WO2014167160A1 - Peptide compounds suitable for use as antibiotic agents - Google Patents

Peptide compounds suitable for use as antibiotic agents Download PDF

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Publication number
WO2014167160A1
WO2014167160A1 PCT/ES2014/070286 ES2014070286W WO2014167160A1 WO 2014167160 A1 WO2014167160 A1 WO 2014167160A1 ES 2014070286 W ES2014070286 W ES 2014070286W WO 2014167160 A1 WO2014167160 A1 WO 2014167160A1
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Prior art keywords
dab
radical
gram
group
bacteria
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PCT/ES2014/070286
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Spanish (es)
French (fr)
Inventor
Francesc Rabanal Anglada
Yolanda C. CAJAL VISA
Ariadna GRAU CAMPISTANY
Jordi VILA ESTAPÉ
Xavier VILA FARRÉS
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Universitat De Barcelona
Fundació Clínic Per A La Recerca Biomèdica
Hospital Clínic De Barcelona
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Publication of WO2014167160A1 publication Critical patent/WO2014167160A1/en

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/04—Linear peptides containing only normal peptide links
    • C07K7/06—Linear peptides containing only normal peptide links having 5 to 11 amino acids
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/50—Cyclic peptides containing at least one abnormal peptide link
    • C07K7/54—Cyclic peptides containing at least one abnormal peptide link with at least one abnormal peptide link in the ring
    • C07K7/60—Cyclic peptides containing at least one abnormal peptide link with at least one abnormal peptide link in the ring the cyclisation occurring through the 4-amino group of 2,4-diamino-butanoic acid
    • C07K7/62—Polymyxins; Related peptides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/04—Peptides having up to 20 amino acids in a fully defined sequence; Derivatives thereof
    • A61K38/12—Cyclic peptides, e.g. bacitracins; Polymyxins; Gramicidins S, C; Tyrocidins A, B or C
    • 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
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides

Definitions

  • the present invention relates to compounds that are active against Gram-positive and Gram-negative bacteria, their preparation process, pharmaceutical compositions containing them, and their use in the treatment of bacterial infections.
  • Antimicrobial peptides constitute a new class of therapeutic agents to which bacteria are not able to develop genetic resistance, since they act primarily on the lipid component of cell membranes.
  • polymyxin B which is approved for clinical use, is acquiring a new therapeutic relevance and is beginning to be considered as a representative of the class of active antibiotics against multiresistant bacteria.
  • Polymyxin B is an antibiotic lipopeptide isolated from Bacillus polymvxa. Its basic structure consists of a polycationic peptide cycle from which a tripeptide linked to a fatty acid chain hangs. Polymyxin B has resurfaced in medical practice in recent years and its use will continue to increase due to the limited development of new antibiotics by pharmaceutical companies and the increasing worldwide prevalence of nosocomial infections caused by multiresistant Gram-negative bacteria ("multidrug resistant ", MDR). Polymyxin B and other members of the polymyxin family are drugs that are used as a last resort to treat infections caused by multiresistant bacteria and are sometimes the only active antibiotic available. In addition, resistance to polymyxin is rare and generally adaptive and therefore reversible.
  • Polymyxin B is also capable of inhibiting the biological activity of bacterial lipopolysaccharide (LPS) through high affinity binding to lipid A, thus being the agent of choice for the treatment of septic shock induced by LPS.
  • LPS bacterial lipopolysaccharide
  • polymyxin B has no activity against Gram-positive or anaerobic bacteria.
  • polymyxins are of limited use because they have some nephrotoxicity and neurotoxicity.
  • the inventors have found new peptide compounds with antibiotic activity that act on the lipid component of bacterial membranes and that are active against both Gram-positive and Gram-negative bacteria. Structurally these compounds are characterized by containing some specific substituents among those mentioned in the application WO201 11 10716 and by the presence of a phenyl modified with fluorine atoms or methyl groups. Unexpectedly, these compounds have a greater activity against certain bacteria than those exhibiting the closest compounds of the state of the art described in the
  • WO201 11 10716 are also active against bacteria resistant to some conventional antibiotics.
  • one aspect of the present invention is related to providing compounds of formula (I),
  • R 0 is a radical CH 3 - (CH 2 ) m-, where m is an integer between 4 and 10;
  • R 2 is a -CH (CH 3 ) (OH) in the formula (I) being the CH of R 2 in either of the two configurations, R or S, giving rise to the corresponding epimers or a mixture thereof;
  • R 5 is a radical - (CH 2 ) -R 10 ;
  • R6 is a linear or branched C 4 -alkyl radical;
  • R 9 is a radical selected from -CONH 2 and -COOH;
  • R 10 is a phenyl radical substituted with one to 5 equal substitu
  • the CH of R 2 is in one of two configurations, R or S,
  • the compounds of formula (I) are those mentioned above where R 10 is a phenyl radical substituted with one to 5 equal substituents selected from the group consisting of F and CH 3.
  • the compounds of formula (1) are those where R 10 is a phenyl substituted with fluorine atoms.
  • the compounds of formula (I) are those where
  • R 10 is a phenyl substituted with methyl groups.
  • the compounds of formula (I) are those where m is an integer between 5 and 10.
  • the compounds of formula (I) are those where m is an integer between 7 and 10; n is an integer between 1 and 3; R 6 is a radical CH 3 - (CH 2 ) 3-; and R 9 is a radical -CONH 2 .
  • the compounds of formula (I) are those where R 0 is a radical CH 3 - (CH 2 ) 8--
  • the compounds of formula (I) are those where R ⁇ R 3 , R 4 , R 7 , and R 8 are equal radicals of formula -CH2-CH2-NH2.
  • D-amino acids The configuration of the D-amino acids has been indicated with a D. When the configuration is not indicated, it is understood that it is an L-amino acid.
  • cycle (S-S) means a macrocycle formed by a disulfide bond between the two cysteines.
  • the compounds of the present invention can be prepared by solid phase synthesis Fmoc / tBu.
  • the preparation procedure for each amino acid comprises the following steps: (i) several resin washes with ⁇ , ⁇ -dimethylformamide (DMF); (ii) treatment with 20% piperidine / DMF; (iii) washing with DMF several times; (iv) acylation with the amino acid Fmoc protected and 2- (1 H-benzotriazol-1-yl) -1, 1, 3,3-tetramethyluronium hexafluorophosphate / diisopropylethylamine (HBTU / DIEA) in DMF; (v) washing with DMF several times and then in dichloromethane (CH 2 CI 2 ) several times; and (vi) washed with DMF several times.
  • DMF ⁇ , ⁇ -dimethylformamide
  • HBTU / DIEA 2- (1 H-benzotriazol-1-yl) -1, 1, 3,3
  • the peptides obtained by the above procedures can be purified by preparative HPLC, whereby a purity equal to or greater than 90%, preferably equal to or greater than 95%, can be obtained.
  • the compounds of the present invention are easily prepared by chemical synthesis according to the procedure mentioned above while commercial polymyxin is obtained by fermentation.
  • Another aspect of the present invention are the compounds of formula (I), for use as antibacterial agents against Gram-positive bacteria.
  • Gram-positive bacteria are several of the well-known genus such as Mycobacterium phlei, Staphylococcus aureus,
  • MRSA methylcycline
  • Gram-positive bacteria are selected from
  • the Gram-positive bacteria is MRSA.
  • This aspect of the invention can also be formulated as the use of a compound as defined above, for the preparation of a medicament for the treatment of a bacterial infection caused by Gram-positive bacteria in a mammal, including a human.
  • the invention is also related to a method for the treatment and / or prophylaxis of a mammal, including a human, who suffers or is susceptible to bacterial infections caused by Gram-positive bacteria, in particular infections. mentioned above, the method comprises the administration to said patient of a therapeutically effective amount of a compound of formula (I), together with pharmaceutically acceptable excipients or carriers.
  • Gram-negative bacteria are selected from
  • the Escherichia coli bacteria is selected from the group consisting of Escherichia coli NDM-1, Escherichia coli VAL-10, Escherichia coli VAL-5 and Escherichia coli MAC21A and the Pseudomonas aeruginosa bacteria is selected from the group consisting of Pseudomonas aeruginosa 38a and Pseudomonas aeruginosa 38b.
  • This aspect of the invention can also be formulated as the use of a compound of formula (I) as defined above for the preparation of a medicament for the treatment of a bacterial infection caused by Gram-negative bacteria in a mammal, including a human.
  • the invention is also related to a method for the treatment and / or prophylaxis of a mammal, including a human who suffers or is susceptible to bacterial infections caused by Gram-negative bacteria, in particular to the infections mentioned above, the method comprises administration to said patient of a therapeutically effective amount of a compound of formula (I) as defined above, together with excipients or carriers
  • the compounds of the present invention can be used analogously to other known antibacterial agents. These can be used alone or in combination with other appropriate bioactive compounds.
  • the compounds of formula (I) can be used in the treatment of bacteremias and / or septicemia resulting from bacterial infections. Gram-negative, administered alone or in combination with conventional antibiotics. In another particular embodiment, the compounds of formula (I) can be used in the treatment of bacteremias and / or septicemia resulting from infections by Gram-positive bacteria, administered alone or in combination with conventional antibiotics.
  • a further aspect of the present invention is related to a pharmaceutical composition
  • a pharmaceutical composition comprising a therapeutically effective amount of the compounds of formula (I), together with appropriate amounts of excipients or carriers.
  • terapéuticaally effective amount refers to the amount of a compound that, when administered, is sufficient to prevent the development of, or relieve to some degree, one or more of the symptoms of the disease a The one that goes.
  • the particular dose of compound administered according to this invention will of course be determined by the particular conditions surrounding the case, including the compound administered, the route of administration, the particular condition being treated, and similar considerations.
  • pharmaceutically acceptable excipients or carriers refers to pharmaceutically acceptable materials, compositions or vehicles. Every
  • component must be pharmaceutically acceptable in the sense of being compatible with the other ingredients of the pharmaceutical composition. It must also be suitable for use in contact with tissues or organs of humans and animals without too much toxicity, irritation, allergic response, immunogenicity or other problems or complications in accordance with a reasonable benefit / risk ratio.
  • compositions may be prepared by combining the compounds of formula (I) of the present invention with pharmaceutically acceptable solid or liquid carriers or excipients, following standard pharmaceutical practices.
  • compositions of the present invention can be administered in a manner suitable for the disease to be treated, for example orally for the decontamination of the digestive tract prior to surgery, parenteral, inhalatory, rectal, transdermal or topical.
  • the pharmaceutical compositions of the present invention are administered parenterally or topically.
  • said pharmaceutical compounds or compositions are preferably administered at a dose to obtain or maintain a concentration that is effective as an antibacterial.
  • said amount or dose that is effective as an antibacterial will be in the approximate range of 0.1 to 100 mg / kg, more preferably about 3.0 to 50 mg / kg body weight / day. It is understood that the doses may vary depending on the requirements of the patient, the severity of the bacterial infection to be treated and the particular compound used.
  • Boc tert-butoxycarbonyl
  • Dab 2,3-diaminobutyric acid
  • DIEA N, N-diisopropylethylamine
  • DIPCDI N.N'-diisopropylcarbodiimide
  • DMF N, N-dimethylformamide
  • DMSO dimethylsulfoxide
  • Fmoc 9-fluorenylmethoxycarbonyl
  • HATU 2- (7-aza-1 H- benzotriazol-1-yl) -1, 1, 3,3-tetramethyluronium hexafluorophosphate
  • HBTU 2- (1 H-benzotriazol-1- and l) -1, 1, 3,3-tetramethyluronium hexafluorophosphate
  • HOBt 1-hydroxybenzotriazole
  • HPLC high performance liquid chromatography
  • MALDI-TOF ionization by laser desorption assisted by matrix-time of flight
  • MS mass ionization by
  • the Fmoc / tBu synthesis protocol for each synthetic cycle consists of the following steps: (i) washing the resin with DMF (5 x 30 s); (ii) treatment with 20% piperidine / DMF (1 x 1 min + 2 x 10 min, Fmoc deprotection); (iii) DMF wash (5 x 30 s); (iv) acylation with the amino acid Fmoc protected (3 times excess) and
  • the general method of de-anchoring and deprotection of the peptides consisted of a treatment with trifluoroacetic acid / 1,2-ethanedithiol / thioanisole / phenol / H 2 0 / triisopropylsilane (68.5 / 10/10/5 / 3.5 / 1, v / v, 3 ml / 100 mg of resin, 3h) in a reactor.
  • the TFA solution was filtered and the resin was collected. It was washed with two additional 0.5 ml portions of TFA mixture (0.5 mL) and the washings were combined.
  • the resulting solution was added over cold ether (TFA: ether in a 1: 20 ratio) to precipitate the peptide.
  • TFA cold ether
  • the peptide crude was washed with ether three times, it was decanted and the solid dried in air.
  • Each peptide crude was then dissolved in a 10% solution of DMSO in water at an approximate concentration of 1 mM or slightly lower for the formation of the disulfide bond.
  • the solution was stirred in an open system for 12-36 h. Delation took place by oxidation and was followed by HPLC and MS.
  • the peptides were characterized by MALDI-TOF mass spectrometry in a VOYAGER-DE (PerSeptive Biosystems) mass spectrometer.
  • the homogeneity of the purified peptides was checked by analytical HPLC using Merck or Kromasil C18 reverse phase columns (4 x 250 mm, 5 ⁇ particle diameter). Elution was carried out at 1 ml min "1 with mixtures of H 2 0- 0.05% TFA and acetonitrile-0.05% TFA and UV detection at 220 nm.
  • the antibacterial activity of synthetic lipopeptides was determined in sterile 96-well plates (Corning Costar 3598 microtiter plates) with a final volume of 200 ⁇ as follows: aliquots (100 ⁇ ) of a suspension of bacteria at a concentration of 10 5 colony forming units / ml_ in culture medium (MH, Muller Hinton Broth, Difco, USA) at pH 7.4, were added to 100 ⁇ _ of lipopeptide solution prepared from a stock solution in water of 1 mg / mL, in dilutions double dilution series in MH at pH 7.4 (Jorgensen & Turnide, 2003).
  • Bacterial growth inhibition was determined from absorbance at 492 nm in an Absorbance Microplate reader ELx 800 instrument (Bio-tek Instruments) after incubation at 37 ° C for 18-20 h.
  • the antibacterial activity was expressed as MIC, the concentration at which no growth is detected after 18-20 h of incubation.
  • the microorganisms were grown in Tryptycase Soy Broth (Pronadisa, Barcelona), incubating at 37 ° C until bacterial growth was observed. They were then seeded in Trypticase Soy Agar (Pronadisa, Barcelona) and incubated at 37 ° C until colony formation was observed. The microorganisms were stored in cryoballs (EAS Victoria, France) at -20 ° C.
  • the strains of the bacteria used to carry out the antibacterial activity test were obtained from: the American Type Culture Collection (ATCC, Rockville, MD, USA):
  • test in resistant and multiresistant bacteria was essentially the same as described above with small changes: samples of final volume of 100 uL were prepared as follows: aliquots (50 uL) of the bacterial suspension at concentration 1.5 x 10 6 forming units of colony per mL in MHB medium were adjusted to pH 7.4 and 50 uL of prepared antibiotic peptide solution was added of a stock solution of 1024 mg / L in water and diluted in factors of 2 serially in pH 7.4 MHB.
  • E. coli strains NDM-1 (multi-resistant); VAL10, VAL5 and MAC21 strains show intermediate resistance to quinolones, namely: MAC21 and VAL10 to nalidixic acid, cotrimoxazole and ampicillin; and VAL 5 to nalidixic acid.
  • P. aeruginosa 38a and 36a strains are highly resistant (to Ceftazidime,
  • Ciprofloxacin Imipenem and Piperacillin-tazobactam
  • NDM-1 New Delhi metallo-beta-lactamase
  • MIC Resistance to (MIC): Amoxicillin 256 mg / L
  • Amoxicillin clavulanate 32 mg / L;

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Abstract

The invention relates to compounds of formula (I), wherein R0 is a CH3-(CH2)m- radical, wherein m is an integer between 4 and 10; R1, R3, R4, R7 and R8 are independently selected radicals which have the following formula: GF-(CH2)n-; wherein n is an integer between 1 and 4; and GF is a radical selected among the group that consists of -NH2 and -NH-C(=NH)-NH2; R2 is a -CH(CH3)(OH), the CH of R2 being in either of the two configurations, R or S in the formula (I), giving rise to the corresponding epimers or to a mixture thereof; R5 is a -(CH2)-R10 radical; R6 is a straight or branched C4-alkyl radical; R9 is a radical selected among -CONH2 and -COOH; and R10 is a phenyl radical substituted with 1 to 5 identical substituents selected from the group that consists of F and (C1-C2)-alkyl, said compounds having been shown to be useful in the treatment of bacterial infections.

Description

Compuestos peptídicos útiles como agentes antibióticos  Peptide compounds useful as antibiotic agents
La presente invención está relacionada con compuestos que son activos contra bacterias Gram-positivas y Gram-negativas, su procedimiento de preparación, composiciones farmacéuticas que los contienen, y su uso en el tratamiento de infecciones bacterianas. The present invention relates to compounds that are active against Gram-positive and Gram-negative bacteria, their preparation process, pharmaceutical compositions containing them, and their use in the treatment of bacterial infections.
ESTADO DE LA TÉCNICA STATE OF THE TECHNIQUE
El hecho de que ciertos microorganismos patógenos se hayan convertido en resistentes a las terapias antibióticas es un problema grave de salud pública. Parte de este problema radica en el hecho de que ciertas bacterias y otros microorganismos infecciosos son extraordinariamente capaces de desarrollar resistencia a los antibióticos. Otra causa principal se debe al uso deficiente de los antibióticos en medicina, veterinaria y agricultura. The fact that certain pathogenic microorganisms have become resistant to antibiotic therapies is a serious public health problem. Part of this problem lies in the fact that certain bacteria and other infectious microorganisms are extraordinarily capable of developing resistance to antibiotics. Another main cause is due to the deficient use of antibiotics in medicine, veterinary medicine and agriculture.
Existe una preocupación a nivel mundial por la creciente prevalencia de infecciones causadas por bacterias multiresistentes, como por ejemplo Staphylococcus aureus resistente a metilicina, los Enterococcus resistentes a vancomincina y ciertas bacterias Gram-negativas como Pseudomonas aeruginosa, Acinetobacter baumannii, Klebsiella pneumoniae y Escherichia coli. Dichas infecciones son muy difíciles de controlar y una causa constante de enfermedad y mortandad. Los antibióticos convencionales actúan habitualmente sobre una o más proteínas o receptores diana y la resistencia genética aparece a una frecuencia que depende de muchos factores, tales como el número de dichas proteínas o receptores diana. There is worldwide concern about the increasing prevalence of infections caused by multiresistant bacteria, such as methylphycin-resistant Staphylococcus aureus, vancomincin-resistant Enterococcus and certain Gram-negative bacteria such as Pseudomonas aeruginosa, Acinetobacter baumannii, Klebsiella pneumoniae and Escherichia coli. Such infections are very difficult to control and a constant cause of disease and death. Conventional antibiotics usually act on one or more proteins or target receptors and genetic resistance appears at a frequency that depends on many factors, such as the number of said proteins or target receptors.
La continua aparición de cepas bacterianas resistentes a los antibióticos convencionales está llevando a enormes esfuerzos dirigidos al desarrollo de nuevos fármacos que actúen en la membrana bacteriana, como los péptidos antimicrobianos ("anti-microbial peptides", AMP). Los péptidos antimicrobianos constituyen una nueva clase de agentes terapéuticos a los cuales las bacterias no son capaces de desarrollar resistencia genética, puesto que actúan principalmente sobre el componente lipídico de las membranas celulares. Entre dichos compuestos, la polimixina B (PxB), que se halla aprobada para su uso clínico, está adquiriendo una nueva relevancia terapéutica y está empezando a ser considerado como un representante de la clase de antibióticos activos contra bacterias multiresistentes. Las polimixinas, en particular la polimixina B, constituyen una familia de antibióticos descubierta en 1947 con una elevada actividad contra bacterias Gram-negativas. La polimixina B es un lipopéptido antibiótico aislado de Bacillus polymvxa. Su estructura básica consiste en un ciclo peptídico policatiónico del cual pende un tripéptido unido a una cadena de ácido graso. La polimixina B ha resurgido en la práctica médica durante los últimos años y su uso continuará en aumento debido al escaso desarrollo de nuevos antibióticos por parte de la compañías farmacéuticas y a la creciente prevalencia mundial de infecciones nosocomiales causadas por bacterias Gram-negativas multiresistentes ("multidrug resistant", MDR). La polimixina B y otros miembros de la familia de las polimixinas son fármacos que se utilizan como último remedio para tratar infecciones causadas por bacterias multiresistentes y algunas veces son el único antibiótico activo disponible. Además, la resistencia a polimixina es rara y en general, adaptativa y, por tanto, reversible. La polimixina B es también capaz de inhibir la actividad biológica del lipopolisacárido (LPS) bacteriano por medio de la unión de alta afinidad al lípido A, siendo así el agente de elección para el tratamiento del shock séptico inducido por LPS. Desgraciadamente, la polimixina B no presenta actividad contra bacterias Gram-positivas o anaeróbicas. Además, las polimixinas son de uso limitado debido a que presentan cierta nefrotoxicidad y neurotoxicidad. The continued emergence of bacterial strains resistant to conventional antibiotics is leading to enormous efforts aimed at the development of new drugs that act on the bacterial membrane, such as antimicrobial peptides ("anti-microbial peptides", AMP). Antimicrobial peptides constitute a new class of therapeutic agents to which bacteria are not able to develop genetic resistance, since they act primarily on the lipid component of cell membranes. Among these compounds, polymyxin B (PxB), which is approved for clinical use, is acquiring a new therapeutic relevance and is beginning to be considered as a representative of the class of active antibiotics against multiresistant bacteria. Polymyxins, in particular polymyxin B, constitute a family of antibiotics discovered in 1947 with high activity against Gram-negative bacteria. Polymyxin B is an antibiotic lipopeptide isolated from Bacillus polymvxa. Its basic structure consists of a polycationic peptide cycle from which a tripeptide linked to a fatty acid chain hangs. Polymyxin B has resurfaced in medical practice in recent years and its use will continue to increase due to the limited development of new antibiotics by pharmaceutical companies and the increasing worldwide prevalence of nosocomial infections caused by multiresistant Gram-negative bacteria ("multidrug resistant ", MDR). Polymyxin B and other members of the polymyxin family are drugs that are used as a last resort to treat infections caused by multiresistant bacteria and are sometimes the only active antibiotic available. In addition, resistance to polymyxin is rare and generally adaptive and therefore reversible. Polymyxin B is also capable of inhibiting the biological activity of bacterial lipopolysaccharide (LPS) through high affinity binding to lipid A, thus being the agent of choice for the treatment of septic shock induced by LPS. Unfortunately, polymyxin B has no activity against Gram-positive or anaerobic bacteria. In addition, polymyxins are of limited use because they have some nephrotoxicity and neurotoxicity.
Más recientemente, se han descrito algunos compuestos lipidíeos basados en la estructura de la polimixina natural que tienen actividad antibiótica y actúan sobre el componente lipídico de las membranas bacterianas y que son activos tanto contra bacterias Gram-postivas como contra Gran negativas (cf. WO2010029196 y More recently, some lipid compounds based on the natural polymyxin structure that have antibiotic activity and act on the lipid component of bacterial membranes and that are active against both Gram-positive and Great negative bacteria have been described (cf. WO2010029196 and
WO2011 110716). WO2011 110716).
De lo que se conoce en el estado de la técnica se deriva que la búsqueda de nuevos agentes antibacterianos activos no sólo contra bacterias Gram-negativas sino también contra bacterias Gram-positivas es un campo activo y que todavía existe la necesidad de encontrar compuestos con actividad contra bacterias resistentes. From what is known in the state of the art it follows that the search for new active antibacterial agents not only against Gram-negative bacteria but also against Gram-positive bacteria is an active field and that there is still a need to find compounds with activity against resistant bacteria.
EXPLICACIÓN DE LA INVENCIÓN EXPLANATION OF THE INVENTION
Los inventores han encontrado nuevos compuestos peptídicos con actividad antibiótica que actúan sobre el componente lipídico de las membranas bacterianas y que son activos tanto contra bacterias Gram-positivas como contra Gram-negativas. Estructuralmente estos compuestos se caracterizan por contener algunos sustituyentes específicos de entre los mencionados en la solicitud WO201 11 10716 y por la presencia de un fenilo modificado con átomos de flúor o grupos metilo. Inesperadamente, estos compuestos presentan una actividad mayor frente a determinadas bacterias que la que presentan los compuestos más cercanos del estado de la técnica descritos en la The inventors have found new peptide compounds with antibiotic activity that act on the lipid component of bacterial membranes and that are active against both Gram-positive and Gram-negative bacteria. Structurally these compounds are characterized by containing some specific substituents among those mentioned in the application WO201 11 10716 and by the presence of a phenyl modified with fluorine atoms or methyl groups. Unexpectedly, these compounds have a greater activity against certain bacteria than those exhibiting the closest compounds of the state of the art described in the
WO201 11 10716 y además son activos frente a bacterias resistentes a algunos antibióticos convencionales. WO201 11 10716 and are also active against bacteria resistant to some conventional antibiotics.
Así, un aspecto de la presente invención está relacionado con proporcionar compuestos de fórmula (I), Thus, one aspect of the present invention is related to providing compounds of formula (I),
Figure imgf000005_0001
Figure imgf000005_0001
(i) donde: R0 es un radical CH3-(CH2)m-, donde m es un número entero entre 4 y 10; R3, R4, R7 y R8 son radicales seleccionados independientemente que tienen la fórmula siguiente: GF-(CH2)n-; donde n es un entero entre 1 y 4; y GF es un radical seleccionado entre el grupo que consiste en -NH2 y -NH-C(=NH)-NH2; R2 es un -CH(CH3)(OH) estando en la fórmula (I) el CH del R2 en cualquiera de las dos configuraciones, R o S, dando lugar a los correspondientes epímeros o a una mezcla de los mismos; R5 es un radical -(CH2)-R10; R6 es una radical C4-alquilo lineal o ramificado; R9 es un radical seleccionado entre -CONH2 y -COOH; y R10 es un radical fenilo sustituido con uno a 5 sustituyentes iguales seleccionados del grupo que consiste en F, (C C2)-alquilo y CF3. (i) where: R 0 is a radical CH 3 - (CH 2 ) m-, where m is an integer between 4 and 10; R 3 , R 4 , R 7 and R 8 are independently selected radicals having the following formula: GF- (CH 2 ) n -; where n is an integer between 1 and 4; and GF is a radical selected from the group consisting of -NH 2 and -NH-C (= NH) -NH 2 ; R 2 is a -CH (CH 3 ) (OH) in the formula (I) being the CH of R 2 in either of the two configurations, R or S, giving rise to the corresponding epimers or a mixture thereof; R 5 is a radical - (CH 2 ) -R 10 ; R6 is a linear or branched C 4 -alkyl radical; R 9 is a radical selected from -CONH 2 and -COOH; and R 10 is a phenyl radical substituted with one to 5 equal substituents selected from the group consisting of F, (CC 2 ) -alkyl and CF 3 .
En una realización particular, en la fórmula (I) el CH del R2 está en una de las dos configuraciones, R o S, En una realización preferida, los compuestos de fórmula (I) son aquéllos anteriormente mencionados donde R10 es un radical fenilo sustituido con uno a 5 sustituyentes iguales seleccionados del grupo que consiste en F y CH3. In a particular embodiment, in formula (I) the CH of R 2 is in one of two configurations, R or S, In a preferred embodiment, the compounds of formula (I) are those mentioned above where R 10 is a phenyl radical substituted with one to 5 equal substituents selected from the group consisting of F and CH 3.
En una realización más preferida, los compuestos de fórmula (l) son aquéllos donde R10 es un fenilo sustituido con átomos de flúor. En una realización todavía más preferida, los compuestos de fórmula (I) son aquéllos donde In a more preferred embodiment, the compounds of formula (1) are those where R 10 is a phenyl substituted with fluorine atoms. In an even more preferred embodiment, the compounds of formula (I) are those where
R10 es un fenilo sustituido con grupos metilo. R 10 is a phenyl substituted with methyl groups.
En otra realización preferida, los compuestos de fórmula (I) son aquéllos donde m es un número entero entre 5 y 10. In another preferred embodiment, the compounds of formula (I) are those where m is an integer between 5 and 10.
En otra realización preferida, los compuestos de fórmula (I) son aquéllos donde m es un número entero entre 7 y 10; n es un entero entre 1 y 3; R6 es una radical CH3-(CH2)3-; y R9 es un radical -CONH2. In another preferred embodiment, the compounds of formula (I) are those where m is an integer between 7 and 10; n is an integer between 1 and 3; R 6 is a radical CH 3 - (CH 2 ) 3-; and R 9 is a radical -CONH 2 .
En otra realización preferida, los compuestos de fórmula (I) son aquéllos donde R0 es un radical CH3-(CH2)8-- In another preferred embodiment, the compounds of formula (I) are those where R 0 is a radical CH 3 - (CH 2 ) 8--
En otra realización preferida, los compuestos de fórmula (I) son aquéllos donde R^ R3, R4, R7, y R8 son radicales iguales de fórmula -CH2-CH2-NH2. In another preferred embodiment, the compounds of formula (I) are those where R ^ R 3 , R 4 , R 7 , and R 8 are equal radicals of formula -CH2-CH2-NH2.
Los compuestos más preferidos de fórmula (I) se seleccionan de entre los siguientes: The most preferred compounds of formula (I) are selected from the following:
Decanoil-Dab-Thr-Dab-ciclo(S-S)[Cys-Dab-(p-fluoro)DPhe-Nle-Dab-Dab-DCys], Decanoyl-Dab-Thr-Dab-cycle (S-S) [Cys-Dab- (p-fluoro) DPhe-Nle-Dab-Dab-DCys],
Decanoil-Dab-Thr-Dab-ciclo(S-S)[Cys-Dab-(pentafluoro)DPhe-Nle-Dab-Dab-DCys], y Decanoyl-Dab-Thr-Dab-cycle (S-S) [Cys-Dab- (pentafluoro) DPhe-Nle-Dab-Dab-DCys], and
Decanoil-Dab-Thr-Dab-ciclo(S-S)[Cys-Dab-(4-metil)DPhe-Nle-Dab-Dab-DCys]. Decanoyl-Dab-Thr-Dab-cycle (S-S) [Cys-Dab- (4-methyl) DPhe-Nle-Dab-Dab-DCys].
La configuración de los D-aminoácidos se ha indicado con una D. Cuando no se indica la configuración se entiende que se trata de un L-aminoácido. The configuration of the D-amino acids has been indicated with a D. When the configuration is not indicated, it is understood that it is an L-amino acid.
Por el término "ciclo(S-S)" se entiende un macrociclo formado por un enlace disulfuro entre las dos cisteínas. The term "cycle (S-S)" means a macrocycle formed by a disulfide bond between the two cysteines.
Los compuestos de la presente invención pueden prepararse por síntesis en fase sólida Fmoc/tBu. El procedimiento de preparación para cada aminoácido comprende las etapas siguientes: (i) varios lavados de la resina con Ν,Ν-dimetilformamida (DMF); (ii) tratamiento con 20% de piperidina/DMF; (iii) lavado con DMF varias veces; (iv) acilación con el aminoácido Fmoc protegido y 2-(1 H-benzotriazol-1-il)-1 , 1 ,3,3-tetrametiluronio hexafluorofosfato/diisopropiletilamina (HBTU/DIEA) en DMF; (v) lavado con DMF varias veces y posteriormente en diclorometano (CH2CI2) varias veces; y (vi) lavado con DMF varias veces. The compounds of the present invention can be prepared by solid phase synthesis Fmoc / tBu. The preparation procedure for each amino acid comprises the following steps: (i) several resin washes with Ν, Ν-dimethylformamide (DMF); (ii) treatment with 20% piperidine / DMF; (iii) washing with DMF several times; (iv) acylation with the amino acid Fmoc protected and 2- (1 H-benzotriazol-1-yl) -1, 1, 3,3-tetramethyluronium hexafluorophosphate / diisopropylethylamine (HBTU / DIEA) in DMF; (v) washing with DMF several times and then in dichloromethane (CH 2 CI 2 ) several times; and (vi) washed with DMF several times.
Los péptidos obtenidos por los procedimientos anteriores pueden purificarse por HPLC preparativa, mediante lo cual puede obtenerse una pureza igual o superior al 90%, preferiblemente igual o superior al 95%. The peptides obtained by the above procedures can be purified by preparative HPLC, whereby a purity equal to or greater than 90%, preferably equal to or greater than 95%, can be obtained.
Así, los compuestos de la presente invención se preparan fácilmente por síntesis química según el procedimiento mencionado anteriormente mientras que la polimixina comercial se obtiene por fermentación. Thus, the compounds of the present invention are easily prepared by chemical synthesis according to the procedure mentioned above while commercial polymyxin is obtained by fermentation.
Otro aspecto de la presente invención son los compuestos de fórmula (I), para uso como agentes antibacterianos contra bacterias Gram-positivas. Another aspect of the present invention are the compounds of formula (I), for use as antibacterial agents against Gram-positive bacteria.
Entre las bacterias Gram-positivas médicamente relevantes se encuentran varias de género bien conocido tales como Micobacterium phlei, Staphylococcus aureus, Among the medically relevant Gram-positive bacteria are several of the well-known genus such as Mycobacterium phlei, Staphylococcus aureus,
Enterococus faecalis y Staphylococcus aureus resistente a metilicina (MRSA). Enterococus faecalis and Staphylococcus aureus resistant to methylcycline (MRSA).
En una realización particular, las bacterias Gram-positivas se seleccionan entre In a particular embodiment, Gram-positive bacteria are selected from
Micobacterium phlei ATCC41423, Staphylococcus aureus ATCC 29213, Mycobacterium phlei ATCC41423, Staphylococcus aureus ATCC 29213,
Enterococus faecalis ATCC 29212 y Staphylococcus aureus resistente a metilicina ATCC 43300. Enterococus faecalis ATCC 29212 and Staphylococcus aureus resistant to methylCCine ATCC 43300.
En una realización preferida, la bacteria Gram-positiva es MRSA. In a preferred embodiment, the Gram-positive bacteria is MRSA.
Este aspecto de la invención se puede formular también como el uso de un compuesto como se ha definido anteriormente, para la preparación de un medicamento para el tratamiento de una infección bacteriana causada por bacterias Gram-positivas en un mamífero, incluyendo un humano. This aspect of the invention can also be formulated as the use of a compound as defined above, for the preparation of a medicament for the treatment of a bacterial infection caused by Gram-positive bacteria in a mammal, including a human.
La invención también está relacionada con un método para el tratamiento y/o la profilaxis de un mamífero, incluyendo un humano, que sufre o es susceptible a infecciones bacterianas causadas por bacterias Gram-positivas, en particular a las infecciones mencionadas anteriormente, el método comprende la administración al mencionado paciente de una cantidad terapéuticamente efectiva de un compuesto de fórmula (I), junto a excipientes o portadores farmacéuticamente aceptables. The invention is also related to a method for the treatment and / or prophylaxis of a mammal, including a human, who suffers or is susceptible to bacterial infections caused by Gram-positive bacteria, in particular infections. mentioned above, the method comprises the administration to said patient of a therapeutically effective amount of a compound of formula (I), together with pharmaceutically acceptable excipients or carriers.
Es también parte de la invención los compuestos de fórmula (I) como se han definido anteriormente para su uso como agentes antibacterianos contra bacterias Gram- negativas. Las bacterias Gram-negativas médicamente relevantes comprenden Compounds of formula (I) as defined above for use as antibacterial agents against Gram-negative bacteria are also part of the invention. Medically relevant Gram-negative bacteria comprise
Pseudomonas aeruginosa, Escherichia coli y Acinetobacter sp. Pseudomonas aeruginosa, Escherichia coli and Acinetobacter sp.
En una realización particular, las bacterias Gram-negativas se seleccionan entre In a particular embodiment, Gram-negative bacteria are selected from
Pseudomonas aeruginosa ATCC 27853, Escherichia coli ATCC 25922 y Acinetobacter sp ATCC 5798. Pseudomonas aeruginosa ATCC 27853, Escherichia coli ATCC 25922 and Acinetobacter sp ATCC 5798.
En una realización preferida, la bacteria Escherichia coli se selecciona del grupo que consiste en Escherichia coli NDM-1 , Escherichia coli VAL-10, Escherichia coli VAL-5 y Escherichia coli MAC21A y la bacteria Pseudomonas aeruginosa se selecciona del grupo que consiste en Pseudomonas aeruginosa 38a y Pseudomonas aeruginosa 38b. In a preferred embodiment, the Escherichia coli bacteria is selected from the group consisting of Escherichia coli NDM-1, Escherichia coli VAL-10, Escherichia coli VAL-5 and Escherichia coli MAC21A and the Pseudomonas aeruginosa bacteria is selected from the group consisting of Pseudomonas aeruginosa 38a and Pseudomonas aeruginosa 38b.
Este aspecto de la invención puede ser también formulado como el uso de un compuesto de fórmula (I) como se ha definido anteriormente para la preparación de un medicamento para el tratamiento de una infección bacteriana causada por bacterias Gram-negativas en un mamífero, incluyendo un humano. This aspect of the invention can also be formulated as the use of a compound of formula (I) as defined above for the preparation of a medicament for the treatment of a bacterial infection caused by Gram-negative bacteria in a mammal, including a human.
La invención también está relacionada con un método para el tratamiento y/o la profilaxis de un mamífero, incluyendo un humano que sufre o es susceptible a infecciones bacterianas causadas por bacterias Gram-negativas, en particular a las infecciones mencionadas anteriormente, el método comprende la administración al mencionado paciente de una cantidad terapéuticamente efectiva de un compuesto de fórmula (I) como se han definido anteriormente, junto a excipientes o portadores The invention is also related to a method for the treatment and / or prophylaxis of a mammal, including a human who suffers or is susceptible to bacterial infections caused by Gram-negative bacteria, in particular to the infections mentioned above, the method comprises administration to said patient of a therapeutically effective amount of a compound of formula (I) as defined above, together with excipients or carriers
farmacéuticamente aceptables. pharmaceutically acceptable.
Los compuestos de la presente invención se pueden utilizar análogamente a otros agentes antibacterianos conocidos. Estos se pueden utilizar solos o en combinación con otros compuestos bioactivos apropiados. The compounds of the present invention can be used analogously to other known antibacterial agents. These can be used alone or in combination with other appropriate bioactive compounds.
En una realización particular, los compuestos de fórmula (I) se pueden utilizar en el tratamiento de bacteremias y/o septicemia consecuencia de infecciones por bacterias Gram-negativas, administrados solos o en combinación con antibióticos convencionales. En otra realización particular, los compuestos de fórmula (I) se pueden utilizar en el tratamiento de bacteremias y/o septicemia consecuencia de infecciones por bacterias Gram-positivas, administrados solos o en combinación con antibióticos convencionales. In a particular embodiment, the compounds of formula (I) can be used in the treatment of bacteremias and / or septicemia resulting from bacterial infections. Gram-negative, administered alone or in combination with conventional antibiotics. In another particular embodiment, the compounds of formula (I) can be used in the treatment of bacteremias and / or septicemia resulting from infections by Gram-positive bacteria, administered alone or in combination with conventional antibiotics.
Un aspecto adicional de la presente invención está relacionado con una composición farmacéutica que comprende una cantidad terapéuticamente efectiva de los compuestos de fórmula (I), junto con cantidades apropiadas de excipientes o portadores A further aspect of the present invention is related to a pharmaceutical composition comprising a therapeutically effective amount of the compounds of formula (I), together with appropriate amounts of excipients or carriers.
farmacéuticamente aceptables. pharmaceutically acceptable.
La expresión "cantidad terapéuticamente efectiva" como se usa aquí, se refiere a la cantidad de un compuesto que, cuando se administra, es suficiente para prevenir el desarrollo de, o aliviar en cierto grado, uno o más de los síntomas de la enfermedad a la que se dirige. La dosis particular de compuesto administrado según esta invención será por supuesto determinada por las condiciones particulares que rodean el caso, incluyendo el compuesto administrado, la ruta de administración, la condición particular que se está tratando, y las consideraciones similares. The term "therapeutically effective amount" as used herein, refers to the amount of a compound that, when administered, is sufficient to prevent the development of, or relieve to some degree, one or more of the symptoms of the disease a The one that goes. The particular dose of compound administered according to this invention will of course be determined by the particular conditions surrounding the case, including the compound administered, the route of administration, the particular condition being treated, and similar considerations.
La expresión "excipientes o portadores farmacéuticamente aceptables" se refiere a materiales farmacéuticamente aceptables, composiciones o vehículos. Cada The term "pharmaceutically acceptable excipients or carriers" refers to pharmaceutically acceptable materials, compositions or vehicles. Every
componente debe ser farmacéuticamente aceptable en el sentido de ser compatible con los otros ingredientes de la composición farmacéutica. Debe ser también apto para el uso en contacto con tejidos u órganos de humanos y animales sin demasiada toxicidad, irritación, respuesta alérgica, immunogenicidad u otros problemas o complicaciones acorde con una relación beneficio/riesgo razonable. component must be pharmaceutically acceptable in the sense of being compatible with the other ingredients of the pharmaceutical composition. It must also be suitable for use in contact with tissues or organs of humans and animals without too much toxicity, irritation, allergic response, immunogenicity or other problems or complications in accordance with a reasonable benefit / risk ratio.
Las composiciones farmacéuticas se pueden preparar por combinación de los compuestos de fórmula (I) de la presente invención con excipientes o portadores sólidos o líquidos farmacéuticamente aceptables, siguiendo prácticas farmacéuticas estándar. Pharmaceutical compositions may be prepared by combining the compounds of formula (I) of the present invention with pharmaceutically acceptable solid or liquid carriers or excipients, following standard pharmaceutical practices.
Las composiciones farmacéuticas de la presente invención se pueden administrar de manera adecuada para la enfermedad que se desea tratar, por ejemplo por vía oral para la descontaminación del tracto digestivo previa a cirugía, parenteral, inhalatoria, rectal, transdérmica o tópica. The pharmaceutical compositions of the present invention can be administered in a manner suitable for the disease to be treated, for example orally for the decontamination of the digestive tract prior to surgery, parenteral, inhalatory, rectal, transdermal or topical.
En una realización preferida, las composiciones farmacéuticas de la presente invención se administran por vía parenteral o por vía tópica. En el uso terapéutico para tratar o combatir infecciones bacterianas dichos compuestos o composiciones farmacéuticas, preferentemente, se administran a una dosis para obtener o mantener una concentración que sea efectiva como antibacteriano. Por ejemplo, por vía parenteral dicha cantidad o dosis que sea efectiva como antibacteriano se encontrará en el intervalo aproximado de 0.1 a 100 mg/Kg, más preferentemente alrededor de 3.0 a 50 mg/Kg de peso corporal/día. Se entiende que las dosis pueden variar dependiendo de los requisitos del paciente, la severidad de la infección bacteriana a tratar y del compuesto particular que se use. In a preferred embodiment, the pharmaceutical compositions of the present invention are administered parenterally or topically. In therapeutic use to treat or combat bacterial infections, said pharmaceutical compounds or compositions are preferably administered at a dose to obtain or maintain a concentration that is effective as an antibacterial. For example, parenterally, said amount or dose that is effective as an antibacterial will be in the approximate range of 0.1 to 100 mg / kg, more preferably about 3.0 to 50 mg / kg body weight / day. It is understood that the doses may vary depending on the requirements of the patient, the severity of the bacterial infection to be treated and the particular compound used.
A lo largo de la descripción y las reivindicaciones la palabra "comprende" y sus variantes no pretenden excluir otras características técnicas, aditivos, componentes o pasos. Para los expertos en la materia, otros objetos, ventajas y características de la invención se desprenderán en parte de la descripción y en parte de la práctica de la invención. Los siguientes ejemplos se proporcionan a modo de ilustración, y no se pretende que sean limitativos de la presente invención. Además, la presente invención cubre todas las posibles combinaciones de realizaciones particulares y preferidas aquí indicadas. Throughout the description and the claims the word "comprises" and its variants are not intended to exclude other technical characteristics, additives, components or steps. For those skilled in the art, other objects, advantages and features of the invention will be derived partly from the description and partly from the practice of the invention. The following examples are provided by way of illustration, and are not intended to be limiting of the present invention. In addition, the present invention covers all possible combinations of particular and preferred embodiments indicated herein.
EJEMPLOS EXAMPLES
Abreviaciones: Boc, tert-butoxicarbonilo; Dab: ácido 2,3-diaminobutírico; DIEA, N,N- diisopropiletilamina; DIPCDI, N.N'-diisopropilcarbodiimida; DMF, N,N-dimetillformamida; DMSO, dimetilsulfoxido; Fmoc, 9-fluorenylmetoxicarbonilo; HATU, 2-(7-aza-1 H- benzotriazol-1-il)-1 ,1 ,3,3-tetrametiluronio hexafluorofosfato; HBTU, 2-(1 H-benzotriazol-1- yl)-1 , 1 ,3,3-tetrametiluronio hexafluorofosfato; HOBt, 1-hidroxibenzotriazol; HPLC, cromatografía líquida de alta resolución; MALDI-TOF, ionización por desorción láser asistida por matriz-tiempo de vuelo; MS, espectrometría de masas; CMI, concentración mínima inhibitoria; MW, peso molecular; tBu, tert-butilo; TFA, ácido trifluoroacético; Trt, trifilo; MHB: Múller-Hinton broth. Abbreviations: Boc, tert-butoxycarbonyl; Dab: 2,3-diaminobutyric acid; DIEA, N, N-diisopropylethylamine; DIPCDI, N.N'-diisopropylcarbodiimide; DMF, N, N-dimethylformamide; DMSO, dimethylsulfoxide; Fmoc, 9-fluorenylmethoxycarbonyl; HATU, 2- (7-aza-1 H- benzotriazol-1-yl) -1, 1, 3,3-tetramethyluronium hexafluorophosphate; HBTU, 2- (1 H-benzotriazol-1- and l) -1, 1, 3,3-tetramethyluronium hexafluorophosphate; HOBt, 1-hydroxybenzotriazole; HPLC, high performance liquid chromatography; MALDI-TOF, ionization by laser desorption assisted by matrix-time of flight; MS, mass spectrometry; MIC, minimum inhibitory concentration; MW, molecular weight; tBu, tert-butyl; TFA, trifluoroacetic acid; Trt, trifilo; MHB: Múller-Hinton Broth.
Métodos generales General methods
Se ha empleado el protocolo general de síntesis en fase sólida Fmoc/tBu para preparar los compuestos de los ejemplos. The general solid phase synthesis protocol Fmoc / tBu has been used to prepare the compounds of the examples.
El protocolo de síntesis Fmoc/tBu para cada ciclo sintético consiste en las etapas siguientes: (i) lavado de la resina con DMF (5 x 30 s); (ii) tratamiento con 20% de piperidina/DMF(1 x 1 min + 2 x 10 min, desprotección Fmoc); (iii) lavado con DMF (5 x 30 s); (iv) acilación con el aminoácido Fmoc protegido (3 veces de exceso) y The Fmoc / tBu synthesis protocol for each synthetic cycle consists of the following steps: (i) washing the resin with DMF (5 x 30 s); (ii) treatment with 20% piperidine / DMF (1 x 1 min + 2 x 10 min, Fmoc deprotection); (iii) DMF wash (5 x 30 s); (iv) acylation with the amino acid Fmoc protected (3 times excess) and
HBTU/DIEA (3:6 veces de exceso, respectivamente) en la cantidad mínima de DMF; (v) lavado con DMF (5 x 30 s) y CH2CI2 (5 x 30 s); (vi) prueba de Kaiser (con una muestra de resina peptídica); (vii) lavado con DMF (5 x 30 s). HBTU / DIEA (3: 6 times excess, respectively) in the minimum amount of DMF; (v) washing with DMF (5 x 30 s) and CH 2 CI 2 (5 x 30 s); (vi) Kaiser test (with a sample of peptide resin); (vii) DMF wash (5 x 30 s).
El método general de desanclaje y desprotección de los péptidos consistió en un tratamiento con ácido trifluoroacético/1 ,2-etanoditiol/tioanisol/fenol/H20 /triisopropilsilano (68.5/10/10/5/3.5/1 , v/v, 3 ml/100 mg de resina, 3h) en un reactor. La solución de TFA se filtró y la resina se recogió. Se lavó con dos porciones adicionales de 0.5 mi de mezcla de TFA (0.5 mL) y los lavados se juntaron. La solución resultante se añadió sobre éter frío (TFA:éter en proporción 1 :20) para precipitar el péptido. El crudo peptídico se lavó con éter tres veces, éste se decantó y el sólido se seco al aire. The general method of de-anchoring and deprotection of the peptides consisted of a treatment with trifluoroacetic acid / 1,2-ethanedithiol / thioanisole / phenol / H 2 0 / triisopropylsilane (68.5 / 10/10/5 / 3.5 / 1, v / v, 3 ml / 100 mg of resin, 3h) in a reactor. The TFA solution was filtered and the resin was collected. It was washed with two additional 0.5 ml portions of TFA mixture (0.5 mL) and the washings were combined. The resulting solution was added over cold ether (TFA: ether in a 1: 20 ratio) to precipitate the peptide. The peptide crude was washed with ether three times, it was decanted and the solid dried in air.
Cada crudo peptídico se disolvió a continuación en una solución de DMSO en agua al 10% a una concentración aproximada de 1 mM o ligeramente inferior para la formación del enlace disulfuro. La solución se agitó en sistema abierto durante 12-36 h. La delación tuvo lugar por oxidación y se siguió por HPLC y MS. Each peptide crude was then dissolved in a 10% solution of DMSO in water at an approximate concentration of 1 mM or slightly lower for the formation of the disulfide bond. The solution was stirred in an open system for 12-36 h. Delation took place by oxidation and was followed by HPLC and MS.
La purificación se llevó a cabo por HPLC preparativa. En los Ejemplos se empleó un equipo a Varían Pro-star 200 prep-system con una columna Varían Load&lock 1 pulgada C18 (250 x 1 pulgada., 10 μηι) y se eluyó en gradiente con mezclas de H20-acetonitrilo- 0.05% TFA y detección UV a 220 nm. Purification was carried out by preparative HPLC. In the Examples a Varian Pro-star 200 prep-system device was used with a Varian Load & lock 1 inch C18 column (250 x 1 inch, 10 μηι) and eluted in gradient with mixtures of H 2 0-acetonitrile- 0.05% TFA and UV detection at 220 nm.
Los péptidos se caracterizaron por espectrometría de masas MALDI-TOF en un espectrómetro de masas VOYAGER-DE (PerSeptive Biosystems). The peptides were characterized by MALDI-TOF mass spectrometry in a VOYAGER-DE (PerSeptive Biosystems) mass spectrometer.
La homogeneidad de los péptidos purificados se comprobó por HPLC analítica empleando columnas Merck o Kromasil C18 de fase reversa (4 x 250 mm, 5 μηι de diámetro de partícula). La elución se llevó a cabo a 1 ml min"1 con mezclas de H20- 0.05% TFA y acetonitrilo-0.05% TFA y detección UV a 220 nm. The homogeneity of the purified peptides was checked by analytical HPLC using Merck or Kromasil C18 reverse phase columns (4 x 250 mm, 5 μηι particle diameter). Elution was carried out at 1 ml min "1 with mixtures of H 2 0- 0.05% TFA and acetonitrile-0.05% TFA and UV detection at 220 nm.
Ejemplo 1 : Preparación de decanoil-Dab-Thr-Dab-ciclo(S-S)[Cvs-Dab-(p-fluoro)DPhe- Nle-Dab-Dab-DCvsl (compuesto de fórmula (I) con Rn = CH¾(CH?)fi-, Ri = -CH?CH?NH? (cadena lateral de Dab), R? = -CH(CH¾)OH, (cadena lateral de Thr), R¾ = -CH?CH?NH? (cadena lateral de Dab), R = -CH?CH?NH? (cadena lateral de Dab), Rs =- CHp para- fluoro)Ph (cadena lateral de D(p-fluoroPhe), RR = -CH?CH?CH?CH¾ (cadena lateral de Nle), R7 = -CH?CH?NH? (cadena lateral de Dab), y Rfi = -CH?CH?NH? (cadena lateral de Dab) Example 1: Preparation of decanoyl-Dab-Thr-Dab-cycle (SS) [Cvs-Dab- (p-fluoro) DPhe-Nle-Dab-Dab-DCvsl (compound of formula (I) with R n = CH¾ (CH ? ) fi -, Ri = -CH ? CH ? NH ? (Dab side chain), R? = -CH (CH ¾ ) OH, (Thr side chain), R ¾ = -CH ? CH ? NH ? ( Dab side chain), R = -CH? CH? NH? (Dab side chain), Rs = - CHp para-fluoro) Ph (D side chain (p-fluoroPhe), R R = -CH ? CH? CH ? CH¾ (side chain of Nle), R7 = -CH ? CH ? NH ? (side chain of Dab), and R fi = -CH ? CH ? NH ? (Dab side chain)
Aminoácidos protegidos: Fmoc-Cys(Trt)-OH, Fmoc-Dab(Boc)-OH, Fmoc-Nle-OH, Fmoc- (para-fluoro)DPhe-OH, Fmoc-Thr(tBu)-OH, Fmoc-DCys(Trt)-OH. Protected amino acids: Fmoc-Cys (Trt) -OH, Fmoc-Dab (Boc) -OH, Fmoc-Nle-OH, Fmoc- (para-fluoro) DPhe-OH, Fmoc-Thr (tBu) -OH, Fmoc-DCys (Trt) -OH.
Se llevo a cabo la síntesis manual según un protocolo estándar Fmoc/'Bu en jeringas de polipropileno con un filtro de polietileno. Se utilizó la resina Fmoc-Rink amida MBHA (300mg, 0.144 mmoles, f= 0.48mmol/g de resina). Los aminoácidos de la secuencia se introdujeron según un protocolo estándar de síntesis en fase sólida Fmoc/'Bu como se ha descrito anteriormente. Una vez la secuencia se completó, se introdujo el ácido graso por medio de cloruro de decanoilo (110 μΙ, 0.576 mmol, 4 veces de exceso) con 3M DIEA (8 veces de exceso) en la cantidad mínima de DMF. Una vez concluida la reacción, la resina se lavó con DMF (5 x 30 s) y CH2CI2 (5 x 30 s). Manual synthesis was carried out according to a standard Fmoc / 'Bu protocol in polypropylene syringes with a polyethylene filter. The FHAc-Rink amide MBHA resin (300mg, 0.144 mmol, f = 0.48mmol / g resin) was used. The amino acids in the sequence were introduced according to a standard Fmoc / 'Bu solid phase synthesis protocol as described above. Once the sequence was completed, the fatty acid was introduced by means of decanoyl chloride (110 μΙ, 0.576 mmol, 4 times excess) with 3M DIEA (8 times excess) in the minimum amount of DMF. After completion of the reaction, the resin was washed with DMF (5 x 30 s) and CH 2 CI 2 (5 x 30 s).
El peso del crudo peptídico tras el desanclaje y desprotección según el método indicado anteriormente fue de 170mg (rendimiento 72%). La purificación por HPLC preparativa rindió 28 mg de péptido puro (rendimiento 16%). The weight of the peptide crude after de-anchoring and deprotection according to the method indicated above was 170mg (72% yield). Purification by preparative HPLC yielded 28 mg of pure peptide (16% yield).
Caracterización del péptido purificado: Homogeneidad (por integración del área de la traza de HPLC) >98%; MALDI-TOF: m/z 1257.29 ([M+H+], 32%), 1279.29 ([M+Na+], 100%). MW 1256.29. Characterization of the purified peptide: Homogeneity (by integration of the HPLC trace area)>98%; MALDI-TOF: m / z 1257.29 ([M + H + ], 32%), 1279.29 ([M + Na + ], 100%). MW 1256.29.
Ejemplo 2: Preparación de decanoil-Dab-Thr-Dab-cyclo(S-S)[Cvs-Dab- (pentafluoro)DPhe-Nle-Dab-Dab-DCysl, (compuesto de fórmula (I) con R„ = CH¾(CH?)g-, Ri =-CH?CH?NH? (cadena lateral de Dab), R? = -CH(CH¾)OH, (cadena lateral de Thr), Ra = -CH?CH?NH? (cadena lateral de Dab), R¿ = -CH?CH?NH? (cadena lateral de Dab), Rfi =-CH?(pentafluoro)Ph (cadena lateral de pentafluoroDPhe), RR = -CH?CH?CH?CH¾ (cadena lateral de Nle), R7 = -CH?CH?NH? (cadena lateral de Dab), y Rfi = -CH?CH?NH? (cadena lateral de Dab) Example 2: Preparation of decanoyl-Dab-Thr-Dab-cyclo (SS) [Cvs-Dab- (pentafluoro) DPhe-Nle-Dab-Dab-DCysl, (compound of formula (I) with R "= CH" (CH? ) g-, Ri = -CH ? CH ? NH ? (Dab side chain), R? = -CH (CH ¾ ) OH, (Thr side chain), Ra = -CH ? CH ? NH ? (side chain of Dab), R ¿= -CH ? CH ? NH ? (Dab side chain), Rfi = -CH? (pentafluoro) Ph (pentafluoroDPhe side chain), R R = -CH? CH ? CH ? CH¾ (chain Nle side), R 7 = -CH ? CH ? NH ? (Dab side chain), and R fi = -CH ? CH ? NH ? (Dab side chain)
Aminoácidos protegidos: Fmoc-Cys(Trt)-OH, Fmoc-Dab(Boc)-OH, Fmoc-Nle-OH, Fmoc- pentafluoroDPhe-OH, Fmoc-Thr(tBu)-OH, Fmoc-DCys(Trt)-OH. Protected amino acids: Fmoc-Cys (Trt) -OH, Fmoc-Dab (Boc) -OH, Fmoc-Nle-OH, Fmoc- pentafluoroDPhe-OH, Fmoc-Thr (tBu) -OH, Fmoc-DCys (Trt) -OH .
Se llevo a cabo la síntesis manual según un protocolo estándar Fmoc/'Bu sobre resina Fmoc-Rink Amida MBHA como se ha descrito anteriormente en el Ejemplo 1. El peso del crudo peptídico tras el desanclaje y formación del enlace disulfuro fue 240 mg (rendimiento 94%). La purificación por HPLC preparativa proporcionó 45 mg de péptido puro (rendimiento 19%). Manual synthesis was carried out according to a standard Fmoc / 'Bu protocol on Fmoc-Rink Amida MBHA resin as described above in Example 1. The weight of the peptide crude after the de-anchoring and formation of the disulfide bond was 240 mg (94% yield). Purification by preparative HPLC provided 45 mg of pure peptide (19% yield).
Caracterización del péptido purificado: Homogeneidad (por integración del área de la traza de HPLC) >99%; MALDI-TOF: m/z 1328.44 ([M+H]+, 100%), 1350.43 ([M+Na]+, 40%). MW 1327.48. Characterization of the purified peptide: Homogeneity (by integration of the HPLC trace area)>99%; MALDI-TOF: m / z 1328.44 ([M + H] + , 100%), 1350.43 ([M + Na] + , 40%). MW 1327.48.
Ejemplo 3: Preparación de decanoil-Dab-Thr-Dab-cyclo(S-S)[Cvs-Dab-(4-metil)DPhe- Nle-Dab-Dab-DCvsl, (compuesto de fórmula (I) con Rn = CHafCH?)»-, Ri =-CH?CH?NH? (cadena lateral de Dab), R? = -CH(CH¾)OH, (cadena lateral de Thr), Ra = -CH?CH?NH? (cadena lateral de Dab), R4 = -CH?CH?NH? (cadena lateral de Dab), Rfi =-CH?(4- methvDPh (cadena lateral de (4-metil)DPhe), RR = -CH?CH?CH?CH¾ (cadena lateral de Nle), R7 = -CH?CH?NH? (cadena lateral de Dab), y Rfi = -CH?CH?NH? (cadena lateral de Dab) Example 3: Preparation of decanoyl-Dab-Thr-Dab-cyclo (SS) [Cvs-Dab- (4-methyl) DPhe-Nle-Dab-Dab-DCvsl, (compound of formula (I) with R n = CHafCH? ) »-, Ri = -CH ? CH ? NH ? (Dab side chain), R? = -CH (CH ¾ ) OH, (Thr side chain), R a = -CH ? CH ? NH ? (Dab side chain), R 4 = -CH? CH? NH? (Dab side chain), R fi = -CH? (4- methvDPh ((4-methyl) DPhe side chain), R R = -CH ? CH? CH ? CH¾ (Nle side chain), R 7 = -CH ? CH ? NH ? (Dab side chain), and R fi = -CH ? CH ? NH ? (Dab side chain)
Aminoácidos protegidos: Fmoc-Cys(Trt)-OH, Fmoc-Dab(Boc)-OH, Fmoc-Nle-OH, Fmoc- (4-metil)DPhe-OH, Fmoc-Thr(tBu)-OH, Fmoc-DCys(Trt)-OH Protected amino acids: Fmoc-Cys (Trt) -OH, Fmoc-Dab (Boc) -OH, Fmoc-Nle-OH, Fmoc- (4-methyl) DPhe-OH, Fmoc-Thr (tBu) -OH, Fmoc-DCys (Trt) -OH
Se llevo a cabo la síntesis manual según un protocolo estándar Fmoc/'Bu sobre resina Fmoc-Rink Amida MBHA como se ha descrito anteriormente en el Ejemplo 1. Manual synthesis was carried out according to a standard Fmoc / 'Bu protocol on Fmoc-Rink Amida MBHA resin as described above in Example 1.
El peso del crudo peptídico tras el desanclaje y formación del enlace disulfuro fue 265 mg (rendimiento 88%). La purificación por HPLC preparativa proporcionó 56 mg de péptido puro (rendimiento 21 %). The weight of the peptide crude after the de-anchoring and formation of the disulfide bond was 265 mg (yield 88%). Purification by preparative HPLC provided 56 mg of pure peptide (yield 21%).
Caracterización del péptido purificado: Homogeneidad (por integración del área de la traza de HPLC) >99%; MALDI-TOF: m/z 1252.66 ([M+H]+, 100%), 1273.95 ([M+Na]+, 20%). MW 1251.58 Characterization of the purified peptide: Homogeneity (by integration of the HPLC trace area)>99%; MALDI-TOF: m / z 1252.66 ([M + H] + , 100%), 1273.95 ([M + Na] + , 20%). MW 1251.58
Ejemplo 4: Ensayo de actividad antibacteriana Example 4: Antibacterial activity test
Cf. Jorgensen J.H., Turnide J.D., Antibacterial susceptibility test: dilutions and diffusion methods. In: Murray PR, Barón, E.J., Jorgensen, J.H. Pfaller, M.A., Yorken, R.H., eds. Manual of Clinical Microbiology, ASM Press Washington DC, p. 1108 (2003). La actividad antibacteriana de los lipopéptidos sintéticos se determinó en placas estériles de 96 pocilios (Corning Costar 3598 microtiter plates) con un volumen final de 200μί como se indica a continuación: alícuotas (100 μί) de una suspensión de bacterias a una concentración de 105 unidades formadoras de colonias/ml_ en medio de cultivo (MH, Muller Hinton Broth, Difco, USA) a pH 7.4, se adicionaron a 100μΙ_ de solución de lipopéptido preparada a partir de una disolución madre en agua de 1 mg/mL, en diluciones seriadas a doble dilución en MH a pH 7.4 (Jorgensen & Turnide, 2003). Cf. Jorgensen JH, Turnide JD, Antibacterial susceptibility test: dilutions and diffusion methods. In: Murray PR, Baron, EJ, Jorgensen, JH Pfaller, MA, Yorken, RH, eds. Manual of Clinical Microbiology, ASM Press Washington DC, p. 1108 (2003). The antibacterial activity of synthetic lipopeptides was determined in sterile 96-well plates (Corning Costar 3598 microtiter plates) with a final volume of 200μί as follows: aliquots (100 μί) of a suspension of bacteria at a concentration of 10 5 colony forming units / ml_ in culture medium (MH, Muller Hinton Broth, Difco, USA) at pH 7.4, were added to 100μΙ_ of lipopeptide solution prepared from a stock solution in water of 1 mg / mL, in dilutions double dilution series in MH at pH 7.4 (Jorgensen & Turnide, 2003).
La inhibición de crecimiento bacteriano se determinó a partir de la absorbancia a 492 nm en un instrumento Absorbance Microplate reader ELx 800 (Bio-tek Instruments) tras incubación a 37 °C durante 18-20 h. La actividad antibacteriana se expresó como CMI, la concentración a la cual no se detecta crecimiento tras las 18-20 h de incubación. Bacterial growth inhibition was determined from absorbance at 492 nm in an Absorbance Microplate reader ELx 800 instrument (Bio-tek Instruments) after incubation at 37 ° C for 18-20 h. The antibacterial activity was expressed as MIC, the concentration at which no growth is detected after 18-20 h of incubation.
Los microorganismos se cultivaron en Tryptycase Soy Broth (Pronadisa, Barcelona), incubando a 37 °C hasta observar crecimiento bacteriano. A continuación, se sembraron en Trypticase Soy Agar (Pronadisa, Barcelona) y se incubaron a 37°C hasta observar la formación de colonias. Los microorganismos se conservaron en criobolas (EAS laboratoire, France) a -20°C. The microorganisms were grown in Tryptycase Soy Broth (Pronadisa, Barcelona), incubating at 37 ° C until bacterial growth was observed. They were then seeded in Trypticase Soy Agar (Pronadisa, Barcelona) and incubated at 37 ° C until colony formation was observed. The microorganisms were stored in cryoballs (EAS laboratoire, France) at -20 ° C.
Las cepas de las bacterias usadas para llevar a cabo el test de actividad antibacteriana se obtuvieron de: the American Type Culture Collection (ATCC, Rockville, MD, USA): The strains of the bacteria used to carry out the antibacterial activity test were obtained from: the American Type Culture Collection (ATCC, Rockville, MD, USA):
Escherichia coli ATCC 25922 Escherichia coli ATCC 25922
Pseudomonas aeruginosa ATCC 27853 Pseudomonas aeruginosa ATCC 27853
Acinetobacter sp ATCC 5798 Acinetobacter sp ATCC 5798
Staphylococcus aureus ATCC 29213 Staphylococcus aureus ATCC 29213
Mycobacterium phlei ATCC 41423 Mycobacterium phlei ATCC 41423
Enterococus faecalis ATCC 29212 Enterococus faecalis ATCC 29212
Staphylococcus aureus resistente a metilicina ATCC 43300  Stacylococcus aureus resistant to methylcycline ATCC 43300
El ensayo en bacterias resistentes y multirresistentes fue esencialemte el mismo que descrito anteriormente con pequeños cambios: las muestras de volumen final de 100 uL se prepararon de la siguiente manera: alícuotas (50 uL) de la suspensión bacteriana a concentración 1.5 x 106 unidades formadoras de colonia por mL en medio MHB se ajustaron a pH 7.4 y se adicionaron 50 uL de solución de péptido antibiótico preparado de un solución madre de 1024 mg/L en agua y se diluyó en factores de 2 de manera seriada en MHB de pH 7.4. The test in resistant and multiresistant bacteria was essentially the same as described above with small changes: samples of final volume of 100 uL were prepared as follows: aliquots (50 uL) of the bacterial suspension at concentration 1.5 x 10 6 forming units of colony per mL in MHB medium were adjusted to pH 7.4 and 50 uL of prepared antibiotic peptide solution was added of a stock solution of 1024 mg / L in water and diluted in factors of 2 serially in pH 7.4 MHB.
Las muestras de bacterias Gram-negativas resistentes y multirresistentes eran clínicas y no estandarizadas: The samples of resistant and multiresistant Gram-negative bacteria were clinical and not standardized:
Cepas E. coli: NDM-1 (multirresistente); cepas VAL10, VAL5 y MAC21 muestran resistencia intermedia a quinolonas, concretamente: MAC21 y VAL10 a ácido nalidíxico, cotrimoxazole y ampicilina; y VAL 5 a ácido nalidíxico. E. coli strains: NDM-1 (multi-resistant); VAL10, VAL5 and MAC21 strains show intermediate resistance to quinolones, namely: MAC21 and VAL10 to nalidixic acid, cotrimoxazole and ampicillin; and VAL 5 to nalidixic acid.
Cepas P. aeruginosa 38a and 36a son altamente resistentes (a Ceftazidime, P. aeruginosa 38a and 36a strains are highly resistant (to Ceftazidime,
Ciprofloxacina, Imipenem y Piperacillin-tazobactam). Ciprofloxacin, Imipenem and Piperacillin-tazobactam).
Panel de resistencia de: metalo-beta-lactamasa New Delhi (NDM-1): Resistencia a (MIC): Amoxicillina 256 mg/L; Amoxicillina clavulanato 32 mg/L ; Resistance panel of: New Delhi metallo-beta-lactamase (NDM-1): Resistance to (MIC): Amoxicillin 256 mg / L; Amoxicillin clavulanate 32 mg / L;
Piperacillina/tazobactam 256 mg/L; Cefoxitina 256 mg/L; Cefotaxime 256 mg/L; Piperacillin / tazobactam 256 mg / L; Cefoxitin 256 mg / L; Cefotaxime 256 mg / L;
Ceftazidime 256 mg/L; Cefepime 256 mg/L; Imipenem 8 mg/L; Meropenem 16 mg/L; Doripenem 6 mg/L; Ertapenem 24 mg/L; Aztreonam 256 mg/L; Gentamicina 8 mg/L; Amikacina 32 mg/L, Tobramicina 8 mg/L; Ciprofloxacina 32 mg/L. (cf: M. Solé et al., "First Description of an Escherichia coli Strain Producing NDM-1 Carbapenemase in Spain" Antimicrobial Agents and Chemotherapy , 2011 , vol. 55, pp. 4402-4404. Ceftazidime 256 mg / L; Cefepime 256 mg / L; Imipenem 8 mg / L; Meropenem 16 mg / L; Doripenem 6 mg / L; Ertapenem 24 mg / L; Aztreonam 256 mg / L; Gentamicin 8 mg / L; Amikacin 32 mg / L, Tobramycin 8 mg / L; Ciprofloxacin 32 mg / L. (cf: M. Solé et al., "First Description of an Escherichia coli Strain Producing NDM-1 Carbapenemase in Spain" Antimicrobial Agents and Chemotherapy, 2011, vol. 55, pp. 4402-4404.
Los resultados de CMI obtenidos contra las bacterias ATCC mencionadas anteriormente se indican en la tabla 1. The MIC results obtained against the ATCC bacteria mentioned above are indicated in Table 1.
Tabla 1 : Table 1 :
Comp. E. faecalis S.aureus M. phlei E. coli P. aeruginosa Acinetobacter (I) _SD_ Comp. E. faecalis S.aureus M. phlei E. coli P. aeruginosa Acinetobacter (I) _SD_
Ejemplo 8-16 8 4 2 2 2  Example 8-16 8 4 2 2 2
3  3
Ejemplo 8-16 8 8 2-4 2 4-8  Example 8-16 8 8 2-4 2 4-8
2  2
Ejemplo 16-32 4 2-4 4 2 4  Example 16-32 4 2-4 4 2 4
1 Los resultados de CMI obtenidos contra bacterias resistentes se indican en la tabla 2 y en la tabla 3. one The MIC results obtained against resistant bacteria are indicated in Table 2 and Table 3.
Tabla 2: Table 2:
Figure imgf000016_0001
Figure imgf000016_0001
Tabla 3: Table 3:
Comp. MRSA Comp. MRSA
(I)  (I)
Ejemplo 4  Example 4
3  3
Ejemplo 4  Example 4
2  2
Ejemplo 4  Example 4
1  one

Claims

REIVINDICACIONES
1. Compuesto de fórmula (I), 1. Compound of formula (I),
Figure imgf000017_0001
Figure imgf000017_0001
(I) donde: (I where:
R0 es un radical CH3-(CH2)m-, donde m es un número entero entre 4 y 10; R 0 is a radical CH 3 - (CH 2 ) m-, where m is an integer between 4 and 10;
Ri , R3, R4, R7 y R8 son radicales seleccionados independientemente que tienen la fórmula siguiente: Ri, R 3 , R 4 , R 7 and R 8 are independently selected radicals that have the following formula:
GF-(CH2)n-; donde n es un entero entre 1 y 4; y GF es un radical seleccionado entre el grupo que consiste en -NH2 y -NH-C(=NH)-NH2; GF- (CH 2 ) n -; where n is an integer between 1 and 4; and GF is a radical selected from the group consisting of -NH 2 and -NH-C (= NH) -NH 2 ;
R2 es un -CH(CH3)(OH), estando en la fórmula (I) el CH del R2 en cualquiera de las dos configuraciones, R o S, dando lugar a los correspondientes epímeros a una mezcla de los mismos; R 2 is a -CH (CH 3 ) (OH), in the formula (I) being the CH of R 2 in either of the two configurations, R or S, giving the corresponding epimers a mixture thereof;
R5 es un radical -(CH2)-R10; R6 es una radical C4-alquilo lineal o ramificado; R 5 is a radical - (CH 2 ) -R 10 ; R 6 is a linear or branched C 4 -alkyl radical;
R9 es un radical seleccionado entre -CONH2 y -COOH; y R 9 is a radical selected from -CONH 2 and -COOH; Y
R10 es un radical fenilo sustituido con uno a 5 sustituyentes iguales seleccionados del grupo que consiste en F, (CrC2)-alquilo y CF3. R 10 is a phenyl radical substituted with one to 5 equal substituents selected from the group consisting of F, (CrC 2 ) -alkyl and CF 3 .
2. Compuesto según la reivindicación 1 , donde R10 es un radical fenilo sustituido con uno a 5 sustituyentes iguales seleccionados del grupo que consiste en F y CH3. 2. A compound according to claim 1, wherein R 10 is a phenyl radical substituted with one to 5 equal substituents selected from the group consisting of F and CH 3 .
3. Compuesto según cualquiera de las reivindicaciones 1-2, donde R10 es un fenilo sustituido con átomos de flúor. 3. Compound according to any of claims 1-2, wherein R 10 is a phenyl substituted with fluorine atoms.
4. Compuesto según cualquiera de las reivindicaciones 1-2, donde R10 es un fenilo sustituido con grupos metilo. 4. Compound according to any of claims 1-2, wherein R 10 is a phenyl substituted with methyl groups.
5. Compuesto según cualquiera de las reivindicaciones 1-4, donde m es un número entero entre 7 y 10; n es un entero entre 1 y 3; R6 es una radical CH3-(CH2)3-; y Rg es un radical -CONH2. 5. Compound according to any of claims 1-4, wherein m is an integer between 7 and 10; n is an integer between 1 and 3; R 6 is a radical CH 3 - (CH 2 ) 3-; and Rg is a radical -CONH 2 .
6. Compuesto según cualquiera de las reivindicaciones 1-5, donde R0 es un radical CH3- (CH2)8-. 6. Compound according to any of claims 1-5, wherein R 0 is a CH 3 - (CH 2 ) 8 - radical.
7. Compuesto según cualquiera de las reivindicaciones 1-6, donde R^ R3, R4, R7 y R8 son radicales iguales de fórmula -CH2-CH2-NH2. 7. Compound according to any of claims 1-6, wherein R ^ R 3 , R 4 , R 7 and R 8 are equal radicals of the formula -CH 2 -CH 2 -NH 2.
8. Compuesto según la reivindicación 1 , que se selecciona de entre los siguientes: Decanoil-Dab-Thr-Dab-ciclo(S-S)[Cys-Dab-(p-fluoro)DPhe-Nle-Dab-Dab-DCys], Decanoil-Dab-Thr-Dab-ciclo(S-S)[Cys-Dab-(pentafluoro)DPhe-Nle-Dab-Dab-DCys], y Decanoil-Dab-Thr-Dab-ciclo(S-S)[Cys-Dab-(4-metil)DPhe-Nle-Dab-Dab-DCys]. 8. Compound according to claim 1, which is selected from the following: Decanoyl-Dab-Thr-Dab-cycle (SS) [Cys-Dab- (p-fluoro) DPhe-Nle-Dab-Dab-DCys], Decanoil -Dab-Thr-Dab-cycle (SS) [Cys-Dab- (pentafluoro) DPhe-Nle-Dab-Dab-DCys], and Decanoil-Dab-Thr-Dab-cycle (SS) [Cys-Dab- (4 -methyl) DPhe-Nle-Dab-Dab-DCys].
9. Uso de un compuesto como se ha definido en cualquiera de las reivindicaciones 1-8, para la preparación de un medicamento para el tratamiento de una infección bacteriana causada por bacterias Gram-positivas en un mamífero, incluyendo un humano. 9. Use of a compound as defined in any of claims 1-8, for the preparation of a medicament for the treatment of a bacterial infection caused by Gram-positive bacteria in a mammal, including a human.
10. Uso según la reivindicación 9, donde las bacterias Gram-positivas se seleccionan entre el grupo que consiste en Micobacterium phlei, Staphylococcus aureus, 10. Use according to claim 9, wherein the Gram-positive bacteria are selected from the group consisting of Micobacterium phlei, Staphylococcus aureus,
Enterococus faecalis y Staphylococcus aureus resistente a metilicina. Enterococus faecalis and Staphylococcus aureus resistant to methylcycline.
1 1. Uso según la reivindicación 10, donde la bacteria Gram-positiva es Staphylococcus aureus resistente a metilicina. 1 1. Use according to claim 10, wherein the Gram-positive bacterium is Staphylococcus aureus resistant to methylcycline.
12. Uso de un compuesto como se ha definido en cualquiera de las reivindicaciones 1-8, para la preparación de un medicamento para el tratamiento de una infección bacteriana causada por bacterias Gram-negativas en una mamífero, incluyendo un humano. 12. Use of a compound as defined in any of claims 1-8, for the preparation of a medicament for the treatment of a bacterial infection caused by Gram-negative bacteria in a mammal, including a human.
13. Uso según la reivindicación 12, donde las bacterias Gram-negativas se seleccionan entre el grupo que consiste en Pseudomonas aeruginosa, Escherichia coli y 13. Use according to claim 12, wherein the Gram-negative bacteria are selected from the group consisting of Pseudomonas aeruginosa, Escherichia coli and
Acinetobacter sp. Acinetobacter sp.
14. Uso según la reivindicación 13, donde la bacteria Escherichia coli se selecciona del grupo que consiste en Escherichia coli NDM-1 , Escherichia coli VAL-10, Escherichia coli VAL-5 y Escherichia coli MAC21A y donde la bacteria Pseudomonas aeruginosa se selecciona del grupo que consiste en Pseudomonas aeruginosa 38a y Pseudomonas aeruginosa 38b. 14. Use according to claim 13, wherein the Escherichia coli bacteria is selected from the group consisting of Escherichia coli NDM-1, Escherichia coli VAL-10, Escherichia coli VAL-5 and Escherichia coli MAC21A and wherein the Pseudomonas aeruginosa bacteria is selected from group consisting of Pseudomonas aeruginosa 38a and Pseudomonas aeruginosa 38b.
15. Composición farmacéutica que comprende una cantidad terapéuticamente efectiva de un compuesto como se ha definido en cualquiera de las reivindicaciones 1-8, junto a excipientes o portadores farmacéuticamente aceptables. 15. Pharmaceutical composition comprising a therapeutically effective amount of a compound as defined in any of claims 1-8, together with pharmaceutically acceptable excipients or carriers.
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EP3173421A1 (en) 2015-11-30 2017-05-31 Universitat de Barcelona Peptidic compounds useful as antibacterial agents
EP3636659A1 (en) 2018-10-08 2020-04-15 Universitat de Barcelona Polymyxin-based compounds useful as antibacterial agents

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Publication number Priority date Publication date Assignee Title
EP3173421A1 (en) 2015-11-30 2017-05-31 Universitat de Barcelona Peptidic compounds useful as antibacterial agents
WO2017093210A1 (en) 2015-11-30 2017-06-08 Universitat De Barcelona Peptidic compounds useful as antibacterial agents
EP3636659A1 (en) 2018-10-08 2020-04-15 Universitat de Barcelona Polymyxin-based compounds useful as antibacterial agents
WO2020074405A1 (en) 2018-10-08 2020-04-16 Universitat De Barcelona Polymyxin-based compounds useful as antibacterial agents

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