Treatment of bacterial infections

Details Treatment of bacterial infections Treatment of bacterial infections

1999-02-26

2001-06-05

Leary, Louise N. (Department: 1623)

Chemistry: molecular biology and microbiology

Measuring or testing process involving enzymes or...

Involving viable micro-organism

C435S013000, C435S036000, C435S024000, C435S244000, C435S252100, C435S252800, C435S253400, C435S848000, C435S849000, C435S851000, C435S885000, C435S882000

Reexamination Certificate

active

06242210

ABSTRACT:

TECHNICAL FIELD OF THE INVENTION
The invention relates to assays for the detection of compounds useful for the treatment of the bacterial infection and in particular conditions associated with infection by curli-expressing gram negative bacteria and streptococci.
BACKGROUND OF THE INVENTION
Some micro-organisms such as Streptococci, Salmonella,
E.coli
and
S.aureus
may cause severe invasive infections such as sepsis or septic shock. Fever, hypotension and bleeding disorders are common symptoms of sepsis and septic shock. Invasive infections caused by these bacteria may result from resistance to antibiotics or defects in the immune system of the infected individual.
The endogenous or intrinsic pathway of inflammation and coagulation is triggered by assembly of the contact phases system. The contact phase system is orchestrated by three serine proteases, factor XI (hereinafter referred to as F XI), factor XII (F XII) and plasma kallikrein (PK) as well as the non-enzymatic co-factor H-kininogen (HK). H-kininogen forms equimolar complexes with F XI and PK. The local activation of this proteolytic system triggers different cascades such as the surface-dependent activation of blood coagulation, fibrinolysis, kinin generation and inflammation reactions.
Events which allow the assembly of the contact phase components lead to conversion of F XII to the active enzyme F XIIa which triggers the contact phase system. Partially activated V XII cleaves PK which is bound to surfaces via HK. By a mechanism of reciprocal activation, PK amplifies the activity of F XII and in addition PK cleaves HK to release the nona peptide bradykinin. Activated F XII cleaves F XI into its active form which leads to initiation of the intrinsic pathway of coagulation.
Bradykinin and the physiologically important related peptides kallidin (Lys-bradykinin) and Met-Lys bradykinin, contract smooth muscle for example to produce diarrheoa and inflammatory bowel disease and asthma, lower blood pressure, mediate inflammation as in allergies, arthritis and asthma, participate in blood clotting and complement-mediated reaction in the body, mediate rhinitis (viral, allergic and non-allergic) and are over produced in pathological conditions such as acute pancreatitis, hereditary angioneurotic edema, post-gastrectomy dumping syndrome, carcinoid syndrome, anaphylactic shock, reduced sperm mobility and certain other conditions.
As a result of the fact that bradykinin is involved in all the above mentioned clinical indications, a large number of bradykinin antagonists have been developed. Such antagonists are disclosed in e.g. U.S. Pat. No. 4,693,993. Wirth et al., Can. J. Physiol. Pharmacol. Vol. 73 pp 797-804 presents clinical studies regarding administering bradykinin antagonists for treating post-operative pain, asthma, anaphylactoid reactions, systemic inflammatory response syndrome, and suspected sepsis, head injury and hantavirus infections. A review on clinical applications of bradykinin antagonists can be found in Cheronis et al. eds: Proteases, Protease Inhibitors and Protease Derived Peptides pp 167-176. Although these citations disclose administration of bradykinin antagonists for treating sepsis, it is evident that the suspected sepsis treated by said antagonists it not primarily caused by bacterial infections. Accordingly the citations are completely silent about using bradykinin antagonists for treating bacterial infections. Moreover it is evident that bradykinin antagonists have been administered to relieve the symptoms of inflammation.
SUMMARY OF THE INVENTION
Some of the symptoms of sepsis may be explained by activation of the contact phase system. However the precise mechanisms involved are not well understood. The inventors have now established the initial stages involved in activation of the contact phase system during bacterial infection. These studies have led to new assays being proposed to identify agents which specifically target the reactions resulting from bacterial activation of the contact phase system. These studies also demonstrate the disruption of the normal coagulation pathway in bacterial infections and thus propose new substances to treat conditions such as sepsis and septic shock.
The invention provides a number of assays to identify useful agents for the treatment of bacterial infections. In a first aspect, the invention provides an assay for compounds useful in the treatment of a bacterial-induced coagulation disorder, which assay comprises the steps of:
a) incubating a plasma sample with a strain of bacteria;
b) adding a compound to be assayed to the plasma sample before, during or after step a);
c) conducting an activated partial thromboplastin time aPTT test on the sample after steps a) and b);
d) determining the clotting time.
In a second aspect, the invention provides an assay for compounds useful in the treatment of a bacterial-induced coagulation or inflammatory disorder, which assay comprises the steps of:
a) incubating a strain of bacteria with one or more contact phase proteins selected from the group consisting of H-kininogen, prekallikrein, factor XII and factor XI;
b) adding a compound to be assay before, during or after step a); and
c) determining the binding of the contact phase proteins to the bacterial surface.
In a further aspect, the invention provides an assay for compounds useful in the treatment of a bacterial induced coagulation or inflammatory disorder, which assay comprises the steps of:
a) incubating a strain of bacteria with one or more contact phase proteins selected from the group consisting of H-kininogen, prekallikrein, factor XII and factor XI
b) adding the compound to be assayed before, during or after step a); and
c) determining the activation of the or each contact phase protein.
By monitoring the interactions between bacteria and the contact phase components, agents can be identified which may be useful in the treatment of shock, sepsis and bleeding disorders seen in more severe bacterial infections.


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