Diagnosis of infection in the lungs of patients

Abstract

The present invention relates to methods for detecting P aeruginosa infection and bacterial burden in the lungs of patients who are at risk for P. aeruginosa infections, especially including patients with Cystic Fibrosis (CF). The present method provides numerous tests (breath, blood, urine) which are readily administered to a patient that will sensitively and specifically detect the presence and extent of lung infection P. aeruginosa (both mucoid and non-mucoid), and allow monitoring of bacterial load as a parameter in monitoring treatment.

The invention claimed is: 
     
       1. A method for diagnosing a presence of Pseudomonas aeruginosa in the lungs of a subject, including the steps of:
 (a) administering an effective amount of at least one isotopically labeled active agent selected from the group consisting of a nitrate salt, a nitrite salt, nitric oxide (NO), a nitric oxide precursor compound, urea and glycine or their pharmaceutically acceptable salts to the subject, said active agent being acted upon by P. aeruginosa to form an isotopically labeled cleavage product or metabolite; and 
 (b) collecting breath, urine, whole blood, plasma or serum samples from the subject; and 
 (c) analyzing said samples to determine a concentration or concentrations of said cleavage product(s) or metabolite(s), said concentration(s) indicating the presence or absence of P. aeruginosa including optionally, a presence or absence of mucoid phenotype P. aeruginosa, in the lungs of said patient. 
 
     
     
       2. The method according to  claim 1  wherein said active agent is selected from the group consisting of sodium nitrate, potassium nitrate, sodium nitrite, potassium nitrite, nitric oxide, a pharmaceutically acceptable S-nitrosothiol compound, urea, glycine, pharmaceutically acceptable salts thereof and mixtures thereof. 
     
     
       3. The method according to  claim 2  wherein said S-nitrosothiol compound is an alkylthionitrile. 
     
     
       4. The method according to  claim 1  wherein said nitric oxide precursor compound is S-glutathione or S-nitroso-N-acetylpenicillamine. 
     
     
       5. The method according to  claim 1  wherein said isotopically labeled active agent is a mixture of urea, glycine or a pharmaceutically acceptable salt thereof, and optionally, at least one compound selected from the group consisting of an isotopically labeled nitrate salt, nitrite salt, nitric oxide and S-nitrosothiol compound. 
     
     
       6. The method according to  claim 1  wherein said sample is the breath of said subject taken for a predetermined time. 
     
     
       7. The method according to  claim 1  wherein said active agent is isotopically labeled with carbon-13, nitrogen-15, oxygen-17, oxygen-18 or mixtures thereof. 
     
     
       8. The method according to  claim 7  wherein said active agent is urea isotopically labeled with carbon-13, nitrogen-15, oxygen-17 or oxygen-18. 
     
     
       9. The method according to  claim 8  wherein said urea is isotopically labeled with carbon-13 or nitrogen-15. 
     
     
       10. The method according to  claim 1  wherein said active agent is glycine isotopically labeled with nitrogen-15 or carbon-13. 
     
     
       11. The method according to  claim 10  wherein said glycine is isotopically labeled with nitrogen-15. 
     
     
       12. The method according to  claim 1  wherein said urea is isotopically labeled with carbon-13 and said glycine is isotopically labeled with nitrogen-15. 
     
     
       13. The method according to  claim 1  wherein said cleavage product or metabolite is a gas. 
     
     
       14. The method according to  claim 13  wherein said gas is N 2 , N 2 O, NO, CO 2 , HCN or mixtures thereof. 
     
     
       15. The method according to  claim 14  wherein said gas is a mixture of isotopically labeled CO 2 , from the action of P. aeruginosa on urea, and HCN, from the action of P. aeruginosa on glycine and optionally, at least one gas selected from the group consisting of N 2 , N 2 O and NO. 
     
     
       16. The method according to  claim 13  wherein a measured concentration of said gas is indicative of a P. aeruginosa infection in the lungs of said patient. 
     
     
       17. The method according to  claim 14  wherein said gas is HCN or includes HCN and a measured concentration of said HCN gas at a radiometric ratio of at least 1.25 is indicative of a mucoid phenotype of P. aeruginosa in the lungs of said patient. 
     
     
       18. The method according to  claim 1  wherein said isotopically labeled cleavage product or metabolite is selected from the group consisting of nitrate, nitrite, dissolved nitric oxide, carbonate/bicarbonate, cyanate, thiocyanate and mixtures thereof, which is measured in the urine, whole blood, serum or plasma of the patient. 
     
     
       19. The method according to  claim 18  wherein said cleavage product or metabolite is cyanate, thiocyanate or mixtures thereof or includes cyanate, thiocyanate or mixtures thereof and a measured concentration of said cleavage product or metabolite in a radiometric ratio of at least about 1.25 is indicative of a mucoid phenotype of P. aeruginosa. 
     
     
       20. The method according to any of  claims 1 - 16  wherein said analyzing step comprises comparing at least one ratio of isotopically labeled element(s) to non-isotopically labeled element(s) in said exhaled breath of said subject to a control ratio of isotopically labeled element(s) to non-isotopically labeled elements in the exhaled breath of said subject or a control group prior to administration of said isotopically labeled active agent. 
     
     
       21. The method according to  claim 1  wherein said isotopically labeled active agent(s) is delivered to the lungs by pulmonary route of administration. 
     
     
       22. The method according to  claim 1  wherein said isotopically labeled active agent(s) is delivered by oral route of administration. 
     
     
       23. The method according to  claim 22  wherein said active agent is or includes urea and said active agent is administered by oral route in combination with a urease inhibitor. 
     
     
       24. The method according to  claim 22  or wherein said active agent is urea or includes urea and is administered by oral route in an enteric capsule. 
     
     
       25. The method according to  claim 1  wherein isotopically labeled glycine and urea are both administered to said subject, alone or in combination with at least one additional isotopically labeled active agent selected from the group consisting of a nitrate salt, a nitrite salt, NO gas and a NO precursor compound, and wherein the absence of glycine cleavage product, urea cleavage and other cleavage products is evidence of the absence of a P. aeruginosa infection in said subject. 
     
     
       26. The method according to  claim 1  wherein isotopically labeled glycine and urea are both administered to said subject, alone or in combination with at least one additional isotopically labeled active agent selected from the group consisting of a nitrate salt, a nitrite salt, NO gas and a NO precursor compound, and wherein the absence of glycine cleavage product and the presence of urea cleavage product and optionally, other cleavage products is evidence of a non-mucoid P. aeruginosa infection in said subject. 
     
     
       27. The method according to  claim 1  wherein isotopically labeled glycine and urea are both administered to said subject, alone or in combination with at least one additional isotopically labeled active agent selected from the group consisting of a nitrate salt, a nitrite salt, NO gas and a NO precursor compound, and wherein the presence of glycine cleavage product and the presence of urea cleavage product and optionally, other cleavage products is evidence of a mucoid P. aeruginosa infection in said subject. 
     
     
       28. The method according to  claim 1  wherein said urea is cleaved to produce carbon dioxide or ammonia. 
     
     
       29. The method according to  claim 1  comprising optional steps of fitting the concentrations of said cleavage products obtained to a curve; and analyzing the curve or a plateau of said curve to determine the extent of infection. 
     
     
       30. A method for diagnosing a urease-positive bacterial infection in the lungs of a subject comprising:
 administrating to the lungs of the subject, an effective amount isotopically labeled urea;   collecting one or more samples of exhaled breath from the subject; and   analyzing said samples to determine a concentration of isotopically labeled CO 2  in said samples;   said concentration indicating the presence or absence of the urease-positive bacterial infection in the lungs of the subject.   
     
     
       31. The method of claim 30, wherein the isotopically labeled urea is  13 C-urea. 
     
     
       32. The method of claim 30, wherein said samples are taken for a predetermined time. 
     
     
       33. The method of claim 30, wherein said gas is  13 CO 2 . 
     
     
       34. The method of claim 31, wherein the analysis comprises comparing the ratio of  13 CO 2  to  12 CO 2  in said samples to the ratio of  13 CO 2  to  12 CO 2  in a sample obtained from the subject prior to the administration of the isotopically labeled urea. 
     
     
       35. The method of claim 31, wherein an increase in the ratio of  13 CO 2  to  12 CO 2  in said samples to the ratio of  13 CO 2  to  12 CO 2  in a sample obtained from the subject prior to the administration of the isotopically labeled urea.