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Helicobacter Pylori by Acetohydroxamic Acid

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Helicobacter Pylori by Acetohydroxamic Acid Treatment ofacute myeloid leukaemia in a renal allograft recipient 695 cyclosporin may not greatly increase the 1 Anonymous. Multidrug resistance in cancer. [Editorial]. Lancet 1989;ii:1075-6. toxicity ofaggressive chemotherapy used in the 2 Holmes J, Jacobs A, Carter G, et al. Multidrug-resistance in treatment of haematological malignancies. haemopoietic cell lines. Myelodysplastic Syndromes and acute myeloblastic leukaemia. Br J Haematol 1989;72: Sonneveld and Nooter reported a patient with 40-4. J Clin Pathol: first published as 10.1136/jcp.44.8.695 on 1 August 1991. Downloaded from resistent AML to whom they administered 3 Case records ofthe Massachusetts General Hospital. Weekly clinico-pathological exercises. Case 18-1983. A young cyclosporin without any excessive toxicity, man with pancytopenia after a renal transplant. N Engi J although their patient had profound marrow Med 1983;308:1081-91. 4 Ellerton JA, deVeber GA, Baker MA. Erythroleukaemia in a hypoplasia for three weeks.9 renal transplant recipient. Cancer 1979;43:1924-6. Septic shock is a well recognised problem in 5 Pikler GM, Say B, Stamper S. Cytogenetic findings in acute monocytic leukaemia in a renal allograft recipient. Cancer severely neutropenic patients. It is difficult to Genet Cytogenet 1986;20:101-7. be sure that our patient's clinical course would 6 Hoy WE, Packman CH, Freeman RB. Evolution of acute leukaemia in a renal allograft recipient:? Relationship to have been any different had he not received azathioprine. Transplantation 1982;33:331-3. cyclosporin. Although cyclosporin and other 7 Ihle BU, Constable J, Gordon S, Mahony JF. Myelodys- plasia in cadaver renal allografts: A report of four cases. inhibitors of P-glycoprotein may improve the Am J Kidney Dis 1985;5:251-7. response to chemotherapy, the use of 8 Butler J, Korb S, Light L. Acute myelogenous leukaemia in a renal allograft recipient receiving cyclosporine therapy. immunosuppressiVe agents during the treat- Transplantation 1990;49:813-15. ment of acute myeloid leukaemia does require 9 Sonneveld P, Nooter K. Reversal of drug-resistance by cyclosporin-A in a patient with acute myelocytic leuk- caution. Non-immunosuppressive cyclosporin aemia. Br J Haematol 1990;75:208-1 1. analogues may produce the desired positive 10 Twentyman PR, Fox NE, White DJG. Cyclosporin A and its analogues as modifiers of adriamycin and vincristine effects of P-glycoprotein inhibition without resistance in a multidrug resistant human lung cancer cell increased infection risk.'" line. Br J Cancer 1987;56:55-7. J Clin Pathol 1991;44:695-697 Inhibition of urease activity but not growth of http://jcp.bmj.com/ Helicobacter pylori by acetohydroxamic acid J Goldie, S J 0 Veldhuyzen van Zanten, S Jalali, H Richardson, R H Hunt on September 27, 2021 by guest. Protected copyright. Abstract infections, associated with struvite stone for- The in vitro effects of acetohydroxamic mation, in which urea splitting organisms are acid (AHA), a potent urease inhibitor, important.7 AHA prevents alkalinisation of Division of were studied to determine the effect on the urine by inhibiting urease, thus prevent- Gastroenterology and Department of the urease activity and growth of 38 ing hydrolysis of urea and subsequent produc- Laboratory Medicine, strains of Helicobacter pylori. AHA in tion of ammonia. McMaster University concentrations of 50-1000 mg/l had a The high urease activity of H pylori might Medical Centre, 1200 Main Street West, noticeably reversible inhibitory effect on be inhibited by AHA and we therefore studied Hamilton, Ontario the urease activity of the organism but this in vitro to determine whether AHA L8N 3Z5, Canada no effect on growth. inhibits urease activity and the growth of H J Goldie S Jalali pylori. H Richardson R H Hunt Helicobacter pylori has a very high urease Division of activity which is thought to be related to its Methods Gastroenterology, pathogenicity, allowing it to colonise and Thirty three recent clinical isolates and five Dalhousie University, Victoria General survive in the harsh gastric environment.- reference strains (obtained from LCDC, Hospital, Halifax There is a need for a more effective treat- Ottawa) of H pylori were grown micro- SJO Veldhuyzen van ment against H pylori because currently aerobically at 35°C for five days. Dense sus- Zanten available treatments are unsatisfactory.2 pensions were made in 3 mmol monobasic Correspondence to: J Goldie Acetohydroxamic acid (AHA) is a potent sodiumphosphate buffer (NaH2PO4) contain- Accepted for publication inhibitor of the enzyme urease.3 AHA has ing a concentration of AHA to approximate a 12 February 1991 been used in the treatment of urinary tract final concentration of 10i organisms/ml when 696 Goldie, Veldhuyzen van Zanten, Jalali, Richardson, Hunt compared with a McFarland 0-5 opacity stan- that inhibition is reversible and does not affect dard. Parallel control suspensions of H pylorn the growth of the organism. We wanted to with buffer but without AHA were included in study urease activity during optimal growth each test series. The pH of the solutions was conditions of the organism, which is at J Clin Pathol: first published as 10.1136/jcp.44.8.695 on 1 August 1991. Downloaded from + 6. pH + 5 0-8-0 and is adversely affected by AHA was prepared in an aqueous solution lower pH. Because the organism lives in the of 10 mg/ml. Dilutions were made in 3 mmol gastric mucus where the pH is neutral, these phosphate buffer (pH 6.0) to final concentra- studies were performed at a pH of 6-7. tions of 1000, 500, 250, 100 and 50 mg/I. The characteristic high preformed urease Twenty five microlitires of Hpylori suspen- activity of H pylori is considered important sions in AHA solutions were inoculated into for its pathogenicity. Pathophysiological bottles containing 1 5 ml ofbuffered urea solu- mechanisms that have been proposed are tion of pH + 6 (solution contains 1 ml of hydrogen ion back-diffusion'0 and interference Sigma urea standard solution and 49 ml of with the trichoracetic acid cycle. The latter 3 mmol phosphate buffer). We tested urease results in a decreased production of adenosine activity in organisms that had grown triphosphate (ATP) in aerobic cells which microaerobically for five days rather than three may inhibit normal cell function." Further- days because we found the urease production more, urease activity allows the organism to to be greatest at five days. Ammonia concen- use urea as an important nitrogen source. trations were measured after 30 minutes at The urease of H pylori differs from other room temperature using the Kodak Ektachem strong urease producers such as Proteus chemical analyser.8 Standard crystalline jack- mirabilis in showing much higher affinity for bean urease (sigma) was used as standard. The urea and higher activity of the enzyme, which urease activity of the AHA solutions (nmol/ may be necessary for the survival of the residual NH3/1) was compared with control organism as urea is less available in the gastric suspension without AHA, and background mucus than in urine.' activity of ammonia was substracted. The Although initially AHA was believed to average results of three to five experiments per cause irreversible inhibition of urease strain are shown (table). The coefficient of activity,3 later studies, which are confirmed by variation of this method was 4%. Urease our study, show that inhibition is temporary.'2 activity of the suspensions was also checked The inhibitory effect of AHA is highly specific after four and 24 hours using our own rapid for urease.6 The optimal pH of the urease urease test.9 activity is about 8 2' and corresponds to Subcultures were made after 24 hours from optimal growth conditions of H pylori. the inoculated AHA suspensions and control (Goldie J, Hollingsworth J, Hunt RH. suspensions using chocolate agar and buffered Campylobacter pylori multidisciplinary work- peptone medium with 5% horse serum. Both shop, Keystone, Colorado, 1987). Hydrox- media were at pH 7 0. Each was incubated amic acids probably do not have clinically microaerobically for five days. important antimicrobial activity, although one http://jcp.bmj.com/ report indicates that AHA is bacteriostatic against several bacteria.4 '3 In vitro, both syn- Results ergy and antagonism have been found when The table shows that AHA inhibited the AHA was combined with other antibiotics urease activity of H pylori. The highest level against Gram negative urease producing of inhibition occurred with AHA concentra- organisms.4"' Furthermore, synergism may be tions of 200-1000 mg/l and as there were only pH dependent, which may be relevant for H on September 27, 2021 by guest. Protected copyright. minimal differences among strains the average pylori, given that the organism must establish result for each AHA concentration is shown. itself in the acidic environment of the The inhibition of urease is only temporary stomach.'3 because results of the rapid urease test were The mechanism by which AHA inhibits positive again after 24 hours. AHA did not urease is unknown, but because growth is not affect the survival of the organism. inhibited it does not inactivate the whole organism. Possible mechanisms include inhibition of enzyme activity or enzyme syn- Discussion thesis, or blocking contact of the enzyme with The results show that AHA is a potent the substrate urea. It has been shown that the inhibitor of the urease activity of H pylori but cell wall of the organism is no barrier for AHA.' Our results suggest that AHA when used Results of average residual ammonia production, rapid urease test, andgrowth of suspensions containing only H pylori and suspensions containing the organism with AHA alone will not result in eradication of the organism because growth is not affected. Average NH, Rapid urease test Inhibition of the urease activity, however, production in Growth of Suspension inpmol/l (SD) 4 hours 24 hours Hpylori could have an adjuvant therapeutic role in the treatment of H pylori by making the organism All H pylori suspensions > 900 + + + without AHA more susceptible to other antimicrobial AHA 1000mg/l 41(8) - + + agents.
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