WO2017194734A1 - Means and methods for treating mycobacterial diseases - Google Patents

Means and methods for treating mycobacterial diseases Download PDF

Info

Publication number
WO2017194734A1
WO2017194734A1 PCT/EP2017/061453 EP2017061453W WO2017194734A1 WO 2017194734 A1 WO2017194734 A1 WO 2017194734A1 EP 2017061453 W EP2017061453 W EP 2017061453W WO 2017194734 A1 WO2017194734 A1 WO 2017194734A1
Authority
WO
WIPO (PCT)
Prior art keywords
substituted
unsubstituted
alkyl
mycobacterium
group
Prior art date
Application number
PCT/EP2017/061453
Other languages
French (fr)
Inventor
Knud ESSER
Jan VOMACKA
Norbert Reiling
Protzer ULRIKE
Stephan Sieber
Matthew Nodwell
Johannes Lehmann
Katharina KOLBE
Patrick RÄMER
Original Assignee
Technische Universität München
Helmholtz Zentrum München - Deutsches Forschungszentrum für Gesundheit und Umwelt (GmbH)
Forschungszentrum Borstel, Leibniz-Zentrum Für Medizin Und Biowissenschaften
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Technische Universität München, Helmholtz Zentrum München - Deutsches Forschungszentrum für Gesundheit und Umwelt (GmbH), Forschungszentrum Borstel, Leibniz-Zentrum Für Medizin Und Biowissenschaften filed Critical Technische Universität München
Publication of WO2017194734A1 publication Critical patent/WO2017194734A1/en

Links

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
    • A61K31/445—Non condensed piperidines, e.g. piperocaine
    • A61K31/4453—Non condensed piperidines, e.g. piperocaine only substituted in position 1, e.g. propipocaine, diperodon
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/535—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with at least one nitrogen and one oxygen as the ring hetero atoms, e.g. 1,2-oxazines
    • A61K31/5375—1,4-Oxazines, e.g. morpholine
    • A61K31/5377—1,4-Oxazines, e.g. morpholine not condensed and containing further heterocyclic rings, e.g. timolol
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/04—Antibacterial agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/04—Antibacterial agents
    • A61P31/06—Antibacterial agents for tuberculosis

Definitions

  • the invention relates to compounds which are suitable for treating mycobacterial diseases and to pharmaceutical compositions containing such compounds. Also encompassed are such compounds for use in medicine.
  • the invention further relates to a kit of parts comprising a pharmaceutical composition containing such compounds and at least one additional pharmaceutically active compound.
  • the mycobacteria are a diverse collection of acid-fast, non-motile, gram-positive bacteria. It comprises several species, which include, Mycobacterium africanum (M. africanum), M. avium, M. bovis, M. bovis-BCG, M. chelonae, M. fortuitum, M. gordonae, M. intracellular, M. kansasii, M. microti, M. scrofulaceum, M. paratuberculosis, M. leprae, M. tuberculosis, and M. ranae. Certain of these organisms are the causative agents of disease.
  • M. africanum Mycobacterium africanum
  • M. avium M. bovis
  • M. bovis-BCG M. chelonae
  • M. fortuitum M. gordonae
  • M. intracellular M. kansasii, M. microti, M. scrofulaceum, M.
  • Tuberculosis and leprosy are the best known mycobacterial diseases. People may also be infected by any of a group of mycobacterial species collectively called non-tuberculous mycobacteria (NTM). In children, NTM cause lymphadenitis, skin and soft tissue infections, and occasionally also lung disease and disseminated infections. Manifestations can be indistinguishable from tuberculosis on the basis of clinical and radiological findings and tuberculin skin testing. Although over 150 different species of NTM have been described, pulmonary infections are most commonly due to Mycobacterium avium complex (MAC), Mycobacterium kansasii, and Mycobacterium abscessus.
  • MAC Mycobacterium avium complex
  • Mycobacterium kansasii Mycobacterium abscessus.
  • Tuberculosis is one of the world's most prevalent infectious diseases responsible for the largest fraction of infection related casualties (before HIV).
  • the disease is caused by bacteria of the Mycobacterium tuberculosis Complex.
  • Species in this complex include M. tuberculosis, M. africanum, M. bovis, M. caprae etc, whereby M. tuberculosis is the main causative agent of tuberculosis.
  • Tuberculosis usually affects the lung and represents the most frequent form of TB.
  • Tuberculosis is usually controlled using extended antibiotic therapy.
  • the preferred mode of treatment for tuberculosis is the short course chemotherapy in which there are two phases.
  • the first phase consists of a daily regimen for two months with isoniazid (300 mg), rifampicin (600 mg), pyrazinamide (3 g) and ethambutol (1.5 g).
  • the second phase or the continuation phase consists of a daily regimen for the next four months with isoniazid and rifampicin.
  • the vast majority of individuals infected with drug susceptible M. tuberculosis strains can be effectively cured when medicines are provided and taken properly.
  • MDR-TB multidrug-resistant tuberculosis
  • WHO Global tuberculosis report. 2014
  • XDR-TB extensively drug-resistant
  • Mycobacterium tuberculosis is transmitted by aerosol and is initially taken up by alveolar lung macrophages, which phagocytose but don't kill the bacterium. Most infected individuals can control the disease for a long time. Estimates believe that one-third of the world's population is latently infected (LTBI; latent tuberculosis infection). The bacteria adapt and survive in diverse environmental niches in vivo, e.g. in solid granulomas, a characteristic feature of latent TB infection. It is presumed that M.
  • LTBI latent tuberculosis infection
  • tuberculosis resides in these regions in a slow growing or non- replicating, phenotypically drug resistant dormant-like state, due to limited availability and supply of oxygen and nutrients (Gengenbacher, M. and S.H. Kaufmann, Mycobacterium tuberculosis: success through dormancy. FEMS Microbiol Rev, 2012. 36(3): p. 514-32).
  • TAGs triacylglycerols
  • tuberculosis growth and survival requiring diverse lipases and hydrolases Moreover, it is believed that these fatty acids are crucial for bacteria to enter and maintain the dormant state by the production of foamy lung macrophages during latent infection (Peyron, P., et al., Foamy macrophages from tuberculous patients' granulomas constitute a nutrient-rich reservoir for M. tuberculosis persistence. PLoS Pathog, 2008. 4(1 1 ): p.
  • Mycobacterium tuberculosis uses host triacylglycerol to accumulate lipid droplets and acquires a dormancy-like phenotype in lipid- loaded macrophages.
  • PLoS Pathog, 201 1. 7(6): p. e1002093 Specifically targeting the bacterial lipid metabolism could represent a viable strategy to limit the growth of M. tuberculosis and open a new opportunity to shorten the long TB therapy (Warner DF, Mizrahi V. Shortening treatment for tuberculosis— to basics. N Engl J Med, 2014. 371 (17):1642-3), when given simultaneously with known first and second line antibiotics.
  • Lalistat is a thiadiazole carbamate initially developed as a specific inhibitor of the mammalian acid lipase, an enzyme located in cellular late endosomes hydrolysing cholesterol esters and triglycerides from incoming lipoproteins.
  • Lalistat covalently binds to the active site serine of lipases and thereby inhibits enzyme activity and is considered as a potential Niemann-Pick Type C disease therapeutic (Rosenbaum, A.I., et al., Thiadiazole carbamates: potent inhibitors of lysosomal acid lipase and potential Niemann-Pick type C disease therapeutics. J Med Chem, 2010. 53(14): p. 5281-9).
  • WO 2015/135652 discloses lalistat and derivatives thereof for the treatment or prevention of viral infections, in particular Hepatitis C virus infection.
  • WO 2014/000058 A1 discloses a method of treating or preventing an abnormality of glucose metabolism in a subject, comprising administering an antagonist of lysosomal acid lipase (LAL) to the subject such that LAL is antagonized in a pancreatic ⁇ cell of the subject.
  • LAL lysosomal acid lipase
  • Tetrahydrolipstatin targets ⁇ / ⁇ -hydrolases, including many lipid esterases (LipD, G, H, I, M, N, O, V, W, and TesA).
  • the underlying problem of the present invention is the provision of new chemical agents having significant therapeutic activity against mycobacterial diseases, allowing inhibiting the extracellular and intracellular growth of mycobacteria.
  • thiadiazole derivatives and five-membered heterocyclic compounds derived from lalistat are suitable as anti-mycobacterial agents. It has been found that those compounds inhibit the growth of M. tuberculosis and in particular the intracellular growth of M. tuberculosis in human macrophage host cells, while not affecting the growth of other gram-positive and gram-negative bacteria as shown for Staphylococcus aureus, Escherichia coli and Listeria monocytogenes.
  • the compounds of the present invention inhibit various members of the lipolytic enzyme family, in particular lipases, hydrolases, carboxylesterases, cutinases and uncharacterized proteins including LipN, Lipl, LipR, LipM, LipG, LipO, LipT, , Hydrolase Rv0183, alpha/beta hydrolase Rv1 192, cutinase Rv1984c, Rv1367c, Rv2715, Rv1730c, RV0293c, rv1399c and Rv0045c, as well as the amidases Rv2888c and Rv2363 and the putative proline iminopeptidase Rv0840c (see also Figure 6).
  • alkyl refers to a monoradical of a saturated straight or branched hydrocarbon.
  • the alkyl group comprises from 1 to 12 (such as 1 to 10) carbon atoms, i.e., 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , or 12 carbon atoms (such as 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10 carbon atoms), more preferably 1 to 8 carbon atoms, such as 1 to 6 or 1 to 4 carbon atoms.
  • Exemplary alkyl groups include methyl, ethyl, propyl, iso-propyl, butyl, iso-butyl, tert-butyl, n-pentyl, iso-pentyl, sec-pentyl, neo-pentyl, 1 ,2-dimethyl-propyl, iso-amyl, n- hexyl, iso-hexyl, sec-hexyl, n-heptyl, iso-heptyl, n-octyl, 2-ethyl-hexyl, n-nonyl, n-decyl, n- undecyl, n-dodecyl, and the like.
  • the alkyl chain is a linear. In some embodiments the alkyl chain is branched. In some embodiments the alkyl chain is substituted. In some embodiment the alkyl chain is unsubstituted. In some embodiments the alkyl chain is linear and substituted or unsubstituted. In some embodiments the alkyl chain is branched and substituted or unsubstituted.
  • alkenyl refers to a monoradical of an unsaturated straight or branched hydrocarbon having at least one carbon-carbon double bond.
  • the maximal number of carbon-carbon double bonds in the alkenyl group can be equal to the integer which is calculated by dividing the number of carbon atoms in the alkenyl group by 2 and, if the number of carbon atoms in the alkenyl group is uneven, rounding the result of the division down to the next integer.
  • the maximum number of carbon-carbon double bonds is 4.
  • the alkenyl group has 1 to 4, i.e., 1 , 2, 3, or 4, carbon-carbon double bonds.
  • the alkenyl group comprises from 2 to 10 carbon atoms, i.e., 2, 3, 4, 5, 6, 7, 8, 9, or 10 carbon atoms, more preferably 2 to 8 carbon atoms, such as 2 to 6 carbon atoms or 2 to 4 carbon atoms.
  • the alkenyl group comprises from 2 to 10 carbon atoms and 1 , 2, 3, 4, or 5 carbon-carbon double bonds, more preferably it comprises 2 to 8 carbon atoms and 1 , 2, 3, or 4 carbon-carbon double bonds, such as 2 to 6 carbon atoms and 1 , 2, or 3 carbon-carbon double bonds or 2 to 4 carbon atoms and 1 or 2 carbon-carbon double bonds.
  • the carbon- carbon double bond(s) may be in cis (Z) or trans (E) configuration.
  • Exemplary alkenyl groups include vinyl, 1-propenyl, 2-propenyl (i.e., allyl), 1-butenyl, 2-butenyl, 3-butenyl, 1 -pentenyl, 2- pentenyl, 3-pentenyl, 4-pentenyl, 1-hexenyl, 2-hexenyl, 3-hexenyl, 4-hexenyl, 5-hexenyl, 1- heptenyl, 2-heptenyl, 3-heptenyl, 4-heptenyl, 5-heptenyl, 6-heptenyl, 1-octenyl, 2-octenyl, 3- octenyl, 4-octenyl, 5-octenyl, 6-octenyl, 7-octenyl, 1 -nonenyl, 2-nonenyl, 3-nonenyl,
  • the alkenyl chain is a linear. In some embodiments the alkenyl chain is branched. In some embodiments the alkenyl chain is substituted. In some embodiment the alkenyl chain is unsubstituted. In some embodiments the alkenyl chain is linear and substituted or unsubstituted. In some embodiments the alkenyl chain is branched and substituted or unsubstituted.
  • alkynyl refers to a monoradical of an unsaturated straight or branched hydrocarbon having at least one carbon-carbon triple bond.
  • the maximal number of carbon-carbon triple bonds in the alkynyl group can be equal to the integer which is calculated by dividing the number of carbon atoms in the alkynyl group by 2 and, if the number of carbon atoms in the alkynyl group is uneven, rounding the result of the division down to the next integer. For example, for an alkynyl group having 9 carbon atoms, the maximum number of carbon-carbon triple bonds is 4.
  • the alkynyl group has 1 to 4, i.e., 1 , 2, 3, or 4, more preferably 1 or 2 carbon-carbon triple bonds.
  • the alkynyl group comprises from 2 to 10 carbon atoms, i.e., 2, 3, 4, 5, 6, 7, 8, 9, or 10 carbon atoms, more preferably 2 to 8 carbon atoms, such as 2 to 6 carbon atoms or 2 to 4 carbon atoms.
  • the alkynyl group comprises from 2 to 10 carbon atoms and 1 , 2, 3, 4, or 5 (preferably 1 , 2, or 3) carbon-carbon triple bonds, more preferably it comprises 2 to 8 carbon atoms and 1 , 2, 3, or 4 (preferably 1 or 2) carbon-carbon triple bonds, such as 2 to 6 carbon atoms and 1 , 2 or 3 carbon-carbon triple bonds or 2 to 4 carbon atoms and 1 or 2 carbon-carbon triple bonds.
  • alkynyl groups include ethynyl, 1 -propynyl, 2- propynyl, 1-butynyl, 2-butynyl, 3-butynyl, 1 -pentynyl, 2-pentynyl, 3-pentynyl, 4-pentynyl, 1- hexynyl, 2-hexynyl, 3-hexynyl, 4-hexynyl, 5-hexynyl, 1-heptynyl, 2-heptynyl, 3-heptynyl, 4- heptynyl, 5-heptynyl, 6-heptynyl, 1-octynyl, 2-octynyl, 3-octynyl, 4-octynyl, 5-octynyl, 6-octynyl, 7-octynyl, 1-nonylyl, 2-nonynyl
  • the alkynyl chain is a linear. In some embodiments the alkynyl chain is branched. In some embodiments the alkynyl chain is substituted. In some embodiment the alkynyl chain is unsubstituted. In some embodiments the alkynyl chain is linear and substituted or unsubstituted. In some embodiments the alkynyl chain is branched and substituted or unsubstituted.
  • aryl or "aromatic ring” refers to a monoradical of an aromatic cyclic hydrocarbon.
  • the aryl group contains 3 to 14
  • aryl groups include cyclopropenylium, cyclopentadienyl, phenyl, indenyl, naphthyl, azulenyl, fluorenyl, anthryl, and phenanthryl.
  • aryl refers to a monocyclic ring containing 6 carbon atoms or an aromatic bicyclic ring system containing 10 carbon atoms. Preferred examples are phenyl and naphthyl. In some embodiments the aryl is unsubstituted. In some embodiments the aryl is substituted.
  • heteroaryl or “heteroaromatic ring” means an aryl group as defined above in which one or more carbon atoms in the aryl group are replaced by heteroatoms of O, S, or N.
  • the heteroaryl group contains 3 to 10 carbon atoms.
  • heteroaryl refers to a five or six-membered aromatic monocyclic ring wherein 1 , 2, or 3 carbon atoms are replaced by the same or different heteroatoms of O, N, or S.
  • it means an aromatic bicyclic or tricyclic ring system wherein 1 , 2, 3, 4, or 5 carbon atoms are replaced with the same or different heteroatoms of O, N, or S.
  • heteroaryl groups include furanyl, thienyl, oxazolyl, isoxazolyl, oxadiazolyl (1 ,2,5- and 1 ,2,3-), pyrrolyl, imidazolyl, pyrazolyl, triazolyl (1 ,2,3- and 1 ,2,4-), tetrazolyl, thiazolyl, isothiazolyl, thiadiazolyl (1 ,2,3- and 1 ,2,5-), pyridyl, pyrimidinyl, pyrazinyl, triazinyl (1 ,2,3-, 1 ,2,4-, and 1 ,3,5-), benzofuranyl (1- and 2-), indolyl, isoindolyl, benzo
  • Exemplary 5- or 6-memered heteroaryl groups include furanyl, thienyl, oxazolyl, isoxazolyl, oxadiazolyl (1 ,2,5- and 1 ,2,3-), pyrrolyl, imidazolyl, pyrazolyl, triazolyl (1 ,2,3- and 1 ,2,4-), thiazolyl, isothiazolyl, thiadiazolyl (1 ,2,3- and 1 ,2,5-), pyridyl, pyrimidinyl, pyrazinyl, triazinyl (1 ,2,3-, 1 ,2,4-, and 1 ,3,5-), and pyridazinyl.
  • the heteroaryl is unsubstituted. In some embodiments the heteroaryl is substituted.
  • arylalkyi and heteroarylalkyi are meant to include those radicals in which an aryl group and heteroaryl group, respectively, is attached to an alkyl group (e.g., benzyl, phenethyl, pyridylmethyl and the like) including those alkyl groups in which a carbon atom (e.g., a methylene group) has been replaced by, for example, an oxygen atom (e.g., phenoxymethyl, 2-pyridyloxymethyl, 3-(1-naphthyloxy)propyl, and the like).
  • alkyl group e.g., benzyl, phenethyl, pyridylmethyl and the like
  • an oxygen atom e.g., phenoxymethyl, 2-pyridyloxymethyl, 3-(1-naphthyloxy)propyl, and the like.
  • the Arylalkyl is a substituted or unsubstituted (C 6 -Ci 0 )aryl(Ci-C 6 )alkyl.
  • the Heteroarylalkyl is a substituted or unsubstituted
  • the alkyl chain is a linear. In some embodiments the alkyl chain is branched. In some embodiments the alkyl chain is substituted. In some embodiments the alkyl chain is unsubstituted. In some embodiments the alkyl chain is linear and substituted or unsubstituted. In some embodiments the alkyl chain is branched and substituted or unsubstituted.
  • the arylalkyl is unsubstituted. In some embodiments the arylalkyl is substituted. In some embodiments the heteroarylalkyl is unsubstituted. In some embodiments the heteroarylalkyl is substituted.
  • cycloalkyi or "cycloaliphatic” represents cyclic non-aromatic versions of “alkyl” and “alkenyl” with preferably 3 to 14 carbon atoms, such as 3 to 10 carbon atoms, i.e., 3, 4, 5, 6, 7, 8, 9, or 10 carbon atoms, more preferably 3 to 8 carbon atoms, even more preferably 3 to 7 carbon atoms.
  • cycloalkyi groups include cyclopropyl, cyclopropenyl, cyclobutyl, cyclobutenyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cycloheptyl, cycloheptenyl, cyclooctyl, cyclooctenyl, cyclononyl, cyclononenyl, cylcodecyl, cylcodecenyl, and adamantyl.
  • cycloalkyi is also meant to include bicyclic and tricyclic versions thereof.
  • bicyclic rings are formed it is preferred that the respective rings are connected to each other at two adjacent carbon atoms, however, alternatively the two rings are connected via the same carbon atom, i.e., they form a spiro ring system or they form "bridged" ring systems.
  • cycloalkyi include C 3 -C 8 - cycloalkyl, in particular cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, spiro[3,3]heptyl, spiro[3,4]octyl, spiro[4,3]octyl, bicyclo[4.1.0]heptyl, bicyclo[3.2.0]heptyl, bicyclo[2.2.1]heptyl, bicyclo[2.2.2]octyl, bicyclo[5.1.0]octyl, and bicyclo[4.2.0]octyl.
  • cycloalkyi is unsubstituted. In some embodiments the cycloalkyi is substituted.
  • cyclopropylene means a cyclopropyl group as defined above in which one hydrogen atom has been removed resulting in a diradical.
  • the cyclopropylene may link two atoms or moieties via the same carbon atom (1 ,1 - cyclopropylene, i.e., a geminal diradical) or via two carbon atoms (1 ,2-cyclopropylene).
  • heterocyclyl or “heterocyclic ring” or “heterocycle” means a cycloalkyi group as defined above in which from 1 , 2, 3, or 4 carbon atoms in the cycloalkyi group are replaced by heteroatoms of O, S, or N.
  • the maximum number of O atoms is 1
  • the maximum number of S atoms is 1
  • the maximum total number of O and S atoms is 2.
  • heterocyclyl is also meant to encompass partially or completely hydrogenated forms (such as dihydro, tetrahydro or perhydro forms) of the above-mentioned heteroaryl groups.
  • exemplary heterocyclyl groups include morpholino, isochromanyl, chromanyl, pyrrolidinyl, imidazolidinyl, pyrazolidinyl, piperidinyl, piperazinyl, indolinyl, isoindolinyl, di- and tetrahydrofuranyl, di- and tetrahydrothienyl, di- and tetrahydrooxazolyl, di- and tetrahydroisoxazolyl, di- and tetrahydrooxadiazolyl (1 ,2,5- and
  • di- and tetrahydropyrrolothiazolyl such as 5,6-dihydro-4H-pyrrolo[3,4-d][1 ,3]thiazolyl
  • di- and tetrahydrophenothiazinyl di- and tetrahydroisobenzofuranyl, di- and tetrahydrochromenyl, di- and tetrahydroxanthenyl, di- and tetrahydrophenoxathiinyl, di- and tetrahydropyrrolizinyl, di- and tetrahydroindolizinyl, di- and tetrahydroindazolyl, di- and tetrahydropurin
  • di- and tetrahydrocinnolinyl di- and tetrahydropteridinyl, di- and tetrahydrocarbazolyl, di- and tetrahydrophenanthridinyl, di- and tetrahydroacridinyl, di- and tetrahydroperimidinyl, di- and tetrahydrophenanthrolinyl (1 ,7-, 1 ,8-, 1 ,10-, 3,8-, and 4,7-), di- and tetrahydrophenazinyl, di- and tetrahydrooxazolopyridinyl, di- and tetrahydroisoxazolopyridinyl, di- and tetrahydropyrrolooxazolyl, and di- and tetrahydropyrrolopyrrolyl.
  • Exemplary 5- or 6-memered heterocyclyl groups include morpholino, pyrrolidinyl, imidazolidinyl, pyrazolidinyl, piperidinyl, piperazinyl, di- and tetrahydrofuranyl, di- and tetrahydrothienyl, di- and tetrahydrooxazolyl, di- and tetrahydroisoxazolyl, di- and tetrahydrooxadiazolyl (1 ,2,5- and 1 ,2,3-), dihydropyrrolyl, dihydroimidazolyl, dihydropyrazolyl, di- and tetrahydrotriazolyl (1 ,2,3- and 1 ,2,4-), di- and tetrahydrothiazolyl, di- and tetrahydroisothiazolyl, di- and tetrahydrothiadiazolyl (1 ,2,3- and
  • heterocyclyl is unsubstituted. In some embodiments the heterocyclyl is substituted.
  • halogen or halo means fluoro, chloro, bromo, or iodo.
  • the term "optionally substituted” or “substituted” indicates that one or more (such as 1 to the maximum number of hydrogen atoms bound to a group, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9, or up to 10, such as between 1 to 5, 1 to 4, or 1 to 3, or 1 or 2) hydrogen atom(s) may be replaced with a group different from hydrogen such as alkyl (preferably, Ci -6 alkyl), alkenyl (preferably, C 2-6 alkenyl), alkynyl (preferably, C 2-6 alkynyl), aryl (preferably, 3- to 14-membered aryl), heteroaryl (preferably, 3- to 14-membered heteroaryl), cycloalkyi (preferably, 3- to 14-membered cycloalkyi), heterocyclyl (preferably, 3- to 14- membered heterocyclyl), halogen, -CN, azido, -N0 2 , -OR 71 , -
  • R 71 , R 72 , and R 73 are independently selected from the group consisting of -H, C 1-6 alkyl, C 2-6 alkenyl, C 2-6 alkynyl, 3- to 7-membered cycloalkyi, 5- or 6-membered aryl, 5- or 6-membered heteroaryl, and 3- to 7-membered heterocyclyl, wherein each of the alkyl, alkenyl, alkynyl, cycloalkyi, aryl, heteroaryl, and heterocyclyl groups is optionally substituted with one, two or three substituents selected from the group consisting of C 1-3 alkyl, halogen, -CF 3 , -CN, azido, -N0 2 , -OH, -0(C 1-3 alkyl), -S(C 1-3 alkyl), -NH 2 , -NH(C 1-3 alkyl), -N(C 1-3 alkyl) 2 ,
  • the present invention relates to a compound for use in the treatment of a mycobacterial disease, said compound having a structure according to Formula I
  • X is a nitrogen containing heterocycle
  • Y is CH 2 , O or NR ; n is an integer between 0 and 3;
  • R and R 2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (C T C 6 )alkyl, substituted or unsubstituted (C 2 -C 6 )alkenyl, substituted or unsubstituted (C 2 -C 6 )alkynyl, substituted or unsubstituted (C 3 -C 8 )cycloalkyl, substituted or unsubstituted (C 6 -C 10 )aryl, substituted or unsubstituted substituted or unsubstituted (C 3 -C 10 )heteroaryl, and substituted or unsubstituted (C3-C 10 )heteroaryl(C T C 6 )alkyl; or a pharmaceutically acceptable salt, solvate or hydrate thereof;
  • mycobacterial disease or “mycobacterial infection” are used interchangeably and refer to any pathological state, including any asymptomatic, acute or chronic mycobacterial infection and any state caused by or associated with such mycobacterial infection.
  • the mycobacterial infection is an infection caused by by bacteria of the Mycobacterium tuberculosis Complex, including M. tuberculosis, M. africanum, M. bovis, M. caprae etc.
  • the mycobacterial infection is an infection caused by Mycobacterium tuberculosis.
  • the mycobacterial infection is multidrug-resistant tuberculosis (MDR-TB).
  • the mycobacterial disease is extensively drug-resistant tuberculosis (XDR-TB). Further specific embodiments defining the mycobacterial disease are described herein below.
  • the term "pharmaceutically acceptable salt” refers to a salt that retains the desired biological activity of the parent compound and does not impart any undesired toxicological effects (see e.g., Berge, S. M., et al. (1977) J. Pharm. Sci. 66: 1 -19).
  • examples of such salts include acid addition salts and base addition salts.
  • Acid addition salts include those derived from nontoxic inorganic acids, such as hydrochloric, nitric, phosphoric, sulfuric, hydrobromic, hydroiodic, phosphorous and the like, as well as from nontoxic organic acids such as aliphatic mono- and dicarboxylic acids, phenyl-substituted alkanoic acids, hydroxy alkanoic acids, aromatic acids, aliphatic and aromatic sulfonic acids and the like.
  • nontoxic inorganic acids such as hydrochloric, nitric, phosphoric, sulfuric, hydrobromic, hydroiodic, phosphorous and the like
  • nontoxic organic acids such as aliphatic mono- and dicarboxylic acids, phenyl-substituted alkanoic acids, hydroxy alkanoic acids, aromatic acids, aliphatic and aromatic sulfonic acids and the like.
  • Base addition salts include those derived from alkaline earth metals, such as sodium, potassium, magnesium, calcium and the like, as well as from nontoxic organic amines, such as ⁇ , ⁇ '- dibenzylethylenediamine, N-methylglucamine, chloroprocaine, choline, diethanolamine, ethylenediamine, procaine and the like.
  • pharmaceutically acceptable may in particular mean approved by a regulatory agency or other generally recognized pharmacopoeia for use in animals, and more particularly in humans.
  • solvate refers to an addition complex of a dissolved material in a solvent (such as an organic solvent (e.g., an aliphatic alcohol (such as methanol, ethanol, n-propanol, isopropanol), acetone, acetonitrile, ether, and the like), water or a mixture of two or more of these liquids), wherein the addition complex exists in the form of a crystal or mixed crystal.
  • a solvent such as an organic solvent (e.g., an aliphatic alcohol (such as methanol, ethanol, n-propanol, isopropanol), acetone, acetonitrile, ether, and the like), water or a mixture of two or more of these liquids), wherein the addition complex exists in the form of a crystal or mixed crystal.
  • the amount of solvent contained in the addition complex may be stoichiometric or non-stoichiometric.
  • a "hydrate” is a solvate wherein the solvent is
  • A is selected from the group consisting of:
  • X is a nitrogen containing heterocycle
  • Y is CH 2 , O or NR
  • n is an integer between 0 and 3
  • R and R 2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C 6 )alkyl, substituted or unsubstituted (C 2 -C 6 )alkenyl, substituted or unsubstituted (C 2 -C 6 )alkynyl, substituted or unsubstituted (C 3 -C 8 )cycloalkyl, substituted or unsubstituted (C 6 -Ci 0 )aryl, substituted or unsubstituted (C 6 -Ci 0 )aryl(Ci-C 6 )alkyl, substituted or unsubstituted (C 3 -Ci 0 )heteroaryl, and substituted or unsubstituted (C 3 -Ci 0 )heteroaryl(
  • the present invention relates to a compound for use in medicine, said compound having a structure according to Formula I
  • A is selected from the group consisting of:
  • X is a nitrogen containing heterocycle
  • Y is CH 2 , O or NR
  • n is an integer between 0 and 3;
  • R 1 and R 2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C 6 )alkyl, substituted or unsubstituted (C 2 -C 6 )alkenyl, substituted or unsubstituted (C 2 -C 6 )alkynyl, substituted or unsubstituted (C 3 -C 8 )cycloalkyl, substituted or unsubstituted (C 6 -Cio)aryl, substituted or unsubstituted (C 6 -Cio)aryl(CrC 6 )alkyl, substituted or unsubstituted (C 3 -Ci 0 )heteroaryl, and substituted or unsubstituted (C 3 -Ci 0 )heteroaryl(Ci-C 6 )alkyl; or a pharmaceutically acceptable salt, solvate or hydrate thereof; with the proviso that
  • the present invention relates to a pharmaceutical composition for use in the treatment of a mycobacterial disease, wherein said composition comprises a compound having a structure according to Formula I
  • A is selected from the group consisting of:
  • X is a nitrogen containing heterocycle
  • Y is CH 2 , O or NR
  • n is an integer between 0 and 3;
  • R 1 and R 2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (C T C 6 )alkyl, substituted or unsubstituted (C 2 -C 6 )alkenyl, substituted or unsubstituted (C 2 -C 6 )alkynyl, substituted or unsubstituted (C 3 -C 8 )cycloalkyl, substituted or unsubstituted (C 6 -Ci 0 )aryl, substituted or unsubstituted (C 6 -Cio)aryl(CrC 6 )alkyl, substituted or unsubstituted (C 3 -Ci 0 )heteroaryl, and substituted or unsubstituted (C 3 -C 10 )heteroaryl(Ci-C 6 )alkyl; or a pharmaceutically acceptable salt, solvate or hydrate thereof;
  • the compound for use in the treatment of a mycobacterial disease as disclosed herein above the compound for use in medicine as disclosed herein above and the compound of the composition for use in the treatment of a mycobacterial disease as disclosed herein above are further defined.
  • the compounds for use or the compound of the pharmaceutical composition for use havin according to Formula I, as erein, are selected from the group consisting of:
  • A is In some embodiments A is . In some bodiments A is I Inn some embodiments A is In some
  • A is ssoommee eemmbbooddiimmeennttss AA iiss ..
  • A is ⁇ ⁇ . In some embodiments A is
  • the compounds for use or the compound of the pharmaceutical composition for use, as disclosed herein are characterized by having a structure according Formula (la)
  • X, Y, n and R 1 are defined as above and wherein the provisos are defined as above for the compounds for use or the compound of the pharmaceutical composition for use of the invention.
  • the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I or Formula la, as disclosed herein, are characterized in that X is wherein
  • Z is CR 3 R 4 , O or NR 5 ;
  • R 3 and R 4 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (C T C 6 )alkyl, substituted or unsubstituted (C 2 -C 6 )alkenyl, substituted or unsubstituted (C 2 -C 6 )alkynyl, substituted or unsubstituted (C 3 -C 8 )cycloalkyl, substituted or unsubstituted (C 6 -Ci 0 )aryl, substituted or unsubstituted (C6-Cio)aryl(CrC 6 )alkyl, substituted or unsubstituted (C 3 -Ci 0 )heteroaryl, substituted or unsubstituted (C 3 -Cio)heteroaryl(Ci-C 6 )alkyl, halogen, -CN, -N0 2 , -OR 11 , -N(R 12 )(R 13
  • R 5 is selected from the group consisting of hydrogen, substituted or unsubstituted (C T C 6 )alkyl, substituted or unsubstituted (C 2 -C 6 )alkenyl, substituted or unsubstituted (C 2 -C 6 )alkynyl, substituted or unsubstituted (C 3 -C 8 )cycloalkyl, substituted or unsubstituted (C 6 -Ci 0 )aryl, substituted or unsubstituted (C 6 -C 10 )aryl(CrC 6 )alkyl, substituted or unsubstituted (C 3 - Cio)heteroaryl, substituted or unsubstituted (C 3 -Ci 0 )heteroaryl(Ci-
  • R 14 is independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, heterocyclyl, and -OR 11 ;
  • R 5 and R 6 are independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, and heterocyclyl.
  • Z is CR 3 R 4 .
  • Z is O or NR 5 .
  • Z is O.
  • Z is NR 5 .
  • the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I or Formula la, as disclosed herein, are characterized in that Y is CH 2 or O. In some embodiments Y is CH 2 . In some embodiments Y is O. In some embodiments Y is NR 1 .
  • the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I or Formula la, as disclosed herein, are characterized in that R 1 and R 2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C 6 )alkyl, substituted or unsubstituted (C 2 -C 6 )alkenyl, substituted or unsubstituted (C 3 -C B )cycloalkyl, substituted or unsubstituted (C 6 -C 10 )aryl, substituted or unsubstituted (C 6 -C 10 )aryl(CrC 6 )alkyl, substituted or unsubstituted (C 3 - Cio)heteroaryl, and substituted or unsubstituted (C 3 -Cio)heteroaryl(Ci-C 6 )alkyl.
  • R 1 and R 2 are independently selected from the group consisting of hydrogen, substituted or unsubsti
  • R and R 2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C 6 )alkyl, substituted or unsubstituted (C 3 -C 8 )cycloalkyl, substituted or unsubstituted (C 6 -Ci 0 )aryl, substituted or unsubstituted (C 6 -Ci 0 )aryl(Ci-C 6 )alkyl, and substituted or unsubstituted (C 3 -C 10 )heteroaryl.
  • R and R 2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (C r C 6 )alkyl, substituted or unsubstituted (C 3 -C 8 )cycloalkyl and substituted or unsubstituted (C 6 - Cio)aryl.
  • R and R 2 are independently selected from the group consisting of hydrogen and substituted or unsubstituted (C C 6 )alkyl.
  • R is hydrogen and R 2 is alkyl.
  • R is alkyl and R 2 is hydrogen.
  • R 1 and R 2 are alkyl.
  • R and R 2 are hydrogen.
  • the compounds for use or the compound of the pharmaceutical composition for use, as disclosed herein are characterized by having a structure according to Formula II
  • R 1 , R 3 , R 4 , R 5 , R 11 , R 12 , R 3 , R 14 , R 5 , R 6 and n are defined as above for Formula I or Formula la;
  • the compounds for use or the compound of the pharmaceutical composition for use, as disclosed herein are characterized by having a structure according to Formula III
  • Z is O or CR 3 R 4 ;
  • R 3 and R 4 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (C T C 6 )alkyl, substituted or unsubstituted (C 2 -C 6 )alkynyl and -OR 11 ;
  • R 1 is, in each case, selected from the group consisting of hydrogen and substituted or unsubstituted (Ci-C 6 )alkyl.
  • R 3 and R 4 are hydrogen or substituted or unsubstituted (C C 6 )alkyl, preferably methyl. In some embodiments at least one of R 3 and R 4 is hydrogen. In some embodiments Z is O. In some embodiments R 3 and R 4 are hydrogen. In some embodiments R 3 is hydrogen and R 4 is substituted or unsubstituted (Ci-C 6 )alkyl, preferably wherein R 4 is tert- butyl. In some embodiments R 3 is hydrogen and R 4 is substituted or unsubstituted (C ⁇ - C 6 )alkynyl, preferably wherein R 4 is ethynyl.
  • R 3 is hydrogen and R 4 is -OR 11 , wherein R is hydrogen and substituted or unsubstituted (CrC 6 )alkyl. In some embodiments R 3 is hydrogen and R 4 is -OR 11 , wherein R 11 is hydrogen. In some embodiments R 3 is hydrogen and R 4 is -OR 11 , wherein R 11 is substituted or unsubstituted (Ci-C 6 )alkyl, preferably methyl.
  • the compounds for use or the compound of the pharmaceutical composition for use ha ure according to Formula I, as disclosed herein, are selected from the group consisting of:
  • A is ;
  • X is ⁇ wherein Z is CH 2 or O;
  • Y is CH 2 or O C H 2 and n is 2.
  • Z is O;
  • Y is N-(2-aminoethyl)
  • the compounds for use or the compound of the pharmaceutical composition for use ha ormula I, as disclosed herein, are selected from the compounds for use or the compound of the pharmaceutical composition for use ha ormula I, as disclosed herein, are selected from the compounds for use or the compound of the pharmaceutical composition for use ha ormula I, as disclosed herein, are selected from the compounds for use or the compound of the pharmaceutical composition for use ha ormula I, as disclosed herein.
  • A is ; ⁇ is wherein Z is CH 2 or O; Y is CH 2 or O
  • n is 2.
  • erein Z is CH 2 ; Y is
  • nts A is wherein Z is CH 2 ; Y
  • A is O and n is 2. In some embodiments A is IS wherein Z is O; Y is CH 2 and n is 2. In some embodiments A is wherein Z is O; Y is O and n is 2.
  • the compounds for use or the compound of the pharmaceutical composition for use ormula I, as disclosed herein are selected from:
  • n is 2.
  • erein Z is CH 2 ; Y is
  • A is wherein Z is O; Y is
  • A is ;
  • X is wherein Z is O;
  • Y is O and n is 2.
  • the compounds for use or the compound of the pharmaceutical composition for use ha ormula I, as disclosed herein, are selected from the compounds for use or the compound of the pharmaceutical composition for use ha ormula I, as disclosed herein, are selected from the compounds for use or the compound of the pharmaceutical composition for use ha ormula I, as disclosed herein, are selected from the compounds for use or the compound of the pharmaceutical composition for use ha ormula I, as disclosed herein.
  • A is wherein Z is CH 2 or O; Y is CH 2 or O and n is 2.
  • A is erein Z is CH 2 ; Y is
  • A is ;
  • X is wherein Z is CH 2 ;
  • Y is wherein Z is CH 2 ;
  • A is wherein Z is O; Y is
  • A is ;
  • X is wherein Z is O;
  • Y is O and n is 2.
  • the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, as disclosed herein, are selected from the group consisting of:
  • A is ;
  • X is wherein Z is O;
  • Y is O and n is 2.
  • X is wherein Z is CH 2 or O; Y is CH 2 or
  • O 2 is hydrogen or substituted or unsubstituted (Ci-C e )alkyl and n is 2. In some embodiments
  • A is ;
  • X is wherein Z is CH 2 ;
  • Y is CH 2 , R 2 is hydrogen or substituted
  • a is X is herein Z is CH 2 ; Y is O, R i substituted or unsubstituted (C C 6 )alkyl and n is 2. In some embodiments A is ; X is wherein Z is
  • Y is CH 2
  • R 2 is hydrogen or substituted or unsubstituted (Ci-C 6 )alkyl and n is 2.
  • A is ;
  • X is wherein Z is O;
  • Y is O,
  • R 2 is hydrogen or substituted or unsubstituted (CrC 6 )alkyl and n is 2.
  • the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, as disclosed herein, are characterized in that A is wherein Z is CH 2 or O; Y is CH 2 or
  • R 2 is hydrogen or substituted or unsubstituted (Ci-C 6 )alkyl and n is 2. In some embodiments
  • A is X is herein Z is CH 2 ; Y is O, R 2 i substituted or unsubstituted (Ci- C 6 )alkyl and n is 2. In some embodiments A is ; X is ⁇ / wherein Z is
  • Y is CH 2
  • R 2 is hydrogen or substituted or unsubstituted (Ci-C 6 )alkyl and n is 2.
  • A is ;
  • X is N / wherein Z is O;
  • Y is O,
  • R 2 is hydrogen or substituted or unsubstituted (C C 6 )alkyl and n is 2.
  • the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, as disclosed herein, are selected from the group consisting of:
  • A is ;
  • X is wherein Z is CH 2 or O;
  • Y is CH 2 or O,
  • R 2 is hydrogen or substituted or unsubstituted (C T C 6 )alkyl and n is 2.
  • A is X is wherein Z is CH 2 ; Y is CH 2 , R is hydrogen or substituted or
  • X is wherein Z is CH 2 ; Y is O, R is hydrogen or substituted or unsubstituted (C r
  • A is ⁇ ⁇ is wherein Z is O;
  • Y is CH 2
  • R 2 is hydrogen or substituted or unsubstituted (CrCeJalkyl and n is 2.
  • A is ;
  • X is wherein Z is O;
  • Y is O,
  • R 2 is hydrogen or substituted or unsubstituted (CrCeJalkyl and n is 2.
  • the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, as disclosed herein, are selected from the group consisting of:
  • A is wherein Z is CH 2 or O; Y is CH 2 or O,
  • R 2 is hydrogen or substituted or unsubstituted (C ! -CeJalkyl and n is 2.
  • A is wherein Z is CH 2 ; Y is CH 2 , R 2 is hydrogen or substituted or
  • (CrC 6 )alkyl and n is 2.
  • A is ;
  • X is wherein Z is CH 2 ;
  • Y is O, R is hydrogen or substituted or unsubstituted (C r
  • A is wherein Z is O; Y is CH 2 , R 2 is hydrogen or substituted or unsubstituted (CrCeJalkyl and n is 2.
  • embodiments A is wherein Z is O; Y is O, R 2 is hydrogen or substituted or unsubstituted (CrCeJalkyl and n is 2.
  • the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, Formula la, Formula II and Formula III, as disclosed herein are useful for the treatment of a mycobacterial disease, wherein the mycobacterial disease is caused by at least one bacteria selected from the group consisting of Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae, Mycobacterium microti, Mycobacterium leprae, Mycobacterium lepromatosis, Mycobacterium avium, Mycobacterium silvaticum, Mycobacterium hominissuis, Mycobacterium paratuberculosis, Mycobacterium kansasii, Mycobacterium xenopi, Mycobacterium simiae, Mycobacterium abcessus, Mycobacterium fortuitum, Mycobacterium chelonae, Mycobacterium ulcerans, Mycobacterium marinum and/or Mycobacterium for
  • Mycobacterial diseases are caused by mycobacteria. Tuberculosis and leprosy (Hansen's disease) are the best known mycobacterial diseases. However, people may also be infected by any of a group of mycobacterial species collectively called non-tuberculous mycobacteria. While tuberculosis and leprosy are most common in resource-limited countries, non-tuberculous mycobacterial infections occur worldwide.
  • Tuberculosis is caused by bacteria of the Mycobacterium tuberculosis Complex.
  • the Mycobacterium tuberculosis Complex includes bacteria selected from the group consisting of Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae and Mycobacterium microti.
  • the mycobacterial disease is caused by at least one bacteria of the Mycobacterium tuberculosis Complex.
  • the mycobacterial disease is caused by Mycobacterium tuberculosis.
  • the mycobacterial disease is caused by Mycobacterium africanum.
  • the mycobacterial disease is caused by Mycobacterium bovis.
  • the mycobacterial disease is caused by Mycobacterium caprae. In some embodiments the mycobacterial disease is caused by Mycobacterium microti. In some embodiments the mycobacterial disease is tuberculosis and/or caused by bacteria of the group consisting of Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae, Mycobacterium microti, preferably Mycobacterium tuberculosis.
  • Leprosy is caused by Mycobacterium leprae.
  • the mycobacterial disease is caused by Mycobacterium leprae.
  • the mycobacterial disease may be also caused by non-tuberculous mycobacteria (NTM).
  • NTM non-tuberculous mycobacteria
  • NTM causes mycobacteria which do not cause tuberculosis or leprosy.
  • NTM cause lymphadenitis, skin and soft tissue infections, and occasionally also lung disease and disseminated infections. Manifestations can be indistinguishable from tuberculosis on the basis of clinical and radiological findings and tuberculin skin testing.
  • non-tuberculosis bacteria are selected from a group consisting of Mycobacterium avium, Mycobacterium silvaticum, Mycobacterium hominissuis, Mycobacterium paratuberculosis, Mycobacterium kansasii, Mycobacterium xenopi, Mycobacterium simiae, Mycobacterium abcessus, Mycobacterium fortuitum, Mycobacterium chelonae, Mycobacterium ulcerans, Mycobacterium marinum, Mycobacterium gordonae and/or Mycobacterium fortuitum.
  • the Mycobacterium avium complex includes bacteria selected from the group consisting of Mycobacterium avium, Mycobacterium silvaticum, Mycobacterium hominissuis and Mycobacterium paratuberculosis.
  • the mycobacterial disease is caused by non-tuberculous mycobacteria.
  • the mycobacterial disease is caused by at least one bacteria of the Mycobacterium avium complex (MAC).
  • the mycobacterial disease is caused by Mycobacterium avium.
  • the mycobacterial disease is caused by Mycobacterium silvaticum.
  • the mycobacterial disease is caused by Mycobacterium hominissuis.
  • the mycobacterial disease is caused by Mycobacterium paratuberculosis. In some embodiments the mycobacterial disease is caused by Mycobacterium kansasii. Mycobacterium kansasii causes chronic pulmonary infection that resembles pulmonary tuberculosis. However, it may also infect other organs. M kansasii infection is the second-most- common nontuberculous opportunistic mycobacterial infection associated with HIV/AIDS. In some embodiments the mycobacterial disease is caused by Mycobacterium xenopi. In some embodiments the mycobacterial disease is caused by Mycobacterium simiae. In some embodiments the mycobacterial disease is caused by Mycobacterium abcessus.
  • the mycobacterial disease is caused by Mycobacterium fortuitum, Mycobacterium chelonae. In some embodiments the mycobacterial disease is caused by Mycobacterium ulcerans. In some embodiments the mycobacterial disease is caused by Mycobacterium marinum. In some embodiments the mycobacterial disease is caused by Mycobacterium fortuitum. In some embodiments the mycobacterial disease is caused by Mycobacterium gordonae.
  • the mycobacterial disease is a pulmonary infection caused by non- tuberculous mycobacteria (NTM), preferably a chronic pulmonary infection.
  • the mycobacterial disease is a skin and/or soft tissue infection caused by non- tuberculous mycobacteria (NTM).
  • the mycobacterial disease is a lung disease caused by non-tuberculous mycobacteria (NTM).
  • the mycobacterial disease is selected from tuberculosis, leprosy (Hansen's disease), lepromatosis infections caused by non-tuberculosis mycobacteria including lymphadenitis and pulmonary infections, skin infections caused by mycobacteria including Buruli ulcer and fish tank granuloma.
  • the mycobacterial disease is tuberculosis.
  • the mycobacterial disease is leprosy (Hansen's disease).
  • the mycobacterial disease is lepromatosis.
  • the mycobacterial disease is a skin infection caused by mycobacteria including Buruli ulcer and fish tank granuloma.
  • the mycobacterial disease is multidrug-resistant tuberculosis (MDR-TB).
  • the mycobacterial disease is extensively drug- resistant tuberculosis (XDR-TB).
  • the compounds of Formula I, Formula la, Formula II and Formula III and pharmaceutically- acceptable salts, solvates and hydrates thereof may be used on their own but will generally be administered in the form of a pharmaceutical composition in which the formula (I) or ( ⁇ ) compound/salt/solvate (active ingredient) is in association with a pharmaceutically-acceptable adjuvant, diluent or carrier.
  • the pharmaceutical composition will preferably comprise from 0.05 to 99 % w (per cent by weight), more preferably from 0.10 to 70 % w, of active ingredient, and, from 1 to 99.95 % w, more preferably from 30 to 99.90 % w, of a pharmaceutically-acceptable adjuvant, diluent or carrier, all percentages by weight being based on total composition.
  • the pharmaceutical composition may additionally contain an additional pharmaceutically active agent, such as an antibiotic, antifungal or anti-HIV compound and/or various other ingredients known in the art, for example, a lubricant, stabilising agent, buffering agent, emulsifying agent, viscosity regulating agent, surfactant, preservative, flavouring or colorant.
  • the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, Formula la, Formula II and Formula III are in particular useful as bactericidal agents and/or bacteriostatic agents.
  • the compounds are also non-hazardous for bacteria of gram-positive bacterial strains other than mycobacterial strains and non-hazardous for gram-negative bacterial strains, in particular they do not inhibit the growth of gram-negative bacterial strains and gram-positive bacterial stains other than mycobacterial strain.
  • the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, Formula la, Formula II and Formula III, as disclosed herein are characterized in that they are bacteriostatic, preferably bacteriostatic for mycobacteria.
  • the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, Formula la, Formula II and Formula III, as disclosed herein are characterized in that they are bactericidal, preferably bactericidal for mycobacteria.
  • the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, Formula la, Formula II and Formula III, as disclosed herein are characterized in that they are non-hazardous for bacteria of gram-positive bacterial strains other than mycobacterial strains and non-hazardous for gram-negative bacterial strains, in particular the growth of gram-negative bacterial strains and gram-positive bacterial stains other than mycobacterial strain is not inhibited.
  • the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, Formula la, Formula II and Formula III, as disclosed herein are characterized in that they are non-hazardous for bacteria of gram-positive bacterial strains other than mycobacterial strains, in particular the growth of gram- positive bacterial stains other than mycobacterial strain is not inhibited.
  • the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, Formula la, Formula II and Formula III, as disclosed herein are characterized in that they are non-hazardous for gram-negative bacterial strains, in particular the growth of gram-negative bacterial strains is not inhibited.
  • the pharmaceutical composition for use as disclosed herein further comprises at least one pharmaceutically acceptable carrier.
  • the compounds according to Formula I, Formula la, Formula II and Formula III or a pharmaceutically acceptable salt, solvate or hydrate thereof may be included in a pharmaceutically acceptable carrier.
  • carrier and “excipient” are used interchangeably herein.
  • Pharmaceutically acceptable carriers or excipients include diluents (fillers, bulking agents, e.g. lactose, microcrystalline cellulose), disintegrants (e.g. sodium starch glycolate, croscarmellose sodium), binders (e.g. PVP, HPMC), lubricants (e.g.
  • glidants e.g. colloidal Si0 2
  • solvents/co-solvents e.g. aqueous vehicle, Propylene glycol, glycerol
  • buffering agents e.g. citrate, gluconates, lactates
  • preservatives e.g. Na benzoate, parabens (Me, Pr and Bu)
  • anti-oxidants e.g. BHT, BHA, Ascorbic acid
  • wetting agents e.g. polysorbates, sorbitan esters
  • anti-foaming agents e.g. Simethicone
  • thickening agents e.g. methylcellulose or hydroxyethylcellulose
  • sweetening agents e.g.
  • sorbitol saccharin, aspartame, acesulfame
  • flavoring agents e.g. peppermint, lemon oils, butterscotch, etc
  • humectants e.g. propylene, glycol, glycerol, sorbitol
  • a non-exhaustive list of exemplary pharmaceutically acceptable carriers or excipients includes (biodegradable) liposomes; microspheres made of the biodegradable polymer poly(D,L)-lactic- coglycolic acid (PLGA), albumin microspheres; synthetic polymers (soluble); nanofibers, protein- DNA complexes; protein conjugates; erythrocytes; or virosomes.
  • Various carrier based dosage forms comprise solid lipid nanoparticles (SLNs), polymeric nanoparticles, ceramic nanoparticles, hydrogel nanoparticles, copolymerized peptide nanoparticles, nanocrystals and nanosuspensions, nanocrystals, nanotubes and nanowires, functionalized nanocarriers, nanospheres, nanocapsules, liposomes, lipid emulsions, lipid microtubules/microcylinders, lipid microbubbles, lipospheres, lipopolyplexes, inverse lipid micelles, dendrimers, ethosomes, multicomposite ultrathin capsules, aquasomes, pharmacosomes, colloidosomes, niosomes, discomes, proniosomes, microspheres, microemulsions and polymeric micelles.
  • the pharmaceutical composition of the invention will generally be designed for specific routes and methods of administration, for specific dosages and frequencies of administration, for specific treatments of specific diseases, with ranges of bio-availability and persistence, among other things.
  • the materials of the composition are preferably formulated in concentrations that are acceptable for the site of administration.
  • Formulations and compositions thus may be designed in accordance with the invention for delivery by any suitable route of administration.
  • the routes of administration include
  • topical routes such as epicutaneous, inhalational, nasal, opthalmic, auricular / aural, vaginal, mucosal
  • enteral routes such as oral, gastrointestinal, sublingual, sublabial, buccal, rectal
  • parenteral routes such as intravenous, intraarterial, intraosseous, intramuscular, intracerebral, intracerebroventricular, epidural, intrathecal, subcutaneous, intraperitoneal, extra-amniotic, intraarticular, intracardiac, intradermal, intralesional, intrauterine, intravesical, intravitreal, transdermal, intranasal, transmucosal, intrasynovial, intraluminal).
  • the administration may be a parenteral route, in particular intravenous or intramuscular.
  • the pharmaceutical composition, as disclosed herein is administered to a subject in need thereof in an amount effective to treat said mycobacterial disease.
  • the subject is preferably a mammal.
  • the subject is more preferably a human subject.
  • the mycobacterial disease can be any mycobacterial disease disclosed herein above and below.
  • Subject means animals, including warm blooded mammals such as humans and primates; avians; domestic household or farm animals such as cats, dogs, sheep, goats, cattle, horses and pigs; laboratory animals such as mice, rats and guinea pigs; fish; reptiles; zoo and wild animals; and the like.
  • the subject is preferably a mammal, more preferably a human.
  • the term "amount effective" in the context of a composition or dosage form for administration to a subject refers to an amount of the composition or dosage form sufficient to provide a benefit in the treatment of mycobacterial disease, to delay or minimize symptoms associated with mycobacterial infection or mycobacterial-induced disease, or to cure or ameliorate the disease or infection or cause thereof.
  • a therapeutically effective amount means an amount sufficient to provide a therapeutic benefit in vivo.
  • the term preferably encompasses a non-toxic amount that improves overall therapy, reduces or avoids symptoms or causes of disease, or enhances the therapeutic efficacy of or synergies with another therapeutic agent.
  • Amounts effective will depend, of course, on the particular subject being treated; the severity of a condition, disease or disorder; the individual patient parameters including age, physical condition, size and weight; the duration of the treatment; the nature of concurrent therapy (if any); the specific route of administration and like factors within the knowledge and expertise of the health practitioner. These factors are well known to those of ordinary skill in the art and can be addressed with no more than routine experimentation. It is generally preferred that a maximum dose be used, that is, the highest safe dose according to sound medical judgment. It will be understood by those of ordinary skill in the art, however, that a patient may insist upon a lower dose or tolerable dose for medical reasons, psychological reasons or for virtually any other reason.
  • the pharmaceutical composition for use, as disclosed herein is administered to a subject in need thereof in an amount effective to treat said mycobacterial disease, wherein said subject is treated with at least one additional pharmaceutically active compound, including an antibiotic, antifungal or anti-HIV compound.
  • the additional pharmaceutically active compound is preferably an antibiotic, antifungal and/or anti-HIV compound.
  • the additional pharmaceutically active compound is more preferably an antibiotic.
  • the antibiotic is preferably an anti-tuberculosis drug or an agent or compound active against Mycobacterium tuberculosis.
  • Anti-tuberculosis (TB) drugs are classified into five groups based on evidence of efficacy, potency, drug class and experience of use. In the United States rifampicin is called rifampin. First-line anti-TB drugs (Group 1 ) are currently recommended in a four-drug combination for the treatment of drug-susceptible TB. Second-line anti-TB drugs (Groups 2, 3 and 4) are reserved for drug-resistant TB. Third-line anti-TB drugs (Group 5) have unclear efficacy or undefined roles.
  • the additional pharmaceutically active compound is selected from the group of First-line agents consisting of Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin and Rrifabutin.
  • the additional pharmaceutically active compound is selected from the group of Second-line agents consisting of Aminoglycosides including Kanamycin and Amikacin, Polypeptides including Capreomycin and Viomycin and Streptomycin.
  • the additional pharmaceutically active compound is selected from the group of Second-line agents consisting of fluoroquinolones including Moxifloxacin, Levofloxacin, Ofloxacin and Gatifloxacin.
  • the additional pharmaceutically active compound is selected from the group of a bacteriostatic Second-line agents consisting of Thioamides including Ethionamide and protionamide, Cycloserine, Terizidone, Thioacetone and p-Aminosalicylic acid.
  • the additional pharmaceutically active compound is selected from the group of Third-line agents consisting of Clofazimine, Linezolid, Amoxicillin/clavulanate, Thioacetazone, Imipenem/cilastatin, high-dose isoniazid and Clarithromycin.
  • the additional pharmaceutically active compound is selected from the group consisting of Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin, Rifabutin, Animoglycosides including Kanamycin and/or Amikacin, Polypetides including Capreomycin, Viomycin and/or Streptomycin, fluoroquinolones including Moxifloxacin, Levofloxacin, Ofloxacin and/or Gatifloxacin, thioamides including Ethionamide and/or Protionamide, Cycloserine, Terizidone, Thioacetone, p-Aminosalicylic acid, Clofazimine, Linezolid, Amoxicillin, Clavulanate, Thioacetazone, Imipenem, Cilastatin, Clarithromycin, Delamanid and/or Bedaquiline.
  • the additional pharmaceutically active compound is selected from the group consisting of Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin, Rifabutin, Animoglycosides including Kanamycin and/or Amikacin, Polypetides including Capreomycin, Viomycin and/or Streptomycin, fluoroquinolones including Moxifloxacin, Levofloxacin, Ofloxacin and/or Gatifloxacin, thioamides including Ethionamide and/or Protionamide, Cycloserine, Terizidone, Thioacetone and/or p-Aminosalicylic acid.
  • the additional pharmaceutically active compound is selected from the group consisting of Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin and/or Rifabutin. In some embodiments the additional pharmaceutically active compound is selected from the group consisting of Isoniazid, Rifampicin, Ethambutol and/or Pyrazinamide. In some embodiments the additional pharmaceutically active compound is Isoniazid. In some embodiments the additional pharmaceutically active compound is Rifampicin. In some embodiments the additional pharmaceutically active compound is Ethambutol. In some embodiments the additional pharmaceutically active compound is Pyrazinamide. In some embodiments the additional pharmaceutically active compound is Rifabutin. In some embodiments the additional pharmaceutically active compound is Delamanid. In some embodiments the additional pharmaceutically active compound is Bedaquiline.
  • the additional pharmceutically active compound an antifungal compound.
  • the antifungal compound selected from the group consisting of Allylamines including Terbinafin and/or Naftifin, Azole-Antimycotics including Bifonazole, Butoconazole, Clotrimazole, Econazole, Fenticonazole, Fluconazole, Isoconazole, Itraconazole, Ketoconazole, Miconazole, Oxiconazole, Posaconazole, Voriconazole, Efinaconazole, Luliconazole, Sertaconazole and/or Tioconazole, Benzylamines including Butenafine, Polyenes including Amphotericin B, Nystatin and/or Pentamycine, Morpholine- Derivates including Amorolfine, Hydroxypyridone derivates including Ciclopirox, Echinocandins including Anidulafungin, Caspo
  • the additional pharmceutically active compound an anti-HIV compound.
  • the anti-HIV compound selected from the group consisting of Nucleoside/Nucleotide Reverse Transcriptase Inhibitors (NRTIs) including Abacavir, Atripla, Combivir, Complera, Didanosine, Emtriva, Entecavir, Epivir, Epzicom, Retrovir, Trizivir, Truvada, Videx, Videx EC, Viread, Zerit and/or Ziagen, Non-Nucleoside Reverse Transcriptase Inhibitors (NNRTIs) including Edurant.lntelence, Rescriptor, Sustiva, Viramune and/or Viramune XR, Protease Inhibitors (Pis) including Aptivus, Crixivan, Evotaz, Invirase, Kaletra, Lexiva, Norvir, Prezcobix, Prezista, Reyataz and/or Viracept, Entry/ Fusion In
  • NRTIs Nu
  • A is selected from the group consisting of:
  • X is a nitrogen containing heterocycle
  • Y is CH 2 , O or NR 1 ; n is an integer between 0 and 3;
  • the present invention also envisions a method of treating in a subject a mycobacterial infection, in particular tuberculosis, comprising administering to said subject an efficient amount of a compound according to formula (I) or a pharmaceutically acceptable salt, solvate or hydrate thereof or a pharmaceutical composition comprising said compound.
  • Said method preferably comprises further administering at least one additional pharmaceutically active compound, including an antibiotic, antifungal or anti-HIV compound.
  • Figure 1 Mycobacterium tuberculosis growth assay with GFP expression readout. Lalistat inhibits Mycobacterium tuberculosis growth at low micromolar concentrations. GFP: Green Fluorescent Protein, RLU: Relative light units.
  • FIG. 1 Activity on human macrophages infected with Mycobacterium tuberculosis strain H37Rv. Lalistat inhibits Mycobacterium tuberculosis growth at low micromolar concentrations. CFU: Colony Forming Units, DMSO: Dimethylsulfoxid 0,1 %.
  • Figure 3 Effect of 40 ⁇ lalistat (La-0) on the growth of Staphylococcus aureus, Escherichia coli and Listeria monocytogenes.
  • Figure 4 A) Synthetic route to Lalistat (La-0) and lalistat probe (La-1 ). B) H NMR of lalistat probe (La-1 ). TFA: Trifluoroacetic acid, DIEA: ⁇ , ⁇ -Diisopropylethylamine, THF: Tetrahydrofurane, Boc: tert-butyloxycarbonyl.
  • Figure 5 A) Chemical structure of lalistat La-0 and lalistat probe La-1.
  • RLU Relative light units.
  • Figure 6 In vivo target identification / deconvolution via chemical proteomics: Activity based protein profiling (ABPP) with 50 ⁇ lalistat probe La-1.
  • DMSO Dimethylsulfoxid.
  • Figure 7 Activity based protein profiling (ABPP) workflow including DMSO control and competition control experimental procedures.
  • DMSO Dimethylsulfoxid.
  • Figure 8 Alignment of selected target protein hits using the Universal Protein Resource (UniProt) platform (Reorganizing the protein space at the Universal Protein Resource (UniProt). Nucleic Acids Res, 2012. 40(Database issue): p. D71-5).
  • P38571 is (SEQ ID No: 1 )
  • P95125 is (SEQ ID No: 2)
  • P71668 is (SEQ ID No: 3)
  • P9WK85 is (SEQ ID No: 4).
  • Figure 9 Validation of protein target LipR identified via ABPP: LipR was overexpressed in E. coli and labelled with lalistat probe La-1. Rhodamine-Azide was attached via Click reaction after cell lysis. The experiment was conducted with and without induction by IPTG (Isopropyl- -D- thiogalactopyranosid). Fluorescence SDS-PAGE analysis and the corresponding coomassie stained gel are shown.
  • ABPP Activity Based Protein Profiling.
  • Figure 10 Competition ABPP experiment visualized via SDS-PAGE and fluorescent scanning.
  • Figure 11 Proposed mechanism of target inhibition by Lalistat.
  • 4-Ethynylpiperidine-TFA salt was prepared according to Braisted et al. (Braisted, A.C., et al., Discovery of a potent small molecule IL-2 inhibitor through fragment assembly. J Am Chem Soc, 2003. 125(13): p. 3714-5).
  • the mobile phase for elution consisted of a gradient mixture of 0.1 % (v/v) TFA in water (buffer A, HPLC grade) and 0.1 % (v/v) TFA in ACN (buffer B, HPLC grade) unless noted otherwise.
  • GFP-expressing Mycobacterium tuberculosis H37Rv (Michelucci, A., et al., Immune-responsive gene 1 protein links metabolism to immunity by catalyzing itaconic acid production).
  • Proc Natl Acad Sci U S A, 2013. 110(19): p. 7820-5) were generated using the plasmid 32362:pMN437 (Addgene), kindly provided by M. Niederweis (University of Alabama, Birmingham, AL) (Song, H., et al., Identification of outer membrane proteins of Mycobacterium tuberculosis. Tuberculosis (Edinb), 2008. 88(6): p. 526-44).
  • Mononuclear cells were isolated from peripheral blood (PBMC) of healthy volunteers by density gradient centrifugation. Monocytes were separated (purity consistently >95%) by counterflow elutriation.
  • Human monocyte-derived Macrophages hMDM
  • M-CSF human macrophage colony-stimulating factor
  • tuberculosis growth in human macrophages was analyzed as described (Reiling, N., et al., Clade-specific virulence patterns of Mycobacterium tuberculosis complex strains in human primary macrophages and aerogenically infected mice. MBio, 2013. 4(4)).
  • 2x10 5 hMDMs were cultured in 500 ⁇ RPMI 1640 with 10% FCS and 4mM L-glutamine in 48-well flat-bottom microtiter plates (Nunc) at 37°C in a humidified atmosphere containing 5% C0 2 .
  • Macrophages were infected with M. tuberculosis strain H37Rv with a multiplicity of infection (MOI) of 1 :1.
  • HBSS Hanks' balanced salt solution
  • Invitrogen Invitrogen
  • 0.5 ml media was added to the macrophage culture.
  • supernatants were completely removed and macrophage cultures were lysed at 4 hours and 7 days post infection by adding 10 ⁇ 10% Saponin solution (Sigma) in HBSS at 37°C for 15 min.
  • Lysates were serially diluted in sterile water containing 0.05% Tween 80 (Merck, Darmstadt, Germany) and plated twice on 7H10 agar containing 0.5% glycerol (Serva) and 10% heat-inactivated bovine calf serum (BioWest, France). After 3 weeks at 37°C the colony forming units (CFUs) were counted.
  • ABPP Activity based protein profiling
  • M. tuberculosis cells were grown to stationary phase and incubated with La-1 or DMSO as a control. After cell lysis a rhodamine tag was attached to the alkyne moiety of La-1 via bioorthogonal click-reaction (R.Huisgen, Proc Chem Soc, 1961 , 357.; V. V. Rostovtsev, et al., Angew Chem Int Ed Engl, 2002, 41 , 2596.; C. W. Tornoe, C. Christensen and M.
  • Bacterial culture was derived from frozen stock (2.5x10 8 bacteria/ml). Homogenous bacterial suspension was prepared in 7H9 medium (50 mL) supplemented with oleic acid-albumin-dextrose-catalase (OADC) (10%), Tween 80 (0.05%), and glycerol (0.2%). 25 mL each was incubated in 30 mL square medium bottles (Nalgene) at 37°C without shaking for three days. Preculture was diluted to 450 mL and incubated for four days. Bacteria were washed with PBS and an optical density at 600 nm of 40 was adjusted.
  • OADC oleic acid-albumin-dextrose-catalase
  • the lysates were mixed by vortexing and 20 ⁇ CuS0 solution (50 rriM in ddH 2 0) (two times distilled water) were added. The lysates were mixed by vortexing again and incubated for 1 h at RT in the dark. After the click-reaction the lysates were transferred to 15 mL falcon tubes and 8 mL of cold acetone (-80°C, MS grade) were added. Proteins were precipitated ON at -80°C. The precipitated proteins were thawed on ice, pelletized (16,900 xg, 15 min, 4°C) and supernatant was disposed.
  • Beads were washed 3 times with 1 mL 0.2 % SDS in PBS, 2 times with 1 mL 6 M urea in water and 3 times with 1 mL PBS (resuspension: carefully inverting 20 times, pelleting: 400 xg, 3 min, RT).
  • the beads were resuspended in 200 ⁇ denaturation buffer (7 M urea, 2 M thiourea in 20 mM pH
  • thermomixer 450 rpm, 2 h, RT, in the dark.
  • TEAB solution 600 pL, 50 mM in water
  • trypsin 1.5 pL, 0.5 pg/pL in 50 mM acetic acid, Promega
  • the reaction was incubated in a thermomixer (450 rpm, 13-15 h, 37 °C).
  • the digest was stopped by adding 6 pL formic acid (FA) and vortexing.
  • Samples were analyzed via HPLC-MS/MS using an UltiMate 3000 nano HPLC system (Dionex, Sunnyvale, California, USA) equipped with Acclaim C18 PepMap100 75 pm ID x 2 cm trap and Acclaim C18 PepMap RSLC, 75 pM ID x 15 cm separation columns coupled to an Orbitrap Fusion mass spectrometer (Thermo Fisher Scientific Inc., Waltham, Massachusetts, USA). Samples were loaded on the trap and washed for 10 min with 0.1 % formic acid, then transferred to the analytical column and separated using a 120 min gradient from 3% to 25% acetonitrile in 0.1 % formic acid and 5% dimethyl sulfoxide (at 200 nL/min flow rate).
  • Orbitrap Fusion was operated in a 3 second top speed data dependent mode. Full scan acquisition was performed in the orbitrap at a resolution of 120000 and an ion target of 4E5 in a scan range of 300 - 1700 m/z. Monoisotopic precursor selection as well as dynamic exclusion for 60 s were enabled. Precursors with charge states of 2 - 7 and intensities greater than 5E3 were selected for fragmentation. Isolation was performed in the quadrupole using a window of 1.6 m/z. Precursors were collected to a target of 1 E2 for a maximum injection time of 250 with "inject ions for all available parallelizable time" enabled. Fragments were generated using higher-energy collisional dissociation (HCD) and detected in the ion trap at a rapid scan rate. Internal calibration was performed using the ion signal of fluoranthene cations (EASY-ETD/IC source)
  • Peptide and protein identifications were performed using MaxQuant 1.5.3.8 software with Andromeda as search engine using following parameters: Carbamidomethylation of cysteines as fixed and oxidation of methionine as well as acetylation of N-termini as dynamic modifications, trypsin/P as the proteolytic enzyme, 4.5 ppm for precursor mass tolerance (main search ppm) and 0.5 Da for fragment mass tolerance (ITMS MS/MS tolerance). Searches were done against the Uniprot database for M. tuberculosis H37Rv (taxon identifier: 83332, downloaded on 19.5.2015).
  • Quantification was performed using dimethyl labeling with the following settings: light: DimethLysO, DimethNterO; medium: Dimethl_ys4, DimethNter4 and heavy: Dimethl_ys8, DimethNter8. Variable modifications were included for quantification.
  • the I L and re-quantify options were used. Identification was done with at least 2 unique peptides and quantification only with unique peptides.
  • Proteins were finally ranked according to the sum of the ranking values from dimethyllabeling ratios and -log-io(p-value) across both experiments (regular ABPP: probe / DMSO, competition experiment: probe / competition). 2% of the identified proteins with the highest final ranking (including regular ABPP and competition experiments dimethyllabeling ratios and p-values) were considered to be hits of lalistat. This cut-off was chosen as by this analysis a visual separation of enriched vs. not enriched proteins in both regular ABPP and competition experiments could be achieved. 1.5) Recombinant expression and labelling of proteins in E. coli
  • a N-terminal His 6 affinity tagged LipR construct was PCR amplified from genomic DNA from M. tuberculosis H37Rv using the primers shown below and cloned in a pDONR201 (Invitrogen) vector and then in a pET300 expression vector via the GATEWAY cloning system. Expression was induced at an OD 600 of 0.6 by addition of lsopropyl-3-D-thiogalactopyranosid (IPTG; final concentration: 0.25 mM) and carried out 4 h at 37°C in E. coli BL21 cells.
  • IPTG lsopropyl-3-D-thiogalactopyranosid
  • Primer 1 ggggacaagtttgtacaaaaagcaggctttgagaatctttattttcagggcAACCTGCGCAAAAACGTCATCC (SEQ ID No: 5)
  • Primer 2 ggggaccactttgtacaagaaagctgggtgTCATTTGACTACTCCCCGTGG (SEQ ID No: 6)
  • Pellets were resuspended in 1 ml_ PBS (4°C) and transferred to a 'Precellys Glass/Ceramic Kit SK38 2.0 ml.' tube. Tubes were cooled on ice for about 5 min or longer and cells were lysed with the Precellys Homogeniser using two times lysis program 3 (5,400 rpm, run number: 1 , run time: 20 sec, pause: 5 sec). After each lysis run the tubes were cooled on ice for 5 min. The ball mill tubes were centrifuged (16,200 xg, 10 min, 4°C) and 86 ⁇ _ of supernatant were transferred to new 1.5 ml.
  • the lysates were mixed by vortexing and 2 ⁇ _ CuS0 4 solution (50 mM in ddH20) were added. The lysates were again mixed by vortexing and incubated for 1 h at RT in the dark. Then 80 ⁇ _ 2x Laemmli Sample Buffer were added, samples were mixed in a thermomixer (300 rpm, 3 min, 96°C) and analyzed via SDS PAGE (10 % agarose gel (PEQLAB Biotechnologie GmbH, Er Weg, PerfectBlue Dual Gel System, the gel was prepared according to the manual), 3.5 h, 300 V, 8 ⁇ _ fluorescent protein standard) and fluorescence imaging (GE Healthcare, ImageQuant LAS- 4000). After fluorescence scanning the gel was Coomassie stained.
  • the lalistat-derived probe (La-1 ) retained activity and reduced M. tuberculosis growth comparable to the parent molecule (La-0) ( Figure 5A and 5B).
  • target identification experiments were conducted. Bacterial cells were grown to stationary phase and incubated with lalistat probe (La-1 ) or DMSO as a control. After cell lysis a biotin or rhodamine tag was attached to the alkyne tag of La-1 via bioorthogonal click-reaction (Huisgen, R., 1,3-Dipolar Cycloadditions. Proc. Chem. Soc, 1961 : p.
  • the combined target spectrum includes 7 proteins from the Lip family among other lypolytic / hydrolytic enzymes.
  • a proposed inhibition mechanism for those enzymes is depicted in figure 1 1.
  • An alignment of the amino acid sequences of the three most enriched Lip proteins (LipN, Lipl and LipR) together with the known human lalistat target lysosomal acid lipase LipA reveals the active site consensus sequence: GDSAGGXI/L (SEQ ID No: 7) or GXSXGXXI/L (SEQ ID No: 8) ( Figure 8).
  • the inventors were able to validate one of the target proteins (LipR) by a heterologous expression labeling experiment in E. coli ( Figure 9).
  • M. tuberculosis LipR was overexpressed in the host and could be labeled also in this strain using chemical tool compound La-1 in combination with rhodamine azide and SDS-PAGE fluorescence visualization.
  • LipN showed the strongest drug binding characteristics. LipN, similar to Lipl, which was the second highest hit among the identified members of the lip family, possesses esterase activity on short chain TGs. In contrast, like many members of the mycobacterial HSL-family, LipR shows no TG hydrolysing activity but rather acts on other short- and mid chain substrates (Delorme, V. et al. MmPPOX Inhibits Mycobacterium tuberculosis Lipolytic Enzymes Belonging to the Hormone-Sensitive Lipase Family and Alters Mycobacterial Growth. PLoS One 7, (2012)).
  • Lipl and LipG Underlining the important function of lipases from the lip family in mycobacterial infection, Lipl and LipG have both been shown to be essential for mycobacterial growth in vitro and in macrophages, respectively(Rengarajan, J., Bloom, B. R. & Rubin, E. J. Genome-wide requirements for Mycobacterium tuberculosis adaptation and survival in macrophages. Proc. Natl. Acad. Sci. U. S. A. 102, 8327-8332 (2005), Griffin, J. E. et al. High-resolution phenotypic profiling defines genes essential for mycobacterial growth and cholesterol catabolism. PLoS Pathog.
  • LipO is indispensable for prevention of endosomal maturation and acidification (Pethe, K. et al. Isolation of Mycobacterium tuberculosis mutants defective in the arrest of phagosome maturation. Proc. Natl. Acad. Sci. U. S. A. 101, 13642-7 (2004)). Furthermore, lipases belonging to the HSL family have been reported to be up-regulated during starvation after TG accumulation (Danelishvili, L, Poort, M. J. & Bermudez, L. E. Identification of Mycobacterium avium genes up-regulated in cultured macrophages and in mice. FEMS Microbiol. Lett. 239, 41- 49 (2004), Fisher, M.
  • LipY is the only member reported to hydrolyse long chain TG and, thus, is regarded as the only "real" lipase. From the pattern of the targets identified by these investigations it can be concluded that lalistat preferentially binds to lipolytic enzymes, which show a wider substrate range including those possessing shorter acyl chains or monoacylglycerides compared to TG-hydrolysing lipases.
  • Rv0183 The second highest hit of our lalistat target profiling, Rv0183, is an among the mycobacterial species highly conserved monoalcylglycerol lipase and, similar to as described for mycobacterial lipases belonging to the HSL family, has been proposed as an important anti-mycobacterial target in persistent TB (Saravanan, P., Dubey, V. K. & Patra, S. Potential Selective Inhibitors against Rv0183 of Mycobacterium tuberculosis Targeting Host Lipid Metabolism. Chem. Biol. Drug Des. 79, 1056-1062 (2012)).
  • Rv1984 represents the only enzyme among the first 10 identified lalistat-targets, which belongs to the cutinase-family. It is located in the bacterial cell wall and is proposed to be involved in exogenous host cell lipid hydrolysis (Dhouib, R., Laval, F., Carriere, F., Daffe, M. & Canaan, S. A monoacylglycerol lipase from Mycobacterium smegmatis involved in bacterial cell interaction. J. Bacteriol. 192, 4776-4785 (2010)). Its lypolytic activity has been reported to be preferentially directed against medium-chain carboxylic esters and monoacylglycerols underlining its potential role in mycobacterial pathogenicity during persistence.
  • cutinases are serine esterases active on a wider panel of substrates including cutins from plants, phospholipids and acylglycerols, it is important to note that other mycobacterial cutinases like RV3452 preferentially targeting non-acylglycerols have not been detected in our target identification experiment (Schue, M. et al. Two cutinase-like proteins secreted by Mycobacterium tuberculosis show very different lipolytic activities reflecting their physiological function. FASEB J. 24, 1893-1903 (2010)).
  • Rv2715 is the fourth strongest lalistat-binding hit. It represents a still uncharacterized protein, which is suggested to participate in lipid hydrolysis in bacterial metabolism. However, its detailed function is still unknown. Further studies have to be performed to investigate its capability to serve as a potential anti-mycobacterial target. Similar to RV2715, Rv1 192, which has also been identified among the 10 strongest lalistat-binding proteins, is a further uncharacterized protein suggested to belong to the lipolytic enzyme family. It has a membranous localisation and contains a PS00120 lipase pattern.
  • the inventors detected two proteins (third and seventh hit of our screen) similar to other Mycobacterium tuberculosis hypothetical penicillin binding proteins and esterases, which are possibly involved in cell wall biosynthesis: Rv1367 and Rv1730.
  • Rv1730 it has been shown that its function is essential for mycobacterial growth in vitro (Sassetti, C. M., Boyd, D. H. & Rubin, E. J. Genes required for mycobacterial growth defined by high density mutagenesis. Mol. Microbiol. 48, 77-84 (2003)) and might represent an interesting anti-mycobacterial target.
  • Rv2888c and Rv2363 belong to the family of putative amidases and Rv0840c is a putative proline iminopeptidase.
  • Rv2888c has been identified in chronic but not early stages of disease when investigated in the guinea pig model of tuberculosis (Kruh, N. A., Troudt, J., Izzo, A., Prenni, J. & Dobos, K. M. Portrait of a pathogen: The Mycobacterium tuberculosis proteome In vivo. PLoS One 5, (2010)).
  • Rv0293c identified as the protein hit 1 1 , is a non-characterised protein with unkown function. It is highly conserved and upregulated during starvation (Sassetti, C. M., Boyd, D. H. & Rubin, E. J. Genes required for mycobacterial growth defined by high density mutagenesis. Mol. Microbiol. 48, 77-84 (2003)). Finally, Rv1 191 representing the target with the lowest affinity for lalistat identified, is a conserved protein with unknown function and distinct similarity to proline iminopeptidases.
  • Multi-targeting is an effective approach to avoid bacterial drug resistance while combating the disease (Saravanan, P. & Patra, S. Discovery of Potential Dual Inhibitors against Lipases
  • the invention is characterized by the following items:
  • a compound for use in the treatment of a mycobacterial disease said compound having a structure according to Formula I wherein
  • X is a nitrogen containing heterocycle
  • Y is CH 2 , O or NR 1 ; n is an integer between 0 and 3;
  • R 1 and R 2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (C T C 6 )alkyl, substituted or unsubstituted (C 2 -C 6 )alkenyl, substituted or unsubstituted (C 2 -C 6 )alkynyl, substituted or unsubstituted (C 3 -C 8 )cycloalkyl, substituted or unsubstituted (C 6 -Ci 0 )aryl, substituted or unsubstituted (C 6 -Ci 0 )aryl(Ci-C 6 )alkyl, substituted or unsubstituted (C 3 -C 10 )heteroaryl, and substituted or unsubstituted (C 3 - Cio)heteroaryl(Ci-C 6 )alkyl; or a pharmaceutically acceptable salt, solvate or hydrate thereof;
  • a compound for use in medicine said compound having a structure according to Formula I
  • X is a nitrogen containing heterocycle
  • Y is CH 2 , O or NR ; n is an integer between 0 and 3;
  • R 1 and R 2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C 6 )alkyl, substituted or unsubstituted (C 2 -C 6 )alkenyl, substituted or unsubstituted (C 2 -C 6 )alkynyl, substituted or unsubstituted (C 3 -C 8 )cycloalkyl, substituted or unsubstituted (C6-C 10 )aryl, substituted or unsubstituted (Ce-C ⁇ JaryliC CeJalkyl, substituted or unsubstituted (C 3 -Ci 0 )heteroaryl, and substituted or unsubstituted (C 3 - or a pharmaceutically acceptable salt, solvate or hydrate thereof;
  • compositions for use in the treatment of a mycobacterial disease comprising a compound having a structure according to Formula I
  • X is a nitrogen containing heterocycle
  • Y is CH 2 , O or NR ; n is an integer between 0 and 3;
  • composition for use according to item 3 wherein A is The compound for use or the pharmaceutical composition for use according to any one of items1-4, wherein
  • Z is CR 3 R 4 , O or NR 5 ;
  • R 3 and R 4 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C 6 )alkyl, substituted or unsubstituted (C 2 -C 6 )alkenyl, substituted or unsubstituted (C 2 -C 6 )alkynyl, substituted or unsubstituted (C 3 -C 8 )cycloalkyl, substituted or unsubstituted (C 6 -Ci 0 )aryl, substituted or unsubstituted (C 6 -Ci 0 )aryl(Ci-C 6 )alkyl, substituted or unsubstituted (C 3 -C 10 )heteroaryl, substituted or unsubstituted (C 3 - CioJheteroaryliCr alkyl,
  • R 5 is selected from the group consisting of hydrogen, substituted or unsubstituted (Cr C 6 )alkyl, substituted or unsubstituted (C 2 -C 6 )alkenyl, substituted or unsubstituted (C 2 - C B )alkynyl, substituted or unsubstituted (C 3 -C 8 )cycloalkyl, substituted or unsubstituted (C 6 -Cio)aryl, substituted or unsubstituted (C 6 -Ci 0 )aryl(Ci-C 6 )alkyl, substituted or unsubstituted (C 3 -C 10 )heteroaryl, substituted or unsubstituted
  • W is independently selected from O, S, and N(R 14 );
  • R 1 is, in each case, selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, and heterocyclyl;
  • R 14 is independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, heterocyclyl, and -OR 11 ;
  • R 15 and R 16 are independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, and heterocyclyl.
  • Z is O or CR 3 R 4 ;
  • R 3 and R 4 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (C T C 6 )alkyl, substituted or unsubstituted (C 2 -C 6 )alkynyl and -OR 11 ;
  • R 11 is, in each case, selected from the group consisting of hydrogen and substituted or unsubstituted (Ci-C 6 )alkyl;
  • R 3 and R 4 are hydrogen or substituted or unsubstituted (C C 6 )alkyl; or wherein at least one of R 3 and R 4 is hydrogen.
  • mycobacterial disease is selected from tuberculosis, leprosy (Hansen's disease), lepromatosis, infections caused by non-tuberculosis mycobacteria including lymphadenitis and pulmonary infections, skin infections caused by mycobacteria including Buruli ulcer and fish tank granuloma.
  • mycobacterial disease is tuberculosis and/or caused by bacteria of the group consisting of Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae, Mycobacterium microti, preferably Mycobacterium tuberculosis.
  • compositions for use according to any one of items 3-10 further comprising at least one pharmaceutically acceptable carrier.
  • the pharmaceutical composition for use according to any one of items 3-1 1 wherein said composition is administered to a subject in need thereof in an amount effective to treat said mycobacterial disease, preferably wherein the subject is a mammal, more preferably a human subject.
  • 13 The pharmaceutical composition for use according to item 12, wherein said subject is treated with at least one additional pharmaceutically active compound, including an antibiotic, antifungal or anti-HIV compound, preferably wherein said additional pharmaceutically active compound is an antibiotic, antifungal and/or anti-HIV compound, more preferably an antibiotic.
  • compositions for use according to item 13 wherein said additional pharmaceutically active compound is selected from the group consisting of Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin, Rifabutin, Animoglycosides including Kanamycin and/or Amikacin, Polypetides including Capreomycin, Viomycin and/or Streptomycin, fluoroquinolones including Moxifloxacin, Levofloxacin, Ofloxacin and/or
  • A is selected from the group consisting of:
  • X is a nitrogen containing heterocycle
  • Y is CH 2 , O or NR ; n is an integer between 0 and 3;
  • R 1 and R 2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C 6 )alkyl, substituted or unsubstituted (C 2 -C 6 )alkenyl, substituted or unsubstituted (C 2 -C 6 )alkynyl, substituted or unsubstituted (C 3 -C 8 )cycloalkyl, substituted or unsubstituted (C 6 -Ci 0 )aryl, substituted or unsubstituted (C 6 -Ci 0 )aryl(Ci-C 6 )alkyl, substituted or unsubstituted (C 3 -Ci 0 )heteroaryl, and substituted or unsubstituted (C 3 - Cio)heteroaryl(Ci-C 6 )alkyl; or a pharmaceutically acceptable salt, solvate or hydrate thereof;
  • said additional pharmaceutically active compound is selected from the group consisting of an antibiotic, antifungal or anti-HIV compound, preferably an antibiotic selected from the group of Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin, Rifabutin, Animoglycosides including Kanamycin and/or Amikacin, Polypetides including Capreomycin, Viomycin and/or Streptomycin, fluoroquinolones including Moxifloxacin, Levofloxacin, Ofloxacin and/or Gatifloxacin, thioamides including Ethionamide and/or Protionamide, Cycloserine, Terizidone, Thioacetone, p-Aminosalicylic acid, Clofazimine, Linezolid, Amoxicillin, Clavulanate, Thioacetazone, Imipenem, Cilastatin, Clarithro

Landscapes

  • Health & Medical Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Veterinary Medicine (AREA)
  • Medicinal Chemistry (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Public Health (AREA)
  • Epidemiology (AREA)
  • General Chemical & Material Sciences (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Oncology (AREA)
  • Communicable Diseases (AREA)
  • Organic Chemistry (AREA)
  • Pulmonology (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)

Abstract

The invention relates to compounds which are suitable for treating mycobacterial diseases and to pharmaceutical compositions containing such compounds. Also encompassed are such compounds for use in medicine. The invention further relates to a kit of parts comprising a pharmaceutical composition containing such compounds and at least one additional pharmaceutically active compound.

Description

MEANS AND METHODS FOR TREATING MYCOBACTERIAL DISEASES
The invention relates to compounds which are suitable for treating mycobacterial diseases and to pharmaceutical compositions containing such compounds. Also encompassed are such compounds for use in medicine. The invention further relates to a kit of parts comprising a pharmaceutical composition containing such compounds and at least one additional pharmaceutically active compound.
The mycobacteria are a diverse collection of acid-fast, non-motile, gram-positive bacteria. It comprises several species, which include, Mycobacterium africanum (M. africanum), M. avium, M. bovis, M. bovis-BCG, M. chelonae, M. fortuitum, M. gordonae, M. intracellular, M. kansasii, M. microti, M. scrofulaceum, M. paratuberculosis, M. leprae, M. tuberculosis, and M. ranae. Certain of these organisms are the causative agents of disease. Tuberculosis and leprosy (Hansen's disease) are the best known mycobacterial diseases. People may also be infected by any of a group of mycobacterial species collectively called non-tuberculous mycobacteria (NTM). In children, NTM cause lymphadenitis, skin and soft tissue infections, and occasionally also lung disease and disseminated infections. Manifestations can be indistinguishable from tuberculosis on the basis of clinical and radiological findings and tuberculin skin testing. Although over 150 different species of NTM have been described, pulmonary infections are most commonly due to Mycobacterium avium complex (MAC), Mycobacterium kansasii, and Mycobacterium abscessus.
Tuberculosis (TB) is one of the world's most prevalent infectious diseases responsible for the largest fraction of infection related casualties (before HIV). The disease is caused by bacteria of the Mycobacterium tuberculosis Complex. Species in this complex include M. tuberculosis, M. africanum, M. bovis, M. caprae etc, whereby M. tuberculosis is the main causative agent of tuberculosis. Tuberculosis usually affects the lung and represents the most frequent form of TB. Worldwide, TB has an estimated incidence of 9.6 million people with 1.5 million annual deaths in 2014. Tuberculosis is usually controlled using extended antibiotic therapy. There are six front-line drugs known to be highly effective against M. tuberculosis and several second-line drugs including streptomycin as well as third-line drugs, which are used when resistance to one or more of the front-line drugs is detected. The preferred mode of treatment for tuberculosis is the short course chemotherapy in which there are two phases. The first phase consists of a daily regimen for two months with isoniazid (300 mg), rifampicin (600 mg), pyrazinamide (3 g) and ethambutol (1.5 g). The second phase or the continuation phase consists of a daily regimen for the next four months with isoniazid and rifampicin. The vast majority of individuals infected with drug susceptible M. tuberculosis strains can be effectively cured when medicines are provided and taken properly. However, inappropriate or incorrect use of antimycobacterial drugs, or use of ineffective formulations of drugs (e.g. use of single drugs, poor quality medicines or bad storage conditions), and premature treatment interruption can cause drug resistance of the bacteria, which can then be transmitted, especially in crowded settings such as prisons and hospitals. According to WHO estimates 0.48 million people were infected with strains resistant to isoniazid and rifampicin (MDR-TB; multidrug-resistant tuberculosis) in 2014 leading to 0.19 million deaths (WHO, Global tuberculosis report. 2014). About 9% of MDR-TB patients were infected with an almost untreatable extensively drug-resistant (XDR-TB) variant. Mycobacterium tuberculosis is transmitted by aerosol and is initially taken up by alveolar lung macrophages, which phagocytose but don't kill the bacterium. Most infected individuals can control the disease for a long time. Estimates believe that one-third of the world's population is latently infected (LTBI; latent tuberculosis infection). The bacteria adapt and survive in diverse environmental niches in vivo, e.g. in solid granulomas, a characteristic feature of latent TB infection. It is presumed that M. tuberculosis resides in these regions in a slow growing or non- replicating, phenotypically drug resistant dormant-like state, due to limited availability and supply of oxygen and nutrients (Gengenbacher, M. and S.H. Kaufmann, Mycobacterium tuberculosis: success through dormancy. FEMS Microbiol Rev, 2012. 36(3): p. 514-32).
During infection it has been shown that M. tuberculosis accumulates triacylglycerols (TAGs) within intracellular inclusion bodies (Garton, N.J., et al., Intracellular lipophilic inclusions of mycobacteria in vitro and in sputum. Microbiology, 2002. 148(Pt 10): p. 2951-8). The hydrolysis of TAGs to free fatty acids is an essential prerequisite for M. tuberculosis growth and survival requiring diverse lipases and hydrolases Moreover, it is believed that these fatty acids are crucial for bacteria to enter and maintain the dormant state by the production of foamy lung macrophages during latent infection (Peyron, P., et al., Foamy macrophages from tuberculous patients' granulomas constitute a nutrient-rich reservoir for M. tuberculosis persistence. PLoS Pathog, 2008. 4(1 1 ): p. e1000204 and Daniel, J., et al., Mycobacterium tuberculosis uses host triacylglycerol to accumulate lipid droplets and acquires a dormancy-like phenotype in lipid- loaded macrophages. PLoS Pathog, 201 1. 7(6): p. e1002093). Specifically targeting the bacterial lipid metabolism could represent a viable strategy to limit the growth of M. tuberculosis and open a new opportunity to shorten the long TB therapy (Warner DF, Mizrahi V. Shortening treatment for tuberculosis— to basics. N Engl J Med, 2014. 371 (17):1642-3), when given simultaneously with known first and second line antibiotics. Lalistat is a thiadiazole carbamate initially developed as a specific inhibitor of the mammalian acid lipase, an enzyme located in cellular late endosomes hydrolysing cholesterol esters and triglycerides from incoming lipoproteins. Lalistat covalently binds to the active site serine of lipases and thereby inhibits enzyme activity and is considered as a potential Niemann-Pick Type C disease therapeutic (Rosenbaum, A.I., et al., Thiadiazole carbamates: potent inhibitors of lysosomal acid lipase and potential Niemann-Pick type C disease therapeutics. J Med Chem, 2010. 53(14): p. 5281-9). In addition, WO 2015/135652 discloses lalistat and derivatives thereof for the treatment or prevention of viral infections, in particular Hepatitis C virus infection.
Schwaid et al., "Development of a selective activity-based probe for glycosylated UPA", Bioorganic & Medicinal Chemistry Letters, vol. 26, no. 8, pages 1993 to 1996, discloses a lysosomal acid lipase (LIPA) activity based probe, which is specific for particular form of glycosylated LIPA in cells.
Jayendra Z. Patel et al. "Optimization of 1,2,5-Thiadiazole Carbamates as Potent and Selective ABHD6 Inhibitors" CHEMMEDCHEM, vol. 10, no. 2, pages 253 to 265, discloses a study directed to potential new inhibitors of α/β-hydrolase domain 6 (ABHD6). Additionally, activity- based protein profiling indicated that JZP-430 displays good selectivity among the serine hydrolases of the mouse brain membrane proteome.
Rosenbaum A. I. et al "Chemical screen to reduce sterol accumulation in Niemann-Pick C disease cells identifies novel lysosomal acid lipase inhibitors" Biochimica and Biophysica Acta. Molecular and Cell Biology of Lipids, vol. 1791 , no. 12, page 1 155 to 1 165 deals with novel lysosomal acid lipase inhibitors, to reduce sterol accumulation in Niemann-Pick C disease cells.
WO 2014/000058 A1 discloses a method of treating or preventing an abnormality of glucose metabolism in a subject, comprising administering an antagonist of lysosomal acid lipase (LAL) to the subject such that LAL is antagonized in a pancreatic β cell of the subject.
Vincent Delorme et al. "MmPPOX Inhibits Mycobacterium tuberculosis Lipolytic Enzymes Belonging to the Hormone-Sensitive Lipase Family and Alters Mycobacterial Growth" PLOS ONE, vol. 7, no. 9, page e46493 discloses a study which shows that MmPPOX inhibits Mycobacterium tuberculosis lipolytic enzymes belonging to the hormone-sensitive lipase family and alters Mycobacterial Growth. M. S. Ravindran et al. "Targeting Lipid Esterases in Mycobacteria Grown Under Different Physiological Conditions Using Activity-based Profiling with Tetrahydroliptatin (THL)", Molecular & Cellular Proteomics, vol. 13, no. 2, pages 435 to 448 discloses that Tetrahydrolipstatin (THL) targets α/β-hydrolases, including many lipid esterases (LipD, G, H, I, M, N, O, V, W, and TesA).
Gurdyal Singh et al. "Lipid hydrolyzing enzymes in virulence: Mycobacterium tuberculosis as a model system" , Critical Reviews in Microbiology, vol. 36, no. 3, pages 259-269 is a review article related to technical background of virulent traits of lipolytic enzymes from bacteria with special emphasis on mycobactehum tuberculosis. Inhibitors for these enzymes are not disclosed.
There is an urgent medical need to identify new means and methods with significant therapeutic activity against mycobacterial diseases in general and in particular against single-or multiple- drug resistant strains of M. tuberculosis and with pharmacokinetic properties that permit reduced dosing resulting also in the reduction of side effects which will in turn encourage better compliance.
The underlying problem of the present invention is the provision of new chemical agents having significant therapeutic activity against mycobacterial diseases, allowing inhibiting the extracellular and intracellular growth of mycobacteria.
The inventors of the present invention have conducted intensive studies and found surprisingly, that thiadiazole derivatives and five-membered heterocyclic compounds derived from lalistat are suitable as anti-mycobacterial agents. It has been found that those compounds inhibit the growth of M. tuberculosis and in particular the intracellular growth of M. tuberculosis in human macrophage host cells, while not affecting the growth of other gram-positive and gram-negative bacteria as shown for Staphylococcus aureus, Escherichia coli and Listeria monocytogenes. Applicant found that the compounds of the present invention inhibit various members of the lipolytic enzyme family, in particular lipases, hydrolases, carboxylesterases, cutinases and uncharacterized proteins including LipN, Lipl, LipR, LipM, LipG, LipO, LipT, , Hydrolase Rv0183, alpha/beta hydrolase Rv1 192, cutinase Rv1984c, Rv1367c, Rv2715, Rv1730c, RV0293c, rv1399c and Rv0045c, as well as the amidases Rv2888c and Rv2363 and the putative proline iminopeptidase Rv0840c (see also Figure 6). Without wishing to be bound by theory, applicant believes that binding to a panel of distinct members of this lipolytic enzyme family mediates the observed anti-bacterial effect of the compound according to the invention. In particular, lipids and fatty acids are crucial for pathogenicity of mycobacteria to survive within the host, inhibition of hydrolytic enzymes involved in the interconversion of this process leads to increased susceptibility of the bacteria. This has the advantages, that the mycobacteria can be killed more efficiently, that drug resistance to the compounds of the present invention is not easily achieved by the targeted mycobacteria and that cross-resistance with other anti-mycobacterial drugs is less likely. Further, the multi-target interaction ability of the compounds makes them to versatile tools and agents for the fight against mycobacterial diseases and in particular for the treatment of MDR-TB and XDR-TB.
Although the present invention is described in detail below, it is to be understood that this invention is not limited to the particular methodologies, protocols and reagents described herein as these may vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to limit the scope of the present invention which will be limited only by the appended claims. Unless defined otherwise, all technical and scientific terms used herein have the same meanings as commonly understood by one of ordinary skill in the art.
In the following, the elements of the present invention will be described. These elements are listed with specific embodiments, however, it should be understood that they may be combined in any manner and in any number to create additional embodiments. The variously described examples and preferred embodiments described throughout the specification should not be construed to limit the present invention to only the explicitly described embodiments. This description should be understood to support and encompass embodiments which combine the explicitly described embodiments with any number of the disclosed and/or preferred elements. Furthermore, any permutations and combinations of all elements described herein should be considered disclosed by the description of the present application unless the context indicates otherwise.
Throughout this specification and the claims which follow, unless the context requires otherwise, the word "comprise", and variations such as "comprises" and "comprising", will be understood to imply the inclusion of a stated member, integer or step or group of members, integers or steps but not the exclusion of any other member, integer or step or group of members, integers or steps although in some embodiments such other member, integer or step or group of members, integers or steps may be excluded, i.e. the subject-matter consists in the inclusion of a stated member, integer or step or group of members, integers or steps. The terms "a" and "an" and "the" and similar reference used in the context of describing the invention (especially in the context of the claims) are to be construed to cover both the singular and the plural, unless otherwise indicated herein or clearly contradicted by context. Recitation of ranges of values herein is merely intended to serve as a shorthand method of referring individually to each separate value falling within the range. Unless otherwise indicated herein, each individual value is incorporated into the specification as if it were individually recited herein.
All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples, or exemplary language (e.g., "such as"), provided herein is intended merely to better illustrate the invention and does not pose a limitation on the scope of the invention otherwise claimed. No language in the specification should be construed as indicating any non-claimed element essential to the practice of the invention.
Unless otherwise indicated, the term "at least" preceding a series of elements is to be understood to refer to every element in the series. Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments of the invention described herein. Such equivalents are intended to be encompassed by the present invention.
When used herein "consisting of" excludes any element, step, or ingredient not specified in the claim element. When used herein, "consisting essentially of" does not exclude materials or steps that do not materially affect the basic and novel characteristics of the claim. In each instance herein any of the terms "comprising", "consisting essentially of" and "consisting of" may be replaced with either of the other two terms.
Several documents are cited throughout the text of this specification. Each of the documents cited herein (including all patents, patent applications, scientific publications, manufacturer's specifications, instructions, etc.), whether supra or infra, are hereby incorporated by reference in their entirety. Nothing herein is to be construed as an admission that the invention is not entitled to antedate such disclosure by virtue of prior invention.
In order that the present invention may be more readily understood, certain terms are first defined. Additional definitions are set forth throughout the description.
As used herein and throughout the entire description, the term "alkyl" refers to a monoradical of a saturated straight or branched hydrocarbon. Preferably, the alkyl group comprises from 1 to 12 (such as 1 to 10) carbon atoms, i.e., 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 1 1 , or 12 carbon atoms (such as 1 , 2, 3, 4, 5, 6, 7, 8, 9, or 10 carbon atoms), more preferably 1 to 8 carbon atoms, such as 1 to 6 or 1 to 4 carbon atoms. Exemplary alkyl groups include methyl, ethyl, propyl, iso-propyl, butyl, iso-butyl, tert-butyl, n-pentyl, iso-pentyl, sec-pentyl, neo-pentyl, 1 ,2-dimethyl-propyl, iso-amyl, n- hexyl, iso-hexyl, sec-hexyl, n-heptyl, iso-heptyl, n-octyl, 2-ethyl-hexyl, n-nonyl, n-decyl, n- undecyl, n-dodecyl, and the like. In some embodiments the alkyl chain is a linear. In some embodiments the alkyl chain is branched. In some embodiments the alkyl chain is substituted. In some embodiment the alkyl chain is unsubstituted. In some embodiments the alkyl chain is linear and substituted or unsubstituted. In some embodiments the alkyl chain is branched and substituted or unsubstituted. As used herein and throughout the entire description, the term "alkenyl" refers to a monoradical of an unsaturated straight or branched hydrocarbon having at least one carbon-carbon double bond. Generally, the maximal number of carbon-carbon double bonds in the alkenyl group can be equal to the integer which is calculated by dividing the number of carbon atoms in the alkenyl group by 2 and, if the number of carbon atoms in the alkenyl group is uneven, rounding the result of the division down to the next integer. For example, for an alkenyl group having 9 carbon atoms, the maximum number of carbon-carbon double bonds is 4. Preferably, the alkenyl group has 1 to 4, i.e., 1 , 2, 3, or 4, carbon-carbon double bonds. Preferably, the alkenyl group comprises from 2 to 10 carbon atoms, i.e., 2, 3, 4, 5, 6, 7, 8, 9, or 10 carbon atoms, more preferably 2 to 8 carbon atoms, such as 2 to 6 carbon atoms or 2 to 4 carbon atoms. Thus, in a preferred embodiment, the alkenyl group comprises from 2 to 10 carbon atoms and 1 , 2, 3, 4, or 5 carbon-carbon double bonds, more preferably it comprises 2 to 8 carbon atoms and 1 , 2, 3, or 4 carbon-carbon double bonds, such as 2 to 6 carbon atoms and 1 , 2, or 3 carbon-carbon double bonds or 2 to 4 carbon atoms and 1 or 2 carbon-carbon double bonds. The carbon- carbon double bond(s) may be in cis (Z) or trans (E) configuration. Exemplary alkenyl groups include vinyl, 1-propenyl, 2-propenyl (i.e., allyl), 1-butenyl, 2-butenyl, 3-butenyl, 1 -pentenyl, 2- pentenyl, 3-pentenyl, 4-pentenyl, 1-hexenyl, 2-hexenyl, 3-hexenyl, 4-hexenyl, 5-hexenyl, 1- heptenyl, 2-heptenyl, 3-heptenyl, 4-heptenyl, 5-heptenyl, 6-heptenyl, 1-octenyl, 2-octenyl, 3- octenyl, 4-octenyl, 5-octenyl, 6-octenyl, 7-octenyl, 1 -nonenyl, 2-nonenyl, 3-nonenyl, 4-nonenyl, 5-nonenyl, 6-nonenyl, 7-nonenyl, 8-nonenyl, 1-decenyl, 2-decenyl, 3-decenyl, 4-decenyl, 5- decenyl, 6-decenyl, 7-decenyl, 8-decenyl, 9-decenyl, and the like. If an alkenyl group is attached to a nitrogen atom, the double bond cannot be alpha to the nitrogen atom. In some embodiments the alkenyl chain is a linear. In some embodiments the alkenyl chain is branched. In some embodiments the alkenyl chain is substituted. In some embodiment the alkenyl chain is unsubstituted. In some embodiments the alkenyl chain is linear and substituted or unsubstituted. In some embodiments the alkenyl chain is branched and substituted or unsubstituted.
As used herein and throughout the entire description, the term "alkynyl" refers to a monoradical of an unsaturated straight or branched hydrocarbon having at least one carbon-carbon triple bond. Generally, the maximal number of carbon-carbon triple bonds in the alkynyl group can be equal to the integer which is calculated by dividing the number of carbon atoms in the alkynyl group by 2 and, if the number of carbon atoms in the alkynyl group is uneven, rounding the result of the division down to the next integer. For example, for an alkynyl group having 9 carbon atoms, the maximum number of carbon-carbon triple bonds is 4. Preferably, the alkynyl group has 1 to 4, i.e., 1 , 2, 3, or 4, more preferably 1 or 2 carbon-carbon triple bonds. Preferably, the alkynyl group comprises from 2 to 10 carbon atoms, i.e., 2, 3, 4, 5, 6, 7, 8, 9, or 10 carbon atoms, more preferably 2 to 8 carbon atoms, such as 2 to 6 carbon atoms or 2 to 4 carbon atoms. Thus, in a preferred embodiment, the alkynyl group comprises from 2 to 10 carbon atoms and 1 , 2, 3, 4, or 5 (preferably 1 , 2, or 3) carbon-carbon triple bonds, more preferably it comprises 2 to 8 carbon atoms and 1 , 2, 3, or 4 (preferably 1 or 2) carbon-carbon triple bonds, such as 2 to 6 carbon atoms and 1 , 2 or 3 carbon-carbon triple bonds or 2 to 4 carbon atoms and 1 or 2 carbon-carbon triple bonds. Exemplary alkynyl groups include ethynyl, 1 -propynyl, 2- propynyl, 1-butynyl, 2-butynyl, 3-butynyl, 1 -pentynyl, 2-pentynyl, 3-pentynyl, 4-pentynyl, 1- hexynyl, 2-hexynyl, 3-hexynyl, 4-hexynyl, 5-hexynyl, 1-heptynyl, 2-heptynyl, 3-heptynyl, 4- heptynyl, 5-heptynyl, 6-heptynyl, 1-octynyl, 2-octynyl, 3-octynyl, 4-octynyl, 5-octynyl, 6-octynyl, 7-octynyl, 1-nonylyl, 2-nonynyl, 3-nonynyl, 4-nonynyl, 5-nonynyl, 6-nonynyl, 7-nonynyl, 8- nonynyl, 1-decynyl, 2-decynyl, 3-decynyl, 4-decynyl, 5-decynyl, 6-decynyl, 7-decynyl, 8-decynyl, 9-decynyl, and the like. If an alkynyl group is attached to a nitrogen atom, the triple bond cannot be alpha to the nitrogen atom. In some embodiments the alkynyl chain is a linear. In some embodiments the alkynyl chain is branched. In some embodiments the alkynyl chain is substituted. In some embodiment the alkynyl chain is unsubstituted. In some embodiments the alkynyl chain is linear and substituted or unsubstituted. In some embodiments the alkynyl chain is branched and substituted or unsubstituted.
As used herein and throughout the entire description, the term "aryl" or "aromatic ring" refers to a monoradical of an aromatic cyclic hydrocarbon. Preferably, the aryl group contains 3 to 14
(e.g., 5 to 10, such as 5, 6, or 10) carbon atoms, more preferably 6 to 10 carbon atoms, which can be arranged in one ring (e.g., phenyl) or two or more condensed rings (e.g., naphthyl).
Exemplary aryl groups include cyclopropenylium, cyclopentadienyl, phenyl, indenyl, naphthyl, azulenyl, fluorenyl, anthryl, and phenanthryl. Preferably, "aryl" refers to a monocyclic ring containing 6 carbon atoms or an aromatic bicyclic ring system containing 10 carbon atoms. Preferred examples are phenyl and naphthyl. In some embodiments the aryl is unsubstituted. In some embodiments the aryl is substituted.
As used herein and throughout the entire description, the term "heteroaryl" or "heteroaromatic ring" means an aryl group as defined above in which one or more carbon atoms in the aryl group are replaced by heteroatoms of O, S, or N. Preferably, the heteroaryl group contains 3 to 10 carbon atoms. Preferably, heteroaryl refers to a five or six-membered aromatic monocyclic ring wherein 1 , 2, or 3 carbon atoms are replaced by the same or different heteroatoms of O, N, or S. Alternatively, it means an aromatic bicyclic or tricyclic ring system wherein 1 , 2, 3, 4, or 5 carbon atoms are replaced with the same or different heteroatoms of O, N, or S. Preferably, in each ring of the heteroaryl group the maximum number of O atoms is 1 , the maximum number of S atoms is 1 , and the maximum total number of O and S atoms is 2. Exemplary heteroaryl groups include furanyl, thienyl, oxazolyl, isoxazolyl, oxadiazolyl (1 ,2,5- and 1 ,2,3-), pyrrolyl, imidazolyl, pyrazolyl, triazolyl (1 ,2,3- and 1 ,2,4-), tetrazolyl, thiazolyl, isothiazolyl, thiadiazolyl (1 ,2,3- and 1 ,2,5-), pyridyl, pyrimidinyl, pyrazinyl, triazinyl (1 ,2,3-, 1 ,2,4-, and 1 ,3,5-), benzofuranyl (1- and 2-), indolyl, isoindolyl, benzothienyl (1- and 2-), 1 H-indazolyl, benzimidazolyl, benzoxazolyl, indoxazinyl, benzisoxazolyl, benzothiazolyl, benzisothiazolyl, benzotriazolyl, quinolinyl, isoquinolinyl, benzodiazinyl, quinoxalinyl, quinazolinyl, benzotriazinyl (1 ,2,3- and 1 ,2,4- benzotriazinyl), pyridazinyl, phenoxazinyl, thiazolopyridinyl, pyrrolothiazolyl, phenothiazinyl, isobenzofuranyl, chromenyl, xanthenyl, phenoxathiinyl, pyrrolizinyl, indolizinyl, indazolyl, purinyl, quinolizinyl, phthalazinyl, naphthyridinyl (1 ,5-, 1 ,6-, 1 ,7-, 1 ,8-, and 2,6-), cinnolinyl, pteridinyl, carbazolyl, phenanthridinyl, acridinyl, perimidinyl, phenanthrolinyl (1 ,7-, 1 ,8-, 1 ,10-, 3,8-, and 4,7- ), phenazinyl, oxazolopyridinyl, isoxazolopyridinyl, pyrrolooxazolyl, and pyrrolopyrrolyl. Exemplary 5- or 6-memered heteroaryl groups include furanyl, thienyl, oxazolyl, isoxazolyl, oxadiazolyl (1 ,2,5- and 1 ,2,3-), pyrrolyl, imidazolyl, pyrazolyl, triazolyl (1 ,2,3- and 1 ,2,4-), thiazolyl, isothiazolyl, thiadiazolyl (1 ,2,3- and 1 ,2,5-), pyridyl, pyrimidinyl, pyrazinyl, triazinyl (1 ,2,3-, 1 ,2,4-, and 1 ,3,5-), and pyridazinyl. In some embodiments the heteroaryl is unsubstituted. In some embodiments the heteroaryl is substituted.
As used herein and throughout the entire description, the terms "arylalkyi" and "heteroarylalkyi" are meant to include those radicals in which an aryl group and heteroaryl group, respectively, is attached to an alkyl group (e.g., benzyl, phenethyl, pyridylmethyl and the like) including those alkyl groups in which a carbon atom (e.g., a methylene group) has been replaced by, for example, an oxygen atom (e.g., phenoxymethyl, 2-pyridyloxymethyl, 3-(1-naphthyloxy)propyl, and the like). Preferably the Arylalkyl is a substituted or unsubstituted (C6-Ci0)aryl(Ci-C6)alkyl. Preferably the Heteroarylalkyl is a substituted or unsubstituted
Figure imgf000011_0001
In some embodiments the alkyl chain is a linear. In some embodiments the alkyl chain is branched. In some embodiments the alkyl chain is substituted. In some embodiments the alkyl chain is unsubstituted. In some embodiments the alkyl chain is linear and substituted or unsubstituted. In some embodiments the alkyl chain is branched and substituted or unsubstituted. In some embodiments the arylalkyl is unsubstituted. In some embodiments the arylalkyl is substituted. In some embodiments the heteroarylalkyl is unsubstituted. In some embodiments the heteroarylalkyl is substituted.
As used herein and throughout the entire description, the term "cycloalkyi" or "cycloaliphatic" represents cyclic non-aromatic versions of "alkyl" and "alkenyl" with preferably 3 to 14 carbon atoms, such as 3 to 10 carbon atoms, i.e., 3, 4, 5, 6, 7, 8, 9, or 10 carbon atoms, more preferably 3 to 8 carbon atoms, even more preferably 3 to 7 carbon atoms. Exemplary cycloalkyi groups include cyclopropyl, cyclopropenyl, cyclobutyl, cyclobutenyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cycloheptyl, cycloheptenyl, cyclooctyl, cyclooctenyl, cyclononyl, cyclononenyl, cylcodecyl, cylcodecenyl, and adamantyl. The term "cycloalkyi" is also meant to include bicyclic and tricyclic versions thereof. If bicyclic rings are formed it is preferred that the respective rings are connected to each other at two adjacent carbon atoms, however, alternatively the two rings are connected via the same carbon atom, i.e., they form a spiro ring system or they form "bridged" ring systems. Preferred examples of cycloalkyi include C3-C8- cycloalkyl, in particular cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, spiro[3,3]heptyl, spiro[3,4]octyl, spiro[4,3]octyl, bicyclo[4.1.0]heptyl, bicyclo[3.2.0]heptyl, bicyclo[2.2.1]heptyl, bicyclo[2.2.2]octyl, bicyclo[5.1.0]octyl, and bicyclo[4.2.0]octyl. In some embodiments the cycloalkyi is unsubstituted. In some embodiments the cycloalkyi is substituted. As used herein and throughout the entire description, the term "cyclopropylene" means a cyclopropyl group as defined above in which one hydrogen atom has been removed resulting in a diradical. The cyclopropylene may link two atoms or moieties via the same carbon atom (1 ,1 - cyclopropylene, i.e., a geminal diradical) or via two carbon atoms (1 ,2-cyclopropylene).
As used herein and throughout the entire description, the term "heterocyclyl" or "heterocyclic ring" or "heterocycle" means a cycloalkyi group as defined above in which from 1 , 2, 3, or 4 carbon atoms in the cycloalkyi group are replaced by heteroatoms of O, S, or N. Preferably, in each ring of the heterocyclyl group the maximum number of O atoms is 1 , the maximum number of S atoms is 1 , and the maximum total number of O and S atoms is 2. The term "heterocyclyl" is also meant to encompass partially or completely hydrogenated forms (such as dihydro, tetrahydro or perhydro forms) of the above-mentioned heteroaryl groups. Exemplary heterocyclyl groups include morpholino, isochromanyl, chromanyl, pyrrolidinyl, imidazolidinyl, pyrazolidinyl, piperidinyl, piperazinyl, indolinyl, isoindolinyl, di- and tetrahydrofuranyl, di- and tetrahydrothienyl, di- and tetrahydrooxazolyl, di- and tetrahydroisoxazolyl, di- and tetrahydrooxadiazolyl (1 ,2,5- and
1 ,2,3-), dihydropyrrolyl, dihydroimidazolyl, dihydropyrazolyl, di- and tetrahydrotriazolyl (1 ,2,3- and 1 ,2,4-), di- and tetrahydrothiazolyl, di- and tetrahydrothiazolyl, di- and tetrahydrothiadiazolyl
(1 ,2,3- and 1 ,2,5-), di- and tetrahydropyridyl, di- and tetrahydropyrimidinyl, di- and tetrahydropyrazinyl, di- and tetrahydrotriazinyl (1 ,2,3-, 1 ,2,4-, and 1 ,3,5-), di- and tetrahydrobenzofuranyl (1 - and 2-), di- and tetrahydroindolyl, di- and tetrahydroisoindolyl, di- and tetrahydrobenzothienyl (1 - and 2), di- and tetrahydro-1 H-indazolyl, di- and tetrahydrobenzimidazolyl, di- and tetrahydrobenzoxazolyl, di- and tetrahydroindoxazinyl, di- and tetrahydrobenzisoxazolyl, di- and tetrahydrobenzothiazolyl, di- and tetrahydrobenzisothiazolyl, di- and tetrahydrobenzotriazolyl, di- and tetrahydroquinolinyl, di- and tetrahydroisoquinolinyl, di- and tetrahydrobenzodiazinyl, di- and tetrahydroquinoxalinyl, di- and tetrahydroquinazolinyl, di- and tetrahydrobenzotriazinyl (1 ,2,3- and 1 ,2,4-), di- and tetrahydropyridazinyl, di- and tetrahydrophenoxazinyl, di- and tetrahydrothiazolopyridinyl (such as 4,5,6-7- tetrahydro[1 ,3]thiazolo[5,4-c]pyridinyl or 4,5,6-7-tetrahydro[1 ,3]thiazolo[4,5-c]pyridinyl, e.g.,
4,5,6-7-tetrahydro[1 ,3]thiazolo[5,4-c]pyridin-2-yl or 4,5,6-7-tetrahydro[1 ,3]thiazolo[4,5-c]pyridin-2- yl), di- and tetrahydropyrrolothiazolyl (such as 5,6-dihydro-4H-pyrrolo[3,4-d][1 ,3]thiazolyl), di- and tetrahydrophenothiazinyl, di- and tetrahydroisobenzofuranyl, di- and tetrahydrochromenyl, di- and tetrahydroxanthenyl, di- and tetrahydrophenoxathiinyl, di- and tetrahydropyrrolizinyl, di- and tetrahydroindolizinyl, di- and tetrahydroindazolyl, di- and tetrahydropurinyl, di- and tetrahydroquinolizinyl, di- and tetrahydrophthalazinyl, di- and tetrahydronaphthyridinyl (1 ,5-, 1 ,6-,
1 ,7-, 1 ,8-, and 2,6-), di- and tetrahydrocinnolinyl, di- and tetrahydropteridinyl, di- and tetrahydrocarbazolyl, di- and tetrahydrophenanthridinyl, di- and tetrahydroacridinyl, di- and tetrahydroperimidinyl, di- and tetrahydrophenanthrolinyl (1 ,7-, 1 ,8-, 1 ,10-, 3,8-, and 4,7-), di- and tetrahydrophenazinyl, di- and tetrahydrooxazolopyridinyl, di- and tetrahydroisoxazolopyridinyl, di- and tetrahydropyrrolooxazolyl, and di- and tetrahydropyrrolopyrrolyl. Exemplary 5- or 6-memered heterocyclyl groups include morpholino, pyrrolidinyl, imidazolidinyl, pyrazolidinyl, piperidinyl, piperazinyl, di- and tetrahydrofuranyl, di- and tetrahydrothienyl, di- and tetrahydrooxazolyl, di- and tetrahydroisoxazolyl, di- and tetrahydrooxadiazolyl (1 ,2,5- and 1 ,2,3-), dihydropyrrolyl, dihydroimidazolyl, dihydropyrazolyl, di- and tetrahydrotriazolyl (1 ,2,3- and 1 ,2,4-), di- and tetrahydrothiazolyl, di- and tetrahydroisothiazolyl, di- and tetrahydrothiadiazolyl (1 ,2,3- and
1 ,2,5-), di- and tetrahydropyridyl, di- and tetrahydropyrimidinyl, di- and tetrahydropyrazinyl, dill and tetrahydrotriazinyl (1 ,2,3-, 1 ,2,4-, and 1 ,3,5-), and di- and tetrahydropyridazinyl. In some embodiments the heterocyclyl is unsubstituted. In some embodiments the heterocyclyl is substituted.
As used herein and throughout the entire description, the term "halogen" or "halo" means fluoro, chloro, bromo, or iodo.
As used herein and throughout the entire description, the term "azido" means N3.
As used herein and throughout the entire description, the term "optionally substituted" or "substituted" indicates that one or more (such as 1 to the maximum number of hydrogen atoms bound to a group, e.g., 1 , 2, 3, 4, 5, 6, 7, 8, 9, or up to 10, such as between 1 to 5, 1 to 4, or 1 to 3, or 1 or 2) hydrogen atom(s) may be replaced with a group different from hydrogen such as alkyl (preferably, Ci-6 alkyl), alkenyl (preferably, C2-6 alkenyl), alkynyl (preferably, C2-6 alkynyl), aryl (preferably, 3- to 14-membered aryl), heteroaryl (preferably, 3- to 14-membered heteroaryl), cycloalkyi (preferably, 3- to 14-membered cycloalkyi), heterocyclyl (preferably, 3- to 14- membered heterocyclyl), halogen, -CN, azido, -N02, -OR71, -N(R72)(R73), -ON(R72)(R73),
-N+(-0 )(R72)(R73),-S(0)o-2R71 , -S(O)0-2OR7 , -OS(O)0-2R71 , -OS(O)0-2OR7 , -S(O)0-2N(R72)(R73),
-OS(O)0-2N(R72)(R73), -N(R71)S(0)o-2R71, -NR71S(O)0-2OR71, -NR7 S(O)0-2N(R72)(R73), -C(=W1)R71, -C(=W )W R71, -W C(=W )R71, and -W C(=W )W R71;
wherein R71, R72, and R73 are independently selected from the group consisting of -H, C1-6 alkyl, C2-6 alkenyl, C2-6 alkynyl, 3- to 7-membered cycloalkyi, 5- or 6-membered aryl, 5- or 6-membered heteroaryl, and 3- to 7-membered heterocyclyl, wherein each of the alkyl, alkenyl, alkynyl, cycloalkyi, aryl, heteroaryl, and heterocyclyl groups is optionally substituted with one, two or three substituents selected from the group consisting of C1-3 alkyl, halogen, -CF3, -CN, azido, -N02, -OH, -0(C1-3 alkyl), -S(C1-3 alkyl), -NH2, -NH(C1-3 alkyl), -N(C1-3 alkyl)2,
-NHS(0)2(C1-3 alkyl), -S(0)2NH2-z(C1-3 alkyl)z, -C(=0)OH, -C(=0)0(C1-3 alkyl), -C(=0)NH2-z(C1-3 alkyl)z, -NHC(=0)(C1-3 alkyl), -NHC(=NH)NHz-2(C1-3 alkyl)z, and -N(C1-3 alkyl)C(=NH)NH2-z(C1-3 alkyl)z, wherein z is 0, 1 , or 2 and C1-3 alkyl is methyl, ethyl, propyl or isopropyl; W is independently selected from O, S, and NR84, wherein R84 is -H or C1-3 alkyl.
In a first aspect the present invention relates to a compound for use in the treatment of a mycobacterial disease, said compound having a structure according to Formula I
Figure imgf000014_0001
Figure imgf000014_0002
X is a nitrogen containing heterocycle;
Y is CH2, O or NR ; n is an integer between 0 and 3;
R and R2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (CTC6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-C10)aryl, substituted or unsubstituted
Figure imgf000014_0003
substituted or unsubstituted (C3-C10)heteroaryl, and substituted or unsubstituted (C3-C10)heteroaryl(CTC6)alkyl; or a pharmaceutically acceptable salt, solvate or hydrate thereof;
with the proviso that when n = 0, Y is not O or NR1.
As used herein and throughout the entire description, the terms "mycobacterial disease" or "mycobacterial infection" are used interchangeably and refer to any pathological state, including any asymptomatic, acute or chronic mycobacterial infection and any state caused by or associated with such mycobacterial infection. In some embodiments the mycobacterial infection is an infection caused by by bacteria of the Mycobacterium tuberculosis Complex, including M. tuberculosis, M. africanum, M. bovis, M. caprae etc. In some embodiments the mycobacterial infection is an infection caused by Mycobacterium tuberculosis. In some embodiments the mycobacterial infection is multidrug-resistant tuberculosis (MDR-TB). In some embodiments, the mycobacterial disease is extensively drug-resistant tuberculosis (XDR-TB). Further specific embodiments defining the mycobacterial disease are described herein below.
As used herein and throughout the entire description, the term "pharmaceutically acceptable salt" refers to a salt that retains the desired biological activity of the parent compound and does not impart any undesired toxicological effects (see e.g., Berge, S. M., et al. (1977) J. Pharm. Sci. 66: 1 -19). Examples of such salts include acid addition salts and base addition salts. Acid addition salts include those derived from nontoxic inorganic acids, such as hydrochloric, nitric, phosphoric, sulfuric, hydrobromic, hydroiodic, phosphorous and the like, as well as from nontoxic organic acids such as aliphatic mono- and dicarboxylic acids, phenyl-substituted alkanoic acids, hydroxy alkanoic acids, aromatic acids, aliphatic and aromatic sulfonic acids and the like. Base addition salts include those derived from alkaline earth metals, such as sodium, potassium, magnesium, calcium and the like, as well as from nontoxic organic amines, such as Ν,Ν'- dibenzylethylenediamine, N-methylglucamine, chloroprocaine, choline, diethanolamine, ethylenediamine, procaine and the like. As used herein and throughout the entire description, the term "pharmaceutically acceptable" may in particular mean approved by a regulatory agency or other generally recognized pharmacopoeia for use in animals, and more particularly in humans.
As used herein and throughout the entire description, the term "solvate" as used herein refers to an addition complex of a dissolved material in a solvent (such as an organic solvent (e.g., an aliphatic alcohol (such as methanol, ethanol, n-propanol, isopropanol), acetone, acetonitrile, ether, and the like), water or a mixture of two or more of these liquids), wherein the addition complex exists in the form of a crystal or mixed crystal. The amount of solvent contained in the addition complex may be stoichiometric or non-stoichiometric. A "hydrate" is a solvate wherein the solvent is water. In another aspect the present invention relates to a compound for use in medicine, said compound having a structure according to Formula I
Figure imgf000016_0001
wherein
A is selected from the group consisting of:
Figure imgf000016_0002
X is a nitrogen containing heterocycle; Y is CH2, O or NR ; n is an integer between 0 and 3; R and R2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-Ci0)aryl, substituted or unsubstituted (C6-Ci0)aryl(Ci-C6)alkyl, substituted or unsubstituted (C3-Ci0)heteroaryl, and substituted or unsubstituted (C3-Ci0)heteroaryl(Ci-C6)alkyl; or a pharmaceutically acceptable salt, solvate or hydrate thereof; with the proviso that
(i) when n = 0, Y is not O or NR1;
(ii) when n = 2, Y is not CH2; or
(iii) the compound is not
Figure imgf000017_0001
ln some embodiments, the present invention relates to a compound for use in medicine, said compound having a structure according to Formula I
Figure imgf000017_0002
wherein
A is selected from the group consisting of:
Figure imgf000018_0001
X is a nitrogen containing heterocycle; Y is CH2, O or NR ; n is an integer between 0 and 3;
R1 and R2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-Cio)aryl, substituted or unsubstituted (C6-Cio)aryl(CrC6)alkyl, substituted or unsubstituted (C3-Ci0)heteroaryl, and substituted or unsubstituted (C3-Ci0)heteroaryl(Ci-C6)alkyl; or a pharmaceutically acceptable salt, solvate or hydrate thereof; with the proviso that
(i) when n = 0, Y is not O or NR1;
(ii) when n = 2, Y is not CH2; or (iii) the compound is not
Figure imgf000018_0002
Figure imgf000019_0001
heterocycle.
In another aspect the present invention relates to a pharmaceutical composition for use in the treatment of a mycobacterial disease, wherein said composition comprises a compound having a structure according to Formula I
Figure imgf000019_0002
wherein A is selected from the group consisting of:
Figure imgf000020_0001
X is a nitrogen containing heterocycle; Y is CH2, O or NR ; n is an integer between 0 and 3;
R1 and R2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (CTC6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-Ci0)aryl, substituted or unsubstituted (C6-Cio)aryl(CrC6)alkyl, substituted or unsubstituted (C3-Ci0)heteroaryl, and substituted or unsubstituted (C3-C10)heteroaryl(Ci-C6)alkyl; or a pharmaceutically acceptable salt, solvate or hydrate thereof;
with the proviso that when n = 0, Y is not O or NR1.
In the following embodiments the compound for use in the treatment of a mycobacterial disease as disclosed herein above, the compound for use in medicine as disclosed herein above and the compound of the composition for use in the treatment of a mycobacterial disease as disclosed herein above are further defined.
In some embodiments the compounds for use or the compound of the pharmaceutical composition for use havin according to Formula I, as erein, are
characterized in that A is
Figure imgf000020_0002
In some embodiments A is . In some bodiments A is I Inn some embodiments A is In some
embodiments A is In some embodiments A is In some
embodiments A is
Figure imgf000021_0001
ssoommee eemmbbooddiimmeennttss AA iiss .. In some
Figure imgf000021_0002
embodiments A is ^ ^ . In some embodiments A is
In some embodiments the compounds for use or the compound of the pharmaceutical composition for use, as disclosed herein, are characterized by having a structure according Formula (la)
Figure imgf000021_0003
wherein X, Y, n and R1 are defined as above and wherein the provisos are defined as above for the compounds for use or the compound of the pharmaceutical composition for use of the invention.
In some embodiments the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I or Formula la, as disclosed herein, are characterized in that X is
Figure imgf000022_0001
wherein
Z is CR3R4, O or NR5;
R3 and R4 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (CTC6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-Ci0)aryl, substituted or unsubstituted (C6-Cio)aryl(CrC6)alkyl, substituted or unsubstituted (C3-Ci0)heteroaryl, substituted or unsubstituted (C3-Cio)heteroaryl(Ci-C6)alkyl, halogen, -CN, -N02, -OR11 , -N(R12)(R13), -N(R11)(OR11), -S(O)0-2R11 , -S(0)1-2OR11 , -OS(0)1-2R11, - OS(0)1-2OR11 , -S(0)1-2N(R12)(R13), -OS(0)1-2N(R12)(R13), -N(R1 )S(0)1-2R1 1 , -NR11S(0)1-2OR1 1 , -N R S(0)1-2N(R 2)(R13), -C(=W)R11, -C(=W)WR11 , -WC(=W)R11, and -WC(=W)WR11 ,
preferably hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2- C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, halogen, -CN, -N02, -OR11 , -N(R 2)(R13), -N(R )(OR11), -S(O)0-2R1 1 , -S(0)1-2OR11 , -S(0)1-2N(R 2)( R 3), -N(R1 )S(0)1-2R11 , -NR 1S(0)1-2OR1 1, -NR1 S(0)1-2N(R 2)(R13), -C(=W)R11 , -C(=W)WR11 , -W C(=W)R11 , and -WC(=W)WR11 , more preferably hydrogen, substituted or unsubstituted (Cr C6)alkyl, substituted or unsubstituted (C2-C6)alkynyl and -OR11 ;
R5 is selected from the group consisting of hydrogen, substituted or unsubstituted (CTC6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-Ci0)aryl, substituted or unsubstituted (C6-C10)aryl(CrC6)alkyl, substituted or unsubstituted (C3- Cio)heteroaryl, substituted or unsubstituted (C3-Ci0)heteroaryl(Ci-
C6)alkyl, -N(R12)(R13), -N(R1 1)(OR11), -S(O)0-2R11 , -S(0)1-2OR11 , -N(R11)S(0)1-2R1 1 , -NR1 1S(0)1-20 R11, -NR11S(0)1-2N(R12)(R13), -C(=W)R11 , -C(=W)WR11, -WC(=W)R11 , and -WC(=W)WR11 , preferably hydrogen, substituted or unsubstituted (CTC6)alkyl, substituted or unsubstituted (C2- C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3- C8)cycloalkyl, substituted or unsubstituted (C6-C10)aryl, substituted or unsubstituted (C6-
Figure imgf000022_0002
substituted or unsubstituted (C3-C10)heteroaryl, substituted or unsubstituted (C3-Cio)heteroaryl(C C6)alkyl and -S(O)0-2R1 1 ; W is independently selected from O, S, and N(R14), preferably O and N(R14); R is, in each case, selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, and heterocyclyl, preferably -H, alkyl, and cycloalkyl;
R12 and R13 are, in each case, independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, and heterocyclyl, or R 2 and R 3 may join together with the nitrogen atom to which they are attached to form the group -N=CR15R16;
R14 is independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, heterocyclyl, and -OR11;
R 5 and R 6 are independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, and heterocyclyl. In some embodiments Z is CR3R4. In some embodiments Z is O or NR5. In some embodiments Z is O. In some embodiments Z is NR5.
In some embodiments the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I or Formula la, as disclosed herein, are characterized in that Y is CH2 or O. In some embodiments Y is CH2. In some embodiments Y is O. In some embodiments Y is NR1.
In some embodiments the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I or Formula la, as disclosed herein, are characterized in that n is 0, 1 , 2 or 3, with the proviso that when n = 0, Y is not O or NR1 In some embodiments n is 1 , 2 or 3. In some embodiments n is 1. In some embodiments n is 2. In some embodiments n is 3.
In some embodiments the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I or Formula la, as disclosed herein, are characterized in that R1 and R2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C3-CB)cycloalkyl, substituted or unsubstituted (C6-C10)aryl, substituted or unsubstituted (C6-C10)aryl(CrC6)alkyl, substituted or unsubstituted (C3- Cio)heteroaryl, and substituted or unsubstituted (C3-Cio)heteroaryl(Ci-C6)alkyl. In some embodiments R and R2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-Ci0)aryl, substituted or unsubstituted (C6-Ci0)aryl(Ci-C6)alkyl, and substituted or unsubstituted (C3-C10)heteroaryl. In some embodiments R and R2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Cr C6)alkyl, substituted or unsubstituted (C3-C8)cycloalkyl and substituted or unsubstituted (C6- Cio)aryl. In some embodiments R and R2 are independently selected from the group consisting of hydrogen and substituted or unsubstituted (C C6)alkyl. In some embodiments R is hydrogen and R2 is alkyl. In some embodiments R is alkyl and R2 is hydrogen. In some embodiments R1 and R2 are alkyl. In some embodiments R and R2 are hydrogen.
In some embodiments the compounds for use or the compound of the pharmaceutical composition for use, as disclosed herein, are characterized by having a structure according to Formula II
Figure imgf000024_0001
wherein
Y, Z, W, R1, R3, R4, R5, R11, R12, R 3, R14, R 5, R 6 and n are defined as above for Formula I or Formula la;
with the proviso that when n = 0, Y is not O or NR1.
In some embodiments the compounds for use or the compound of the pharmaceutical composition for use, as disclosed herein, are characterized by having a structure according to Formula III
Figure imgf000025_0001
wherein
Z is O or CR3R4; R3 and R4 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (CTC6)alkyl, substituted or unsubstituted (C2-C6)alkynyl and -OR11;
R 1 is, in each case, selected from the group consisting of hydrogen and substituted or unsubstituted (Ci-C6)alkyl.
In some embodiments R3 and R4 are hydrogen or substituted or unsubstituted (C C6)alkyl, preferably methyl. In some embodiments at least one of R3 and R4 is hydrogen. In some embodiments Z is O. In some embodiments R3 and R4 are hydrogen. In some embodiments R3 is hydrogen and R4 is substituted or unsubstituted (Ci-C6)alkyl, preferably wherein R4 is tert- butyl. In some embodiments R3 is hydrogen and R4 is substituted or unsubstituted (C^- C6)alkynyl, preferably wherein R4 is ethynyl. In some embodiments R3 is hydrogen and R4 is -OR11, wherein R is hydrogen and substituted or unsubstituted (CrC6)alkyl. In some embodiments R3 is hydrogen and R4 is -OR11, wherein R11 is hydrogen. In some embodiments R3 is hydrogen and R4 is -OR11, wherein R11 is substituted or unsubstituted (Ci-C6)alkyl, preferably methyl.
In some embodiments the compounds for use or the compound of the pharmaceutical composition for use ha ure according to Formula I, as disclosed herein, are
characterized in that A is
Figure imgf000025_0002
; X is \ wherein Z is CH2 or O; Y is CH2 or O C
Figure imgf000026_0001
H2 and n is 2. In some IS wherein Z is O; Y is
O and n is 2.
In some embodiments the compounds for use or the compound of the pharmaceutical composition for use ha ormula I, as disclosed herein, are
characterized in that A is
Figure imgf000026_0002
; χ is wherein Z is CH2 or O; Y is CH2 or O
and n is 2. In some embodiments erein Z is CH2; Y is
CH2 and n is 2. In some embodime
Figure imgf000026_0003
nts A is is wherein Z is CH2; Y
is O and n is 2. In some embodiments A is
Figure imgf000026_0004
IS wherein Z is O; Y is CH2 and n is 2. In some embodiments A
Figure imgf000027_0001
is wherein Z is O; Y is O and n is 2.
In some embodiments the compounds for use or the compound of the pharmaceutical composition for use ormula I, as disclosed herein, are
characterized in that
Figure imgf000027_0002
CH2 or O; Y is CH2 or O
and n is 2. In some embodiments erein Z is CH2; Y is
CH2 and n is 2. In some embodime
Figure imgf000027_0003
nts A is ; X is wherein Z is CH2; Y
is O and n is 2. In some embodiments A is wherein Z is O; Y is
CH2 and n is 2. In some embodiments A is
Figure imgf000027_0004
; X is wherein Z is O; Y is O and n is 2.
In some embodiments the compounds for use or the compound of the pharmaceutical composition for use ha ormula I, as disclosed herein, are
characterized in that A is
Figure imgf000027_0005
wherein Z is CH2 or O; Y is CH2 or O and n is 2. In some embodiments A is erein Z is CH2; Y is
CH2 and n is 2. In some embodiments A
Figure imgf000028_0001
is ; X is wherein Z is CH2; Y
is O and n is 2. In some embodiments A is wherein Z is O; Y is
CH2 and n is 2. In some embodiments A is
Figure imgf000028_0002
; X is wherein Z is O; Y is O and n is 2.
In some embodiments the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, as disclosed herein, are
Figure imgf000028_0003
CH2 and n is 2. In some embodiments A is
Figure imgf000029_0001
; X is wherein Z is O; Y is O and n is 2.
In some embodiments the compounds for use or the compound of the pharmaceutical composition for use according to Formula I, as disclosed herein, are
characterized in that
Figure imgf000029_0002
X is wherein Z is CH2 or O; Y is CH2 or
O, 2 is hydrogen or substituted or unsubstituted (Ci-Ce)alkyl and n is 2. In some embodiments
A is
Figure imgf000029_0003
; X is wherein Z is CH2; Y is CH2, R2 is hydrogen or substituted
(Ci-C6)alkyl and n is 2. In some embodiments A is X is herein Z is CH2; Y is O, R i substituted or unsubstituted (C
Figure imgf000029_0004
C6)alkyl and n is 2. In some embodiments A is ; X is wherein Z is
O; Y is CH2, R2 is hydrogen or substituted or unsubstituted (Ci-C6)alkyl and n is 2. In some
embodiments A is
Figure imgf000029_0005
; X is wherein Z is O; Y is O, R2 is hydrogen or substituted or unsubstituted (CrC6)alkyl and n is 2.
In some embodiments the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, as disclosed herein, are characterized in that A is
Figure imgf000030_0001
wherein Z is CH2 or O; Y is CH2 or
O, R2 is hydrogen or substituted or unsubstituted (Ci-C6)alkyl and n is 2. In some embodiments
A IS
Figure imgf000030_0002
; X IS wherein Z is CH2; Y is CH2, R2 is hydrogen or substituted
(d-CeJalkyl and n is 2. In some embodiments A is X is herein Z is CH2; Y is O, R2 i substituted or unsubstituted (Ci-
Figure imgf000030_0003
C6)alkyl and n is 2. In some embodiments A is ; X is \ / wherein Z is
O; Y is CH2, R2 is hydrogen or substituted or unsubstituted (Ci-C6)alkyl and n is 2. In some
embodiments A is
Figure imgf000030_0004
; X is N / wherein Z is O; Y is O, R2 is hydrogen or substituted or unsubstituted (C C6)alkyl and n is 2.
In some embodiments the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, as disclosed herein, are
characterized in that A is
Figure imgf000030_0005
; X is wherein Z is CH2 or O; Y is CH2 or O,
R2 is hydrogen or substituted or unsubstituted (CTC6)alkyl and n is 2. In some embodiments A is
Figure imgf000031_0001
X is wherein Z is CH2; Y is CH2, R is hydrogen or substituted or
(CrC6)alkyl and n is 2. In some embodiments X is
Figure imgf000031_0002
wherein Z is CH2; Y is O, R is hydrogen or substituted or unsubstituted (Cr
C6)alkyl and n is 2. In some embodiments A is
Figure imgf000031_0003
■ χ is wherein Z is O;
Y is CH2, R2 is hydrogen or substituted or unsubstituted (CrCeJalkyl and n is 2. In some
embodiments A is
Figure imgf000031_0004
; X is wherein Z is O; Y is O, R2 is hydrogen or substituted or unsubstituted (CrCeJalkyl and n is 2.
In some embodiments the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, as disclosed herein, are
characterized in that A is
Figure imgf000031_0005
wherein Z is CH2 or O; Y is CH2 or O,
R2 is hydrogen or substituted or unsubstituted (C!-CeJalkyl and n is 2. In some embodiments A is
Figure imgf000032_0001
wherein Z is CH2; Y is CH2, R2 is hydrogen or substituted or
(CrC6)alkyl and n is 2. In some embodiments A is ; X is
Figure imgf000032_0002
wherein Z is CH2; Y is O, R is hydrogen or substituted or unsubstituted (Cr
C6)alkyl and n is 2. In some embodiments A is
Figure imgf000032_0003
wherein Z is O; Y is CH2, R2 is hydrogen or substituted or unsubstituted (CrCeJalkyl and n is 2. In some
embodiments A is
Figure imgf000032_0004
wherein Z is O; Y is O, R 2 is hydrogen or substituted or unsubstituted (CrCeJalkyl and n is 2.
An exemplary synthetic route for preparing a non-limiting embodiment of a compound according to Formula I, Formula la, Formula II and Formula III as provided herein is described below. It will be understood that this illustrative embodiment is provided to show a possible synthetic route for preparing a non-limiting embodiment of the present invention. It should also be understood that other synthetic routes are possible for preparing the compounds according to Formula I, Formula la, Formula II and Formula III provided herein, as will be known to the person of skill in the art. In this non-limiting embodiment, a possible synthetic route for preparing lalistat (La-0) having a piperidine group at positions 3 and 4 of the 1 ,2,5-thiadiazol core is provided. A synthetic route for preparing this derivative is shown in Figure 4 A. All other compounds encompassed by Formula I, Formula la, Formula II and Formula III can be prepared in an analogous manner. Starting from dichloride 1 , piperidine was condensed and the resulting monochloride 2 was converted to 4-(Piperidin-1-yl)-1 ,2,5-thiadiazol-3-ol (3). Compound 3 was reacted with piperidine- carbonyl chloride under basic conditions to yield lalistat (La-0) (4), according to the synthetic route of Rosenbaum et al. (Rosenbaum, A.I., et al., Thiadiazole carbamates: potent inhibitors of lysosomal acid lipase and potential Niemann-Pick type C disease therapeutics. J Med Chem, 2010. 53(14): p. 5281-9). Alternatively, compound 3 may be reacted with phosgene under basic conditions and the in-situ formed chloroformate treated with piperidine to yield lalistat (La-0) (4).
In some embodiments the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, Formula la, Formula II and Formula III, as disclosed herein, are useful for the treatment of a mycobacterial disease, wherein the mycobacterial disease is caused by at least one bacteria selected from the group consisting of Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae, Mycobacterium microti, Mycobacterium leprae, Mycobacterium lepromatosis, Mycobacterium avium, Mycobacterium silvaticum, Mycobacterium hominissuis, Mycobacterium paratuberculosis, Mycobacterium kansasii, Mycobacterium xenopi, Mycobacterium simiae, Mycobacterium abcessus, Mycobacterium fortuitum, Mycobacterium chelonae, Mycobacterium ulcerans, Mycobacterium marinum and/or Mycobacterium fortuitum, preferably Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae, Mycobacterium microti, Mycobacterium leprae, Mycobacterium lepromatosis and/or Mycobacterium kansasii, more preferably Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae, Mycobacterium microti and/or Mycobacterium kansasii, even more preferably Mycobacterium tuberculosis.
Mycobacterial diseases are caused by mycobacteria. Tuberculosis and leprosy (Hansen's disease) are the best known mycobacterial diseases. However, people may also be infected by any of a group of mycobacterial species collectively called non-tuberculous mycobacteria. While tuberculosis and leprosy are most common in resource-limited countries, non-tuberculous mycobacterial infections occur worldwide.
Tuberculosis is caused by bacteria of the Mycobacterium tuberculosis Complex. The Mycobacterium tuberculosis Complex includes bacteria selected from the group consisting of Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae and Mycobacterium microti. In some embodiments the mycobacterial disease is caused by at least one bacteria of the Mycobacterium tuberculosis Complex. In some embodiments the mycobacterial disease is caused by Mycobacterium tuberculosis. In some embodiments the mycobacterial disease is caused by Mycobacterium africanum. In some embodiments the mycobacterial disease is caused by Mycobacterium bovis. In some embodiments the mycobacterial disease is caused by Mycobacterium caprae. In some embodiments the mycobacterial disease is caused by Mycobacterium microti. In some embodiments the mycobacterial disease is tuberculosis and/or caused by bacteria of the group consisting of Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae, Mycobacterium microti, preferably Mycobacterium tuberculosis.
Leprosy (also known as Hansen's disease) is caused by Mycobacterium leprae. In some embodiments the mycobacterial disease is caused by Mycobacterium leprae. The mycobacterial disease may be also caused by non-tuberculous mycobacteria (NTM). As used herein and throughout the entire description, the term "non-tuberculous mycobacteria" means mycobacteria which do not cause tuberculosis or leprosy. In children, NTM cause lymphadenitis, skin and soft tissue infections, and occasionally also lung disease and disseminated infections. Manifestations can be indistinguishable from tuberculosis on the basis of clinical and radiological findings and tuberculin skin testing. Although over 150 different species of NTM have been described, pulmonary infections are most commonly due to Mycobacterium avium complex (MAC), Mycobacterium kansasii, and Mycobacterium abscessus. In some embodiments the non-tuberculosis bacteria are selected from a group consisting of Mycobacterium avium, Mycobacterium silvaticum, Mycobacterium hominissuis, Mycobacterium paratuberculosis, Mycobacterium kansasii, Mycobacterium xenopi, Mycobacterium simiae, Mycobacterium abcessus, Mycobacterium fortuitum, Mycobacterium chelonae, Mycobacterium ulcerans, Mycobacterium marinum, Mycobacterium gordonae and/or Mycobacterium fortuitum.
The Mycobacterium avium complex (MAC) includes bacteria selected from the group consisting of Mycobacterium avium, Mycobacterium silvaticum, Mycobacterium hominissuis and Mycobacterium paratuberculosis. In some embodiments the mycobacterial disease is caused by non-tuberculous mycobacteria. In some embodiments the mycobacterial disease is caused by at least one bacteria of the Mycobacterium avium complex (MAC). In some embodiments the mycobacterial disease is caused by Mycobacterium avium. In some embodiments the mycobacterial disease is caused by Mycobacterium silvaticum. In some embodiments the mycobacterial disease is caused by Mycobacterium hominissuis. In some embodiments the mycobacterial disease is caused by Mycobacterium paratuberculosis. In some embodiments the mycobacterial disease is caused by Mycobacterium kansasii. Mycobacterium kansasii causes chronic pulmonary infection that resembles pulmonary tuberculosis. However, it may also infect other organs. M kansasii infection is the second-most- common nontuberculous opportunistic mycobacterial infection associated with HIV/AIDS. In some embodiments the mycobacterial disease is caused by Mycobacterium xenopi. In some embodiments the mycobacterial disease is caused by Mycobacterium simiae. In some embodiments the mycobacterial disease is caused by Mycobacterium abcessus. In some embodiments the mycobacterial disease is caused by Mycobacterium fortuitum, Mycobacterium chelonae. In some embodiments the mycobacterial disease is caused by Mycobacterium ulcerans. In some embodiments the mycobacterial disease is caused by Mycobacterium marinum. In some embodiments the mycobacterial disease is caused by Mycobacterium fortuitum. In some embodiments the mycobacterial disease is caused by Mycobacterium gordonae.
In some embodiments the mycobacterial disease is a pulmonary infection caused by non- tuberculous mycobacteria (NTM), preferably a chronic pulmonary infection. In some embodiments the mycobacterial disease is a skin and/or soft tissue infection caused by non- tuberculous mycobacteria (NTM). In some embodiments the mycobacterial disease is a lung disease caused by non-tuberculous mycobacteria (NTM).
In some embodiments the mycobacterial disease is selected from tuberculosis, leprosy (Hansen's disease), lepromatosis infections caused by non-tuberculosis mycobacteria including lymphadenitis and pulmonary infections, skin infections caused by mycobacteria including Buruli ulcer and fish tank granuloma. In some embodiments the mycobacterial disease is tuberculosis. In some embodiments the mycobacterial disease is leprosy (Hansen's disease). In some embodiments the mycobacterial disease is lepromatosis. In some embodiments the mycobacterial disease is a skin infection caused by mycobacteria including Buruli ulcer and fish tank granuloma. In some embodiments the mycobacterial disease is multidrug-resistant tuberculosis (MDR-TB). In some embodiments the mycobacterial disease is extensively drug- resistant tuberculosis (XDR-TB).
The compounds of Formula I, Formula la, Formula II and Formula III and pharmaceutically- acceptable salts, solvates and hydrates thereof may be used on their own but will generally be administered in the form of a pharmaceutical composition in which the formula (I) or (Γ) compound/salt/solvate (active ingredient) is in association with a pharmaceutically-acceptable adjuvant, diluent or carrier.
Depending on the mode of administration, the pharmaceutical composition will preferably comprise from 0.05 to 99 % w (per cent by weight), more preferably from 0.10 to 70 % w, of active ingredient, and, from 1 to 99.95 % w, more preferably from 30 to 99.90 % w, of a pharmaceutically-acceptable adjuvant, diluent or carrier, all percentages by weight being based on total composition. The pharmaceutical composition may additionally contain an additional pharmaceutically active agent, such as an antibiotic, antifungal or anti-HIV compound and/or various other ingredients known in the art, for example, a lubricant, stabilising agent, buffering agent, emulsifying agent, viscosity regulating agent, surfactant, preservative, flavouring or colorant.
The compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, Formula la, Formula II and Formula III are in particular useful as bactericidal agents and/or bacteriostatic agents. The compounds are also non-hazardous for bacteria of gram-positive bacterial strains other than mycobacterial strains and non-hazardous for gram-negative bacterial strains, in particular they do not inhibit the growth of gram-negative bacterial strains and gram-positive bacterial stains other than mycobacterial strain. In some embodiments the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, Formula la, Formula II and Formula III, as disclosed herein, are characterized in that they are bacteriostatic, preferably bacteriostatic for mycobacteria. In some embodiments the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, Formula la, Formula II and Formula III, as disclosed herein, are characterized in that they are bactericidal, preferably bactericidal for mycobacteria. In some embodiments the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, Formula la, Formula II and Formula III, as disclosed herein, are characterized in that they are non-hazardous for bacteria of gram-positive bacterial strains other than mycobacterial strains and non-hazardous for gram-negative bacterial strains, in particular the growth of gram-negative bacterial strains and gram-positive bacterial stains other than mycobacterial strain is not inhibited. In some embodiments the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, Formula la, Formula II and Formula III, as disclosed herein, are characterized in that they are non-hazardous for bacteria of gram-positive bacterial strains other than mycobacterial strains, in particular the growth of gram- positive bacterial stains other than mycobacterial strain is not inhibited. In some embodiments the compounds for use or the compound of the pharmaceutical composition for use having a structure according to Formula I, Formula la, Formula II and Formula III, as disclosed herein, are characterized in that they are non-hazardous for gram-negative bacterial strains, in particular the growth of gram-negative bacterial strains is not inhibited. In some embodiments, the pharmaceutical composition for use as disclosed herein, further comprises at least one pharmaceutically acceptable carrier. In some embodiments, the compounds according to Formula I, Formula la, Formula II and Formula III or a pharmaceutically acceptable salt, solvate or hydrate thereof may be included in a pharmaceutically acceptable carrier. As used herein and throughout the entire description, the terms "carrier" and "excipient" are used interchangeably herein. Pharmaceutically acceptable carriers or excipients include diluents (fillers, bulking agents, e.g. lactose, microcrystalline cellulose), disintegrants (e.g. sodium starch glycolate, croscarmellose sodium), binders (e.g. PVP, HPMC), lubricants (e.g. magnesium stearate), glidants (e.g. colloidal Si02), solvents/co-solvents (e.g. aqueous vehicle, Propylene glycol, glycerol), buffering agents (e.g. citrate, gluconates, lactates), preservatives (e.g. Na benzoate, parabens (Me, Pr and Bu), BKC), anti-oxidants (e.g. BHT, BHA, Ascorbic acid), wetting agents (e.g. polysorbates, sorbitan esters), anti-foaming agents (e.g. Simethicone), thickening agents (e.g. methylcellulose or hydroxyethylcellulose), sweetening agents (e.g. sorbitol, saccharin, aspartame, acesulfame), flavoring agents (e.g. peppermint, lemon oils, butterscotch, etc), humectants (e.g. propylene, glycol, glycerol, sorbitol). The person skilled in the art will readily be able to choose suitable pharmaceutically acceptable carriers or excipients, depending, e.g., on the formulation and administration route of the pharmaceutical composition.
A non-exhaustive list of exemplary pharmaceutically acceptable carriers or excipients includes (biodegradable) liposomes; microspheres made of the biodegradable polymer poly(D,L)-lactic- coglycolic acid (PLGA), albumin microspheres; synthetic polymers (soluble); nanofibers, protein- DNA complexes; protein conjugates; erythrocytes; or virosomes. Various carrier based dosage forms comprise solid lipid nanoparticles (SLNs), polymeric nanoparticles, ceramic nanoparticles, hydrogel nanoparticles, copolymerized peptide nanoparticles, nanocrystals and nanosuspensions, nanocrystals, nanotubes and nanowires, functionalized nanocarriers, nanospheres, nanocapsules, liposomes, lipid emulsions, lipid microtubules/microcylinders, lipid microbubbles, lipospheres, lipopolyplexes, inverse lipid micelles, dendrimers, ethosomes, multicomposite ultrathin capsules, aquasomes, pharmacosomes, colloidosomes, niosomes, discomes, proniosomes, microspheres, microemulsions and polymeric micelles. Other suitable pharmaceutically acceptable excipients are inter alia described in Remington's Pharmaceutical Sciences, 15th Ed., Mack Publishing Co., New Jersey (1991 ) and Bauer et al., Pharmazeutische Technologie, 5th Ed., Govi-Verlag Frankfurt (1997).
The pharmaceutical composition of the invention will generally be designed for specific routes and methods of administration, for specific dosages and frequencies of administration, for specific treatments of specific diseases, with ranges of bio-availability and persistence, among other things. The materials of the composition are preferably formulated in concentrations that are acceptable for the site of administration.
Formulations and compositions thus may be designed in accordance with the invention for delivery by any suitable route of administration. In the context of the present invention, the routes of administration include
• topical routes (such as epicutaneous, inhalational, nasal, opthalmic, auricular / aural, vaginal, mucosal);
• enteral routes (such as oral, gastrointestinal, sublingual, sublabial, buccal, rectal); and · parenteral routes (such as intravenous, intraarterial, intraosseous, intramuscular, intracerebral, intracerebroventricular, epidural, intrathecal, subcutaneous, intraperitoneal, extra-amniotic, intraarticular, intracardiac, intradermal, intralesional, intrauterine, intravesical, intravitreal, transdermal, intranasal, transmucosal, intrasynovial, intraluminal). In some embodiments the administration may be a parenteral route, in particular intravenous or intramuscular.
In some embodiments, the pharmaceutical composition, as disclosed herein, is administered to a subject in need thereof in an amount effective to treat said mycobacterial disease. The subject is preferably a mammal. The subject is more preferably a human subject. The mycobacterial disease can be any mycobacterial disease disclosed herein above and below.
As used herein and throughout the entire description, the term "Subject" means animals, including warm blooded mammals such as humans and primates; avians; domestic household or farm animals such as cats, dogs, sheep, goats, cattle, horses and pigs; laboratory animals such as mice, rats and guinea pigs; fish; reptiles; zoo and wild animals; and the like. The subject is preferably a mammal, more preferably a human. As used herein and throughout the entire description, the term "amount effective" in the context of a composition or dosage form for administration to a subject refers to an amount of the composition or dosage form sufficient to provide a benefit in the treatment of mycobacterial disease, to delay or minimize symptoms associated with mycobacterial infection or mycobacterial-induced disease, or to cure or ameliorate the disease or infection or cause thereof. In particular, a therapeutically effective amount means an amount sufficient to provide a therapeutic benefit in vivo. Used in connection with an amount of a compound of the invention, the term preferably encompasses a non-toxic amount that improves overall therapy, reduces or avoids symptoms or causes of disease, or enhances the therapeutic efficacy of or synergies with another therapeutic agent.
Amounts effective will depend, of course, on the particular subject being treated; the severity of a condition, disease or disorder; the individual patient parameters including age, physical condition, size and weight; the duration of the treatment; the nature of concurrent therapy (if any); the specific route of administration and like factors within the knowledge and expertise of the health practitioner. These factors are well known to those of ordinary skill in the art and can be addressed with no more than routine experimentation. It is generally preferred that a maximum dose be used, that is, the highest safe dose according to sound medical judgment. It will be understood by those of ordinary skill in the art, however, that a patient may insist upon a lower dose or tolerable dose for medical reasons, psychological reasons or for virtually any other reason.
In some embodiments, the pharmaceutical composition for use, as disclosed herein, is administered to a subject in need thereof in an amount effective to treat said mycobacterial disease, wherein said subject is treated with at least one additional pharmaceutically active compound, including an antibiotic, antifungal or anti-HIV compound. The additional pharmaceutically active compound is preferably an antibiotic, antifungal and/or anti-HIV compound. The additional pharmaceutically active compound is more preferably an antibiotic. The antibiotic is preferably an anti-tuberculosis drug or an agent or compound active against Mycobacterium tuberculosis.
Anti-tuberculosis (TB) drugs are classified into five groups based on evidence of efficacy, potency, drug class and experience of use. In the United States rifampicin is called rifampin. First-line anti-TB drugs (Group 1 ) are currently recommended in a four-drug combination for the treatment of drug-susceptible TB. Second-line anti-TB drugs (Groups 2, 3 and 4) are reserved for drug-resistant TB. Third-line anti-TB drugs (Group 5) have unclear efficacy or undefined roles.
In some embodiments the additional pharmaceutically active compound is selected from the group of First-line agents consisting of Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin and Rrifabutin.
In some embodiments the additional pharmaceutically active compound is selected from the group of Second-line agents consisting of Aminoglycosides including Kanamycin and Amikacin, Polypeptides including Capreomycin and Viomycin and Streptomycin. In some embodiments the additional pharmaceutically active compound is selected from the group of Second-line agents consisting of fluoroquinolones including Moxifloxacin, Levofloxacin, Ofloxacin and Gatifloxacin. In some embodiments the additional pharmaceutically active compound is selected from the group of a bacteriostatic Second-line agents consisting of Thioamides including Ethionamide and protionamide, Cycloserine, Terizidone, Thioacetone and p-Aminosalicylic acid.
In some embodiments the additional pharmaceutically active compound is selected from the group of Third-line agents consisting of Clofazimine, Linezolid, Amoxicillin/clavulanate, Thioacetazone, Imipenem/cilastatin, high-dose isoniazid and Clarithromycin.
In some embodiments the additional pharmaceutically active compound is selected from the group consisting of Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin, Rifabutin, Animoglycosides including Kanamycin and/or Amikacin, Polypetides including Capreomycin, Viomycin and/or Streptomycin, fluoroquinolones including Moxifloxacin, Levofloxacin, Ofloxacin and/or Gatifloxacin, thioamides including Ethionamide and/or Protionamide, Cycloserine, Terizidone, Thioacetone, p-Aminosalicylic acid, Clofazimine, Linezolid, Amoxicillin, Clavulanate, Thioacetazone, Imipenem, Cilastatin, Clarithromycin, Delamanid and/or Bedaquiline.
In some embodiments the additional pharmaceutically active compound is selected from the group consisting of Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin, Rifabutin, Animoglycosides including Kanamycin and/or Amikacin, Polypetides including Capreomycin, Viomycin and/or Streptomycin, fluoroquinolones including Moxifloxacin, Levofloxacin, Ofloxacin and/or Gatifloxacin, thioamides including Ethionamide and/or Protionamide, Cycloserine, Terizidone, Thioacetone and/or p-Aminosalicylic acid. In some embodiments the additional pharmaceutically active compound is selected from the group consisting of Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin and/or Rifabutin. In some embodiments the additional pharmaceutically active compound is selected from the group consisting of Isoniazid, Rifampicin, Ethambutol and/or Pyrazinamide. In some embodiments the additional pharmaceutically active compound is Isoniazid. In some embodiments the additional pharmaceutically active compound is Rifampicin. In some embodiments the additional pharmaceutically active compound is Ethambutol. In some embodiments the additional pharmaceutically active compound is Pyrazinamide. In some embodiments the additional pharmaceutically active compound is Rifabutin. In some embodiments the additional pharmaceutically active compound is Delamanid. In some embodiments the additional pharmaceutically active compound is Bedaquiline.
In some embodiments is the additional pharmceutically active compound an antifungal compound. In some embodiments is the antifungal compound selected from the group consisting of Allylamines including Terbinafin and/or Naftifin, Azole-Antimycotics including Bifonazole, Butoconazole, Clotrimazole, Econazole, Fenticonazole, Fluconazole, Isoconazole, Itraconazole, Ketoconazole, Miconazole, Oxiconazole, Posaconazole, Voriconazole, Efinaconazole, Luliconazole, Sertaconazole and/or Tioconazole, Benzylamines including Butenafine, Polyenes including Amphotericin B, Nystatin and/or Pentamycine, Morpholine- Derivates including Amorolfine, Hydroxypyridone derivates including Ciclopirox, Echinocandins including Anidulafungin, Caspofungin and/or Micafungin, Pyrimidines including Flucytosine, and Thiocarbamates including Tolnaftat, Oxaboroles including Tavaborole. In some embodiments the additional pharmaceutically active compound is Methylrosaniline. In some embodiments the additional pharmaceutically active compound is Griseofulvin.
In some embodiments is the additional pharmceutically active compound an anti-HIV compound. In some embodiments is the anti-HIV compound selected from the group consisting of Nucleoside/Nucleotide Reverse Transcriptase Inhibitors (NRTIs) including Abacavir, Atripla, Combivir, Complera, Didanosine, Emtriva, Entecavir, Epivir, Epzicom, Retrovir, Trizivir, Truvada, Videx, Videx EC, Viread, Zerit and/or Ziagen, Non-Nucleoside Reverse Transcriptase Inhibitors (NNRTIs) including Edurant.lntelence, Rescriptor, Sustiva, Viramune and/or Viramune XR, Protease Inhibitors (Pis) including Aptivus, Crixivan, Evotaz, Invirase, Kaletra, Lexiva, Norvir, Prezcobix, Prezista, Reyataz and/or Viracept, Entry/ Fusion Inhibitors including Fuzeon, Integrase Strand Transfer Inhibitors (INSTIs) including Isentress, Tivicay and/or Vitekta, Chemokine Co receptor Antagonists (CCR5 Antagonists) including Seizentry, Pharmacokinetic Enhancer including Cytochrome P4503A (CYP3A) Inhibitors including Tybost, and Immune- Based Therapeutics including Plaquenil. Another aspect of the present invention is a kit of parts comprising a pharmaceutical composition and at least one additional pharmaceutically active compound, wherein said pharmaceutical composition comprises a compound having a structure according to Formula I
Figure imgf000042_0001
wherein
A is selected from the group consisting of:
Figure imgf000042_0002
X is a nitrogen containing heterocycle;
Y is CH2, O or NR1; n is an integer between 0 and 3;
R1 and R2 are independently selected from the group consisting of hydrogen, substituted unsubstituted (CTC6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted unsubstituted (C6-Ci0)aryl, substituted or unsubstituted (C6-Ci0)aryl(Ci-C6)alkyl, substituted or unsubstituted (C3-C10)heteroaryl, and substituted or unsubstituted (Ca-C-ioJheteroaryliCi-CeJalkyl; or a pharmaceutically acceptable salt, solvate or hydrate thereof; with the proviso that when n = 0, Y is not O or NR1; and wherein said additional pharmaceutically active compound is selected from the group consisting of an antibiotic, antifungal or anti-HIV compound, preferably an antibiotic selected from the group of Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin, Rifabutin, Animoglycosides including Kanamycin and/or Amikacin, Polypetides including Capreomycin, Viomycin and/or Streptomycin, fluoroquinolones including Moxifloxacin, Levofloxacin, Ofloxacin and/or Gatifloxacin, thioamides including Ethionamide and/or Protionamide, Cycloserine, Terizidone, Thioacetone, p-Aminosalicylic acid, Clofazimine, Linezolid, Amoxicillin, Clavulanate, Thioacetazone, Imipenem, Cilastatin, Clarithromycin, Delamanid and/or Bedaquiline, preferably Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin, Rifabutin, Animoglycosides including Kanamycin and/or Amikacin, Polypetides including Capreomycin, Viomycin and/or Streptomycin, fluoroquinolones including Moxifloxacin, Levofloxacin, Ofloxacin and/or Gatifloxacin, thioamides including Ethionamide and/or Protionamide, Cycloserine, Terizidone, Thioacetone and/or p-Aminosalicylic acid, more preferably Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin and/or Rifabutin, even more preferably Isoniazid, Rifampicin, Ethambutol and/or Pyrazinamide. In some embodiments the compound of the pharmaceutical composition of the kit, is a compound according to Formula I, Formula la, Formula II and Formula III as disclosed herein, wherein X, Y, Z, W, R1, R2, R3, R4, R5, R11, R 2, R 3, R 4, R 5, R 6 and n are defined as herein above; with the proviso that when n = 0, Y is not O or NR1. In some embodiments is the kit of parts useful for the treatment of mycobacterial diseases, as disclosed herein above, in particular for the treatment of tuberculosis.
The present invention also envisions a method of treating in a subject a mycobacterial infection, in particular tuberculosis, comprising administering to said subject an efficient amount of a compound according to formula (I) or a pharmaceutically acceptable salt, solvate or hydrate thereof or a pharmaceutical composition comprising said compound. Said method preferably comprises further administering at least one additional pharmaceutically active compound, including an antibiotic, antifungal or anti-HIV compound. The above described aspects, embodiments, definitions, ect. are also applicable to said method of treatment, mutatis mutandis. Brief description of the drawings:
Figure 1 : Mycobacterium tuberculosis growth assay with GFP expression readout. Lalistat inhibits Mycobacterium tuberculosis growth at low micromolar concentrations. GFP: Green Fluorescent Protein, RLU: Relative light units.
Figure 2: Activity on human macrophages infected with Mycobacterium tuberculosis strain H37Rv. Lalistat inhibits Mycobacterium tuberculosis growth at low micromolar concentrations. CFU: Colony Forming Units, DMSO: Dimethylsulfoxid 0,1 %. Figure 3: Effect of 40 μΜ lalistat (La-0) on the growth of Staphylococcus aureus, Escherichia coli and Listeria monocytogenes.
Figure 4: A) Synthetic route to Lalistat (La-0) and lalistat probe (La-1 ). B) H NMR of lalistat probe (La-1 ). TFA: Trifluoroacetic acid, DIEA: Ν,Ν-Diisopropylethylamine, THF: Tetrahydrofurane, Boc: tert-butyloxycarbonyl.
Figure 5: A) Chemical structure of lalistat La-0 and lalistat probe La-1. B) Mycobacterium tuberculosis growth assay with GFP expression readout after 7 days. RLU: Relative light units. Figure 6: In vivo target identification / deconvolution via chemical proteomics: Activity based protein profiling (ABPP) with 50 μΜ lalistat probe La-1. DMSO: Dimethylsulfoxid.
Figure 7: Activity based protein profiling (ABPP) workflow including DMSO control and competition control experimental procedures. DMSO: Dimethylsulfoxid.
Figure 8: Alignment of selected target protein hits using the Universal Protein Resource (UniProt) platform (Reorganizing the protein space at the Universal Protein Resource (UniProt). Nucleic Acids Res, 2012. 40(Database issue): p. D71-5). P38571 is (SEQ ID No: 1 ), P95125 is (SEQ ID No: 2), P71668 is (SEQ ID No: 3) and P9WK85 is (SEQ ID No: 4).
Figure 9: Validation of protein target LipR identified via ABPP: LipR was overexpressed in E. coli and labelled with lalistat probe La-1. Rhodamine-Azide was attached via Click reaction after cell lysis. The experiment was conducted with and without induction by IPTG (Isopropyl- -D- thiogalactopyranosid). Fluorescence SDS-PAGE analysis and the corresponding coomassie stained gel are shown. ABPP: Activity Based Protein Profiling.
Figure 10: Competition ABPP experiment visualized via SDS-PAGE and fluorescent scanning.
Figure 11 : Proposed mechanism of target inhibition by Lalistat.
Further, the invention shall be explained in more detail by the following Examples.
Examples
1) Materials and methods
1.1) Synthesis of 4-(Piperidin-1-yl)-1 ,2,5-thiadiazol-3-yl 4-ethynylpiperidine-1-carboxylate (La-1) 4-(Piperidin-1-yl)-1 ,2,5-thiadiazol-3-ol (3) was prepared according to Rosenbaum et al. (Rosenbaum, A.I., et al., Thiadiazole carbamates: potent inhibitors of lysosomal acid lipase and potential Niemann-Pick type C disease therapeutics. J Med Chem, 2010. 53(14): p. 5281-9).
4-Ethynylpiperidine-TFA salt was prepared according to Braisted et al. (Braisted, A.C., et al., Discovery of a potent small molecule IL-2 inhibitor through fragment assembly. J Am Chem Soc, 2003. 125(13): p. 3714-5).
4-(Piperidin-1-yl)-1 ,2,5-thiadiazol-3-ol (76.7 mg, 0.41 mmol) (3) was dissolved in THF (10 mL). NaH (16.4 mg, 60% in mineral oil, 0.41 mmol) was added and the resulting suspension was stirred until recovery of a solution indicating complete deprotonation. A phosgene solution (234 μΙ_, 20% in toluene, 1.75 M, 0.41 mmol) was added and the resulting solution was stirred at RT for 10 min. 4-Ethynylpiperidine-TFA salt (144 mg, 0.70 mmol) (7) and DIPEA (120 μΙ_, 0.70 mmol) were combined in THF (2 mL) and stirred for 5 min. This solution was added to the in-situ formed chloroformate solution and stirred at RT for 20 min. The reaction was stopped by addition of 10 mL 1 M HCI and diluted with EtOAc. After separation of phases the organic layer was washed three times with saturated NaHC03 solution, dried over MgS04 and concentrated in-vacuo. The product was purified by RP-HPLC as the HCI salt to yield as a clear oil (Figure 4) RP-HPLC (analytical setup method: gradient 2% B→ 98% B over 10 min): tR = 8.73 min.
Reverse-phase HPLC analysis was performed on a Waters 2695 separation module, equipped with a Waters PDA 2996 and a Waters XBridge C18 column (3.5 mm, 4.6 x 100 mm, flow = 1.2 mL/min). For preparative scale RP-HPLC separation a Waters 2545 quarternary gradient module in combination with a Waters PDA 2998 and a Waters XBridge C18 (5.0 pm, 30 x 150 mm, flow = 50 mL/min) column or a YMC Triart C18 (3.5 pm, 10 x 250 mm, flow = 10 mL/min) column was used. The mobile phase for elution consisted of a gradient mixture of 0.1 % (v/v) TFA in water (buffer A, HPLC grade) and 0.1 % (v/v) TFA in ACN (buffer B, HPLC grade) unless noted otherwise. 1H-NMR (360 MHz, CDCI3): δ [ppm] = 3.89-3.73 (m, 2 H), 3.59-3.43 (m, 2 H), 3.42-3.35 (m, 4 H), 2.73 (tq, J = 3.7, 7.0 Hz, 1 H), 2.15 (d, J = 2.4 Hz, 1 H), 1.95-1.83 (m, 2 H), 1.78-1.68 (m, 2 H), 1.67-1.57 (m, 6 H).
13C-N R (91 MHz, CDCI3): δ [ppm] = 153.8, 150.9, 146.6, 85.5, 70.5, 49.1 , 43.2, 42.8, 31.4, 20.9, 26.4, 25.5, 24.3. HRMS (ESI): calc. for C15H25N202 [M+H]+: 321.1380; found: 321.1372.
1.2) Mycobacterium tuberculosis Growth Analysis
GFP-expressing Mycobacterium tuberculosis H37Rv (Michelucci, A., et al., Immune-responsive gene 1 protein links metabolism to immunity by catalyzing itaconic acid production). Proc Natl Acad Sci U S A, 2013. 110(19): p. 7820-5) were generated using the plasmid 32362:pMN437 (Addgene), kindly provided by M. Niederweis (University of Alabama, Birmingham, AL) (Song, H., et al., Identification of outer membrane proteins of Mycobacterium tuberculosis. Tuberculosis (Edinb), 2008. 88(6): p. 526-44). 1x106 bacteria were cultured in 7H9 medium supplemented with oleic acid-albumin-dextrose-catalase (OADC) (10%), Tween 80 (0.05%), and glycerol (0.2%) in a total volume of 100 μΙ in a black 96 well plate with clear bottom (Corning Inc, Corning, NY) sealed with an air-permeable membrane (Porvair Sciences, Dunn Labortechnik, Asbach, Germany). Growth was as measured as RLU: Relative light units, at 528 nm after excitation at 485 nm in a fluorescence microplate reader (Synergy 2, Biotek, Winooski, VT) at indicated time points. 1.3) Analysis of M. tuberculosis growth in human macrophages
Mononuclear cells were isolated from peripheral blood (PBMC) of healthy volunteers by density gradient centrifugation. Monocytes were separated (purity consistently >95%) by counterflow elutriation. Human monocyte-derived Macrophages (hMDM) were generated in the presence of 10 ng/ml recombinant human macrophage colony-stimulating factor (M-CSF) from highly purified monocytes as described (Reiling, N., et al., Mycobacteria-induced TNF-alpha and IL-10 formation by human macrophages is differentially regulated at the level of mitogen-activated protein kinase activity. J Immunol, 2001. 167(6): p. 3339-45). M. tuberculosis growth in human macrophages was analyzed as described (Reiling, N., et al., Clade-specific virulence patterns of Mycobacterium tuberculosis complex strains in human primary macrophages and aerogenically infected mice. MBio, 2013. 4(4)). In brief 2x105 hMDMs were cultured in 500 μΙ RPMI 1640 with 10% FCS and 4mM L-glutamine in 48-well flat-bottom microtiter plates (Nunc) at 37°C in a humidified atmosphere containing 5% C02. Macrophages were infected with M. tuberculosis strain H37Rv with a multiplicity of infection (MOI) of 1 :1. Four hours post infection, non- phagocytosed bacteria were removed by washing three times with 0.5 ml Hanks' balanced salt solution (HBSS; Invitrogen) at 37°C. After washing and after 3 days of cultivation, 0.5 ml media was added to the macrophage culture. At day 7 supernatants were completely removed and macrophage cultures were lysed at 4 hours and 7 days post infection by adding 10 μΙ 10% Saponin solution (Sigma) in HBSS at 37°C for 15 min. Lysates were serially diluted in sterile water containing 0.05% Tween 80 (Merck, Darmstadt, Germany) and plated twice on 7H10 agar containing 0.5% glycerol (Serva) and 10% heat-inactivated bovine calf serum (BioWest, France). After 3 weeks at 37°C the colony forming units (CFUs) were counted.
1.4) Activity based protein profiling (ABPP) A general scheme describing activity based protein profiling (ABPP) experiments is shown in figure 7. M. tuberculosis cells were grown to stationary phase and incubated with La-1 or DMSO as a control. After cell lysis a rhodamine tag was attached to the alkyne moiety of La-1 via bioorthogonal click-reaction (R.Huisgen, Proc Chem Soc, 1961 , 357.; V. V. Rostovtsev, et al., Angew Chem Int Ed Engl, 2002, 41 , 2596.; C. W. Tornoe, C. Christensen and M. Meldal, J Org Chem, 2002, 67, 3057.) to visualize target proteins on SDS-PAGE gel by fluorescence scanning. Different probe concentrations were applied and the intensities of target bands evaluated (Figure 10). A concentration between 30 μΜ and 60 μΜ was sufficient to achieve saturated labelling. The inventors thus proceeded with gel-free proteomics and performed labeling studies with 30 μΜ La-1 in M. tuberculosis H37Rv to unravel the identities of target proteins. For these studies a biotin tag served as a handle to enrich labeled proteins on avidin beads (Figure 7). After on- bead digestion primary amines were modified by heavy, medium and light isotopes using dimethyl labeling (DL; J. L. Hsu, et al., Anal Chem, 2003, 75, 6843.). Labelling ratios of probe versus DMSO were normalized, z-score and log2(x) transformed. Target proteins with a high enrichment and statistical significance are located in the upper right area of the enrichment volcano plot in figure 6. To validate these enriched hits, the inventors performed analytical competition experiments where cells were pre-incubated with La-0 (60 μΜ) and subsequently treated with 30 μΜ La-1 probe (Figure 7).
The detailed ABPP procedure is described below. Bacterial culture was derived from frozen stock (2.5x108 bacteria/ml). Homogenous bacterial suspension was prepared in 7H9 medium (50 mL) supplemented with oleic acid-albumin-dextrose-catalase (OADC) (10%), Tween 80 (0.05%), and glycerol (0.2%). 25 mL each was incubated in 30 mL square medium bottles (Nalgene) at 37°C without shaking for three days. Preculture was diluted to 450 mL and incubated for four days. Bacteria were washed with PBS and an optical density at 600 nm of 40 was adjusted. For regular ABPP experiments (probe / DMSO control) 1 mL suspension was supplemented with 30 μΜ lalistat-1 (La-1 ; lalistat probe) or DMSO as a control and incubated for 30 min, vortexed and incubated another 30 min. For competition experiments (probe / competition control) 1 mL suspension was supplemented with 60 μΜ lalistat (La-0) or DMSO control and incubated for 30 min, vortexed and incubated for another 30 min. Both samples were now additionally supplemented with 30 μΜ lalistat-1 (La-1 ; lalistat probe), incubated for 30 min, vortexed and incubated for another 30 min. For both regular and competition experiments bacteria were washed with PBS and the pellet was stored at -80°C over night. Pellet was suspended in 1 mL PBS supplemented with 80 μί protease inhibitor (stock: 1 tablet solved in 2 ml ddH20) (two times distilled water). Samples were sonicated (duty cycle: 50; output 10 (100%)) at 4°C for 20 min each, centrifuged (15,000 xg, 4°C, 30 min) and the supernatant was centrifuged over 0.22 μιη Spin-X Centrifuge tube filter (Costar) (15,000 xg, 4°C, 15 min). Samples were stored at -80°C. Two identical samples each derived from 1 mL OD60o=40 cultures were combined and treated with 120 μί gel-free ABPP Mix (40 μί Biotin-PEG3-N3 (Jena Bioscience, CLK-AZ104P4-100; 10 mM in DMSO), 20 μί fresh TCEP (50 mM in ddH20), 60 μί TBTA Ligand (1.667 mM in 80 % tBuOH and 20 % DMSO)). The final concentrations were 233 μΜ Biotin-PEG3-N3, 581 μΜ TCEP and 58.2 μΜ TBTA Ligand. The lysates were mixed by vortexing and 20 μί CuS0 solution (50 rriM in ddH20) (two times distilled water) were added. The lysates were mixed by vortexing again and incubated for 1 h at RT in the dark. After the click-reaction the lysates were transferred to 15 mL falcon tubes and 8 mL of cold acetone (-80°C, MS grade) were added. Proteins were precipitated ON at -80°C. The precipitated proteins were thawed on ice, pelletized (16,900 xg, 15 min, 4°C) and supernatant was disposed. Falcon tubes were stored on ice during the following washing procedure: The proteins were washed two times with 1 mL cold methanol (-80°C). Resuspension was achieved by sonication (15 sec at 10 % intensity) and proteins were pelletized via centrifugation (16,900 xg, 10 min, 4°C). Only MS grade water was used for the following procedures. After two washing steps supernatant was disposed and the pellet was resuspended in 500 pL 0.2 % SDS in PBS at RT by sonication (15 sec at 10 % intensity). Avidin beads were thawed on ice and resuspended by carefully inverting. Then 50 pL of bead suspension were transferred into Protein LoBind Eppendorf tubes using wide bore pipette tips and washed three times with 1 mL 0.2 % SDS in PBS (resuspension: carefully inverting 10 times, pelleting: 400 xG, 3 min, RT). 500 pL protein solution from the 15 mL falcon tubes were transferred to the Protein LoBind Eppendorf tubes with washed avidin beads and incubated under continuous inverting (20 rpm, 1 h, RT). Beads were washed 3 times with 1 mL 0.2 % SDS in PBS, 2 times with 1 mL 6 M urea in water and 3 times with 1 mL PBS (resuspension: carefully inverting 20 times, pelleting: 400 xg, 3 min, RT).
The beads were resuspended in 200 μΙ denaturation buffer (7 M urea, 2 M thiourea in 20 mM pH
7.5 HEPES buffer). Proteins were reduced through addition of dithiothreitol (DTT, 1 M, 0.2 pL), the tubes were mixed by vortexing shortly and incubated in a thermomixer (450 rpm, 45 min,
RT). Then 2-iodoacetamide (IAA, 550 mM, 2 pL) was added for alkylation, the tubes were mixed by vortexing shortly and incubated in a thermomixer (450 rpm, 30 min, RT, in the dark).
Remaining IAA was quenched by the addition of dithiothreitol (DTT, 1 M, 0.8 pL). The tubes were shortly mixed by vortexing and incubated in a thermomixer (450 rpm, 30 min, RT). LysC
(0.5 pg/pL, Wako) was thawed on ice and 1 pL was added to each microcentrifuge tube, the tubes were shortly mixed by vortexing and incubated in a thermomixer (450 rpm, 2 h, RT, in the dark). TEAB solution (600 pL, 50 mM in water) and then trypsin (1.5 pL, 0.5 pg/pL in 50 mM acetic acid, Promega) were added and tubes were shortly vortexed after each addition. The reaction was incubated in a thermomixer (450 rpm, 13-15 h, 37 °C). The digest was stopped by adding 6 pL formic acid (FA) and vortexing. After centrifugation (100 xg, 1 min, RT), the supernatant was transferred to a new Protein LoBind Eppendorf tube. FA (50 pL, aqueous 0.1 % solution) was added to the beads and after vortexing and centrifugation (100 xg, 1 min, RT) the supernatant was added to the supernatant collected before. Again FA (50 μΙ_, aqueous 0.1 % solution) was added to the beads and after vortexing and centrifugation (16,200 xg, 3 min, RT) the supernatant was transferred to the combined supernatants.
50 mg SepPak C18 columns (Waters) were equilibrated by gravity flow with 1 ml_ acetonitrile, 1 mL elution buffer (80% ACN, 0.5% FA) and 3 mL aqueous 0.5% FA solution. Subsequently the samples were loaded by gravity flow, washed with 5 mL aqueous 0.5% FA solution and labeled with 5 mL of the respective dimethyl labeling solution. The following solutions were used: light (L): 30 mM NaBH3CN, 0.2 % CH20, 10 mM NaH2P04, 35 mM Na2HP04, pH 7.5; medium (M): 30 mM NaBH3CN, 0.2 % CD20, 10 mM NaH2P04> 35 mM Na2HP04, pH 7.5; heavy (H): 30 mM NaBHD3CN, 0.2 % 13CD20, 10 mM NaH2P04, 35 mM Na2HP04, pH 7.5. Labeled peptides were eluted into new 2.0 mL Protein LoBind Eppendorf tubes using two times 250 pL elution buffer. The eluates were lyophilized and stored at -20°C.
For analytical gel-based experiments (figure 10) no avidin bead enrichment was performed. After the click reaction samples were analyzed via SDS-PAGE. Instead of 120 pL gel-free ABPP Mix, 100 pL gel-based ABPP Mix was used: (20 pL RhN3 (Tetramethylrhodamine (TAMRA) Azide (Tetramethylrhodamine 5- Carboxamido-(6-Azidohexanyl)), 5-isomer (life technologies, T10182); 5 mM in DMSO), 20 pL fresh TCEP (50 mM in ddH20), 60 pL TBTA Ligand (1.667 mM in 80% tBuOH and 20% DMSO)).
Before MS measurement the samples from gel-free experiments were dissolved in 30 pL 1 % FA by pipetting up and down, vortexing and sonication for 15 min (brief centrifugation after each step). Differentially labeled samples were mixed. 0.45 pm centrifugal filter units (VWR) were equilibrated with two times 500 pL water, 500 pL 0.05 N NaOH and two times 500 pL 1 % FA (centrifugation: 16,200 xg, 1 min, RT). Reconstituted and mixed peptide samples were filtered through the equilibrated filters (centrifugation: 16,200 xg, 2 min, RT). Samples were analyzed via HPLC-MS/MS using an UltiMate 3000 nano HPLC system (Dionex, Sunnyvale, California, USA) equipped with Acclaim C18 PepMap100 75 pm ID x 2 cm trap and Acclaim C18 PepMap RSLC, 75 pM ID x 15 cm separation columns coupled to an Orbitrap Fusion mass spectrometer (Thermo Fisher Scientific Inc., Waltham, Massachusetts, USA). Samples were loaded on the trap and washed for 10 min with 0.1 % formic acid, then transferred to the analytical column and separated using a 120 min gradient from 3% to 25% acetonitrile in 0.1 % formic acid and 5% dimethyl sulfoxide (at 200 nL/min flow rate). Orbitrap Fusion was operated in a 3 second top speed data dependent mode. Full scan acquisition was performed in the orbitrap at a resolution of 120000 and an ion target of 4E5 in a scan range of 300 - 1700 m/z. Monoisotopic precursor selection as well as dynamic exclusion for 60 s were enabled. Precursors with charge states of 2 - 7 and intensities greater than 5E3 were selected for fragmentation. Isolation was performed in the quadrupole using a window of 1.6 m/z. Precursors were collected to a target of 1 E2 for a maximum injection time of 250 with "inject ions for all available parallelizable time" enabled. Fragments were generated using higher-energy collisional dissociation (HCD) and detected in the ion trap at a rapid scan rate. Internal calibration was performed using the ion signal of fluoranthene cations (EASY-ETD/IC source)
Peptide and protein identifications were performed using MaxQuant 1.5.3.8 software with Andromeda as search engine using following parameters: Carbamidomethylation of cysteines as fixed and oxidation of methionine as well as acetylation of N-termini as dynamic modifications, trypsin/P as the proteolytic enzyme, 4.5 ppm for precursor mass tolerance (main search ppm) and 0.5 Da for fragment mass tolerance (ITMS MS/MS tolerance). Searches were done against the Uniprot database for M. tuberculosis H37Rv (taxon identifier: 83332, downloaded on 19.5.2015). Quantification was performed using dimethyl labeling with the following settings: light: DimethLysO, DimethNterO; medium: Dimethl_ys4, DimethNter4 and heavy: Dimethl_ys8, DimethNter8. Variable modifications were included for quantification. The I = L and re-quantify options were used. Identification was done with at least 2 unique peptides and quantification only with unique peptides.
Statistical analysis was performed with Perseus 1.5.1.6. Putative contaminants, reverse peptides and peptides only identified by site were omitted from further processing. Dimethyl labeling ratios were log2(x) transformed and z-score normalized. The average values of technical replicates were calculated and -log-io(p-values) were obtained by a two sided one sample t-test over six biological replicates for standard ABPP with DMSO control or 4 biological replicates for competition experiments. Proteins were ranked from highest to lowest log2(x) transformed and z-score normalized dimethyllabeling ratios. They were also ranked from highest to lowest -log10(x) transformed p- values. Proteins were finally ranked according to the sum of the ranking values from dimethyllabeling ratios and -log-io(p-value) across both experiments (regular ABPP: probe / DMSO, competition experiment: probe / competition). 2% of the identified proteins with the highest final ranking (including regular ABPP and competition experiments dimethyllabeling ratios and p-values) were considered to be hits of lalistat. This cut-off was chosen as by this analysis a visual separation of enriched vs. not enriched proteins in both regular ABPP and competition experiments could be achieved. 1.5) Recombinant expression and labelling of proteins in E. coli
A N-terminal His6 affinity tagged LipR construct was PCR amplified from genomic DNA from M. tuberculosis H37Rv using the primers shown below and cloned in a pDONR201 (Invitrogen) vector and then in a pET300 expression vector via the GATEWAY cloning system. Expression was induced at an OD600 of 0.6 by addition of lsopropyl-3-D-thiogalactopyranosid (IPTG; final concentration: 0.25 mM) and carried out 4 h at 37°C in E. coli BL21 cells.
Construct: N-His6-attB1 -TEV-LipR-Stop-attB2
Primer 1 : ggggacaagtttgtacaaaaaagcaggctttgagaatctttattttcagggcAACCTGCGCAAAAACGTCATCC (SEQ ID No: 5)
Primer 2: ggggaccactttgtacaagaaagctgggtgTCATTTGACTACTCCCCGTGG (SEQ ID No: 6)
After centrifugation (5 min, 6,200 xg, 4°C) and removal of the supernatant bacteria were resuspended in PBS to get an OD60o of 40. To 1 ml_ of this suspension in a microcentrifuge tube 30 μΙ_ of La-1 solution in DMSO (or just DMSO as a control) were added. After 30 min incubation at RT in the dark the microcentrifuge tube was again mixed by vortexing and incubated for another 30 min at RT in the dark. After centrifugation (6,200 xg, 2 min, 4°C) the supernatant was removed and the pellets were stored at -80°C.
Pellets were resuspended in 1 ml_ PBS (4°C) and transferred to a 'Precellys Glass/Ceramic Kit SK38 2.0 ml.' tube. Tubes were cooled on ice for about 5 min or longer and cells were lysed with the Precellys Homogeniser using two times lysis program 3 (5,400 rpm, run number: 1 , run time: 20 sec, pause: 5 sec). After each lysis run the tubes were cooled on ice for 5 min. The ball mill tubes were centrifuged (16,200 xg, 10 min, 4°C) and 86 μΙ_ of supernatant were transferred to new 1.5 ml. microcentrifuge tubes and treated with 10 μΙ_ gel-based ABPP Mix (2 μΙ_ RhN3 (Tetramethylrhodamine (TAMRA) Azide (Tetramethylrhodamine 5-Carboxamido-(6- Azidohexanyl)), 5-isomer (life technologies, T10182); 5 mM in DMSO), 2 μΙ_ fresh TCEP (50 mM in ddH20), 6 μΙ_ TBTA Ligand (1.667 mM in 80% tBuOH and 20% DMSO)). The final concentrations were: 104 μΜ RhN3, 1.04 mM TCEP and 104 μΜ TBTA Ligand. The lysates were mixed by vortexing and 2 μΙ_ CuS04 solution (50 mM in ddH20) were added. The lysates were again mixed by vortexing and incubated for 1 h at RT in the dark. Then 80 μΙ_ 2x Laemmli Sample Buffer were added, samples were mixed in a thermomixer (300 rpm, 3 min, 96°C) and analyzed via SDS PAGE (10 % agarose gel (PEQLAB Biotechnologie GmbH, Erlangen, PerfectBlue Dual Gel System, the gel was prepared according to the manual), 3.5 h, 300 V, 8 μΙ_ fluorescent protein standard) and fluorescence imaging (GE Healthcare, ImageQuant LAS- 4000). After fluorescence scanning the gel was Coomassie stained.
2) Results
2.1) Lalistat impairs Mycobacterium tuberculosis growth
The effect of lalistat on M. tuberculosis growth was analysed by two independent approaches. First, growth of GFP expressing bacteria was monitored for 7 days in presence of lalistat or the control antibiotic Rifampicin as described by Michelucci et al. (Michelucci, A., et al., Immune- responsive gene 1 protein links metabolism to immunity by catalyzing itaconic acid production. Proc Natl Acad Sci U S A, 2013. 110(19): p. 7820-5). Interestingly, dose dependent growth inhibition with an EC50 of around 4 μΜ was observed for lalistat (Figure 1 ). Second, the effect of lalistat on intracellular growth of M. tuberculosis in human macrophage host cells was determined. To this end the inventors infected human monocyte derived macrophages with strain M. tuberculosis H37Rv (multiplicity of infection 1 :1 ) for 4h and subsequently cultured the cells for 7 days in the absence or presence of lalistat. Cells were then lysed and CFUs were determined as described previously (Reiling, N., et al., Clade-specific virulence patterns of Mycobacterium tuberculosis complex strains in human primary macrophages and aerogenically infected mice. MBio, 2013. 4(4)). Again lalistat substantially reduced bacterial load by 55% compared to the untreated control (Figure 2). In contrast, no growth inhibition was observed in other bacterial strains including gram-positive Staphylococcus aureus, and Listeria monocytogenes as well as gram-negative Escherichia coli (Figure 3) suggesting a mode of action specific for Mycobacteria.
2.2) Mode of action analysis: Protein target identification and validation via ABPP
To analyse the mode of action the inventors identified lalistat protein targets in live bacterial cells via activity based protein profiling (ABPP). For this technology a chemical probe bearing an alkyne tag was synthesized following established protocols, as descried under 1.1 ) (Figure 4A).
The lalistat-derived probe (La-1 ) retained activity and reduced M. tuberculosis growth comparable to the parent molecule (La-0) (Figure 5A and 5B). Next, target identification experiments were conducted. Bacterial cells were grown to stationary phase and incubated with lalistat probe (La-1 ) or DMSO as a control. After cell lysis a biotin or rhodamine tag was attached to the alkyne tag of La-1 via bioorthogonal click-reaction (Huisgen, R., 1,3-Dipolar Cycloadditions. Proc. Chem. Soc, 1961 : p. 357-96; Rostovtsev, V.V., et al., A stepwise huisgen cycloaddition process: copper(l)-catalyzed regioselective "ligation" of azides and terminal alkynes. Angew Chem Int Ed Engl, 2002. 41 (14): p. 2596-9; Tornoe, C.W., C. Christensen, and M. Meldal, Peptidotriazoles on solid phase: [1,2,3]-triazoles by regiospecific copper(i)-catalyzed 1 ,3-dipolar cycloadditions of terminal alkynes to azides. J Org Chem, 2002. 67(9): p. 3057-64). While the rhodamine tag was used to visualize target proteins on a SDS-PAGE gel by fluorescence scanning, the biotin tag served as a handle to enrich target proteins with avidin beads. After on-bead digestion primary amines were modified by heavy, medium and light isotopes using a dimethyl labeling. Labeling ratios of probe (La-1 ) versus DMSO were normalized, z-score and log2(x) transformed. Target proteins with a high enrichment and a high statistical significance can be found in the upper right area of the volcano plots in Figure 6. To verify these targets for lalistat binding, the inventors performed competition experiments where cells were pre-incubated with La-0 and subsequently treated with La-1. Strikingly the combined target spectrum includes 7 proteins from the Lip family among other lypolytic / hydrolytic enzymes. A proposed inhibition mechanism for those enzymes is depicted in figure 1 1. An alignment of the amino acid sequences of the three most enriched Lip proteins (LipN, Lipl and LipR) together with the known human lalistat target lysosomal acid lipase LipA reveals the active site consensus sequence: GDSAGGXI/L (SEQ ID No: 7) or GXSXGXXI/L (SEQ ID No: 8) (Figure 8). In addition, the inventors were able to validate one of the target proteins (LipR) by a heterologous expression labeling experiment in E. coli (Figure 9). Here M. tuberculosis LipR was overexpressed in the host and could be labeled also in this strain using chemical tool compound La-1 in combination with rhodamine azide and SDS-PAGE fluorescence visualization.
3) Discussion
Among the enzymes identified in this study that are targeted by lalistat, LipN showed the strongest drug binding characteristics. LipN, similar to Lipl, which was the second highest hit among the identified members of the lip family, possesses esterase activity on short chain TGs. In contrast, like many members of the mycobacterial HSL-family, LipR shows no TG hydrolysing activity but rather acts on other short- and mid chain substrates (Delorme, V. et al. MmPPOX Inhibits Mycobacterium tuberculosis Lipolytic Enzymes Belonging to the Hormone-Sensitive Lipase Family and Alters Mycobacterial Growth. PLoS One 7, (2012)). Underlining the important function of lipases from the lip family in mycobacterial infection, Lipl and LipG have both been shown to be essential for mycobacterial growth in vitro and in macrophages, respectively(Rengarajan, J., Bloom, B. R. & Rubin, E. J. Genome-wide requirements for Mycobacterium tuberculosis adaptation and survival in macrophages. Proc. Natl. Acad. Sci. U. S. A. 102, 8327-8332 (2005), Griffin, J. E. et al. High-resolution phenotypic profiling defines genes essential for mycobacterial growth and cholesterol catabolism. PLoS Pathog. 7, (201 1 )), while LipO is indispensable for prevention of endosomal maturation and acidification (Pethe, K. et al. Isolation of Mycobacterium tuberculosis mutants defective in the arrest of phagosome maturation. Proc. Natl. Acad. Sci. U. S. A. 101, 13642-7 (2004)). Furthermore, lipases belonging to the HSL family have been reported to be up-regulated during starvation after TG accumulation (Danelishvili, L, Poort, M. J. & Bermudez, L. E. Identification of Mycobacterium avium genes up-regulated in cultured macrophages and in mice. FEMS Microbiol. Lett. 239, 41- 49 (2004), Fisher, M. a, Plikaytis, B. B. & Shinnick, T. M. Microarray analysis of the Mycobacterium tuberculosis transcriptional response to the acidic conditions found in phagosomes. J Bacteriol 184, 4025-32 (2002), Betts, J. C, Lukey, P. T., Robb, L. C, McAdam, R. A. & Duncan, K. Evaluation of a nutrient starvation model of Mycobacterium tuberculosis persistence by gene and protein expression profiling. Mol. Microbiol. 43, 717-731 (2002)), emphasizing their important role during dormancy. However, it must be taken in account that to date it is not clear whether mycobacterial members of the lip family like LipR hydrolysing a wide range of short-chain substrates also participate in metabolic processes like signalling, membrane support and regulation due to a still unknown additional phospholipase, thioesterase or protease activity (Delorme, V. et al. MmPPOX Inhibits Mycobacterium tuberculosis Lipolytic Enzymes Belonging to the Hormone-Sensitive Lipase Family and Alters Mycobacterial Growth. PLoS One 7, (2012)). It can therefore not be excluded that other metabolic pathways distinct from neutral lipid hydrolysis contribute to mycobycterial growth inhibition mediated by lalistat targeting distinct lip activities. In this context it is surprising that among all targets characterized no LipY could be detected . In contrast to other members of the lip family, LipY is the only member reported to hydrolyse long chain TG and, thus, is regarded as the only "real" lipase. From the pattern of the targets identified by these investigations it can be concluded that lalistat preferentially binds to lipolytic enzymes, which show a wider substrate range including those possessing shorter acyl chains or monoacylglycerides compared to TG-hydrolysing lipases.
The second highest hit of our lalistat target profiling, Rv0183, is an among the mycobacterial species highly conserved monoalcylglycerol lipase and, similar to as described for mycobacterial lipases belonging to the HSL family, has been proposed as an important anti-mycobacterial target in persistent TB (Saravanan, P., Dubey, V. K. & Patra, S. Potential Selective Inhibitors against Rv0183 of Mycobacterium tuberculosis Targeting Host Lipid Metabolism. Chem. Biol. Drug Des. 79, 1056-1062 (2012)). It has been identified in chronic but not early stages of disease when investigated in the guinea pig model of tuberculosis, emphasizing its hypothetic role in persistant mycobacterial infection (Kruh, N. A., Troudt, J., Izzo, A., Prenni, J. & Dobos, K. M. Portrait of a pathogen: The Mycobacterium tuberculosis proteome in vivo. PLoS One 5, (2010)).
Rv1984 represents the only enzyme among the first 10 identified lalistat-targets, which belongs to the cutinase-family. It is located in the bacterial cell wall and is proposed to be involved in exogenous host cell lipid hydrolysis (Dhouib, R., Laval, F., Carriere, F., Daffe, M. & Canaan, S. A monoacylglycerol lipase from Mycobacterium smegmatis involved in bacterial cell interaction. J. Bacteriol. 192, 4776-4785 (2010)). Its lypolytic activity has been reported to be preferentially directed against medium-chain carboxylic esters and monoacylglycerols underlining its potential role in mycobacterial pathogenicity during persistence. Since in contrast to lipases, cutinases are serine esterases active on a wider panel of substrates including cutins from plants, phospholipids and acylglycerols, it is important to note that other mycobacterial cutinases like RV3452 preferentially targeting non-acylglycerols have not been detected in our target identification experiment (Schue, M. et al. Two cutinase-like proteins secreted by Mycobacterium tuberculosis show very different lipolytic activities reflecting their physiological function. FASEB J. 24, 1893-1903 (2010)).
Rv2715 is the fourth strongest lalistat-binding hit. It represents a still uncharacterized protein, which is suggested to participate in lipid hydrolysis in bacterial metabolism. However, its detailed function is still unknown. Further studies have to be performed to investigate its capability to serve as a potential anti-mycobacterial target. Similar to RV2715, Rv1 192, which has also been identified among the 10 strongest lalistat-binding proteins, is a further uncharacterized protein suggested to belong to the lipolytic enzyme family. It has a membranous localisation and contains a PS00120 lipase pattern.
Regarding the putative non-lipolytic enzymes bound by lalistat, the inventors detected two proteins (third and seventh hit of our screen) similar to other Mycobacterium tuberculosis hypothetical penicillin binding proteins and esterases, which are possibly involved in cell wall biosynthesis: Rv1367 and Rv1730. For Rv1730 it has been shown that its function is essential for mycobacterial growth in vitro (Sassetti, C. M., Boyd, D. H. & Rubin, E. J. Genes required for mycobacterial growth defined by high density mutagenesis. Mol. Microbiol. 48, 77-84 (2003)) and might represent an interesting anti-mycobacterial target. Three further non-lipolytic enzymes were detected among the hits 12-16 of our target identification which all represent hydrolases: Rv2888c and Rv2363 belong to the family of putative amidases and Rv0840c is a putative proline iminopeptidase. Rv2888c has been identified in chronic but not early stages of disease when investigated in the guinea pig model of tuberculosis (Kruh, N. A., Troudt, J., Izzo, A., Prenni, J. & Dobos, K. M. Portrait of a pathogen: The Mycobacterium tuberculosis proteome In vivo. PLoS One 5, (2010)). Rv0293c, identified as the protein hit 1 1 , is a non-characterised protein with unkown function. It is highly conserved and upregulated during starvation (Sassetti, C. M., Boyd, D. H. & Rubin, E. J. Genes required for mycobacterial growth defined by high density mutagenesis. Mol. Microbiol. 48, 77-84 (2003)). Finally, Rv1 191 representing the target with the lowest affinity for lalistat identified, is a conserved protein with unknown function and distinct similarity to proline iminopeptidases.
Taken together, all characterized targets of lalistat identified in Mycobacterium tuberculosis represent known or putative members of the hydrolase family of which most are lipolytic enzymes, underlining the specificity of the inhibitor lalistat and its preference for lipolytic hydrolases. Most of these targets are highly conserved and have been reported to be upregulated during later chronic stages of infection, suggesting that lalistat has a high potential to target dormant bacteria and to largely improve current TB therapy.
Although the inventors cannot exclude that inhibition of one specific identified target by lalistat is mainly responsible for the anti-mycobacterial activity observed, it is likely that binding to a panel of distinct members of the lipolytic enzyme family mediates the anti-bacterial effect described.
This mode would be in agreement with recent strategies to target not only one but selected groups of mycobacterial proteins for improved future therapeutic applications, including lipases.
Multi-targeting is an effective approach to avoid bacterial drug resistance while combating the disease (Saravanan, P. & Patra, S. Discovery of Potential Dual Inhibitors Against Lipases
Rv0183 and Rv3802c for Tuberculosis Therapeutics. Lett. Drug Des. Discov. 13, 185-195
(2015)).
Moreover, the invention is characterized by the following items:
1. A compound for use in the treatment of a mycobacterial disease, said compound having a structure according to Formula I
Figure imgf000058_0001
wherein
Figure imgf000058_0002
X is a nitrogen containing heterocycle;
Y is CH2, O or NR1; n is an integer between 0 and 3;
R1 and R2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (CTC6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-Ci0)aryl, substituted or unsubstituted (C6-Ci0)aryl(Ci-C6)alkyl, substituted or unsubstituted (C3-C10)heteroaryl, and substituted or unsubstituted (C3- Cio)heteroaryl(Ci-C6)alkyl; or a pharmaceutically acceptable salt, solvate or hydrate thereof;
with the proviso that when n = 0, Y is not O or NR . A compound for use in medicine, said compound having a structure according to Formula I
Figure imgf000059_0001
wherein
Figure imgf000059_0002
X is a nitrogen containing heterocycle;
Y is CH2, O or NR ; n is an integer between 0 and 3;
R1 and R2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-C10)aryl, substituted or unsubstituted (Ce-C^JaryliC CeJalkyl, substituted or unsubstituted (C3-Ci0)heteroaryl, and substituted or unsubstituted (C3-
Figure imgf000059_0003
or a pharmaceutically acceptable salt, solvate or hydrate thereof;
with the proviso that
(iv) when n = 0, Y is not O or NR1;
(v) when n = 2, Y is not CH2; or
Figure imgf000060_0001
 A pharmaceutical composition for use in the treatment of a mycobacterial disease, wherein said composition comprises a compound having a structure according to Formula I
Figure imgf000061_0001
X is a nitrogen containing heterocycle;
Y is CH2, O or NR ; n is an integer between 0 and 3;
R1 and R2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-C10)aryl, substituted or unsubstituted (Cs-CioJaryliC-i-CeJalkyl, substituted or unsubstituted (C3-C10)heteroaryl, and substituted or unsubstituted (C3- C10)heteroaryl(C1-C6)alkyl; or a pharmaceutically acceptable salt, solvate or hydrate thereof; with the proviso that when n = 0, Y is not O or NR1. The compound for use according to any one of the pharmaceutical
composition for use according to item 3, wherein A is
Figure imgf000062_0001
The compound for use or the pharmaceutical composition for use according to any one of items1-4, wherein
X is
Figure imgf000062_0002
wherein
Z is CR3R4, O or NR5;
R3 and R4 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-Ci0)aryl, substituted or unsubstituted (C6-Ci0)aryl(Ci-C6)alkyl, substituted or unsubstituted (C3-C10)heteroaryl, substituted or unsubstituted (C3- CioJheteroaryliCr alkyl,
halogen, -CN, -N02, -OR11, -N(R 2)(R13), -N(R11)(OR11), -S(O)0-2R11, -S(0)1-2OR11, -OS(O) 1-2R11, -OS(0)1-2OR11, -S(0)1-2N(R 2)(R13), -OS(0)1-2N(R 2)(R13), -N(R 1)S(0)1-2R11, -NR11 S(0)1-2OR11, -NR S(0)1-2N(R 2)(R13), -C(=W)R11, -C(=W)WR11, -WC(=W)R11, and -WC(=W)WR11;
R5 is selected from the group consisting of hydrogen, substituted or unsubstituted (Cr C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2- CB)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-Cio)aryl, substituted or unsubstituted (C6-Ci0)aryl(Ci-C6)alkyl, substituted or unsubstituted (C3-C10)heteroaryl, substituted or unsubstituted
Figure imgf000062_0003
CB)alkyl, -N(R 2)(R13), -N(R 1)(OR11), -S(O)0-2R11, -S(0)1-2OR11, -N(R )S(0)1-2R11, -NR S (0)1-2OR11, -NR S(0)1-2N(R 2)(R13), -C(=W)R11, -C(=W)WR11, -WC(=W)R11, and -WC(=W)WR11;
W is independently selected from O, S, and N(R14);
R1 is, in each case, selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, and heterocyclyl;
R12 and R13 are, in each case, independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, and heterocyclyl, or R12 and R13 may join together with the nitrogen atom to which they are attached to form the group -N=CR 5R16;
R14 is independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, heterocyclyl, and -OR11;
R15 and R16 are independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, and heterocyclyl.
The compound for use or the pharmaceutical composition for use according to any one of items 1-5, having a structure according to Formula II
Figure imgf000063_0001
wherein
Y, Z, W, R1, R3, R4, R5, R 1, R12, R 3, R14, R15, R16 and n are defined as above;
with the proviso that when n = 0, Y is not O or NR . The compound for use or the pharmaceutical composition for use according to any one of items 1-6, having a structure according to Formula III
Figure imgf000064_0001
wherein
Z is O or CR3R4;
R3 and R4 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (CTC6)alkyl, substituted or unsubstituted (C2-C6)alkynyl and -OR11;
R11 is, in each case, selected from the group consisting of hydrogen and substituted or unsubstituted (Ci-C6)alkyl;
preferably wherein
R3 and R4 are hydrogen or substituted or unsubstituted (C C6)alkyl; or wherein at least one of R3 and R4 is hydrogen. The compound for use or the pharmaceutical composition for use according to any one of items 1 , 3-7, wherein the mycobacterial disease is caused by at least one bacteria selected from the group consisting of Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae, Mycobacterium microti, Mycobacterium leprae, Mycobacterium lepromatosis, Mycobacterium avium, Mycobacterium silvaticum, Mycobacterium hominissuis, Mycobacterium paratuberculosis, Mycobacterium kansasii, Mycobacterium xenopi, Mycobacterium simiae, Mycobacterium abcessus, Mycobacterium fortuitum, Mycobacterium chelonae, Mycobacterium ulcerans, Mycobacterium marinum and/or Mycobacterium fortuitum, preferably Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae, Mycobacterium microti, Mycobacterium leprae, Mycobacterium lepromatosis and/or Mycobacterium kansasii, more preferably Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae, Mycobacterium microti and/or Mycobacterium kansasii, even more preferably Mycobacterium tuberculosis. 9. The compound for use or the pharmaceutical composition for use according to any one of items 1 , 3-8, wherein the mycobacterial disease is selected from tuberculosis, leprosy (Hansen's disease), lepromatosis, infections caused by non-tuberculosis mycobacteria including lymphadenitis and pulmonary infections, skin infections caused by mycobacteria including Buruli ulcer and fish tank granuloma.
10. The compound for use or the pharmaceutical composition for use according to any one of items 1 , 3-9, wherein the mycobacterial disease is tuberculosis and/or caused by bacteria of the group consisting of Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae, Mycobacterium microti, preferably Mycobacterium tuberculosis.
1 1. The pharmaceutical composition for use according to any one of items 3-10, further comprising at least one pharmaceutically acceptable carrier. 12. The pharmaceutical composition for use according to any one of items 3-1 1 , wherein said composition is administered to a subject in need thereof in an amount effective to treat said mycobacterial disease, preferably wherein the subject is a mammal, more preferably a human subject. 13. The pharmaceutical composition for use according to item 12, wherein said subject is treated with at least one additional pharmaceutically active compound, including an antibiotic, antifungal or anti-HIV compound, preferably wherein said additional pharmaceutically active compound is an antibiotic, antifungal and/or anti-HIV compound, more preferably an antibiotic.
14. The pharmaceutical composition for use according to item 13, wherein said additional pharmaceutically active compound is selected from the group consisting of Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin, Rifabutin, Animoglycosides including Kanamycin and/or Amikacin, Polypetides including Capreomycin, Viomycin and/or Streptomycin, fluoroquinolones including Moxifloxacin, Levofloxacin, Ofloxacin and/or
Gatifloxacin, thioamides including Ethionamide and/or Protionamide, Cycloserine, Terizidone, Thioacetone, p-Aminosalicylic acid, Clofazimine, Linezolid, Amoxicillin, Clavulanate, Thioacetazone, Imipenem, Cilastatin, Clarithromycin, Delamanid and/or Bedaquiline, preferably Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin, Rifabutin, Animoglycosides including Kanamycin and/or Amikacin, Polypetides including Capreomycin, Viomycin and/or Streptomycin, fluoroquinolones including Moxifloxacin, Levofloxacin, Ofloxacin and/or Gatifloxacin, thioamides including Ethionamide and/or Protionamide, Cycloserine, Terizidone, Thioacetone and/or p-Aminosalicylic acid, more preferably Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin and/or Rifabutin, even more preferably Isoniazid, Rifampicin, Ethambutol and/or Pyrazinamide. Kit of parts comprising a pharmaceutical composition and at least one additional pharmaceutically active compound, wherein said pharmaceutical composition comprises a compound having a structure according to Formula I
Figure imgf000066_0001
wherein
A is selected from the group consisting of:
Figure imgf000066_0002
X is a nitrogen containing heterocycle;
Y is CH2, O or NR ; n is an integer between 0 and 3;
R1 and R2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-Ci0)aryl, substituted or unsubstituted (C6-Ci0)aryl(Ci-C6)alkyl, substituted or unsubstituted (C3-Ci0)heteroaryl, and substituted or unsubstituted (C3- Cio)heteroaryl(Ci-C6)alkyl; or a pharmaceutically acceptable salt, solvate or hydrate thereof;
with the proviso that when n = 0, Y is not O or NR ; and wherein said additional pharmaceutically active compound is selected from the group consisting of an antibiotic, antifungal or anti-HIV compound, preferably an antibiotic selected from the group of Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin, Rifabutin, Animoglycosides including Kanamycin and/or Amikacin, Polypetides including Capreomycin, Viomycin and/or Streptomycin, fluoroquinolones including Moxifloxacin, Levofloxacin, Ofloxacin and/or Gatifloxacin, thioamides including Ethionamide and/or Protionamide, Cycloserine, Terizidone, Thioacetone, p-Aminosalicylic acid, Clofazimine, Linezolid, Amoxicillin, Clavulanate, Thioacetazone, Imipenem, Cilastatin, Clarithromycin, Delamanid and/or Bedaquiline, preferably Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin, Rifabutin, Animoglycosides including Kanamycin and/or Amikacin, Polypetides including Capreomycin, Viomycin and/or Streptomycin, fluoroquinolones including Moxifloxacin, Levofloxacin, Ofloxacin and/or Gatifloxacin, thioamides including Ethionamide and/or Protionamide, Cycloserine, Terizidone, Thioacetone and/or p-Aminosalicylic acid, more preferably Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin and/or Rifabutin, even more preferably Isoniazid, Rifampicin, Ethambutol and/or Pyrazinamide.
It will be clear that the invention may be practiced otherwise than as particularly described in the foregoing description and examples. Numerous modifications and variations of the present invention are possible in light of the above teachings and, therefore, are within the scope of the appended claims.

Claims

Claims
1. A compound for use in the treatment of a mycobacterial disease, said compound having a structure according to Formula I
Figure imgf000069_0001
Figure imgf000069_0002
wherein
Z is CR3R4, 0 or NR5;
R3 and R4 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl/ substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-Ci0)aryl, substituted or unsubstituted (C6-Ci0)aryl(Ci-C6)alkyl, substituted or unsubstituted (C3-C10)heteroaryl, substituted or unsubstituted (C CKjjheteroaryllCrCejalkyl, halogen, -CN, -N02, -OR11, -N(R12)(R13), -N(Rn)(ORu), -S(O)0.2Rn, -S(0)1 2ORn, -OSfOJ^R11, -OS(Oh _2ORu, -S(0)!.2N(R12)(R13), -OS(0)!_2N(R12)(R13), -NIR^SIOj^R11, -NR^IOJ^OR11, -N R^IOJ^NIR1 2)(R13), -C(=W)RU, -C(=W)WR , -WC(=W)Rn, and -WC(=W)WRn;
R5 is selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-Ci0)aryl, substituted or unsubstituted (C6-Ci0)aryl(Ci-C6)alkyl, substituted or unsubstituted (C3- CiQ)heteroaryl, substituted or unsubstituted (C3-Ci0)heteroaryl(Ci-
C6)alkyl, -N(R12)(R13), -N(R11)(OR11), -S(O)0.2Rn, -SfO^OR11, -NIR^SIOj^R11, -NR^OJ^OR11, -NR "SfOl^NlR^KR13), -C(=W)R , -C(=W)WRn, -WC(=W)RU, and -WC(=W)WR ;
W is independently selected from 0, S, and N(R14);
R11 is, in each case, selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, and heterocyclyl;
R12 and R13 are, in each case, independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, and heterocyclyl, or R12 and R13 may join together with the nitrogen atom to which they are attached to form the group -N=CR15R16;
R14 is independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, heterocyclyl, and -OR11;
R15 and R16 are independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, and heterocyclyl
Y is CH2, 0 or NR1; n is an integer between 0 and 3; R1 and R2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-Ci0)aryl, substituted or unsubstituted (C6-Ci0)aryl(Ci-C6)alkyl, substituted or unsubstituted (C3-C10)heteroaryl, and substituted or unsubstituted (C3-C10)heteroaryl(Ci-C6)alkyl; or a pharmaceutically acceptable salt, solvate or hydrate thereof;
with the proviso that when n = 0, Y is not O or NR1.
2. A compound for use in medicine, said compound having a structure according to Formula I
Figure imgf000071_0001
X is
Figure imgf000071_0002
wherein
Z is CR3R4, 0 or NR5;
R3 and R4 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-Ci0)aryl, substituted or unsubstituted (C6-Ci0)aryl(Ci-C6)alkyl, substituted or unsubstituted (C3-Ci0)heteroaryl, substituted or unsubstituted (CVCiojheteroaryllCrCejalkyl, halogen, -CN, -N02, -OR11, -N(R12)(R13), -N(Ru)(ORu), -S(O)0.2Rn, -S(0)1 2ORu, -OSfOJ^R11, -OS(Oh _2ORu, -S(0)!.2N(R12)(R13), -OS(0)!_2N(R12)(R13), -NIR^SIOj^R11, -NR^IOJ^OR11, -N R^IOl^NI 1 2)(R13), -C(=W)RU, -C(=W)WR , -WC(=W)Rn, and -WC(=W)WRn;
R5 is selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C5-Ci0)aryl, substituted or unsubstituted (Ce-Cu arylld-Cejalkyl, substituted or unsubstituted (C3- Ci0)heteroaryl, substituted or unsubstituted (C3-Ci0)heteroaryl(Ci-
C6)alkyl, -N(R12)(R13), -N(R11)(OR11), -S(O)0.2Rn, -SfO^OR11, -NIR^SIOj^R11, -NR^Oj^OR11, -NR "SlOj^Nf R^KR13), -C(=W)R , -C(=W)WRn, -WC(=W)Rn, and -WC(=W)WR ;
W is independently selected from 0, S, and N(R14);
R11 is, in each case, selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, and heterocyclyl;
R12 and R13 are, in each case, independently selected from the grou p consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, and heterocyclyl, or R12 and R13 may join together with the nitrogen atom to which they are attached to form the group -N=CR15R16;
R14 is independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, heterocyclyl, and -OR11; R and R are independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, and heterocyclyl
Y is CH2, 0 or NR1; n is an integer between 0 and 3;
R1 and R2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted (C6-Ci0)aryl, substituted (C6-Ci0)aryl(Ci- C6)alkyl, substituted or unsubstituted (C3-Ci0)heteroaryl, and substituted or unsubstituted (C3- CiojheteroaryllC Cgjalkyl; or a pharmaceutically acceptable salt, solvate or hydrate thereof;
with the proviso that
(i) when n = 0, Y is not 0 or NR1;
(ii) when n = 2, Y is not CH2; or
(iii) the compound is not
Figure imgf000073_0001
Figure imgf000074_0001
73
3. A pharmaceutical composition for use in the treatment of a mycobacterial disease, wherein said composition comprises a compound having a structure according to Formula I
Figure imgf000075_0001
Figure imgf000075_0002
wherein
Z is CR3R4, 0 or NR5;
R3 and R4 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-Ci0)aryl, substituted or unsubstituted (C6-Ci0)aryl(Ci-C6)alkyl, substituted or unsubstituted (C3-C10)heteroaryl, substituted or unsubstituted (Cs-Ciojheteroarylld-Cejalkyl, halogen, -CN, -N02, -OR11, -N(R12)(R13), -N(Rn)(ORn), -S(O)0.2Rn, -S(0)1 2ORn, -OSfOJ^R11, -OS(Oh _2ORu, -S(0)!.2N(R12)(R13), -OS(0)!_2N(R12)(R13), -NlR^SlOj^R11, -NR^OJ^OR11, -N R^Oj^NlR1 2)(R13), -C(=W)RU, -C(=W)WR , -WC(=W)Rn, and -WC(=W)WRn;
R5 is selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C3)cycloalkyl, substituted or unsubstituted (C6-C10)aryl, substituted or unsubstituted
Figure imgf000076_0001
substituted or unsubstituted (C3- CiQ)heteroaryl, substituted or unsubstituted (C3-Ci0)heteroaryl(Ci-
C6)alkyl, -N(R1 )(R13), -N(R11)(OR11), -S(O)0.2Rn, -SfO^OR11, -NIR^SIOj^R11, -NR^IOl^OR11, -NR "SlOh^Nf R12)^1 ), -C(=W)R , -C(=W)WRn, -WC(=W)Rn, and -WC(=W)WR ;
W is independently selected from O, S, and N(R14);
R11 is, in each case, selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyi, aryl, heteroaryl, and heterocyclyl;
R12 and R13 are, in each case, independently selected from the grou p consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyi, aryl, heteroaryl, and heterocyclyl, or R12 and R13 may join together with the nitrogen atom to which they are attached to form the group -N=CR15R16;
R14 is independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyi, aryl, heteroaryl, heterocyclyl, and -OR11;
R15 and R16 are independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyi, aryl, heteroaryl, and heterocyclyl
Y is CH2, 0 or NR1; n is an integer between 0 and 3;
R1 and R2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (CL-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl/ substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-Ci0)aryl, substituted or unsubstituted (C6-Ci0)aryl(Ci-C6)alkyl, substituted or unsubstituted (C3-C10)heteroaryl, and substituted or unsubstituted
Figure imgf000077_0001
or a pharmaceutically acceptable salt, solvate or hydrate thereof;
with the proviso that when n = 0, Y is not 0 or N R1.
4. The compound for use according to any one of claims 1-2 or the pharmaceutical composition for
use according to claim 3, wherein A is
Figure imgf000077_0002
5. The compound for use or the pharmaceutical composition for use according to any one of claims 1-4, having a structure according to Formula I I
Figure imgf000077_0003
wherein
Y, Z, W, R1, R3, R4, R5, R11, R12, R13, R14, R15, R16 and n are defined as above;
with the proviso that when n = 0, Y is not 0 or N R1.
6. The compound for use or the pharmaceutical composition for use according to any one of claims 1 and 3-5, having a structure according to Formula I I I
Figure imgf000078_0001
wherein
Z is 0 or CR3R4;
R3 and R4 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkynyl and -OR11;
R11 is, in each case, selected from the group consisting of hydrogen and substituted or unsubstituted (Ci-C6)alkyl;
preferably wherein
R3 and R4 are hydrogen or substituted or unsubstituted (Ci-C6)alkyl; or wherein
at least one of R3 and R4 is hydrogen.
7. The compound for use or the pharmaceutical composition for use according to any one of claims 1, 3-6, wherein the mycobacterial disease is caused by at least one bacteria selected from the group consisting of Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae, Mycobacterium microti, Mycobacterium leprae, Mycobacterium lepromatosis, Mycobacterium avium, Mycobacterium silvaticum, Mycobacterium bominissuis, Mycobacterium paratuberculosis, Mycobacterium kansasii, Mycobacterium xenopi, Mycobacterium simiae, Mycobacterium abcessus, Mycobacterium fortuitum, Mycobacterium chelonae, Mycobacterium ulcerans, Mycobacterium marinum and/or Mycobacterium fortuitum, preferably Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae, Mycobacterium microti, Mycobacterium leprae, Mycobacterium lepromatosis and/or Mycobacterium kansasii, more preferably Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae, Mycobacterium microti and/or Mycobacterium kansasii, even more preferably Mycobacterium tuberculosis.
8. The compound for use or the pharmaceutical composition for use according to any one of claims 1, 3-7, wherein the mycobacterial disease is selected from tuberculosis, leprosy (Hansen's disease), lepromatosis, infections caused by non-tuberculosis mycobacteria including lymphadenitis and pulmonary infections, skin infections caused by mycobacteria including Buruli ulcer and fish tank granuloma.
9. The compound for use or the pharmaceutical composition for use according to any one of claims 1, 3-8, wherein the mycobacterial disease is tuberculosis and/or caused by bacteria of the group consisting of Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium bovis, Mycobacterium caprae, Mycobacterium microti, preferably Mycobacterium tuberculosis.
10. The pharmaceutical composition for use according to any one of claims 3-9, further comprising at least one pharmaceutically acceptable carrier.
11. The pharmaceutical composition for use according to any one of claims 3-10, wherein said composition is administered to a subject in need thereof in an amount effective to treat said mycobacterial disease, preferably wherein the subject is a mammal, more preferably a human subject.
12. The pharmaceutical composition for use according to claim 11, wherein said subject is treated with at least one additional pharmaceutically active compound, including an antibiotic, antifungal or anti-HIV compound, preferably wherein said additional pharmaceutically active compound is an antibiotic, antifungal and/or anti-HIV compound, more preferably an antibiotic.
13. The pharmaceutical composition for use according to claim 12, wherein said additional pharmaceutically active compound is selected from the group consisting of Isoniazid, ifampicin, Ethambutol, Pyrazinamide, Rifapentin, Rifabutin, Animoglycosides including Kanamycin and/or Amikacin, Polypetides including Capreomycin, Viomycin and/or Streptomycin, fluoroquinolones including Moxifloxacin, Levofloxacin, Ofloxacin and/or Gatifloxacin, thioamides including Ethionamide and/or Protionamide, Cycloserine, Terizidone, Thioacetone, p-Aminosalicylic acid, Clofazimine, Linezolid, Amoxicillin, Clavulanate, Thioacetazone, Imipenem, Cilastatin, Clarithromycin, Delamanid and/or Bedaquiline, preferably Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin, Rifabutin, Animoglycosides including Kanamycin and/or Amikacin, Polypetides including Capreomycin, Viomycin and/or Streptomycin, fluoroquinolones including Moxifloxacin, Levofloxacin, Ofloxacin and/or Gatifloxacin, thioamides including Ethionamide and/or Protionamide, Cycloserine, Terizidone, Thioacetone and/or p-Aminosalicylic acid, more preferably Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin and/or Rifabutin, even more preferably Isoniazid, Rifampicin, Ethambutol and/or Pyrazinamide.
Kit of parts comprising a pharmaceutical composition and at least one additional pharmaceutically active compound, wherein said pharmaceutical composition comprises a compound having a structure according to Formula I
Figure imgf000080_0001
X is
Figure imgf000080_0002
wherein Z is CR3R4, 0 or NR5;
R3 and R4 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-Ca)cycloalkyl, substituted or unsubstituted (C6-Ci0)aryl, substituted or unsubstituted
Figure imgf000081_0001
substituted or unsubstituted (C3-Ci0) heteroaryl, substituted or unsubstituted (Cs-CiojheteroaryllCrCejalkyl, halogen, -CN, -N02, -OR11, -N(R12)(R13), -N(Ru)(ORn), -S(O)0.2Rn, -S(0)1 2ORu, -OSfOJ^R11, -OS(O)! _2OR , -S(0)!.2N(R1 )(R13), -OS(0)!.2N(R12)(R13),
Figure imgf000081_0002
2)(R13), -C(=W)R , -C(=W)WR , -WC(=W)Rn, and -WC(=W)WRn;
R5 is selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-Ci0)aryl, substituted or unsubstituted (C5-Cio)aryl(Ci-C5)alkyl, substituted or unsubstituted (C3- C10)heteroaryl, substituted or unsubstituted (Cs-Ciojheteroarylfd-
C6)alkyl, -N(R12)(R13), -N(R11)(OR11), -S(O)0.2Rn, -SfOJ^OR11, -NlR^SIOj^R11, - R'^IOl^OR11, -NR "SfOh^Nf R12)^13), -C(=W)R , -C(=W)WRn, -WC(=W)Rn, and -WC(=W)WR ;
W is independently selected from O, S, and N(R14);
R11 is, in each case, selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, and heterocyclyl;
R12 and R13 are, in each case, independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, and heterocyclyl, or R12 and R13 may join together with the nitrogen atom to which they are attached to form the group -N=CR15R16;
R14 is independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, heterocyclyl, and -OR11;
R15 and R16 are independently selected from the group consisting of -H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heteroaryl, and heterocyclyl Y is CH2, 0 or NR1; n is an integer between 0 and 3;
R1 and R2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted (Ci-C6)alkyl, substituted or unsubstituted (C2-C6)alkenyl, substituted or unsubstituted (C2-C6)alkynyl, substituted or unsubstituted (C3-C8)cycloalkyl, substituted or unsubstituted (C6-CiQ)aryl, substituted or unsubstituted (CE-Cia)aryl(Ci-C8)alkyl, substituted or unsubstituted (C3-Ci0)heteroaryl, and substituted or unsubstituted (C3-Ci0)heteroaryl(Ci-C6)alkyl; or a pharmaceutically acceptable salt, solvate or hydrate thereof;
with the proviso that when n = 0, Y is not 0 or N R1; and wherein said additional pharmaceutically active compound is selected from the group consisting of an antibiotic, antifungal or anti-HIV compound, preferably an antibiotic selected from the group of Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin, Rifabutin, Animoglycosides including Kanamycin and/or Amikacin, Polypetides including Capreomycin, Viomycin and/or Streptomycin, fluoroquinolones including Moxifloxacin, Levofloxacin, Ofloxacin and/or Gatifloxacin, thioamides including Ethionamide and/or Protionamide, Cycloserine, Terizidone, Thioacetone, p-Aminosalicylic acid, Clofazimine, Linezolid, Amoxicillin, Clavulanate, Thioacetazone, Imipenem, Cilastatin, Clarithromycin, Delamanid and/or Bedaquiline, preferably Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin, Rifabutin, Animoglycosides including Kanamycin and/or Amikacin, Polypetides including Capreomycin, Viomycin and/or Streptomycin, fluoroquinolones including Moxifloxacin, Levofloxacin, Ofloxacin and/or Gatifloxacin, thioamides including Ethionamide and/or Protionamide, Cycloserine, Terizidone, Thioacetone and/or p-Aminosalicylic acid, more preferably Isoniazid, Rifampicin, Ethambutol, Pyrazinamide, Rifapentin and/or Rifabutin, even more preferably Isoniazid, Rifampicin, Ethambutol and/or Pyrazinamide.
PCT/EP2017/061453 2016-05-13 2017-05-12 Means and methods for treating mycobacterial diseases WO2017194734A1 (en)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
EP16169499 2016-05-13
EP16169499.7 2016-05-13

Publications (1)

Publication Number Publication Date
WO2017194734A1 true WO2017194734A1 (en) 2017-11-16

Family

ID=56008495

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/EP2017/061453 WO2017194734A1 (en) 2016-05-13 2017-05-12 Means and methods for treating mycobacterial diseases

Country Status (1)

Country Link
WO (1) WO2017194734A1 (en)

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN113271930A (en) * 2019-01-09 2021-08-17 詹森药业有限公司 Combinations for treating nontuberculous mycobacterial diseases
WO2022003421A3 (en) * 2020-06-30 2022-03-17 Redhill Biopharma Ltd. Triple antibiotic fixed-dose combination products, dosing regimen, methods, and kits for treating pulmonary non-tuberculosis mycobacterial infections

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2014000058A1 (en) * 2012-06-29 2014-01-03 Garvan Institute Of Medical Research Method of treating glucose metabolism disorders
WO2015135652A1 (en) * 2014-03-12 2015-09-17 Technische Universität München Antagonists of acid lipase for preventing virus infection

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2014000058A1 (en) * 2012-06-29 2014-01-03 Garvan Institute Of Medical Research Method of treating glucose metabolism disorders
WO2015135652A1 (en) * 2014-03-12 2015-09-17 Technische Universität München Antagonists of acid lipase for preventing virus infection

Non-Patent Citations (8)

* Cited by examiner, † Cited by third party
Title
GURDYAL SINGH ET AL: "Lipid hydrolizing enzymes in virulence: Mycobacterium tuberculosis as a model system", CRITICAL REVIEWS IN MICROBIOLOGY, vol. 36, no. 3, 25 May 2010 (2010-05-25), pages 259 - 269, XP055309932, ISSN: 1040-841X, DOI: 10.3109/1040841X.2010.482923 *
J. LEHMANN ET AL: "Human lysosomal acid lipase inhibitor lalistat impairs Mycobacterium tuberculosis growth by targeting bacterial hydrolases", MEDCHEMCOMM, vol. 7, no. 9, 1 January 2016 (2016-01-01), United Kingdom, pages 1797 - 1801, XP055309841, ISSN: 2040-2503, DOI: 10.1039/C6MD00231E *
JAYENDRA Z. PATEL ET AL: "Optimization of 1,2,5-Thiadiazole Carbamates as Potent and Selective ABHD6 Inhibitors", CHEMMEDCHEM, vol. 10, no. 2, 11 December 2014 (2014-12-11), DE, pages 253 - 265, XP055310052, ISSN: 1860-7179, DOI: 10.1002/cmdc.201402453 *
M. S. RAVINDRAN ET AL: "Targeting Lipid Esterases in Mycobacteria Grown Under Different Physiological Conditions Using Activity-based Profiling with Tetrahydrolipstatin (THL)", MOLECULAR & CELLULAR PROTEOMICS, vol. 13, no. 2, 17 December 2013 (2013-12-17), US, pages 435 - 448, XP055310181, ISSN: 1535-9476, DOI: 10.1074/mcp.M113.029942 *
ROSENBAUM A I ET AL: "Chemical screen to reduce sterol accumulation in Niemann-Pick C disease cells identifies novel lysosomal acid lipase inhibitors", BIOCHIMICA AND BIOPHYSICA ACTA. MOLECULAR AND CELL BIOLOGY OF LIPIDS, ELSEVIER, AMSTERDAM, NL, vol. 1791, no. 12, 1 December 2009 (2009-12-01), pages 1155 - 1165, XP026754993, ISSN: 1388-1981, [retrieved on 20091111], DOI: 10.1016/J.BBALIP.2009.08.005 *
ROSENBAUM A I ET AL: "Thiadiazole Carbamates: Potent Inhibitors of Lysosomal Acid Lipase and Potential Niemann-Pick Type C Disease Therapeutics", JOURNAL OF MEDICINAL CHEMISTRY, AMERICAN CHEMICAL SOCIETY, US, vol. 53, no. 14, 22 July 2010 (2010-07-22), pages 5281 - 5289, XP002727816, ISSN: 0022-2623, [retrieved on 20100617], DOI: 10.1021/JM100499S *
SCHWAID ADAM G ET AL: "Development of a selective activity-based probe for glycosylated LIPA", BIOORGANIC & MEDICINAL CHEMISTRY LETTERS, PERGAMON, AMSTERDAM, NL, vol. 26, no. 8, 2 March 2016 (2016-03-02), pages 1993 - 1996, XP029470643, ISSN: 0960-894X, DOI: 10.1016/J.BMCL.2016.02.089 *
VINCENT DELORME ET AL: "MmPPOX Inhibits Mycobacterium tuberculosis Lipolytic Enzymes Belonging to the Hormone-Sensitive Lipase Family and Alters Mycobacterial Growth", PLOS ONE, vol. 7, no. 9, 28 September 2012 (2012-09-28), pages e46493, XP055310183, DOI: 10.1371/journal.pone.0046493 *

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN113271930A (en) * 2019-01-09 2021-08-17 詹森药业有限公司 Combinations for treating nontuberculous mycobacterial diseases
WO2022003421A3 (en) * 2020-06-30 2022-03-17 Redhill Biopharma Ltd. Triple antibiotic fixed-dose combination products, dosing regimen, methods, and kits for treating pulmonary non-tuberculosis mycobacterial infections
US11590154B2 (en) 2020-06-30 2023-02-28 Redhill Biopharma Ltd. Triple antibiotic fixed-dose combination products, dosing regimen, methods, and kits for treating pulmonary non-tuberculosis mycobacterial infections

Similar Documents

Publication Publication Date Title
US10849982B2 (en) C3-carbon linked glutarimide degronimers for target protein degradation
US10646575B2 (en) Heterocyclic degronimers for target protein degradation
US20230095223A1 (en) Spirocyclic degronimers for target protein degradation
EP0866793B1 (en) Nitroimidazole antibacterial compounds and methods of use thereof
US20190307760A1 (en) Acc inhibitors for use in treating mycobacterial diseases
US20210171514A1 (en) Proteasome inhibitors and uses thereof
EP3464260B1 (en) Urea motif containing compounds and derivatives thereof as antibacterial drugs
AU2014268477A1 (en) Compounds for treatment of drug resistant and persistent tuberculosis
JP2015511601A (en) The broad-spectrum antibiotic allylomycin analog
KR20170113529A (en) Non-beta lactam antibiotics
EP3013818B1 (en) (s)-enantiomer of mepazine as paracaspase (malt1) inhibitor for treating cancer
WO2017194734A1 (en) Means and methods for treating mycobacterial diseases
ES2833929T3 (en) Anti-tuberculosis agent
Nandi et al. Exploring targets of cell wall protein synthesis and overexpression mediated drug resistance for the discovery of potential M. tb inhibitors
US10787442B2 (en) Modulators of indoleamine 2,3-dioxygenase
US20210290629A1 (en) Acc inhibitors for use in treating mycobacterial diseases
CN109071511B (en) Zwitterionic propargyl-linked antifolates useful for the treatment of bacterial infections
US10035771B2 (en) Mercaptonicotinic acid compound and preparation method and use thereof
US8975288B2 (en) F2 derivatives as antibacterial agents
Amruta Adhikrao et al. Tackling Nontuberculous Mycobacteria by Repurposable Drugs and Potential Leads from Natural Products
Bakker Discovery of Antibiotics and Their Targets in Multidrug-Resistant Bacteria
JP6326048B2 (en) Combined anti-mycobacterial agent and activity inhibitor of WecA or its ortholog
WO2023230547A2 (en) Cilagicin compounds and methods of use thereof
Kling Mode of action and resistance mechanism of griselimycin
Antic Identification and Characterization of Ras and Rap1A Specific Protease Domain from Vibrio vulnificus MARTX toxin

Legal Events

Date Code Title Description
NENP Non-entry into the national phase

Ref country code: DE

121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 17725908

Country of ref document: EP

Kind code of ref document: A1

122 Ep: pct application non-entry in european phase

Ref document number: 17725908

Country of ref document: EP

Kind code of ref document: A1