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Treatment of Bacterial Infections

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Treatment of Bacterial Infections (19) & (11) EP 2 510 944 A1 (12) EUROPEAN PATENT APPLICATION (43) Date of publication: (51) Int Cl.: 17.10.2012 Bulletin 2012/42 A61K 38/44 (2006.01) A61K 38/47 (2006.01) A61K 38/54 (2006.01) A61K 33/40 (2006.01) (2006.01) (2006.01) (21) Application number: 11162678.4 A61K 33/18 A61K 33/14 A61K 31/327 (2006.01) A61K 31/7016 (2006.01) (2006.01) (22) Date of filing: 15.04.2011 A61P 31/02 (84) Designated Contracting States: (72) Inventors: AL AT BE BG CH CY CZ DE DK EE ES FI FR GB • O’Flaherty, Vincent GR HR HU IE IS IT LI LT LU LV MC MK MT NL NO Co. Galway (IE) PL PT RO RS SE SI SK SM TR • McCay, Paul Designated Extension States: Galway (IE) BA ME (74) Representative: Gates, Marie Christina Esther et al (71) Applicant: National University of Ireland, Galway Tomkins & Co. Galway (IE) 5 Dartmouth Road Dublin 6 (IE) (54) Treatment of bacterial infections (57) The present application relates to an anti-bac- tuberculosis, cystic fibrosis and the contamination that terial system for use in the treatment of bacterial infec- may result from biofilm formation on medical devices. tions or control of bacterial contamination, which avoids the use of antibiotics. Such infections include mastitis, EP 2 510 944 A1 Printed by Jouve, 75001 PARIS (FR) EP 2 510 944 A1 Description Field of the Invention 5 [0001] The present application relates to an anti-bacterial system for use in the treatment of bacterial infections or control of bacterial contamination, which avoids the use of antibiotics. Such infections include mastitis, tuberculosis, cystic fibrosis and the contamination that may result from biofilm formation on medical devices. Background to the Invention 10 [0002] Mastitis is a persistent inflammatory condition of the udder of cows and other milk- producing animals. It is one of the most common diseases in dairy cattle in the United States and is also the most costly to the dairy industry. Mastitis occurs when white blood cells are released into the mammary gland, usually in response to invasion of bacteria in the teat canal. Milk from cows with mastitis has a higher somatic cell count, and the higher the somatic cell count, the lower 15 the quality of the milk. [0003] Normal treatment for mastitis is with antibiotics, but milk from antibiotic treated cows is not marketable, until the drug has left the cows’ system. The antibiotics used may be systemic and injected into the body, or they may be forced into the teat through the teat canal by intra-mammary infusion. Mastitis can be clinical, whereby visible signs of infection are noted or sub-clinical, where the presence of infection is noted only by an increase in somatic count in the 20 resulting milk. In some clinical situations, cows are often left untreated, though revenue is lost to the dairy farmer through a reduction in the amount of money paid for the milk, which occur where there is an elevated somatic cell count in the milk. [0004] There are a number of other uses of the antibacterial system of the present invention. Cystic fibrosis and tuberculosis are two diseases that are, at present, extremely difficult to treat. Tuberculosis symptoms are caused by infection in the lungs and require long-term antibiotic treatment. Cystic fibrosis (CF) is a condition wherein the sufferer 25 cannot regulate the transfer of chloride ions across their membranes, particularly in the lungs. The condition invariably results in numerous, chronic, lung infections. Antibiotic treatment for either condition can lead to serious drug resistance, minimising their effectivness. At present, antibiotics are delivered through the blood stream intra-venously, or by oral suspension/tablets, or by inhalation. Drug delivery is a big problem for CF sufferers as the antibiotic cannot efficiently transverse the lung membrane to where it is required. This leads to problems wherein resistance to the drug, through 30 the introduction of sub-inhibitory concentrations, may become a serious issue. This makes any further treatment with the drug obsolete. [0005] Bums patients, or patients with open wounds, are extremely susceptible to bacterial infections, notably those due to Staphlycoccal species or Pseudomonad species of bacteria. Treatment of such infections will invariably be by a regimen of antibiotics, either oral or intra-venously. These may be given prophalactically, or when infection is apparent. 35 Such use of antibiotics will often lead to resistance to the drug and an ineffective treatment outcome. The researchers envision a new method of treating bums patients with the present technology. [0006] In addition, large numbers of antibiotic treatments each year are due to the result of medical devices that have become infected whilst in use by a patient. A large number of organisms are responsible for such infections, both Gram- positive and Gram-negative. Infections, on such items as urinary or intra-venous catheters, are often the result of the 40 non-sterile installation of such devices. Over the course of a number of days, any bacterial cells present on the surface of the device will proliferate, leading to the production of biofilms. Such biofilms are extemely difficult to treat with anitbiotics, due to the poor transfer of the drug across to the inner cells of the biofilm mass, leading often to even greater levels of tolerance of the biofilms to the antibiotic. Infection of the medical device will often require its removal and replacement, to the discomfort of the patient. Although the infection will often be noted a number of days after installation 45 of the medical device, it will be typically incurred as the result of bacteria being present very early in the installation. [0007] It is generally known that a bacteriostatic effect is caused by the reaction between hydrogen peroxide and thiocyanate, catalysed by lactoperoxidase. In certain instances, the source of peroxide is a reaction between glucose and glucose oxidase, which results in the production of gluconic acid and peroxide. This process is used during the transport of milk. 50 [0008] Antibacterial treatments for the control of infections, such as mastitis are also known. For example, PCT Ap- plication W02008/041218 discloses a preparation with an antimicrobial and immunostimulatory effect, which comprises an oxidoreductase enzyme, an appropriate substrate for that enzyme to produce hydrogen peroxide, and endogenous hydrogen peroxide preparations. The preparation produces 2-stage hydrogen peroxide release; the endogenous form of peroxide ensuring that there is instantly available hydrogen peroxide and further hydrogen peroxide is produced by 55 the oxidoreductase enzyme. [0009] US Patent No. 6312687 describes a stabilised aqueous enzyme concentration comprising lactoperoxidase, glucose oxidase, an alkaline metal halide salt and a buffering agent, for use as an antimicrobial composition. US Patent No. 5607681 describes antimicrobial compositions, which comprise iodide anions and thiocyanate anions together with 2 EP 2 510 944 A1 D-glucose and either glucose oxidase or glucose oxidase together with and at least one antioxidant. The composition may additionally comprise lactoperoxidase. [0010] The basic process for the bacteriostatic effect of lactoperoxidase in milk is based on thiocyanate and a source of hydrogen peroxide, together with lactoperoxidase to produce a bacteriostatic effect. The thiocyanate ion is oxidised 5 in the presence of hydrogen peroxide by lactoperoxidase, to produce hypothiocyanous acid. This acid reacts with free sulphydryl groups in bacterial proteins, inactivating several metabolic enzymes, thus resulting in a bacteriostatic effect. In certain embodiments, the peroxide ion is produced from glucose by the action of glucose oxidase rather than by using a solution of hydrogen peroxide or its release from a suitable perhydrate (such as sodium percarbonate). The present application suggests the use of further substrates to help bring about the reaction to produce hydrogen peroxide by 10 means of supplemented enzymatic catalysis. [0011] While the lactoperoxidase system has been widely described, it is thought by those skilled in the art that this is an ineffective system to be utilised of the treatment of baterial infection (Rainard & Riollet 2006, "Innate immunity of the bovine mammary gland", Veterinary Research 37, 369-400; Sakai et al., 2008, "Generation of hydrogen peroxide by a low molecular weight compound in whey of Holstein dairy cows", Journal of dairy Research, Journal of Dairy 15 Research, 75, 257-261; Sakai et al., 2008, "Production of hydrogen peroxide by a small molecular mass compound in milk from Holstein cows with high and low milk somatic cell count", Journal of Dairy Research, 75, 335-339). [0012] As discussed elsewhere within this application, the broad spectrum and numerous potential targets of the antimicrobial species of the invention are unlikely to induce the proliferation of resistance genes. Drug reactions would not be a problem either, being that components of the system are found naturally in the mammalian body (thiocyanate, 20 lactoperoxidase, glucose) or are generally regarded as safe and unlikely to stimulate an immune reaction (glucose oxidase). Reaction to drugs is a current problem, with 20% of patients reacting to B- lactam drugs ("Rapid de- sensitization for non-immediate reactions in patients with cystic fibrosis", Whitaker et al., 2011, Journal of Cystic Fibrosis, in press). [0013] The present technology, based on the reaction of a haloperoxidase with a substrate, in the presence of hydrogen peroxidde, would provide a unique way to limit bacterial infection and biofilm growth on the surface of medical devices. 25 Object of the Invention [0014] A first object of the invention is to provide an improved composition for the treatment of bacterial infections. [0015] There is a need for a composition for the treatment of both clinical and sub-clinical mastitis, without the use of 30 antibiotics.
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