JP6611842B2 - Antituberculosis agent - Google Patents

Description

  The present invention relates to an antituberculosis agent.

In recent years, tuberculosis and tuberculosis caused by tuberculosis infection are increasing.
Furthermore, infectious diseases caused by MAC disease caused by M. avium, which is a nontuberculous mycobacteria, are increasing worldwide. In addition, the emergence of antibiotic-resistant bacteria has become a problem (Non-Patent Document 1).

Furthermore, in recent years, the number of patients suffering from tuberculosis is gradually increasing again, and multidrug-resistant tuberculosis (MDR-TB) and super multidrug-resistant tuberculosis (XDR-TB) have emerged. For this reason, an anti-tuberculosis agent effective for them is desired.
Furthermore, development of antibiotics having a novel mechanism is desired.

Skolnik K et al., Curr Treat Options Infect Dis., 8 (4): 259-274

  An object of the present invention is to provide a novel anti-tuberculosis agent, and further, to provide a novel anti-tuberculosis agent that exhibits antibacterial activity against multidrug-resistant tuberculosis bacteria and super-drug-resistant tuberculosis bacteria. It is to provide.

  As a result of intensive studies to solve the above problems, the present inventor has found that a specific compound exhibits antibacterial activity against Mycobacterium tuberculosis, leading to the present invention.

［式（１）中、Ｒ^１は置換基を有していてもよい炭素数が７、８又は９のアシル基を示し、Ｒ^２はメチル基又は水素原子を示し、Ｒ^３はエチル基又はメチル基を示す。］
とする。 That is, the present invention provides an anti-tuberculosis agent characterized by containing a cyclic peptide compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient and for tuberculosis. It is to provide.

[In the formula (1), R ¹ represents an optionally substituted acyl group having 7, 8, or 9 carbon atoms, R ² represents a methyl group or a hydrogen atom, and R ³ represents an ethyl group or Indicates a methyl group. ]
And

  The present invention also provides an anti-tuberculosis agent characterized by containing a compound derived from the deposit number NITE BP-870 belonging to the genus Lysobacter as an active ingredient and for tuberculosis.

  The present invention also provides an anti-tuberculosis agent characterized by containing as an active ingredient a compound derived from a microorganism belonging to the genus Lysobacter having a deposit number of NITE BP-870. is there.

  According to the present invention, it is possible to provide a novel anti-tuberculosis agent effective for diseases caused by M. tuberculosis. In addition, an anti-tuberculosis agent exhibiting antibacterial activity can be provided against multi-drug resistant tuberculosis (MDR-TB) and super multi-drug resistant tuberculosis (XDR-TB).

  With the anti-tuberculosis agent of the present invention, various diseases such as “tuberculosis including pulmonary tuberculosis and extrapulmonary tuberculosis” caused by tuberculosis bacteria including multi-drug resistant tuberculosis or super multi-drug resistant tuberculosis can be treated.

Compared to the treatment of tuberculosis, there is a problem that the treatment success rate of multi-drug resistant tuberculosis, especially the treatment of super multi-drug resistant tuberculosis is low. In addition, the treatment of multidrug-resistant tuberculosis and ultra-multidrug-resistant tuberculosis has a longer treatment period than the treatment of tuberculosis and the burden of treatment costs is also a problem. Early development of agents is desired.
According to this example, the anti-tuberculosis agent of the present invention exhibits good antibacterial activity against tuberculosis, multi-drug resistant tuberculosis (MDR-TB) and super multi-drug resistant tuberculosis (XDR-TB). , Has a noticeable effect.

It is a figure which shows the three-dimensional structure of the amino acid of lysosine E.

  Hereinafter, the present invention will be described, but the present invention is not limited to the following specific embodiments, and can be arbitrarily modified within the scope of the technical idea.

［式（１）中、Ｒ^１は置換基を有していてもよい炭素数が７、８又は９のアシル基を示し、Ｒ^２はメチル基又は水素原子を示し、Ｒ^３はエチル基又はメチル基を示す。］ The anti-tuberculosis agent of the present invention contains a cyclic peptide compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient, and is characterized by being for tuberculosis.

[In the formula (1), R ¹ represents an optionally substituted acyl group having 7, 8, or 9 carbon atoms, R ² represents a methyl group or a hydrogen atom, and R ³ represents an ethyl group or Indicates a methyl group. ]

  In addition, the anti-tuberculosis agent of the present invention contains a compound derived from a microorganism belonging to the genus Lysobacter of the deposit number NITE BP-870 as an active ingredient, and is characterized by being for tuberculosis.

The cyclic peptide compound represented by the above formula (1) (hereinafter may be referred to as “lysosine”) has an accession number at the Patent Microorganism Deposit Center (NPMD) of the National Institute of Technology and Evaluation (NITE). Culturing a microorganism belonging to the genus Lysobacter of NITE BP-870 (hereinafter, this microorganism may be referred to as “RH2180-5”) or a mutant strain capable of producing a compound similar to the microorganism, It is preferable that it is obtained from the culture.
Therefore, the anti-tuberculosis agent of the present invention is characterized by containing, as an active ingredient, a compound derived from a microorganism belonging to the genus Lysobacter having a deposit number of NITE BP-870.
For example, lysosine is described in detail in International Publication No. 2011/148959.

The above-mentioned “RH2180-5” is a patent of Room No. 2-5-8 Kazusa-Kamashita, Kisarazu City, Chiba Prefecture, National Institute of Technology and Evaluation (hereinafter abbreviated as “NITE”). It is a microorganism deposited domestically at the Microorganism Deposit Center (NPMD) and deposited under the deposit number: NITE P-870 (deposit date: January 25, 2010).
“RH2180-5” was then submitted to the Patent Microorganisms Depositary Center (NPMD) of 2-5-8 Kazusa-Kamasashi, Kisarazu City, Chiba Prefecture, and the National Institute for Product Evaluation and Technology (NITE). Submit an application for transfer from domestic deposit (original deposit date: January 25, 2010) to deposit under the Budapest Treaty (transfer date (international deposit date): May 20, 2011) As a result of receiving an application for transfer to a deposit based on the Budapest Treaty (international deposit), it has received the deposit number “NITE BP-870”.

R ¹ in the above formula (1) represents an acyl group having 7, 8 or 9 carbon atoms which may have a substituent. The number of carbon atoms of the acyl group includes the number of carbon atoms of “C═O” (one). The “acyl group having 7, 8 or 9 carbon atoms” excluding the substituent is represented by “R′—C (═O) —”, where R ′ has 6, 7 or 7 carbon atoms. 8 represents an alkyl group. R ′ may be linear or branched, but preferably branched. The branched portion is preferably a methyl group, and there is no particular limitation. However, the end of R ′ opposite to “C═O” is “CH ₃ (CH ₃ ) CH—”. It is particularly preferable. In addition, when R ′, that is, R ¹ has a branch, the carbon number (7, 8 or 9) of R ¹ includes the carbon number of the branched portion. Moreover, it is preferable that the substituent of R < ¹ > in the said Formula (1) is a hydroxyl group.

Specifically, R ¹ in the above formula (1) is a 3-hydroxy-5-methyl-hexanoyl group, a 3-hydroxy-6-methyl-heptanoyl group or a 3-hydroxy-7-methyl-octanoyl group. It is preferable.

In the “cyclic peptide compound or a pharmaceutically acceptable salt thereof” represented by the above formula (1), R ¹ is a 3-hydroxy-5-methyl-hexanoyl group and R ² is a methyl group. R ³ is preferably an ethyl group, particularly preferably R ¹ is a 3-hydroxy-7-methyl-octanoyl group, R ² is a methyl group, and R ³ is an ethyl group.
In the present invention, the “cyclic peptide compound or a pharmaceutically acceptable salt thereof” represented by the above formula (1), wherein R ¹ is a 3-hydroxy-5-methyl-hexanoyl group and R ² is A methyl group and R ³ is an ethyl group is referred to as “lysosine E”.
FIG. 1 shows the three-dimensional structure of lysosine E amino acid.

The anti-tuberculosis agent of the present invention exhibits antibacterial activity also against multidrug-resistant tuberculosis bacteria, and further exhibits antibacterial activity against super multidrug-resistant tuberculosis bacteria.
That is, the anti-tuberculosis agent of the present invention is both an anti-multidrug-resistant tuberculosis agent and an anti-super multidrug-resistant tuberculosis agent.
Multi-drug resistant tuberculosis refers to tuberculosis that is resistant to at least both isoniazid and rifampicin, which are first-line drugs in the treatment of tuberculosis.
In addition to isoniazid and rifampicin, the super multi-drug resistant tuberculosis is a tuberculosis that is resistant to at least one fluoroquinolone and at least one second-line drug in tuberculosis treatment (kanamycin, streptomycin, amikacin or capreomycin). say.

With the anti-tuberculosis agent of the present invention, various diseases caused by tuberculosis including multi-drug resistant tuberculosis and super multi-drug resistant tuberculosis can be treated.
Treating various diseases including “tuberculosis composed of pulmonary tuberculosis and extrapulmonary tuberculosis” caused by “multi-drug resistant tuberculosis, tuberculosis including super multidrug-resistant tuberculosis” by the antituberculous agent of the present invention. Can do. The present invention is also a therapeutic agent for tuberculosis or a tuberculosis disease characterized by containing the above-mentioned antituberculous agent.

  Examples of the above diseases are pulmonary tuberculosis; extrapulmonary tuberculosis such as intestinal tuberculosis, osteoarticular tuberculosis, cutaneous tuberculosis, pharyngeal tuberculosis, spinal caries; tuberculosis tumor, tuberculous pleurisy, tuberculous meningitis, tuberculosis pericarditis, tuberculosis Osteoarthritis, tuberculosis bronchitis, tuberculosis bronchitis, tuberculous tendonitis, tuberculosis osteomyelitis, tuberculosis nephritis, tuberculosis enteritis, tuberculosis spondylitis, tuberculosis lymphadenitis, tuberculosis rheumatism, tuberculous cystitis, Tuberculous peritonitis, tuberculosis pneumonia, tuberculous empyema, tuberculous abscess, tuberculous diseases such as tuberculous skin adenopathies; and the like.

  There is no limitation on the method for obtaining the lysosine, and it may be obtained by any method. For example, it may be obtained by chemical synthesis, may be expressed using a microorganism, may be isolated from a living organism, or may be obtained by combining them. Good.

  Further, the anti-tuberculosis agent of the present invention can contain “other components” in addition to lysosine, which is an active ingredient.

The “other ingredients” in the anti-tuberculosis agent are not particularly limited and can be appropriately selected depending on the purpose within a range not impairing the effects of the present invention. For example, pharmaceutically acceptable. Examples are carriers and excipients to be obtained.
There is no restriction | limiting in particular as this support | carrier, For example, it can select suitably according to the dosage form etc. which are mentioned later. Further, the content of the “other components” in the anti-tuberculosis agent is not particularly limited and can be appropriately selected depending on the purpose.

There is no restriction | limiting in particular as a dosage form of the antituberculous agent of this invention, For example, it can select suitably according to the desired administration method which is mentioned later.
Specifically, for example, oral solids (tablets, coated tablets, granules, powders, capsules, etc.), oral liquids (internal solutions, syrups, elixirs, etc.), injections (solvents, suspensions, etc.) , Ointments, patches, gels, creams, powders for external use, sprays, inhalation sprays and the like.

Examples of the oral solid preparation include, for example, excipients and further additives such as binders, disintegrants, lubricants, colorants, flavoring and flavoring agents as necessary, in addition to the active ingredients. Can be manufactured.
Examples of the excipient include lactose, sucrose, sodium chloride, glucose, starch, calcium carbonate, kaolin, microcrystalline cellulose, silicic acid and the like.
Examples of the binder include water, ethanol, propanol, simple syrup, glucose solution, starch solution, gelatin solution, carboxymethylcellulose, hydroxypropylcellulose, hydroxypropyl starch, methylcellulose, ethylcellulose, shellac, calcium phosphate, polyvinylpyrrolidone and the like. It is done.
Examples of the disintegrant include dry starch, sodium alginate, agar powder, sodium bicarbonate, calcium carbonate, sodium lauryl sulfate, stearic acid monoglyceride, and lactose.
Examples of the lubricant include purified talc, stearate, borax, and polyethylene glycol.
Examples of the colorant include titanium oxide and iron oxide.
Examples of the flavoring / flavoring agent include sucrose, orange peel, citric acid, tartaric acid and the like.

The oral solution can be produced by a conventional method, for example, by adding additives such as a flavoring / flavoring agent, a buffering agent, and a stabilizer to the active ingredient.
Examples of the flavoring / flavoring agent include sucrose, orange peel, citric acid, tartaric acid and the like. Examples of the buffer include sodium citrate. Examples of the stabilizer include tragacanth, gum arabic, and gelatin.

As the injection, for example, a pH adjuster, a buffer, a stabilizer, an isotonic agent, a local anesthetic, etc. are added to the active ingredient, and subcutaneous, intramuscular and intravenous use are performed by a conventional method. Etc. can be manufactured.
Examples of the pH adjusting agent and the buffering agent include sodium citrate, sodium acetate, sodium phosphate and the like. Examples of the stabilizer include sodium pyrosulfite, EDTA, thioglycolic acid, thiolactic acid, and the like. Examples of the isotonic agent include sodium chloride and glucose. Examples of the local anesthetic include procaine hydrochloride and lidocaine hydrochloride.

As the ointment, for example, a known base, stabilizer, wetting agent, preservative and the like may be blended with the active ingredient and mixed by a conventional method.
Examples of the base include liquid paraffin, white petrolatum, white beeswax, octyldodecyl alcohol, and paraffin. Examples of the preservative include methyl paraoxybenzoate, ethyl paraoxybenzoate, propyl paraoxybenzoate, and the like.

  As the patch, for example, a cream, gel or paste as the ointment can be applied to a known support by a conventional method. Examples of the support include woven fabric, nonwoven fabric, soft vinyl chloride, polyethylene, polyurethane and other films made of cotton, suf, and chemical fibers, and foam sheets.

  The content of lysine, which is an active ingredient in the antituberculosis agent of the present invention, with respect to the whole antituberculosis agent is not particularly limited and can be appropriately selected according to the purpose. When 100 parts by mass, the total amount of lysosine is preferably blended in a content of 0.001 to 100 parts by mass, more preferably 0.01 to 99 parts by mass, and particularly preferably 0.1 to 95 parts by mass. Parts, more preferably 1 to 90 parts by weight.

  The animal to be administered with the anti-tuberculosis agent of the present invention is not particularly limited, but for example, humans; mice; rats; monkeys; horses; livestock such as cows, pigs, goats and chickens; Etc.

Moreover, there is no restriction | limiting in particular as the administration method of the said antituberculous agent, For example, according to the dosage form etc. of the said antituberculous agent, it can select suitably, Oral administration, intraperitoneal administration, Respiratory organs are possible. Examples include inhalation, injection into blood, and infusion into the intestine.
The dose of the anti-tuberculosis agent is not particularly limited and may be appropriately selected depending on the age, weight, desired degree of effect, etc. of the individual to be administered. The daily dose is preferably 1 mg to 30 g, more preferably 10 mg to 10 g, and particularly preferably 100 mg to 3 g as the amount of the active ingredient.
Moreover, there is no restriction | limiting in particular also as an administration time of the said antituberculous agent, According to the objective, it can select suitably, For example, you may administer prophylactically and may administer therapeutically.
The antituberculous agent of the present invention may be used in combination with other known antituberculous agents.

  Hereinafter, the present invention will be described in more detail based on examples, test examples, and examination examples, but the present invention is not limited to the specific scope of the following examples and the like.

<Deposit>
As described above, “RH2180-5” used in the examples is the room 2-5-8 Kazusa Kamashitsu, Kisarazu City, Chiba Prefecture, 122, National Institute of Technology and Evaluation; The microorganism was deposited in Japan at the Patent Microorganism Deposit Center (NPMD) of NITE (abbreviated as “NITE”) and deposited under the deposit number: NITE P-870 (deposit date: January 25, 2010).
“RH2180-5” was then submitted to the Patent Microorganisms Depositary Center (NPMD) of 2-5-8 Kazusa-Kamasashi, Kisarazu City, Chiba Prefecture, and the National Institute for Product Evaluation and Technology (NITE). Submit an application for transfer from domestic deposit (original deposit date: January 25, 2010) to deposit under the Budapest Treaty (transfer date (international deposit date): May 20, 2011) As a result of receiving an application for transfer to a deposit based on the Budapest Treaty (international deposit), it has received the deposit number “NITE BP-870”.

Example 1
<Preparation of lysosine E>
As lysosine E, RH2180-5 strain was cultured and the culture solution was purified. The culture and purification procedures followed the procedures described in Japanese Patent Application Laid-Open No. 2012-006917 and International Publication No. 2011/148959.

Study example 1
<Antimicrobial activity of lysosine E against Mycobacterium tuberculosis>
So far, the inventors have studied the antibacterial spectrum of lysosine E against the acid-fast bacterium Mycobacterium smegmatis (International Publication No. 2011/148959).
The MIC value for Mycobacterium smegmatis was higher than that of MSSA, MRSA, Bacillus subtilis, etc. as measured by a micro sample dilution method based on CLSI (formerly NCCLS US Clinical Laboratory Standard Committee). Therefore, lysosine E was thought to have weak antibacterial activity against Mycobacterium.
On the other hand, with the advent of multi-drug resistant tuberculosis (MDR-TB) and super multi-drug resistant tuberculosis (XDR-TB), development of antibacterial agents having a novel mechanism different from conventional antibacterial agents is desired. Lysosine E may have a novel mechanism (Hamamoto H et al., Nat Chem Biol. 2015 Feb; 11 (2): 127-33), multidrug-resistant Mycobacterium tuberculosis (MDR-TB) and It was examined whether there was antibacterial activity against multidrug-resistant tuberculosis bacteria (XDR-TB).

Antibacterial activity against standard strains of Mycobacterium tuberculosis was examined. In Middlebrook 7H9 medium containing 10% ADC and 10 μg / mL kanamycin, Mycobacterium tuberculosis H37Rv-Luc, Mycobacterium tuberculosis MDR066 (resistant to isoniazid and rifampicin), tuberculosis resistant tuberculosis Mycobacterium tuberculosis MDR005 and MRD044 (both isoniazid, rifampicin, streptomycin, ethambutol, kanamycin, levofloxacin, sparfloxacin and ciprofloxacin resistance) were added to each so that OD ₆₀₀ = 0.0005.
Furthermore, lysosine E was added, and the cells were cultured at 37 ° C. under 5% CO _{2 for} 9 days, and the growth of the bacteria was visually confirmed.

  As a result, the growth of Mycobacterium tuberculosis H37Rv-Luc, a tuberculosis bacterium, could not be confirmed when 0.25 μg / mL lysosine E was added. Therefore, it was found that lysosine E exhibits a good antibacterial activity against Mycobacterium tuberculosis. In addition, lysosine E was thought to have weak antibacterial activity against Mycobacterium spp. Until now, but it was surprisingly found that Mycobacterium tuberculosis shows antibacterial activity.

  In addition, 1 μg / mL for Mycobacterium tuberculosis MDR066, which is a multidrug-resistant tuberculosis, and <0.13 μg / mL and 0.5 μg / mL, for Mycobacterium tuberculosis MDR005 and MRD044, which are extremely multidrug-resistant Mycobacterium tuberculosis. When lysosine E was added, the growth of each bacterium could not be confirmed. Therefore, it was found that lysosine E exhibits good antibacterial activity against multi-drug resistant tuberculosis (MDR-TB) and super multi-drug resistant tuberculosis (XDR-TB).

Study example 2
<Antimicrobial activity of lysosine E against various bacteria>
1.0 × 10 ⁶ of the bacteria described in Table 1 was added to 100 μL of a Middlebrook 7H9 medium containing 0.5% Tween 80 and 10% ADC enrichment. Furthermore, lysosine E was added, and after incubation at 37 ° C. for 30 hours, the minimum concentration that inhibited the growth of 90% of the control by the MTT assay was defined as MIC. The results are shown in Table 1.

  As a result of Table 1, it was found that lysosine E does not exhibit antibacterial activity against some bacteria.

<Summary of Study Examples 1-2>
Lysosine E has been shown to exhibit antibacterial activity against Mycobacterium tuberculosis. In addition, since lysosine E has a completely different mechanism from conventional anti-tuberculosis drugs, it has been clarified that lysosine E exhibits antibacterial activity against multi-drug resistant tuberculosis bacteria and super multi-drug resistant tuberculosis bacteria.

  In addition, lysosine E was shown to have extremely rapid and powerful bactericidal properties in the examination example 1 within a few minutes after addition, and the activity of antibacterial activity was inhibited by the lung extract. It is considered that it shows a good therapeutic effect because it is not received.

Example 2
<Formulation of antituberculous agent>
<< Tablet >>
After uniformly mixing 20.0 mg of lysosine E, 40 mg of lactose, 20 mg of starch, and 5 mg of low-substituted hydroxypropylcellulose, granules for tableting were produced by a wet granulation method using 8% by mass aqueous solution of hydroxypropylmethylcellulose as a binder. . To this was added 0.5 mg to 1 mg of magnesium stearate necessary to give lubricity, and then tableted using a tableting machine to obtain tablets.

  10.0 mg of lysosine E was dissolved in 10 mL of a 2% by mass 2-hydroxypropyl-β-cyclodextrin aqueous solution to prepare an injection solution.

  The anti-tuberculosis agent of the present invention can be widely used in the pharmaceutical industry and related industries.

  NITE BP-870

Claims (4)

An anti-tuberculosis fungus agent characterized by containing a cyclic peptide compound represented by the following formula (1) or a pharmaceutically acceptable salt thereof as an active ingredient and for tuberculosis.

[In the formula (1), R ¹ represents an optionally substituted acyl group having 7, 8, or 9 carbon atoms, R ² represents a methyl group or a hydrogen atom, and R ³ represents an ethyl group or Indicates a methyl group. ]   The anti-tuberculosis agent according to claim 1, wherein the tuberculosis bacterium is a multidrug-resistant tuberculosis bacterium.   The anti-tuberculosis agent according to claim 1, wherein the tuberculosis bacterium is a super multidrug-resistant tuberculosis bacterium. A therapeutic agent for tuberculosis or a tuberculosis disease, comprising the anti-tuberculosis agent according to any one of claims 1 to 3 .

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