s, 1H), 7.88 (d, J= 8.0 Hz, 1H), 7.35 (s, 1H), 7.14 (s, 1H), 7.05 (s, 1H), 6.89 (s, 1H), 4.64 (s, 2H), 4.51 (s, 2H), 3.97 (s, 3H), 3.87 (s, 3H) ), 1.50 (s, 3H). LC/MS: (ESI) (M +H)+= 517.1.
[0189] 2627 was synthesized using sodium methoxide in anhydrous MeOH following General Procedure 3. NMR (500 MHz, CDCh) 5 11.31 (s, 1H), 7.89 (d, J= 8.0 H2, 1H), 7.29 (s, 1H), 7.20 (d, J= 8.5 Hz, 1H), 7.06 (s, 1H), 6.87 (s, 1H), 4.44 (s, 2H), 4.26 (s, 3H), 3.97 (s, 3H) ), 3.86 (s, 3H). LC/MS. (ESI) (M +H) = 503.1.
[0190] Reagents and conditions (a) 2-chloro-4,5-dimethoxyphenylboronic acid, K2CO3, Pd(dppf)Ch-DCM, dioxane, H2O, 110°C, Ih (b) NBS, chloroform, 0°C 2h; (c) ethyl 4- chloroacetoacetate, MeCN, 80°C, overnight; (d) LiOH, dioxane/water; (e) EDC, 6- chloropyridine-2,3-diamine, pyridine; (f) 2,4-dimethoxybenzylamine, MeCN/ethanol, microwave irradiation; (g) HOAc, microwave irradiation; (h) Py HF, NaNO2.
[0191] 2-Amino-6-bromopyrazine (337mg, 1.94 mmol), 2-chloro-4,5-dimethoxyphenylboronic acid (501mg, 2.32 mmol), K2CO3 (500mg, 3.62 mmol) and Pd(dppf)Ch-DCM (lOOmg, 0.123 mmol) in water (1.5ml) and dioxane (15ml) was microwave irradiated at 100 °C for Ih. After the solvents were removed, the residue was purified by flash chromatography on silica gel to obtain 6-(2-chloro-4,5-dimethoxy-phenyl)-pyrazin-2-ylamine (89%).
[0192] NBS (265mg, 1.49 mmol) was added in portion to 6-(2-chloro-4,5-dimethoxy-phenyl)- pyrazin-2-ylamine (415mg, 1.57 mmol) in 25ml of anhydrous chloroform at 0°C for 2h. After the solution was removed, the residue was purified by flash chromatography on silica gel eluted with EtOAc/hexane to obtain 3-bromo-6-(2-chloro-4,5-dimethoxy-phenyl)-pyrazin-2-ylamine (71%).
[0193] Ethyl 4-chloroacetoacetate (8ml) was added to 3-bromo-6-(2-chloro-4,5-dimethoxy- phenyl)-pyrazin-2-ylamine (220mg, 0.63mmol) solution in anhydrous MeCN (25ml). The mixture was heated at 80 °C overnight. After the solution was concentrated, the residue was stirred with hexane (20ml) and hexane was decanted. The procedure was repeated twice and the oil residue was purified by flash chromatography on silica gel eluted with EtOAc/hexane to obtain 5-(2-chloro-4,5-dimethoxy-phenyl)-8-chloro-imidazo[l,2-a]pyrazin-2-carboxylic acid ethyl ester, (79%).
[0194] 5-(2-chloro-4,5-dimethoxy-phenyl)-8-chloro-imidazo[l,2-tif]pyrazin-2-caiboxylic acid ethyl ester (187mg, 0.46 mmol) was added in 4 ml of dioxane and 4ml of water, mixed with LiOH (44mg, l.Smmol) in 1ml of water and stirred for Ih at room temperature. The solution was acidified with 1 N HC1 and the solvent was completely removed in vacuo. The residue was dissolved in 10 ml of pyridine, then 6-chloropyridine-2,3-diamine (lOOmg, 0.69 mmol) and EDC hydrochloride (133 mg, 0.69 mmol) were added. The mixture was stirred at room temperature overnight. The solvent was removed on a rotary evaporator. The residue was added in dichloromethane (20 ml) and washed with water (20ml). The organic layer was collected. The aqueous layer was extracted with dichloromethane (20ml) twice. The organic layer was combined and washed with brine. The organic layer was dried over Na2SO4 and concentrated in vacuo. The solvent was removed in vacuo, the residue was recrystallized in MeOH. The light- yellow solid was collected and washed with small amount of cold methanol to obtain 132 mg of Intermediate D. The mother liquid was purified by flash chromatography on silica gel to get 38 mg of Intermediate D.
[0195] 2,4-Dimethoxybenzylamine (68.5mg, 0.41mmol) was added to Intermediate D (10.3mg, 0.020 mmol) in 3ml of anhydrous MeCN and 1ml of anhydrous ethanol. The solution was microwave irradiated at 110 °C for 2h. After the solution was concentrated on the rotary evaporator, the residue was dissolved in 20 ml of EtOAc and washed with 10 ml of water, 20ml of brine and dried ove The solvent was removed in vacuo, the residue was purified by flash chromatography on silica gel. The oily compound was dissolved in 4 ml of acetic acid and microwave irradiated at 125 °C for 45min. The solvent was removed in vacuo, the residue was purified by flash chromatography on silica gel to obtain Intermediate E in 82% yield in two steps. Sodium nitrite (1.38mg, 0.02 mol) was added to Intermediate E (7.84mg, 0.0167 mmol) in 1 ml of Py HF solution at -5 °C. The temperature will gradually increase to room temperature. The reaction was performed for 2 days at room temperature. The solvent was removed in vacuo, the residue was purified by flash chromatography on silica gel to obtain 1 .22 mg of 2620. LC/MS: (ESI) (M +H)+= 473.1.
Reagents and conditions (a) LH-l,2,4-triazole, ethanol; microwave irradiation, 150°C 3h; (b) LiOH, dioxane/water; (c) EDC, 6-chloropyridine-2,3-diamine, pyridine; (d) HOAc, microwave irradiation.
General Procedure 4 (2695-2697)
[0196] The mixture of Intermediate B (5.1 mg, 0.012mmol), LH-l,2,4-triazole (16.6 mg, 0.24 mmol), in 1ml of ethanol was microwave irradiated at 150 °C for 3h. The solution was diluted in 20 ml of EtOAc and washed with 10 ml of water twice, 20ml of brine and dried over Na2SO4. The solvent was removed in vacuo, the residue was purified by flash chromatography on silica gel to obtain 3.2mg of 5-(2-chloro-4,5-dimethoxy-phenyl)-6-fluoro- yl)imidazo[l,2-a]pyrazin-2-carboxylic acid ethyl ester. [0197] 5-(2-chloro-4,5-dimethoxy-phenyl)-6-fluoro-8-(1H-I-l,2,4-triazole-l-yI)imidazo[l,2- a]pyrazin-2-caiboxylic acid ethyl ester (3.2mg , 0.007 mmol) was added in 1 ml of dioxane and 0.2ml of water, mixed with LiOH (0.67mg) and stirred for 40min at room temperature. The solution was acidified with 1 N HC1 and the solvent was completely removed in vacuo. The residue was dissolved in 1 ml of pyridine, then 6-chloropyridine-2,3-diamine (2.0 mg, 0.014 mmol) and EDC hydrochloride (2.0 mg, 0.011 mmol) were added. The mixture was stirred at room temperature overnight. The solvent was removed on a rotary evaporator. The residue was added in ethyl acetate (10 ml) and washed with water (10ml), 10ml of brine and dried over Na2SO4. The organic layer was dried over NazSO* and concentrated in vacuo. The residue was purified by flash chromatography on silica gel. The purified compound was dissolved in 1 ml of acetic acid and microwave irradiated at 125 °C for 45min. The solvent was removed in vacuo, the residue was purified by flash chromatography on silica gel to obtain 2.8mg of 2695. LC/MS: (ESI) (M +H)+= 540.2.
[0198] 2696 was synthesized using 3-methyl-LH-l,2,4-triazole following General Procedure
4. LC/MS: (ESI) (M +H)+= 554.4.
[0199] 2697 was synthesized using 3-ethyl-LH--l,2,4-triazole following General Procedure 4.
LC/MS: (ESI) (M +H) = 568.4.
Example 2; Preparation of Compounds of Formula (ID
Preparation of 2492
Reagents and conditions (a) DMF, K2CO3, 30% yield; (b) 2-chloro-4,5-dimethoxyphenylboronic acid, K3PO4, Pd(PPh3)4, dioxane, H2O, 110°C 3h; (c) LiOH, ethanoVwater; (d) EDC, 6- chloropyridine-2,3-diamine, pyridine; (e) HOAc, microwave irradiation.
[0200] To a solution of 4-bromo-lh-pyrazolo[3,4-c]pyridine (515mg, 2.6mmol) in anhydrous DMF (15ml) was added K2CO3 (538mg, 3.9mmol). The mixture was stirred at room temperature for 30min. Ethyl 2-bromoacetate (270μl, 2.47mmol) was added dropwise at room temperature. The reaction mixture was stirred at room temperature for 1.5 hours and the solvent was removed in vacuo. The residue was dissolved with EtOAc and washed successively with water and brine. The organic layer was dried over Na2SO4 and concentrated in vacuo. The residue was purified by flash column chromatography, giving ethyl (4-bromo-pyrazolo[3?4-c.’]pyridin-2-yl) acetate, in 30% yield (222 mg).
[0201] Ethyl (4-bromo-pyrazolo[3,4-c]pyridin-2-yl) acetate (113.6 mg, 0.40 mmol), 2-chloro- 4,5-dimethoxyphenylboronic acid (112mg, 0.52mmol), K3PO4 (254mg, 1.20 mmol) and tetrakis(triphenylphosphine)palladium(0) (23 mg, 0.02nunol) in water (1.2 ml) and dioxane (7.2 ml) was heated at 110 °C for 3h under N2. After most of organic solvent was removed in vacuo, the mixture was extracted with EtOAc and washed successively with water and brine. The organic layer was dried over Na2SO4 and concentrated in vacuo. The residue was purified by flash chromatography on silica gel to obtain ethyl (4-(2-chloro-4,5-dimethoxyphenyl)- pyrazolo[3,4-c]pyridin-2-yl) acetate (128mg).
[0202] Ethyl (4-(2-chloro-4,5-dimethoxyphenyl)-pyrazolo[3,4-c]pyridin-2-yl) acetate (50.3mg, 0.134 mmol) was added in 2 ml of ethanol and 6 ml of water, mixed with LiOH (12.8mg, 0.54 mmol) and stirred for Ih at room temperature. The solution was acidified with 1 N hydrochloric acid and the solvent was completely removed in vacuo. The residue was dissolved in 10 ml of pyridine, then 6-chloropyridine-2,3-diamine (38.6mg, 0.268 mmol) and EDC hydrochloride (38.6 mg, 0.20 mmol) were added. The mixture was stirred at room temperature overnight. The solvent was removed on a rotary evaporator. The residue was dissolved in EtOAc (30 ml) and washed with water and brine. The organic layer was dried over Na2SO4 and concentrated in vacuo. The residue was purified by flash chromatography on silica gel and further dissolved in 2 ml of acetic acid. The solution was microwave irradiated at 125 °C for 1 hour. After the solvent was removed on the rotary evaporator, the residue was purified by flash column chromatography to give 2492 (43mg). *H NMR (500 MH2, DMSO) 5 8.29 (s, 1H), 7.75 (s, 1H), 7.23 (s, 1H), 7.12 (s, 1H), 6.42 (d, J= 8.4 H2, 1H), 6.34 (s, 1H), 6.23 (s, 1H), 5.18 (s, 2H), 2.98 (s, 3H), 2.92 (s, 3H). LC/MS: (ESI) (M +H)+= 456.2.
Preparation of 2493
[0203] 2493 was synthesized using 6-fluoropyridine-2,3-diamine following General Procedure 5.
LC/MS: (ESI) (M +H)+= 439.6.
[0204] To a stirred solution of 2492 (188mg, 0.41 mmol) in CHCb (10ml) was added dropwise 77% mCPBA (55.5mg, 0.497mmol) in 2ml of CHCb at 0°C. The resulting mixture was stirred at 0°C for 3h. The reaction mixture was diluted with The solution was washed with 10% in saturated NaHCCb and brine. The resulting material was dried over and concentrated in vacuo. The residue was purified by flash column chromatography to give 2500 (90%) as light yellow solid. LC/MS: (ESI) (M +Hf= 472.2
Preparation of 2501 and 2540
[0205] POCb (10 ml) was added to 2500 (174mg, 0.37mmol). The mixture was stirred overnight at 50°C. After most solvent was removed in vacuo, the ice water was added. The mixture was extracted with EtOAc. dried over and concentrated in vacuo. The residue was purified by flash chromatography on silica gel to give 2501 in 90% yield TH NMR (500 MHz, DMSO) 5 7.89 (s, IH), 7.14 (d, J= 8.3 H2, IH), 7.04 (s, IH), 6.43 (d, J= 8.4 H2, IH), 6.35 (s, IH), 6.24 (s, IH), 5.22 (s, 2H), 2.98 (s, 3H), 2.92 (s, 3H). LC/MS: (ESI) (M +H)+= 490.5 and 2540 in 3% yield, LC/MS: (ESI) (M +H)+= 490.5.
[0206] 2513 was synthesized using 2493 following General Procedure 6-7. LC/MS: (ESI) (M +H) = 474.2.
Preparation of 2521
Scheme 9. General Procedure 8
H
[0207] To a solution of 2501 (lOmg, 0.02mmol) in 2ml of anhydrous ethanol was added 2- piperazinone (lOOmg, l.OOmmol). The mixture was microwave irradiated at 120 °C for 4 hour. After the solvent was removed on a rotary evaporator, the residue was extracted with EtOAc and washed successively with water and brine. The organic layer was dried over Na2SO4 and concentrated in vacuo. The residue was purified by flash chromatography on silica gel to obtain 2521 (90%). *H NMR (500 MHz, DMSO) 57.48 (d, J= 9.7 H2, 1H), 7.14 (s, 2H), 7.08 (s, 1H), 6.69 (s, 1H), 6.41 (d, J= 8.4 H2, 1H), 6.26 (s, 1H), 6.12 (s, 1H), 5.10 (d, J= 22.1 Hz, 2H), 3.69 (s, 2H), 3.41 (t, J= 5.3 Hz, 2H), 2.94 (s, 3H), 2.89 (s, 3H). LC/MS: (ESI) (M +H)+= 554.3.
[0208] 2502 was synthesized using Piperazine following General Procedure 8. LC/MS: (ESI) (M +H)+= 540.3.
Preparation of 2505
[0209] 2505 was synthesized using Ethanolamine following General Procedure 8. LC/MS: (ESI) (M +H)+= 515.2.
Preparation of 2517
[0210] 2517 was synthesized using morpholine following General Procedure 8. LC/MS: (ESI) (M +H)+= 541.5. Preparation of 2518
[0211] 2518 was synthesized using l-amino-2-methyl-2-propanol following General Procedure
8. LC/MS: (ESI) (M +H)+= 543.3.
Preparation of 2519
[0212] 2519 was synthesized l,2-diamino-2-methylpropane following General Procedure 8.
LC/MS: (ESI) (M +H)+= 542.3.
Preparation of 2520
[0213] 2520 was synthesized using 2,2-difluoroethylamine following General Procedure 8.
LC/MS: (ESI) (M +H)+= 535.1.
[0214] 2522 was synthesized using 2-(piperazin-l-yl)pyrazine following General Procedure 8.
LC/MS: (ESI) (M +H)4= 618.5.
Preparation of 2533
[0215] 2533 was synthesized using 3-oxa-8-azabicyclo[3.2.1]octane following General Procedure 8. >H NMR (500 MHz, CDCb) 57.97 (s, 1H), 7.92 (s, 1H), 7.62 (s, 1H), 7.28 (s, 1H), 6.94 (s, 1H), 6.85 (s, 1H), 5.95 (s, 2H), 5.29 (s, 2H), 3.91 (s, 3H), 3.88 (d, J= 10.8 Hz, 2H), 3.81 (s, 3H), 3.57 (d, J= 10.8 H2, 2H), 2.05 (d, J= 7.1 H2, 2H), 1.97 (d, J= 5.0 H2, 2H).
LC/MS: (ESI) (M +H)+= 567.2.
[0216] 2534 was synthesized using 4-difluoropiperidine following General Procedure 8.
LC/MS: (ESI) (M +H)+= 575.7.
Preparation of 2535
[0217] 2535 was synthesized using 8-oxa-3-azabicyclo[3.2.1]octane following General Procedure 8. LC/MS: (ESI) (M +H)+= 567.2.
Preparation of 2536
[0218] 2536 was synthesized using 2,6-dimethylpiperazine following General Procedure 8.
NMR (500 MHz, CDCh) 5 7.92 (d, J= 8.4 H2, 1H), 7.80 (s, 1H), 7.62 (s, 1H), 7.21 (d, J= 8.4 Hz, 1H), 6.92 (s, 1H), 6.81 (s, 1H), 5.90 (s, 2H), 5.07 (d, J= 12.2 Hz, 2H), 3.88 (s, 3H), 3.79 (s, 3H), 2.87 (m, 2H), 2.53 (t, J= 16.1 Hz, 2H), 1.05 (d, J= 6.3 H2, 6H). LC/MS: (ESI) (M +H) = 568.2.
Preparation of 2537
[0219] 2537 was synthesized using 4-aminotetrahydropyran following General Procedure 8.
LC/MS: (ESI) (M +H)+= 555.6.
Preparation of 2576
[0220] 2576 was synthesized using oxetan-3-amine following General Procedure 8.
LC/MS: (ESI) (M +H)+= 527.2.
[0221] 2584 was synthesized using 1,1-dioxide thiomorpholine following General Procedure 8.
LC/MS: (ESI) (M +H)+= 589.6.
Preparation of 2587
[0222] 2587 was synthesized using 4-piperidinemethanol following General Procedure 8.
LC/MS: (ESI) (M +H)+= 569.2.
Preparation of 2588
[0223] 2588 was synthesized using 2-oxo-2-(piperazin-l-yl)ethyl acetate following General Procedure 8. LC/MS: (ESI) (M +H)*= 640.1.
Preparation of 2597
[0224] 2597 was synthesized using 1-methanesulfonyl-piperazine following General Procedure
8. LC/MS: (ESI) (M +H)+= 618.6.
Preparation of 2598
[0225] 2598 was synthesized using 1-Boc-ethylenediamine following General Procedure 8.
LC/MS: (ESI) (M +H)+= 614.5.
Preparation of 2589
[0226] 2588 (10.3mg, 0.017 mmol) was added in 1 ml of methanol and 1 ml of water, mixed with LiOH (1.6mg, 0.067 mmol) and stirred for Ih at room temperature. The solution was acidified with 1 N hydrochloric acid and the solvent was completely removed in vacuo. The residue was purified by flash chromatography to give 2589 (8.6mg). LC/MS: (ESI) (M +H)+= 598.2. Preparation of 2585 y
[0227] To a solution of 2502 (5.4mg, O.Olmmol) and DIPEA (0.015mmol) in 2ml of anhydrous at 0°C was added chloromethanesulfonyl chloride (0.01 Inunol). The mixture was stirred at 0°C for Ih and room temperature overnight. The mixture was diluted with EtOAc and washed successively with saturated aqueous solution, water and brine. The organic layer was dried over and concentrated in vacuo. The residue was purified by flash chromatography on silica gel to obtain 2585 (62%). LCZMS: (ESI) (M +H) * = 653.1.
Preparation of 2586
(a) chlorosulphonylisocyanic acid/tert-butanol, DCM [0228] To a solution of chlorosulphonylisocyanic acid in 1ml of anhydrous at 0°C was added ice-cooling tert-butanol (5.5 pl). The mixture was stirred at 0°C for Ih. To the reaction mixture, 2502 (15.6mg, 0.029mmol) and DIPEA (0.0724mmol) were added and stirred overnight at room temperature. The mixture was diluted with EtOAc and washed with saturated dried over Na2SO4 and concentrated in vacuo. The residue was purified by flash chromatography on silica gel to obtain Boc-2586. The solution of Boc-2586 in 2ml of DCM was treated with TFA(lml) for Ih. After the solvent was removed, the residue was purified by flash chromatography on silica gel to obtain 2586 (30%). LC/MS: (ESI) (M +H)+= 619.4.
Preparation of 2539
[0229] To a solution of EtONa (O.Smmol) in EtOH (1ml) was added 2501 (8.0mg, 0.016mmol). The mixture was stirred at room temperature for 2h. The mixture was diluted with EtOAc and washed successively with 0.1 N HC1 solution, water and brine. The organic layer was dried over Na2SO* and concentrated in vacuo. The residue was purified by flash chromatography on silica gel to obtain 2539 (82%). LC/MS: (ESI) (M +H)+= 500.2.
Preparation of 2575 [0230] 2575 was synthesized using MeONa/MeOH following General Procedure 9.
LC/MS: (ESI) (M +H)+= 486.3.
Preparation of 2616
[0231] 2616 was synthesized using Sodium thiomethoxide/DMF following General Procedure 9.
LC/MS: (ESI) (M +H)+= 502.0.
Preparation of 2617
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[0232] To a stirred solution of 2616 (22.3mg, 0.045mmol) in CHCb (3ml) was added dropwise 77% mCPBA (9.98mg, 0.045mmol) in 1ml of at 0°C. The resulting mixture was stirred at 0°C for 3h. The reaction mixture was diluted with The solution was washed with 10% in saturated and brine. The resulting material was dried ove and concentrated in vacuo. The residue was purified by flash column chromatography to give 2617 (88%) as light yellow solid. LC/MS: (ESI) (M +H)+= 518.2. Preparation of 2618
[0233] To a stirred solution of 2617 (lO.Omg, 0.019nunol) i (3ml) was added dropwise 77% mCPBA (4.78mg, 0.021 mmol) in 1ml of at 0°C. The resulting mixture was stirred at 0°C for Ih, then room temperature for 2h. The reaction mixture was diluted with The solution was washed with 10% in saturated and brine. The resulting material was dried over and concentrated in vacuo. The residue was purified by flash column chromatography to give 2618 (90%) as light yellow solid. LC/MS: (ESI) (M +H)+= 534.4.
Preparation of 2579
[0234] To a solution of 1,2-ethanediol (0.5ml) in anhydrous dioxane (1.0ml) was added sodium (15mg) and stirred at room temperature until sodium is consumed in the fume hood. The above solution was added into the solution of 2501 (6.4mg, 0.013mmol) in 0.5ml of anhydrous dioxane. The mixture was microwave irradiated at 80°C for 20min. The mixture was diluted with EtOAc and washed with water and brine. The organic layer was dried over NazSO* and concentrated in vacuo. The residue was purified by flash chromatography on silica gel to obtain 2579 (30%). LC/MS: (ESI) (M +H)+ = 516.4.
Preparation of 2580
[0235] 2580 was synthesized using 2-methyl-propane-l,2-diol following General Procedure 10.
LC/MS: (ESI) (M +H)+= 544.4.
Example 3: Biological Assays
[0236] Media and culture conditions. Mueller Hinton broth (MHB), cation adjusted Mueller Hinton broth (CA-MHB), Todd Hewitt Broth (THB) and Brain Heart Infusion broth (BHI) were purchased from Becton Dickinson (Franklin Lakes, NJ). Tryptic soy agar (TSA) plates and TSA with 5% sheep blood plates were purchased from Remel (San Diego, CA). MHB was used to assay all Staphylococcus aureus strains. CA-MHB was used for Staphylococcus epidermidis, Enterococcus faecalis, Enterococcus faecium, Escherichia coli, and Pseudomonas aeruginosa. THB broth supplemented with 1% yeast extract was used for Streptococcus pneumoniae and Streptococcus pyogenes. Staphylococcus, Enterococcus, Escherichia, and Pseudomonas strains were cultured at 37°C with ambient air. Streptococcus strains were cultured in 37°C with 5% Separate conditions for radiolabeled precursor uptake assays are described below.
[0237] Enzyme assays. Inhibition of SaMetRS is measured using the ATP depletion assay as previously described with some modifications. Compounds are pre-incubated for 15 minutes at room temperature in a 96-well plate with 400 μg/mL bulk E. coli tRNA, 25 nM SaMetRS, 0.1 U/mL pyrophosphatase, 0.2 mM spermine, 0.1 mg/mL bovine serum albumin, 2.5 mM dithiothreitol, 25 mM HEPES-KOH pH 7.9, 10 mM MgCh, 50 mM KC1, and 2% DMSO.
Reagents are purchased from Sigma-Aldrich or Roche. The reaction is started with the addition of 150 nM ATP and 20 μM L-methionine and after 120 minute incubation is stopped by the addition of an equal volume (50 μL) of Kinase-Glo® (Promega). Percent inhibition = 100 x (test compound - AVG low control) / (AVG high control - AVG low control) where the low control is all reagents except the compound and the high control is all reagents except the compounds and L-methionine. IC6o values are calculated by non-linear regression, sigmoidal-dose response, in Prism 3.0.
[0238] Macromolecular synthesis assays: Methods for measuring uptake of radiolabeled precursors by S. aureus (ATCC strain 29213) are adapted from previous publications. For these assays, bacteria are grown in defined media (DM): RPM1-1640 pH 7.3 ± 0.1 without phenol red or L-glutamine (Lonza, Basel, Switzerland) supplemented with 4mM L-glutamine (Lonza, Basel, Switzerland), lOmM HEPES (Lonza, Basel, Switzerland), and 1% (w/v) D-glucose (Sigma Aldrich, St. Louis, MO). Fresh overnight cultures grown in DM at 37°C are diluted 1:50 in pre- warmed DM and grown at 37°C with shaking (150 rpm) until reaching an OD600 of 0.420 correlating to ~1* 109 CFUs/mL in mid-log phase. Each compound is assayed in quadruplicate with an 11 -point three-fold serial dilution per radioisotope. A pre-warmed 96-well V-bottom plate (Coming 3894; Coming, Coming, MA) containing 25 μL of 4x final concentration of test compound is inoculated with 65 μL of mid-log phase bacteria (OD600 of 0.420). Both positive and negative control wells received 25 μL untreated DM and 65 μL of inoculum at the same time. After one minute, 10 μL of radiolabeled precursor (10* final concentration in DM) is added to samples and positive control wells. Final isotope concentrations for assay of [3H]-lysine (protein), [3H] -thymidine (DNA), and [3H]-uridine (RNA) are 10 μCi/mL, 2 μCi/mL, and 2μCiAnL respectively. The plates are incubated at 37°C for 25 minutes and terminated by the addition of 50 μL of 30% trichloroacetic acid (TCA)/70% ethanol to all test and control wells. After termination, 10 μL of 10x radiolabeled precursor is added to negative control wells. The negative control consisted of adding radiolabeled precursors after termination of the bacterial incubation in order to represent background measurement of the isotope. Plates are sealed with plate tape (Thermo Fischer Scientific, Waltham, MA) and shaken at 250 rpm for one hour at room temperature. Aliquots of 125 μL are transferred from the 96-well V-bottom plates to 96- well filter plates (Merck Millipore, Billerica, MA). To bind macromolecules, the samples are passed through the filter membrane (0.45μM hydrophilic Durapore PVDF membrane) with a vacuum manifold, then the filter is washed with 4 x 200 μL 10% TCA and 1 x 150 μL of 95% ethanol, and dried overnight in vacuum at room temperature. 25 μL Ultima Gold scintillation fluid (Perkin Elmer, Waltham, MA) is added to each well and DPM is quantified using a MicroBeta2-2450 (Perkin Elmer, Waltham, MA) scintillation counter. The percent incorporation is determined by subtracting each well by the average negative background and dividing by the average positive incorporation x 100. Error bars represent SEM between replicates. The assay is run twice with similar results.
[0239] Susceptibility testing: Minimum inhibitory concentration (MIC) determinations are performed in triplicate in 96-well round bottom microtiter plates (Coming, Coming NY) as described by the Clinical and Laboratory Standards Institute (CLSI). Serial two-fold dilutions of compounds are added to plates in 50 μL volumes. An additional 50 μL of media containing bacterial cells (lx 106 CFUs/mL) is then added to each well. Maximum DMSO concentrations are 0.5%. Plates are incubated at 37°C for at least 18 h before reading the susceptibility result by optical absorbance (OD600) using a BioTek ELx800. The lowest concentration causing >90% growth inhibition compared to the untreated control is recorded as the MIC (and also corresponded to the visual MIC). MIC6 are measured at least twice and the higher value (if different) is recorded herein.
[0240] Cytotoxicity testing on mammalian cells: Compounds are assayed for cytotoxicity against CRL-8155 (human lymphoblasts) and HepG2 cells (human hepatocellular carcinoma). C6lls are exposed to serial dilutions of compounds for 48 hours and toxicity is quantified using AlamarBlue (ThermoFisher Scientific, Waltham, MA). Assays are performed in quadruplicate and EC6o values are calculated with non-linear regression methods using software by the Collaborative Drug Database (Burlingame, CA. www.collaborativedrug.com)
[0241] Resistance frequency rates determination: The spontaneous resistance frequency rates to test compounds is determined according to published methods. Agar selection plates are made by adding compound from DMSO stocks into molten Mueller Hinton agar in a 55°C water bath. Each compound used four plates (P5981-100EA, 150 x 15mm; Sigma Aldrich, St. Louis, MO) containing multiples of the MIC (16x, 8x, and 4x) of the compound. The final DMSO concentration is < 0.1 % per plate. Plates are dried in a sterile hood for 30 minutes prior to overnight storage at 4°C, and pre-warmed in the 37°C incubator for 1 hour prior to assay.
[0242] A fresh overnight culture is diluted 1:50 in MHB and grown at 37°C with shaking (150 rpm) until reaching an OD600 of 0.4 correlating to ~2x 109 CFUs/mL. Approximately 3 mL for a total of 6* 109 CFUs are distributed onto 4 plates for each compound. Plates are incubated at 37°C for 72 h prior to counting of colonies. The starting inoculum is also serially diluted and plated to quantify initial bacterial load. The resistance frequency is determined as the number of compound-resistant colonies divided by the total colonies plated.
[0243] Serum shift assays: To assess the role of protein binding on compound susceptibility, MIC determinations are performed in triplicate in the presence and absence of 50% human serum. Serial threefold dilutions of 2* compound are generated in MHB or THY (depending on organism) and aliquoted onto 96 well plates with a DMSO limit of 0.5%. Bacteria are adjusted to 1*10* CFUs/mL in media, then further diluted 1 : 100 in media and 100% heat deactivated filter sterilized pooled human serum. Fifty' microliters are added to each well of the corresponding plates, and the plates are incubated at 37°C for ~20 h. The lowest concentration causing >90% growth inhibition is recorded as the MIC.
[0244] Protein binding assays: Compound binding to mouse plasma proteins is determined using 96-well equilibrium dialyzer plates (SDIS 9610EN, Nest Group, Inc.). Mouse plasma (BioreclamationIVT, Westbury, NY) containing compound (final concentration 1 μM) is added to a donor chamber as a 150 μL volume. The buffer solution (0.2 mM phosphate buffer, 150 μL) is added to the reciprocal acceptor chamber. Each compound is tested in triplicate. To prepare calibration solution for compound quantifications, blank wells are prepared containing only mouse plasma in a donor well and buffer solution in its acceptor well. The equilibrium dialysis is carried out by rocking the plate for 22 hours in 37 °C. Once equilibrium is reached, the plasma and buffer solution from both wells are carefully removed for further analysis with liquid chromatography-tandem mass spectrometry. Plasma solution and internal standard are mixed in the presence of 80% acetonitrile. After centrifuging the solution, the supernatant is transferred to an insert. Similarly, the buffer solution from the acceptor side is prepared containing 40% acetonitrile. Calibration standards for donor and acceptor sides are prepared with compound concentrations of 50 nM, 100 nM, 250 nM, 500 nM, and IμM. The compound concentrations from each well are calculated from the calibration curves using Microsoft Excel. The percentage of the test compound bound is determined as follows:
% Free = (Concentration buffer chamber/Concentration plasma chamber) x 100%
% Bound = 100% - % Free
[0245] Microsome stability. Liver microsome stability assays are done by contract research laboratory, Wuxi AppTec Co. (Hubei, China). Briefly, compounds at 1 μM concentration are incubated in singlet with human or CD-I mouse liver microsomes for 6 time points (0, 5, 10, 20, 30, and 60 min). Loss of parent compound is quantified by liquid chromatography /tandem mass spectrometry.
[0246] Murine pharmacokinetics studies. The methods are performed as previously described. Briefly, test compounds are administered to mice by oral gavage (3 mice per compound) followed by tail blood sampling at intervals of 30, 60, 120, 240, 360, 480, and 1440 min. Blood samples are analyzed by extracting dried blood spots in acetonitrile for measurements of compound concentrations by liquid chromatography /tandem mass spectrometry.
[0247] Murine thigh infection model. Animal studies are approved by the Institutional Animal Care and Use Committee at the University of Washington, Seattle. Female specific pathogen free CD1 mice are obtained from Charles River (Wilmington, MA) weighing 23-27 grams and allowed at least 3 days to acclimate prior to study. Mice had access to food and water ad libitum. Neutropenia is induced by administering cyclophosphamide monohydrate (Sigma Aldrich C7397; St. Louis, MO) via IP injection 4 days (at 150 mg/kg) and 1 day (at 100 mg/kg) prior to infection. Neutropenic status is confirmed by neutrophil count < 100 cells/mm3.
[0248] Overnight culture of luminescent MRSA (strain AH4827 is diluted 1:100 in MHB and incubated until reaching mid-log phase ( OD600 < 0.750). The inoculum is prepared by pelleting log-phase culture and re-suspending in sterile dPBS. The culture is adjusted to OD600 of 0.200 and diluted 1 : 100 in sterile dPBS correlating to an inoculum of ~2* 105 CFU/100 μL. The mice are infected by an intramuscular injection of 100 μL in the right posterior thigh while under isoflurane gas anesthesia. At 1 h post infection, one control group is sacrificed for determination of initial inoculum (status level of infection) after all mice are imaged by IVIS to confirm infection. Mice are dosed at 2 and 14 h post-infection or 2, 10, and 18 h post-infection with test compounds (below); they received a SC dose of 5mg/kg ketoprofen at 2 h post-infection for pain management. Mice are imaged by IVIS at 24 h post-infection then sacrificed; the thigh muscle is sterilely removed, weighed, homogenized in 5mLs dPBS, serially diluted, plated on tryptic soy- agar in duplicate, and incubated overnight at 37 °C. Colonies are counted to quantity the bacterial load in CPUs per gram of thigh tissue.
[0249] Linezolid as positive control is administered 50 mg/kg PO according to dosing regimen used for experimental compounds.
[0250] Test compounds and linezolid control are administered PO (orally) or SC (by subcutaneous injection) in 200μL of vehicle containing 7%Tween80, 5%DMSO, 3%EtOH in saline. [0251] Trypanosoma brucei growth inhibition assay: Compounds are tested for anti- trypanosomal activity against Trypanosoma brucei cells (BSF 427 parasites) in HMI-9 media containing 10% heat inactivated FBS 100 U mL 1 penicillin/100 μg mL 1 streptomycin at 37 °C with 5% CCh. Test compounds are assayed in a 96-well format in triplicate with serial 3-fold dilutions of compound or a pentamidine control against an initial inoculum of 10000 cells/well and quantified at 48 hours with Alamar Blue (Invitrogen, Waltham, MA).
[0252] Trypanosoma cruzi growth inhibition assay: The Tulahuen strain of T. cruzi that expresses the Escherichia coli P-galactosidase gene was utilized (PMID: 8913471) . Mammalian stages of parasites were grown at 37°C on monolayers of murine 3T3 fibroblasts in RPMI 1640 (Biowhittaker Inc., Walkersville, Md.) with 10% fetal bovine serum, penicillin, streptomycin, and glutamine. Growth inhibition assays were performed in 96-well tissue culture plates (Costar, Cambridge, Mass.). First, 3T3 fibroblasts were inoculated at 103/well using RPMI 1640 without phenol red (Biowhittaker Inc.) plus 10% fetal bovine serum and glutamine. The next day, the plates were infected with T. cruzi trypomastigotes at 104/well. After 4 h, drugs were added in serial dilutions to give a final volume of 200 plAvell. The plates were incubated at 37°C in 5% CO2 atmosphere for 6 days. At this time, CPRG (100 μM final) and Nonidet P-40 (0.1% final) (Sigma Chemical Co ., St. Louis, Mo.) were added and the plates were incubated at 37°C for approximately 4 h. Wells with p-galactosidase activity turned the media from yellow to red, and this was quantified on an enzyme-linked immunosorbent assay reader at A570.
[0253] In vivo acute T. cruzi infection model. Swiss Webster mice were infected with 2* 104 Trypanosoma cruzi trypomastigotes expressing a red-shifted luciferase gene IP on day zero (M.D. Lewis et al. J Biomol Screen 2015; 20(l):36-43). Mice were dosed orally with the test compounds, vehicle, or the control drug (benznidazole) for 5 days (days 7-11 post- infection). The parasites were imaged and quantified using an IVIS imaging system at multiple time points. The substrate D-luciferin potassium salt was injected SC at 150 mg/kg into mice, followed by imaging 10-15 minutes post injection. Images were taken for each set of mice both dorsally and ventrally. Auto-exposure settings are used with a maximum exposure time set to 5 minutes. The signal from these images is combined and standardized by converting the readouts to radiance (photons/sec/cm2/sr). The decrease in luminescence compared to vehicle control was quantified on day 13 post-infection.
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[0254] Results from Table 1 indicated useful in vivo efficacy of fluorinated 2541 when compared to other non-fluorinated analog compounds despite their similar MIC6 in serum shift assays.
[0255] The results of additional in vitro anti-bacterial testing of more examples of R2 variations are provided in Table 2.
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[0256] Table 2 shows modifications of R2 in Formula (I) with either aromatic or non-aromatic groups can retain the the antibiotic activities with the specific 6-position fluorination on the imidazopyrazine ring.
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[0257] The results of the in vitro growth inhibition and the in vivo acute T.cruzi infection model for compounds of Formula (I) are shown in Table 3 above. The results again revealed that specifically fluorinated (6-F) analogs such as 2541 or 2624 have useful in vivo efficacy compared to non-fluorinated analogs, including close pairwise analogs such as 2503 and 2602. In addition, in contrast to the beneficial specific fluorination at the 6-position on the imidazopyrazine ring as demonstrated in 2541, fluorination at another position, such as in 2620 with 8-F on the imidazopyrazine ring, led to more than an order of magnitute loss of in vitro potentcy when compared to either 2541 or the parent 2503. As in the case of in vitro antibiotic activities, modifications of R2 with 6-F on the imidazopyrazine can maintain in vitro anti-7’. cruzi activities.
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[0258] The results in Table 4 shows that various modifications according to Formula (II) demonstrate antibitotic and antiparasitic activities in vitro.
Examole 4: Svnereism of 2541 with fusidic acid for anti-staohvlococcal activity
[0259] Methods for Time-kill synergy assays: 2541 and fusidic acid were tested against MRSA strain (ATCC 33591). Combination solutions were made by adding both compounds from DMSO stocks into 5ml of CA-MHB at 0.25X, and 0.125X the MIC concentration of the compounds respectively. Individual test solutions were also made for each compound at 0.25X, and 0.125X their MIC concentrations. All of the test solutions were made in 12ml snap cap falcon tubes and were vortexed for a minute to ensure the compound was folly mixed. The test organism was then taken from a fresh overnight culture and added to each tube at a starting OD of 0.04, which is equivalent to 1X105 CFU/ml. A no drug control was also started at this time by adding the test organism to 5ml of CA-MHB that at the same starting concentration. Tubes were incubated at 37°C on a shaker, 200uL samples of the test solutions were taken at 0, 1, 3, 5, 7, and 24 hours after the bacteria was added and the OD600 reading was recorded at these times. At each of these time points the 200ul sample was also plated in serial 10 dilutions on fresh TSA plates. The plates were incubated at 37°C for 18 hours and then the colonies were counted to determine the CFU/ml at each of the time points. The resulting growth curves were graphed and synergy was determined to have occurred if the combination of two test compounds resulted in a greater than 2-log drop from the growth control curve when compared to the individual compound solution as described elsewhere (Zhang, R., et al., Synergy Between Beta-Lactams and Lipo-. Glyco-, and Lipoglycopeptides, Is Independent of the Seesaw Effect in Methicillin- Resistant Staphylococcus aureus. Front Mol Biosci, 2021. 8: p. 688357).
[0260] 2541 and the commercial antibiotic, fosidic acid, have synergistic antibacterial activity against the MRSA strain (ATCC 33591) (FIG. 1). This is demonstrated by decrease in CFU counts at 24 hr by >2 log units by the combination compared to the drugs by themselves (Id.). These results demonstrate the potential for 2541 to enhance the activity of other commercial antibiotics and vice versa. Multiple potential benefits can arise from combining antibiotics, particularly with synergistic activity, including 1) useful efficacy over individual compounds (Tyers, M. and G.D. Wright, Drug combinations: a strategy to extend the life of antibiotics in the 21st century. Nat Rev Microbiol, 2019. 17(3): p. 141-155), 2) potential for dose reduction of one or both antibiotics to reduce risk of side effects or drug interactions, 3) broaden the spectrum of activity (Huber, J., et al., Chemical genetic identification of peptidoglycan inhibitors potentiating carbapenem activity against methicillin-resistant Staphylococcus aureus. Chem Biol, 2009. 16(8): p. 837-48), and 4) reduced risk of antibiotic resistance developing (Tyers et al. (2019); Coates, A.R.M., et al., Antibiotic combination therapy against resistant bacterial infections: synergy, rejuvenation and resistance reduction. Expert Rev Anti Infect Ther, 2020. 18(1): p. 5- 15).
[0261] It is understood that the examples and embodiments described herein are for illustrative purposes only and that various modifications or changes in light thereof will be suggested to persons skilled in the art and are to be incorporated within the spirit and purview of this application and scope of the appended claims. All pubheations, patents, and patent applications cited herein are hereby incorporated herein by reference for all purposes.

Claims

We claim:
1. A compound of the formula (I): or a pharmaceutically acceptable salt thereof, wherein
Ri is halogen;
X is Cl or F; and R2 is hydrogen, halogen, -NO2, -CN, C1-C6 alkyl, C1-C6 haloalkyl, -NH2, -(C1-C6 alkyl)NH2, -NH(C1-C6 alkyl), -NH(C2-C6 alkyl)OH, -NH(C2-G6 alkyl)NH2, -N(C1-C6 alkyl)2, -OH, hydroxy(C1-C6 alkyl), C1-C6 alkoxy, C1-C6 haloalkoxy, -O(C2-C6 alkyl)OH, and -0(C2- C6 alkyl)NH2, R4C(0)(Co-C6 alkyl)-, R4C(0)0(Co-C6 alkyl)-, R4C(0)NH(Co-C6 alkyl)-, R4S(0)(Co-C6 alkyl)-, R4S(0)0(Co-C6 alkyl)-, R4S(0)NH(Co-C6 alkyl)-, R4S(0)2(Co-C6 alkyl)- R, 4 S(0)20(Co-C6 alkyl)-, R4S(0)2NH(Co-C6 alkyl)-, aryl(Co-C6 alkyl)-, heteroaryl(Co- C6 alkyl)-, or heterocyclyl(Co-C6 alkyl)-, and wherein each aryl, heteroaryl, or heterocyclyl moiety is optionally substituted with one or more R3; wherein each R3 is independently selected from halogen, -NO2, -CN, oxo, thioxo, C1-C6 alkyl, C1-C6 haloalkyl, -NH2, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, amino(C1-C6 alkyl), -NH(C2-C6 alkyl)OH, -NH(C2-C6 alkyl)NH2, -OH, hydroxy(C1-G» alkyl), C1-G6 alkoxy, C1-C5 haloalkoxy, -O(C2-C6 alkyl)OH, -O(C2-C6 alkyl)NH2, R4C(0)(Co-C6 alkyl)-, R4C(0)0(Co-C6 alkyl)-, R4C(0)NH(Co-C<5 alkyl)-, R4S(0)(Co-C6 alkyl)-, R4S(0)0(Co-C6 alkyl)-, R4S(0)NH(Co-C6 alkyl)-, R4S(0)2(Co-C6 alkyl)-, R4S(0)20(Co- C6 alkyl)-, and R4S(0)2NH(Co-C6 alkyl)-; and each Rt is independently selected from C1-C6 alkyl, C1-C6 haloalkyl, -NH2, -NH(C1-C6 alkyl), -NH(C2-C6 alkyl)OH, -NH(C2-C6 alkyl)NH2, -N(C1-C6 alkyl)2, -OH, -(C1-C6 alkyl)OH, C1-C6 alkoxy, C1-C6 haloalkoxy, -O(C2-C6 alkyl)OH, and -O(C2-C6 alkyl)NH2.
2. The compound of claim 1, wherein Ri is chloro or fluoro.
3. The compound of claim 1, wherein Ri is chloro.
4. The compound of claim 1, wherein Ri is fluoro.
5. The compound according to any of claims 1 to 4, wherein R2 is hydrogen.
6. The compound according to any of claims 1 to 4, wherein R2 is hydrogen, halogen, -NOa, -CN, C1-C6 alkyl, C1-C6 haloalkyl, -NHa, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, -OH, C1-C6 alkoxy, C1-C6 haloalkoxy, aryl(Co-C6 alkyl), heteroaryl(Co-C6 alkyl), or heterocyclyl(Co-C6 alkyl), and wherein each aryl, heteroaryl, or heterocyclyl moiety is optionally substituted with one or more R3; wherein each each R3 is independently selected from halogen, -NOa, -CN, C1-C6 alkyl, C1-C6 haloalkyl, -NHa, -NH(C1-G6 alkyl), -N(C1-C6 alkyl)2, -OH, C1-C6 alkoxty, and C1-C6 haloalkoxy.
7. The compound according to any of claims 1 to 4, wherein R2 is hydrogen, halogen, -CN, C1-C6 alkyl, C1-C6 haloalkyl, aryl(Co-C6 alkyl), heteroaryl(Co-C6 alkyl), or heterocyclyl(Co-C6 alkyl), and wherein each aryl, heteroaryl, or heterocyclyl moiety is optionally substituted with one or more R3.
8. The compound according to claim 7, wherein R2 is hydrogen, halogen, -CN, C1-C6 alkyl, or C1-C6 haloalkyl.
9. The compound according to any of claims 1 to 4, wherein R2 is hydrogen, aryl(Co-C6 alkyl), heteroaryl(Co-C6 alkyl), or heterocyclyl(Co-Q alkyl), and wherein each aryl, heteroaryl, or heterocyclyl moiety is optionally substituted with one or more R3.
10. The compound of any of claims 1-4, wherein R2 is hydrogen or heteroaryl(Co-C6 alkyl).
11. The compound according to any of claims 1-4, wherein X is F.
12. The compound according to any of claims 1-4, wherein X is Cl.
13. The compound according to claim 1, which is:
or a pharmaceutically acceptable salt thereof.
14. The compound of claim 1, which is:
5-chloro-2-((5-(2-chloro-4,5-dimethoxyphenyl)-6-fluoroimidazo[l,2-a]pyrazin-2- yl)methyl)-lH-imidazo[4,5-b]pyridine, or a pharmaceutically acceptable salt thereof.
15. The compound of claim 1, which is:
5-chloro-2-((5-(2-chloro-4,5-dimethoxyphenyl)-6-fluoro-8-(l,3,5-trimethyl-lH-pyrazol-4- yl)imidazo[l,2-a]pyrazin-2-yl)methyl)-lH-imidazo[4,5-b]pyridine or a pharmaceutically acceptable salt thereof.
16. A pharmaceutical composition comprising one or more compounds according claim 1 and a pharmaceutically acceptable carrier, diluent, or excipient.
17. A phmarmaceutical composition according to claim 16 further comprising one or more additional antibacterial compounds.
18. A compound of the formula (II): or a pharmaceutically acceptable salt thereof, wherein m is an integer 0, 1, or 2; n is an integer 2, 3, 4, or 5;
A is a heteroaiyl group optionally substituted with one or two R2; each Ri is independently selected from halogen, -CN, C1-C6 alkyl, C1-C6 haloalkyl, -NH2, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, -OH C1-C6 alkoxy, C1-C6 haloalkoxy, -SH -S(C1-C6 alkyl), hydroxy(C1-C6 alkyl), C1-C6 alkoxy(C1-C6 alkyl), and amino(C1-C6 alkyl); and
R3 is halogen, -NO2, -CN, C1-C6 alkyl, C1-C6 haloalkyl, -NH2, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, -OH C1-C6 alkoxy, C1-C6 haloalkoxy, hydroxy(C1-C6 alkyl), C1-C6 alkoxy(C1-C6 alkyl), amino(C1-C6 alkyl), -C1-C6 alkyl-NH(C1-C6 alkyl), -C1-C6 alkyl-N(C1-C6 alkyl)2, -CONH2, -CONH(C1-C6 alkyl), -CON(C1-C6 alkyl)2, -CO(C1-C6 alkyl), -NHCO(C1- C6 alkoxy), -NHCO(C1-C6 alkyl), -NHCONH2, -NHCONH(C1-C6 alkyl), -S(0)o-2-(C1-C6 alkyl), -S(O)1-2NH(Co-C6 alkyl), -S(O)1-2N(Co-C6 alkyl)2, -NH-S(O)1-2-(C1-C6 alkyl), -NH- S(O)i-2NH(C1-C6 alkyl), -NH-S(O)i-2-aryl, -NH-S(O)i-2-heteroaryl, aryl(Co-C6 alkyl), heteroaryl(Co-C6 alkyl), or heterocyclyl(Co-C6 alkyl), and wherein each alkjd, aryl, heteroaryl, or heterocyclyl moiety is optionally substituted with one or more Rt; wherein each R2 is independently selected from halogen, -NO2, -CN, C1-C6 alkyl, C1-C6 haloalkyl, -NH2, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, -OH, C1-C6 alkoxy, and C1-C6 haloalkoxy. each R* is independently selected from independently selected from halogen, -NO2, -CN, C1- C6 alkyl, C1-C6 haloalkyl, -NH2, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, -OH oxo, C1-C6 alkoxy, C1-C6 haloalkoxy, -SH thioxo, -S(C1-C6 alkyl), hydroxy(C1-C6 alkyl), C1-C6 alkoxy(C1-C6 alkyl), amino(C1-C6 alkyl), -CH2-NH(C1-C6 alkyl), -CH2-N(C1-C6 alkyl)2, -CONH2, -CONH(C1-C6 alkyl), -CON(C1-C6 alkyl)2, -CONH-OH, -COH, -CO2H, -CO2(C1-C6 alkyl), -CO(C1-C6 alkyl), -OCO(C1-C6 alkyl), -OCONH(C1-C6 alkyl), -NHCO(C1-C6 alkoxy), -NHCO(C1-C6 alkyl), -NHCONH2, -NHCONH(C1-C6 alkyl), -NH-S(O)1-2-(C1-C6 alkyl), -NH-S(O)1-2-aryl, -NH-S(O)1-2-heteroaryl, -CH2- NHCONH2, -CH2-NHCONH(C1-C6 alkyl), and -CH2
19. The compound of claim 18, wherein each Ri is independently selected from halogen, C1- C6 alkyl, C1-C6 haloalkyl, -NH2, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, -OH, C1-C6 alkoxy, and C1- C6 haloalkoxy.
20. The compound of claim 18, wherein each Ri is independently selected from halogen, C1- C6 alkyl, C1-C6 haloalkyl, -OH, C1-C6 alkoxy, and C1-C6 haloalkoxy.
21. The compound of claim 18, wherein each Ri is independently selected from halogen, C1- C6 haloalkyl, -OH, and C1-C6 alkoxy.
22. The compound of claim 18, wherein each Ri is independently selected from halogen and C1-C6 alkoxy.
23. The compound according to any of claims 18 to 22, wherein n is 2 or 3.
24. The compound according to any of claims 18 to 22, wherein n is 2.
25. The compound according to any of claims 18 to 22, wherein n is 3.
26. The compound according to any of claims 18 to 22, which is of formula:
27. The compound according to any of claims 18 to 26, wherein A is a 9-member heteroaryl group optionally substituted with one or two R2.
28. The compound according to any of claims 18 to 26, wherein A is indole, azaindole, benzimidazole, or imidazopyridine, each optionally substituted with one or two R2.
29. The compound according to any of claims 18 to 26, wherein A is indole, azaindole, benzimidazole, or imidazopyridine, each optionally substituted with one or two R2.
30. The compound according to any of claims 18 to 26, wherein A is imidazopyridine, optionally substituted with one or two R2.
31. The compound according to any of claims 18 to 22, which is of formula:
32. The compound according to any of claims 18 to 22, which is of formula:
33. The compound according to any of claims 18 to 32, wherein each R2 is independently selected from halogen, C1-C6 alkyl, C1-C6 haloalkyl, -OH, C1-C6 alkoxy, and C1-C6 haloalkoxy.
34. The compound according to any of claims 18 to 32, wherein each R2 is independently selected from halogen, C1-C6 haloalky 1, -OH, and C1-C6 alkoxy.
35. The compound according to any of claims 18 to 32, wherein each R2 is independently selected from halogen and C1-C6 alkoxy.
36. The compound according to any of claims 18 to 32, wherein each R2 is independently halogen.
37. The compound according to any of claims 18 to 32, wherein A is substituted with one R2, which is chloro.
38. The compound according to any of claims 18 to 22, which is of formula:
39. The compound according to any of claims 18 to 38, wherein m is 0.
40. The compound according to any of claims 18 to 38, wherein m is 0 or 1.
41. The compound according to any of claims 18 to 38, wherein m is 1.
42. The compound according to claim 41, which is of formula:
43. The compound according to any of claims 18 to 38, wherein m is 2.
44. The compound according to claim 43, which is of formula:
45. The compound according to any of claims 18 to 44, wherein R3 is halogen, -NH(C1-C6 alkyl), -N(C1-C6 alkyl)2, -OH, C1-C6 alkoxy, C1-C6 haloalkoxy, hydroxy(C1-C6 alkyl), C1-C6 alkoxy(C1-C6 alkyl), amino(C1-C6 alkyl), -C1-C6 alkyl-NH(C1-C6 alkyl), -C1-C6 alkyl-N(C1-C6 alkyl)2, -S(O)i-2-(C1-C6 alkyl), aiyl(Co-C6 alkyl), heteroaryl(Co-C6 alkyl), or heterocyclyl(Co-C6 alkyl), and wherein each alkyl, aryl, heteroaryl, orheterocyclyl moiety is optionally substituted with one or more R4.
46. The compound according to claim 18, which is:
or a pharmaceutically acceptable salt thereof.
47. A pharmaceutical composition comprising one or more of compounds according to any of claims 18-46 and a pharmaceutically acceptable carrier, diluent, or excipient.
48. A phmarmaceutical composition according to claim 47 further comprising one or more additional antibacterial compounds.
49. A method for inhibiting MetRS in a pathogen, the method comprising exposing the pathogen to an effective amount of a compound according to any of claims 1-15 or claims 18-46.
50. The method of claim 49, wherein the pathogen is a bacteria or a protozoa.
51. A method for treating diseases that are ameliorated by the inhibition of MetRS, the method comprising administering to a patient in need thereof a therapeutically effective amount of a compound according to any of claims 1-15 or claims 18-46 or a pharmaceutical composition of any of claims 16-17 or claims 47-48.
52. The method of claim 51 further comprising administering one or more additional antibacterial compounds in combination with the compound or the pharmaceutical composition.
53. The method of claim 51 further comprising administering a second antibacterial compound in combination with the compound or the pharmaceutical composition.
54. The method of claim 53, wherein the compoun d i the second antibacterial compound is fusidic acid.
55. The method of claim 51, wherein the disease is a protozoan infection.
56. The method of claim 55, wherein the protozoan infection is selected from the group consisting of Cryptosporidia, Cyclospora, Giardia, Leishmania, Trichomonas, and Trypanosoma.
57. The method of claim 51, wherein the disease is a bacterial infection.
58. The method of claim 57, wherein the bacteria is a Gram positive bacteria.
59. The method of claim 58, wherein the Gram positive bacteria is selected from one of Staphylococcus, Streptococcus, Enterococcus, Clostridia, Bacillus, Listeria, Corynebacteria, Arcanobacteria, Rothia, and Rhodococcus.
60. The method of claim 57, wherein the bacteria is a Gram negative bacteria.
61. The method of claim 60, wherein the Gram negative bacteria is Brucella, Campylobacter, and Helicobacter.
62. The method of claim 57, wherein the bacteria is Mycobacteria.
63. The method of claim 62, wherein the Mycobacteria is selected from M. tuberculosis, M. avium, M. abscessus, M. kansasii, M. chelonae, M. marinum, M. ulcerans, and M. haemophilum.
64. The method of claim 57, wherein the bacteria is Mycoplasma.
65. The method of claim 51, wherein the disease is a Trypanosomatid protozoa infection.
66. The method of claim 65, wherein the Trypanosomatid protozoa is one of Trypanosoma brucei, Trypanosoma cruzi, and Leishmania species.
67. The method of claim 57, wherein the bacteria is selected from Staphylococcus aureus, methicillin resistant Staphylococcus aureus (MRSA), Enterococcus faecalis, Enterococcus faecium, and vancomycin resistant Enterococcus (VRE).
68. A compound selected from the group consisting of or a pharmaceutically acceptable salt thereof.
69. A pharmaceutical composition comprising one or more compounds according to claim 68 and a pharmaceutically acceptable carrier, diluent, or excipient.
70. A method for inhibiting MetRS in a pathogen, the method comprising exposing the pathogen to an effective amount of a compound according to claim 68.
71. A method for treating diseases that are ameliorated by the inhibition of MetRS, the method comprising administering to a patient in need thereof a therapeutically effective amount of a compound according to claim 68 or a pharmaceutical composition of claim 69.
EP24811541.2A 2023-05-24 2024-02-07 Inhibitors of type 1 methionyl-trna synthetase and methods of using them Pending EP4719382A1 (en)

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