Drug – bio-affecting and body treating compositions – Designated organic active ingredient containing – Heterocyclic carbon compounds containing a hetero ring...
Reexamination Certificate
1998-07-23
2001-09-25
Peselev, Elli (Department: 1623)
Drug, bio-affecting and body treating compositions
Designated organic active ingredient containing
Heterocyclic carbon compounds containing a hetero ring...
C514S197000, C514S210030, C514S310000, C514S450000, C514S451000
Reexamination Certificate
active
06294526
ABSTRACT:
TECHNICAL FIELD
The invention relates to a novel screening system for detecting chemical compounds which induce sensitivity to &bgr;-lactams in &bgr;-lactam resistant bacteria such as methicillin-resistant
Staphlococcus aurous
(MRSA) and, a pharmaceutical composition having anti-MRSA activity which comprises a flavone derivative selected by the screening system and a &bgr;-lactam, a method of increasing antibacterial activity of a &bgr;-lactam and a method of preventing and treating bacterial infectious diseases.
BACKGROUND OF THE INVENTION
The first antibiotic, penicillin, has a &bgr;-lactam ring and showed an eminent efficacy against staphylococci. However, staphylococci can easily produce penicillinase (i.e., &bgr;-lactamase) to inactivate penicillin. The staphylococci is called as penicillin-resistant bacteria. Against penicillin-resistant bacteria, for example, penicillinase-resistant penicillin- and cephem- antibiotics were developed and the problem involving penicillin-resistant bacteria seemed to be almost solved. However, a MRSA against which all &bgr;-lactams have no effect developed.
The MRSA is widely resistant not only to penicillin- antibiotics but also to cephem- and aminoglycoside- antibiotics. Recently, as a result of overuse of cephem antibiotics of the third generation cephems which are weakly antibacterial to
Staphylococcus aureus
, the cephem-resistant
Staphylococcus aureus
selectively proliferated and spread in hospitals, and therefore, the cephem-resistant
Staphylococcus aureus
has become a serious object of public concern as infectious bacteria in hospitals. Vancomycin (VCM) or the like is currently used as an anti-bacterial agent against MRSA-infectious diseases; however, problems are that the sterilizing power of VCM is not very potent over a short duration and VCM has severe side effects such as ototoxicity and nephrotoxicity.
In order to promote anti-bacterial activity against MRSA, combinations of plural anti-bacterial agents have been studied, for example, a combination of an aminoglycoside and &bgr;-lactam or a phosphomycin and a &bgr;-lactam has been tried, however the effectiveness of the combinations is not sufficient.
Novel anti-bacterial agents are urgently needed to be developed that are based on a novel mechanism of function different from conventional mechanisms and effective against MRSA.
SUMMARY
While the present inventors were researching an herb medicine having no or less side effects and also anti-MRSA activity, they found that apigenin, luteolin and flavone derivatives having chemical structures similar thereto have anti-MRSA activity. It has also been found that the flavone derivatives inhibit &bgr;-lactam-resistance of MRSA and induce sensitivity to &bgr;-lactam in MRSA. The present invention has been achieved based on the above findings.
The invention relates to a novel screening system for detecting chemical compounds which induce sensitivity to &bgr;-lactams in &bgr;-lactam resistant bacteria such as MRSA.
Many combinations of plural antibiotics were studied, however as yet no combination of a non-antibiotic and a &bgr;-lactam has been reported.
Therefore, in a first embodiment, the present invention provides a novel screening system for detecting a non-antibiotic compound having &bgr;-lactam-sensitivity-inducing activity comprising a &bgr;-lactam antibiotic and a non-antibiotic compound suspected to be effective to induce sensitivity.
In a second embodiment, the present invention provides a pharmaceutical composition having anti-MRSA activity which comprises a &bgr;-lactam antibiotic and the non-antibiotic compound detected by the screening system to be effective to induce sensitivity or a pharmaceutically acceptable salt thereof. The non-antibiotic compounds may be liavone derivatives. The pharmaceutical composition may increase anti-bacterial activity of a &bgr;-lactam antibiotic. The anti-bacterial activity may be against MRSA.
A third embodiment is a method for increasing antibacterial activity of a &bgr;-lactam antibiotic which comprises adding a flavone derivative to the &bgr;-lactam antibiotic.
A fourth embodiment is a method for treating or preventing an infectious disease of which the source of infection is
Staphylococcus aureus
or methicillin-resistant
Staphylococcus aureus
which comprises administering a flavone derivative together with the &bgr;-lactam antibiotic.
Apigenin or luteolin, which were selected by the screening system of the present invention, are found in the whole grass of
Scutellaria barbata
D. Don.
Scutellaria barbata
is a perennial plant of (Labiatae) and further includes alkaloids, flavonoid glycosides, polyphenol and furalols.
The dried whole grasses of Scutellaria barbata were almost stripped of leaves down to stems with flower petals and shoots. They are 5-25 cm in length, square. The surface of the stems is yellowish green or purplish brown, lustrous and smooth. Their medical activities have been mentioned as being antipyretic, antidotal, hemostyptic, and analgetic activities. However, It has not been reported that Scutellaria barbata has anti-MRSA activity or that flavone derivatives such as apigenin, and luteolin have anti-MRSA activity.
Further scope of applicability of the present invention will become apparent from the detailed description given hereinafter. However, it should be understood that the detailed description and specific examples, while indicating preferred embodiments of the invention, are given by way of illustration only, since various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description.
DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS
Examples of &bgr;-lactam antibiotics useful herein include methicillin (DMPPC), oxacillin (MIPC), penicillin G (PCG), ampicillin (ABPC), cephalothin (CET), cefoxitin (CFX), cefuroxime (CXM) and cefotaxime (CTX). Examples of flavone derivatives useful herein include flavone, apigenin, luteolin, kaempferol, quercetin, sakuranetin, eriodictyol, (+)-taxifolin and phloretin.
The generic name or an abbreviation of an antibiotic referred to herein include both the free acid and pharmaceutically acceptable salts thereof. “Pharmaceutically acceptable salt” means a pharmaceutically acceptable salt of a &bgr;-lactam antibiotic which is conventionally used. As “a pharmaceutically acceptable salt”, there may be exemplified salts such as sodium, potassium and calcium, an amine salt such as procaine, dibenzyl amine, and an addition salt such as hydrochloric acid salt.
The screening of the present invention may be carried out according to the agar plate dilution method designated by the Japan Society of Chemotherapy (Chemotherapy 29(1), 76-79 (1981)). Mueller-Hinton agar base semi-synthetic medium may be used for a plate culture medium for a sensitivity test together with an inoculating bacteria solution which may be prepared by culturing bacteria to be tested at 37° C. for 20 hours diluted with 8.5% by isotonic sodium chloride solution to 10
6
CFU/ml. Diluted &bgr;-lactam solutions are prepared by two times serial dilution. 1 ml of the diluted &bgr;-lactam solution and 1 ml of a predetermined concentration of a sample to be tested are added to 8 ml of the plate culture medium for a sensitivity test. To the resultant plate culture medium for sensitivity test, the bacteria are inoculated using a microplanter (SAKUMASEISAKUSYO Co.), cultured for 20 hours at 37° C. followed by assessing for a minimum inhibition concentration (MIC). A minimum inhibition concentration means a minimum concentration at which growth of bacteria is completely inhibited.
Alternatively, the screening of the present invention may be carried out according to the broth microdilution method designated by the Japan Society of Chemotherapy (Chemotherapy 38(1), 102-105 (1990)). Muellar-Hinton broth is used as a culture medium for the sensitivity test, and 50 mg/ml of calcium(Ca) ion, 25 mg/ml of magnesium(Mg) ion, and 2% sodium chloride (NaCl) are added thereto. An inoculating
Higuchi Tomihiko
Murasugi Shoji
Sato Yoichi
Alps Pharmaceutical Ind. Co., Ltd.
Peselev Elli
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