The activities of rifapentine and rifampin against Mycobacterium tuberculosis residing in human monocytederived macrophages were determined. The MICs and MBCs of rifapentine for intracellular bacteria were twoto fourfold lower than those of rifampin. For extracellular bacteria, this difference was less noticeable. Nevertheless, the more favorable pharmacokinetics of rifapentine over rifampin was addressed in other experimental models. These models showed substantial differences after short pulsed exposures of the infected macrophages to the drugs and when the infected macrophages were exposed to changing drug concentrations that imitated the pharmacokinetic curves observed in blood. Once-a-week exposures to rifapentine concentrations equivalent to those attained in blood after one 600-mg dose resulted during the first week in a dramatic decline in the number of bacteria, and this decline was maintained at a minimal level for a period of four weeks. The results of this study have shown the suitability of rifapentine for intermittent-treatment regimens. The prolonged effect of rifapentine found in this study may be associated with high ratios of intracellular accumulation, which were four-to fivefold higher than those found for rifampin. Further studies on the intracellular distribution of rifamycins and on the sites of actual interaction between the drugs and bacteria residing in macrophages are necessary.The combined use of isoniazid, rifampin, and pyrazinamide provided the basis for a successful short course of tuberculosis therapy. The discovery of long-lasting rifamycins, particularly of rifapentine (RPT), created the possibility for intermittent drug regimens (5,7,18). In the early studies, RPT was reported to have an in vitro activity the same as or higher than that of rifampin (RMP) (19,20). Subsequent studies have shown that the broth-determined MICs of RPT were two-to threefold lower than those of RMP (8, 10). The elimination half-life of RPT in animals and humans was about five times longer than that of RMP, and the levels of RPT in plasma substantially exceeded the MICs for a period of up to 72 h after a single oral dose (1)(2)(3)12). The advantage of RPT over RMP has been shown in murine models (1,5,11,18). Besides its more favorable pharmacokinetic parameters in blood, RPT accumulates to higher concentrations than does RMP in polymorphonuclear leukocytes and macrophages (6,9,16).In anticipation of a controlled clinical trial for the comparison of RPT and RMP in the therapy of tuberculosis, the present study compares the potential activities of these rifamycins against extracellular and intracellular Mycobacterium tuberculosis. Inhibitory and bactericidal activities (MICs and MBCs) against intracellular bacteria in human monocyte-derived macrophages were determined and compared with MICs and MBCs against extracellular bacteria. The actual accumulation of these drugs in human monocytes was also determined. Special attention was given to the evaluation of the long-lasting effect of RPT after pulsed expo...

Aycobacterium avium strains susceptible to clarithromycin and azithromycin contain mutants resistant to these macrolides with a frequency of 1.1 x 10-10 to 1.2 x 10-6. Cross-resistance between clarithromycin and azithromycin was demonstrated with mutants selected in the laboratory as well as with resistant strains isolated from patients. The susceptibility-resistance patterns of the macrolide-resistant strains with drugs other than macrolides were the same as those of the onrginal susceptible strains. The emergence of clarithromycin resistance in patients was a result of multiplication of the preexisting resistant mutants that survived the elimination of bacteria during the initial period of treatment and was an exclusive cause of the relapse of bacteremia.Clarithromycin, a new macrolide, has been shown to have activity against Mycobacterium avium in various models including experiments in culture media (1,2,4,7,9,12,17,22), macrophages (16,18,19,21,24), and mice (7,13,14). The effectiveness of this drug in the elimination of M. avium bacteremia in patients with AIDS has been shown in controlled clinical trials (3, 6, 10). Another macrolide, azithromycin, has also been shown to be effective in the therapy of M. avium infections in patients with AIDS (23,26). Clinical trials with both macrolides have had limited periods of observation, usually less than 12 weeks. Recent reports on prolonged monotherapy with clarithromycin or azithromycin showed that after a period of elimination of the bacteria, a relapse of bacteremia took place in many patients (3,5,23,26). Those reports also implied that prolonged monotherapy may lead to the emergence of drug resistance.The goals of the present study were to address the following various aspects of drug resistance of M. avium to new macrolides, especially clarithromycin: (i) the frequency of clarithromycin-and azithromycin-resistant mutants in susceptible M. avium strains and the degree of resistance of these mutants, (ii) cross-resistance between clarithromycin and azithromycin, (iii) comparison of the drug susceptibilityresistance patterns between the original strains and their resistant mutants in experiments with other drugs, and (iv) drug susceptibilities of strains isolated from patients with AIDS and bacteremia at the beginning of therapy and at the time of relapse. In the present study, we analyzed the bacteriological response to clarithromycin monotherapy and presented a general description of the phenomenon of the emerging resistance of M. avium to this drug. The genetic basis of macrolide resistance in M. avium was not among the goals of the present study. Neither did we make an attempt to address such issues as the rate of mutation or mechanisms of resistance.MATERIALS AND METHODS Antimicrobial agents. Clarithromycin was obtained from Abbott Laboratories (Abbott Park, Ill.). The drug was dissolved in dimethyl sulfoxide (DMSO; 100 mg/10 ml). This stock solution was diluted further with DMSO to make * Corresponding author. appropriate working solutions, which were ke...

Levofloxacin exhibited twofold greater inhibitory and bactericidal activities than ofloxacin against either extracellular or intracellular tubercle bacilli. The activities of both drugs against extracellular and intracellular bacteria were about the same, despite the accumulation of the drugs in macrophages at a level four-to fivefold greater than that in the extracellular medium. The activities of both drugs against intracellular bacteria were largely associated with the short, 2-h pulsed exposures of the infected macrophages to the concentrations which correspond to those attainable in blood during the period of the maximum concentration of drug in serum.Among the quinolones, ofloxacin was the first agent used to treat patients with tuberculosis (16). Comparison of the in vitro activities of various quinolones indicated that ofloxacin and ciprofloxacin had better potentials than norfloxacin, amifloxacin, perfloxacin, and other compounds of this class for the treatment of tuberculosis (1, 2, 5, 7, 9, 13-15, 17, 18). The MICs of both ofloxacin and ciprofloxacin in various types of liquid media and in 7H10 or 7H11 agar ranged from 0.12 to 2.0 ,ug/ml (10). The MBCs of these two drugs determined in 7H12 broth were 2.0 ,ug/ml, giving MBC/MIC ratios of 2 to 4 (10, 15). Levofloxacin, the optically active L-isomer of ofloxacin, has shown high levels of antibacterial activity both in vitro and in mice against a number of bacterial species, being either as active as or up to fourfold more active than ofloxacin (8).The aim of the study described here was the preclinical evaluation of the activity of levofloxacin in comparison with that of ofloxacin against Mycobacterium tuberculosis in human monocyte-derived macrophages as well as in cell-free medium.Antimicrobial agents. Ofloxacin and levofloxacin were obtained from the R. W. Johnson Pharmaceutical Research Institute (Springhouse, Pa.). Stock solutions were made in phosphate buffer (pH 6.5). These stock solutions were diluted further to make appropriate working solutions.Test strains. Studies in macrophages were conducted with two M. tuberculosis strains: H37Rv, which is susceptible to all antituberculosis drugs, and a Vertullo strain (2937-0), one of the quality control strains in our laboratory, which is resistant to isoniazid, rifampin, streptomycin, and ethambutol. The same two strains and an additional nine clinical isolates were used for determination of the MICs in vitro. Subcultures in 7H9 broth were preserved in aliquots frozen at -70°C. For each experiment, a subculture of frozen stock was made in 7H9 broth. After S to 6 days of cultivation at 37°C, the culture was forced through a 27-gauge needle and was then centrifuged at 250 x g for 5 min to remove large clumps of bacteria.Model of the M. tuberculosis-infected macrophages. The technique for preparation of the human monocyte-derived macrophage monolayers was as described previously by Crowle and colleagues (3, 4), with some modifications given in our * Corresponding author. Mailing address: National Jewish C...

Early bactericidal activity (EBA) of antituberculosis drugs is the rate of decrease in the concentration of tubercle bacilli sputum during the initial days of therapy. The study reported here was designed to optimize the methodology for obtaining precise EBA measurements. The study compared the results with two versus five treatment days; overnight sputum collections with early morning collections; and quantitative smears for acid-fast bacilli (AFB) with quantitative cultures. Isoniazid (INH) was used as a model drug. Among 28 smear-positive patients enrolled in the study in five cities in the United States, 16 were evaluable (INH-susceptible tuberculosis [TB] and adequate sputum collections). The mean baseline bacterial load was 6.69 log10 cfu/ml (SE = 0.24). Quantitative culture of 10- or 12-h sputum collections obtained on two baseline days and treatment Day 5 was the optimal method for EBA measurement. The mean 5-d EBA was 0.21 log10 cfu/ml/d (SE = 0.03; p < 0.001) and the EBA appeared to be constant during the first five treatment days. On the basis of these data, multiarm studies of investigational drugs will require 25 evaluable subjects per arm to detect (80% power and two-tailed alpha of 0.05) an EBA at least 50% as large as the EBA of INH. In countries with a low incidence of TB, the usefulness of this methodology for rapidly assessing new antituberculosis agents may be limited by the relatively large number of subjects required to compare EBA values across treatment arms.