There are currently renewed efforts to develop drugs that could shorten the duration of antituberculosis therapy. This is best achieved by optimizing the sterilizing effect. However, the current pathway for the development of new molecules with the potential to have a sterilizing effect is inefficient. We designed an in vitro pharmacokinetic-pharmacodynamic model in which Mycobacterium tuberculosis replicating slowly at pH 5.8 was exposed to pyrazinamide by use of the concentration-time profiles encountered in patients. The sterilizing effect rates and the time to the emergence of drug resistance were examined. Daily pyrazinamide dosing for 28 days accurately achieved (i) the pyrazinamide pharmacokinetic parameters, (ii) the lack of early bactericidal activity, (iii) a sterilizing effect rate of 0.10 log 10 CFU/ml per day starting on day 6 of therapy, and (iv) a time to the emergence of resistance of the from 2 to 3 weeks of monotherapy encountered in patients with tuberculosis. Next, dose-scheduling studies were performed. Current therapy for tuberculosis (TB) consists of rifampin (rifampicin), isoniazid, and pyrazinamide (5). In order to improve therapy, several studies have examined the pharmacokinetic (PK)-pharmacodynamic (PD) relationship between isoniazid and rifampin treatment and the response of Mycobacterium tuberculosis (22,25,29,30). However, those studies were of short duration and mainly reflected the bactericidal activities of these agents, defined as the killing of bacilli in log-phase growth. No similar studies have examined the sterilizing activities of anti-TB drugs. An anti-TB drug is considered to have a sterilizing effect when it is able to kill one of two subpopulations of M. tuberculosis: slowly growing bacilli in an acidic environment (slowly replicating bacilli [SRB]) and nonreplicating persistent bacilli (5, 26). The sterilizing activity of a drug also reflects its ability to shorten anti-TB therapy duration (41). Pyrazinamide has unique sterilizing activity against SRB, which are estimated to constitute the majority of bacilli in the pulmonary cavities of patients with TB (31,35). A measure of the size of the SRB population is that the addition of pyrazinamide to isoniazid-and rifampin-containing regimens led to a one-third reduction in the duration of therapy and a two-thirds reduction in the rate of TB relapse (2, 48). Given the size of this population and the central role of pyrazinamide in eradicating it, pyrazinamide PK-PD studies could help with the further optimization of therapy.After oral administration, pyrazinamide is almost completely systematically absorbed and eventually enters the pulmonary cavities containing M. tuberculosis. Pyrazinamide works in these cavities under acidic conditions (38). There is conflicting evidence as to whether pyrazinamide enters the bacilli by simple diffusion or by active uptake (44, 56). However, once the bacilli are in the cell, bacillary nicotinamidase deaminates the pyrazinamide to pyrazinoic acid (POA), which is transported to the extrace...

These data, based on a preclinical model, demonstrate that nonadherence alone is not a sufficient condition for MDR-tuberculosis emergence.

ABSTRACTWe hypothesize that low-level efflux pump expression is the first step in the development of high-level drug resistance in mycobacteria. We performed 28-day azithromycin dose-effect and dose-scheduling studies in our hollow-fiber model of disseminatedMycobacterium avium-M. intracellularecomplex. Both microbial kill and resistance emergence were most closely linked to the within-macrophage area under the concentration-time curve (AUC)/MIC ratio. Quantitative PCR revealed that subtherapeutic azithromycin exposures over 3 days led to a 56-fold increase in expression of MAV_3306, which encodes a putative ABC transporter, and MAV_1406, which encodes a putative major facilitator superfamily pump, inM. avium. By day 7, a subpopulation ofM. aviumwith low-level resistance was encountered and exhibited the classic inverted U curve versus AUC/MIC ratios. The resistance was abolished by an efflux pump inhibitor. While the maximal microbial kill started to decrease after day 7, a population with high-level azithromycin resistance appeared at day 28. This resistance could not be reversed by efflux pump inhibitors. Orthologs of pumps encoded by MAV_3306 and MAV_1406 were identified inMycobacterium tuberculosis,Mycobacterium leprae,Mycobacterium marinum,Mycobacterium abscessus, andMycobacterium ulcerans. All had highly conserved protein secondary structures. We propose that induction of several efflux pumps is the first step in a general pathway to drug resistance that eventually leads to high-level chromosomal-mutation-related resistance in mycobacteria as ordered events in an “antibiotic resistance arrow of time.”

Background In the treatment of tuberculosis, ethambutol is used in case there is isoniazid resistance. We examined for the emergence of drug resistance to ethambutol monotherapy in pharmacokinetic-pharmacodynamic studies in the hollow fiber system. Methods Dose–effect studies and dose-scheduling studies were carried out with ethambutol and log-phase growth Mycobacterium tuberculosis to identify exposures and schedules linked to optimal kill and resistance suppression. In one study, after 7 days of ethambutol 300 mg/day isoniazid was administered to each system to determine its early bactericidal activity (EBA). Results Efflux-pump blockage reduced the mutation frequency to ethambutol 64-fold. In dose-effect studies, ethambutol had a maximal EBA of 0.22 log10 CFU/ml/day, as encountered in patients. By day 7, resistance had arisen to both ethambutol and isoniazid. Prior exposure to ethambutol abolished isoniazid EBA. Daily therapy, as opposed to more intermittent therapy, was associated with the least proportion of efflux-pump driven resistance, consistent with time driven effect. Microbial kill was best explained by the ratio of area under the concentration-time curve to minimal inhibitory effect (r2=0.90). Conclusion Induction of an efflux-pump that reduces effect to multiple drugs provides an alternative pathway to sequential acquisition of mutations in the development of multiple drug resistance.

Background & Aims: The therapeutic mechanisms of ribavirin for hepatitis C are unclear. Microarray analyses have shown that ribavirin increases induction of interferon-stimulated genes (ISGs). We evaluated viral kinetics, serum cytokine expression, and viral mutagenesis during early stages of peginterferon therapy with and without ribavirin.

ATV/RTV significantly increases the plasma concentrations of rosuvastatin, most likely by increasing rosuvastatin's oral bioavailability. Dose limitations of RSV with ATV/RTV may be needed.

Ethambutol, together with a macrolide, is the backbone for treatment of disseminated Mycobacterium avium disease. However, at the standard dose of 15 mg/kg of body weight/day, ethambutol efficacy is limited. In addition, susceptibility breakpoints have consistently failed to predict clinical outcome. We performed doseeffect studies with extracellular M. avium as well as with bacilli within human macrophages. The maximal kill rate (E max ) for ethambutol against extracellular bacilli was 5.54 log 10 CFU/ml, compared to 0.67 log 10 CFU/ml for intracellular M. avium, after 7 days of exposure. Thus, extracellular assays demonstrated high efficacy. We created a hollow-fiber system model of intracellular M. avium and performed microbial pharmacokineticpharmacodynamic studies using pharmacokinetics similar to those of ethambutol for humans. The E max in the systems was 0.79 log 10 CFU/ml with 7 days of daily therapy, so the kill rates approximated those encountered in patients treated with ethambutol monotherapy. Ratio of peak concentration to MIC (C max /MIC) was linked to microbial kill rate. The C max /MIC ratio needed to achieve the 90% effective concentration (EC 90 ) in serum was 1.23, with a calculated intramacrophage C max /MIC ratio of 13. In 10,000 patient Monte Carlo simulations, doses of 15, 50, and 75 mg/kg achieved the EC 90 in 35.50%, 76.81%, and 86.12% of patients, respectively. Therefore, ethambutol doses of >50 mg/kg twice a week would be predicted to be better than current doses of 15 mg/kg for treatment of disseminated M. avium disease. New susceptibility breakpoints and critical concentrations of 1 to 2 mg/liter were identified for the determination of ethambutol-resistant M. avium in Middlebrook broth. Given that the modal MIC of clinical isolates is around 2 mg/liter, most isolates should be considered ethambutol resistant.Bacteria of the Mycobacterium avium complex (MAC) are not only a cause of important opportunistic infections in patients with AIDS and cancer but also an important cause of pulmonary disease in "immunocompetent" patients (5,17,26,31). In immunocompromised patients, disseminated disease is common and lethal (8, 10). Current therapy is based on a backbone of ethambutol, concurrent with either azithromycin or clarithromycin, administered for many months to several years (17,18). This therapy fails in up to half of patients and therefore still needs to be optimized.The first test used to examine the therapeutic potential of ethambutol, as for all other antibiotics, is MIC determination. With this test, it is hoped that discrimination can be made as to whether ethambutol should be used in the patient from whom M. avium was isolated. Unfortunately, susceptibility studies have failed to predict the effectiveness of ethambutol in patients with disseminated M. avium disease in the past (5, 34, 39). We hypothesize that part of the problem has been an arbitrarily chosen susceptibility breakpoint, which did not take into account population pharmacokinetics and microbial pharmacokinetics-ph...

Background NAD(P)H:quinone oxidoreductase 1 (NQO1) is a two-electron oxidoreductase expressed in multiple tumour types. ARQ 761 is a β-lapachone (β-lap) analogue that exploits the unique elevation of NQO1 found in solid tumours to cause tumour-specific cell death. Methods We performed a 3+3 dose escalation study of 3 schedules (weekly, every other week, 2/3 weeks) of ARQ 761 in patients with refractory advanced solid tumours. Tumour tissue was analysed for NQO1 expression. After 20 patients were analysed, enrolment was restricted to patients with NQO1-high tumours ( H -score ≥ 200). Results A total of 42 patients were treated. Median number of prior lines of therapy was 4. Maximum tolerated dose was 390 mg/m 2 as a 2-h infusion every other week. Dose-limiting toxicity was anaemia. The most common treatment-related adverse events were anaemia (79%), fatigue (45%), hypoxia (33%), nausea (17%), and vomiting (17%). Transient grade 3 hypoxia, reflecting possible methemoglobinaemia, occurred in 26% of patients. Among 32 evaluable patients, best response was stable disease ( n = 12); 6 patients had tumour shrinkage. There was a trend towards improved efficacy in NQO1-high tumours ( P = 0.06). Conclusions ARQ 761 has modest single-agent activity, which appears associated with tumour NQO1 expression. Principal toxicities include anaemia and possible methemoglobinaemia.