https://orcid.org/0000-0001-7592-8012
Abstract
dcycloserine is used to treat multidrug-resistant tuberculosis. Its efficacy, contribution in combination therapy, and best clinical dose are unclear, also data on thedcycloserine minimum inhibitory concentration (MIC) distributions is scant.
We performed a systematic search to identify pharmacokinetic and pharmacodynamic studies performed withdcycloserine. We then performed a combined exposure-effect and dose fractionation study ofdcycloserine in the hollow fiber system model of tuberculosis (HFS-TB). In parallel, we identifieddcycloserine MICs in 415 clinicalMycobacterium tuberculosis(Mtb) isolates from patients. We utilized these results, including intracavitary concentrations, to identify the clinical dose that would be able to achieve or exceed target exposures in 10000 patients using Monte Carlo experiments (MCEs).
There were no publisheddcycloserine pharmacokinetics/pharmacodynamics studies identified. Therefore, we performed new HFS-TB experiments. Cyloserine killed 6.3 log10colony-forming units (CFU)/mL extracellular bacilli over 28 days. Efficacy was driven by the percentage of time concentration persisted above MIC (%TMIC), with 1.0 log10CFU/mL kill achieved by %TMIC= 30% (target exposure). The tentative epidemiological cutoff value with the Sensititre MYCOTB assay was 64 mg/L. In MCEs, 750 mg twice daily achieved target exposure in lung cavities of 92% of patients whereas 500 mg twice daily achieved target exposure in 85% of patients with meningitis. The proposed MCE-derived clinical susceptibility breakpoint at the proposed doses was 64 mg/L.
Cycloserine is cidal againstMtb. The susceptibility breakpoint is 64 mg/L. However, the doses likely to achieve the cidality in patients are high, and could be neurotoxic.
dcycloserine was discovered by 2 independent teams as a secondary metabolite ofStreptomyces orchidaceusin 1954 [1,2]. Results of its first clinical use were published a year later [3].dcycloserine is a World Health Organization (WHO) group C second-line agent, whose main role is in treatment of multidrug-resistant tuberculosis (MDR-TB). Neuropsychiatric toxicity is common, especially psychosis and seizures, which are encountered in 10%–50% of patients [4]. Because these adverse events are possibly concentration-dependent, it will be imperative to identify doses that could killMycobacterium tuberculosis(Mtb) at concentrations below those associated with toxicity. Here, we performed a systematic analysis to identify the pharmacokinetics (PK) and pharmacodynamics (PD) ofdcycloserine as related to dosing, and if there were gaps to design studies to fill them.
The mechanisms of action ofdcycloserine, an analogue ofd-alanine, are unclear, but there are likely multiple targets, and several resistance mechanisms have been described to date [5–10]. However, the question of whatdcycloserine adds to the current MDR-TB treatment regimens remains. In one murine MDR-TB study,dcycloserine 300 mg/kg/day for 5 months had no effect on lung or spleenMtbburden as monotherapy, and in combination with moxifloxacin did not add any effect to moxifloxacin monotherapy [11]. The possible lack of effect in animal models has been attributed by others to the rapid elimination ofdcycloserine from mice, and potentially to antagonism from naturally abundantd-alanine in mice and guinea pigs [12]. To avoid use of animal sera that may containd-alanine, we examined the efficacy ofdcycloserine in the hollow fiber system model of tuberculosis (HFS-TB), an extracellular model, which utilizes Middlebrook 7H9 broth and relies onl-glutamic acid as the α-amino acid. The HFS-TB has been used extensively to study many first- and second-line agents, with good translational accuracy to patient outcomes [13–16].
METHODS
Systematic Review
Details and steps on our systematic review ofd我们环丝氨酸药物动力学和PK / PD研究re as given in the introduction to this supplement [17]. The search terms were used to query PubMed and Web of Science, with date of last search of 13 September 2017. In the search terms detailed in the introduction, “drug name” was “cycloserine” OR “dcycloserine”, while “alternative drug name” was “SC-49088.”
Materials, Organisms, and Reagents
MtbH37Ra (American Type Culture Collection [ATCC] 25177) was utilized in HFS-TB experiments, as explained in detail elsewhere [18].dcycloserine and niacin (internal standard) were purchased from Sigma-Aldrich (St Louis, Missouri). Hollow fiber cartridges were obtained from FiberCell (Frederick, Maryland). The BACTEC 960 mycobacterial growth indicator tube (MGIT) system (BD, Franklin Lakes, New Jersey) was utilized for monitoring growth and time-to-positivity (TTP).
Minimum Inhibitory Concentrations of Laboratory Strains
Thedcycloserine minimum inhibitory concentrations (MICs) forMtbH37Ra (ATCC 25177) and H37Rv (ATCC 27294) were determined using 4 methods: Sensititre MYCOTB plate, macrobroth dilution, as well as the 1% indirect proportion method using Middlebrook 7H10, and MGIT [19–21]. For the latter 3 methods, the concentrations 0, 0.5, 1, 2, 4, 8, 16, 32, and 64 mg/L were tested. Next, we examined the microbial kill effect of differentdcycloserine concentrations against extracellularMtbin test tubes and intracellularMtbin infected THP-1 cells that were activated for 72 hours with 100 nM of phorbol 12-myristate 13-acetate 12-well plates, and coincubated with drug, using protocols described previously [19,20].
MICs in MDR-TB and Extensively Drug-resistant TB Clinical Isolates From 4 Countries
First, we performed a literature search to identify anydcycloserine MIC distributions. Next, a total of 415 pretreatmentMtbisolates cultured from patients enrolled in observational cohorts or from laboratory surveillance studies were cultured and species confirmed by DNA probe [22–30]. MIC testing was performed with the Sensititre MYCOTB plate (Trek Diagnostic Systems, Cleveland, Ohio) at the mycobacterial laboratories of Siriraj Hospital, Mahidol University in Bangkok, Thailand; the International Centre for Diarrhoeal Disease Research in Dhaka, Bangladesh; Kilimanjaro Clinical Research Institute in Moshi, Tanzania; and the Irkutsk Clinical Tuberculosis Hospital in Irkutsk, Russian Federation. MIC data from theseMtb隔离之前发表的研究of comparative drug-susceptibility methodologies [22–30]. MYCOTB plate results were performed in batches, as previously described [31]. Growth was evaluated visually with a manual viewer at 10–21 days by 2 independent technicians. The MIC was recorded as the lowest antibiotic concentration that prevented visible growth. The H37Rv laboratory strain ATCC 27294 was used for quality control, in each batch. The upper end of the phenotypically wild-type MIC distribution was identified as the tentative epidemiological cutoff (ECOFF) [32].
Hollow Fiber System Model of Tuberculosis
The construction details of the HFS-TB have been described before [16,33–35]. HFS-TB conditions for log-phase growthMtbfordcycloserine are described in detail in the introduction to this supplement [17].Mtbwas inoculated into 10 HFS-TB units, and treated withd环丝氨酸剂量开始24小时后,心肌梗死mic a half-life of 10 hours.dcycloserine was administered daily to 7 HFS-TB units to achieve peak concentrations of 0, 13, 55, 96, 180, 219, and 257 mg/L whereas 3 of the HFS-TB units were dosed once every week with the lowest, third-lowest, and fourth-lowest daily doses given cumulatively (ie, daily dose times 7 given as single dose each week) to break the co-linearity between the PK/PD indices that would otherwise occur with dose escalation. Treatment duration was for 28 days. The central compartment was sampled fordcycloserine concentrations at 0, 1, 6, 11, 21, 23.5, 48, 72, 96, 120, 144, and 168 hours after the last dose.dcycloserine concentrations in these samples were measured using the assay described in theSupplementary Methods. The peripheral compartment was sampled on days 0, 3, 5, 7, 10, 14, 21, and 28 for estimation ofMtbburden using MGIT TTP and colony-forming units (CFU) on Middlebrook 7H10 agar [19,20,35].dcycloserine–resistant colonies were captured by cultures on agar supplemented with 3 times thedcycloserine MIC.
Pharmacokinetic and Pharmacodynamic Modeling
Drug concentrations measured in the central compartments of HFS-TB were modeled using ADAPT-5 software. The pharmacokinetic models were used to calculate the 0- to 24-hour area under the concentration–time curve (AUC0–24) and percentage of the 24-hour dosing interval that concentration persisted above MIC (%TMIC), peak concentration to MIC (Cmax/MIC), and AUC0–24/MIC, which were modeled for microbial kill and resistance as outlined in the introduction to this supplement [17]. Optimal exposures were defined as either (1) the exposure associated with 80% of maximal kill (EC80), (2) the exposure associated with cidal effect (1.0 log10CFU/mL kill below day 0), or (3) the lowest exposure associated with suppression of acquired drug resistance (ADR), which are standard PK/PD definitions [36,37].
Monte Carlo Experiments for Dose Selection
The rationale and steps for Monte Carlo experiments (MCEs) are described in the introduction to this supplement [17]. We utilized MCEs to determine the proportion of 10000 patients treated withdcycloserine doses of 250 mg, 500 mg, 750 mg, 1 g, and 1.5 g each day who would achieve the target exposure [17,18]. Fordcycloserine population pharmacokinetics, we used the results of Alsultan et al (contributed to us by Dr Charles Peloquin) based on 130 patients who had MDR-TB, as well by Chang et al, shown inTable 1[38,39]. For lung cavity penetration ratios ofdcycloserine, we used the mean ± standard deviation lung cavity-to-serum penetration ratios of 0.063 ± 0.026 among those who had detectable cycloserine cavitary concentrations [40]. The penetration ofdcycloserine into cerebrospinal fluid (CSF) is about 80%–100% of concurrent serum concentrations in inflamed meninges; case reports suggest that the clearance from subarachnoid space may be slower than in serum [41,42]. Thus, we utilized a CSF-to-serum penetration ratio of 1.0, essentially the same concentrations as in the blood.
d-Cycloserine Pharmacokinetic Parameters and Variances
| Parameter | Parameters Used in Subroutine PRIOR, Mean ± SD |
10000 Simulated Subjects, Mean ± SD |
|---|---|---|
| Clearance, L/h | 1.16 ± 0.73 | 1.14 ± 0.82 |
| Volume, L | 10.50 ± 3.15 | 10.50 ± 1.79 |
| Absorption constant, h-1 | 5.40 ± 2.11 | 5.40 ± 1.58 |
| TLag, h | 0.46 ± 0.45 | 0.47 ± 0.67 |
| Peak, mg/L, for 250 mg | … | 22.25 ± 3.73 |
| Parameter | Parameters Used in Subroutine PRIOR, Mean ± SD |
10000 Simulated Subjects, Mean ± SD |
|---|---|---|
| Clearance, L/h | 1.16 ± 0.73 | 1.14 ± 0.82 |
| Volume, L | 10.50 ± 3.15 | 10.50 ± 1.79 |
| Absorption constant, h-1 | 5.40 ± 2.11 | 5.40 ± 1.58 |
| TLag, h | 0.46 ± 0.45 | 0.47 ± 0.67 |
| Peak, mg/L, for 250 mg | … | 22.25 ± 3.73 |
Abbreviations: SD, standard deviation; TLag, Time lag.
d-Cycloserine Pharmacokinetic Parameters and Variances
| Parameter | Parameters Used in Subroutine PRIOR, Mean ± SD |
10000 Simulated Subjects, Mean ± SD |
|---|---|---|
| Clearance, L/h | 1.16 ± 0.73 | 1.14 ± 0.82 |
| Volume, L | 10.50 ± 3.15 | 10.50 ± 1.79 |
| Absorption constant, h-1 | 5.40 ± 2.11 | 5.40 ± 1.58 |
| TLag, h | 0.46 ± 0.45 | 0.47 ± 0.67 |
| Peak, mg/L, for 250 mg | … | 22.25 ± 3.73 |
| Parameter | Parameters Used in Subroutine PRIOR, Mean ± SD |
10000 Simulated Subjects, Mean ± SD |
|---|---|---|
| Clearance, L/h | 1.16 ± 0.73 | 1.14 ± 0.82 |
| Volume, L | 10.50 ± 3.15 | 10.50 ± 1.79 |
| Absorption constant, h-1 | 5.40 ± 2.11 | 5.40 ± 1.58 |
| TLag, h | 0.46 ± 0.45 | 0.47 ± 0.67 |
| Peak, mg/L, for 250 mg | … | 22.25 ± 3.73 |
Abbreviations: SD, standard deviation; TLag, Time lag.
RESULTS
Systematic Analysis Findings
Figure 1shows the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) diagram summarizing literature search findings fordcycloserine, which shows that hitherto there have been no PK/PD studies performed in preclinical models. There were 9 pharmacokinetic studies; 2 were duplicates, leading to 7 studies shown inSupplementary Table 1[26,28,38,43–46]. Doses of cycloserine used in the pharmacokinetic studies ranged from 250 mg twice a day to 500 mg twice a day. Only 3 studies assessed multiple drug concentration measurements over 1 dosing interval (ie, >1 sample) [38,47,48]. In 2 studies, only the mean concentration time curves were shown, which could not be analyzed further. Only Chang et al performed a population PK analysis in 98 time–concentration data points in 14 patients treated with 250–500 mg twice daily [38]. The mean parameter estimates (interindividual variability as percentage coefficient of variation) of 1.38L/h (22.3%) for clearance, and 10.5L (35.1%) for volume of distribution [38]. As regards to formulation, 2 noncompartmental pharmacokinetic analyses reported concentrations of cycloserine after administration of terizidone [46,47]. No study has yet explored the relationship between concentrations such as peak or AUC0–24of cycloserine and clinical outcomes such as cure or relapse.
Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) flow diagram. Abbreviations: EBA, early bactericidal activity; PD, pharmacodynamic; PK, pharmacokinetic.
d-Cycloserine MICs in MDR-TB and Extensively Drug-resistant TB Clinical Isolates
In the literature, and from current studies, the MICs of the H37Ra laboratory stain varied between 4 and 8 mg/L depending on the method used, which was comparable to the figures for H37Rv, shown inSupplementary Table 2. However, because only the Sensititre assay had been used in 4 laboratories to measure the MICs of a total of 415 clinical isolates (Figure 2), we adopted the MIC of 8 mg/L for our PK/PD work and simulations. The tentative ECOFF for Sensititre was found to be 64 mg/L [32,49].
![Sensititre d-cycloserine minimum inhibitory concentration (MIC) distribution for 415 clinical isolates. Using the MIC distribution of 415 clinical isolates from 4 countries, including rifampicin-susceptible and -resistant isolates, 64 mg/L was chosen as the tentative epidemiological cutoff for d-cycloserine using the Sensititre method based on visual inspection [49]. No (tentative) ECOFFs are available for other media [32]. Abbreviations: ECOFF, epidemiological cutoff; MIC, minimum inhibitory concentration; RIF-R, rifampicin resistant; RIF-S, rifampicin susceptible.](https://oup.silverchair-cdn.com/oup/backfile/Content_public/Journal/cid/67/suppl_3/10.1093_cid_ciy624/1/m_ciy62402.jpeg?Expires=1686093358&Signature=VldMA0-KTu00G-T97bN4Ck7xf~0BzVjsMpyUHscLvzRAhm-d8ek-1OmgRfI6-ba4OGSl5Z6~UUZd2uv5R342-yEewofHIG1cKM3KM~SNf2Y4F267onMlScIfmUu6nJ25XW2TZXLcm9zw4wcHGjK1lUBv94JVyTEc9-f~YhQMicDDe0Y2qqYLBf2GOi6MmsHX~QkEm7vJqIIxlIvJFZj3tulbGcfeSWJ52hcLuSKC2pDwQDfQtiekt7swp3273SRIA~WrAhuO1-66y36WD-5VPy0i3ei3ltmaBJRazzu6PNdNyPiUpx4x3yfH5Ae3nJ7Kg2xfDJPSFxnCa2pAxEhC2A__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
Sensititredcycloserine minimum inhibitory concentration (MIC) distribution for 415 clinical isolates. Using the MIC distribution of 415 clinical isolates from 4 countries, including rifampicin-susceptible and -resistant isolates, 64 mg/L was chosen as the tentative epidemiological cutoff fordcycloserine using the Sensititre method based on visual inspection [49]. No (tentative) ECOFFs are available for other media [32]. Abbreviations: ECOFF, epidemiological cutoff; MIC, minimum inhibitory concentration; RIF-R, rifampicin resistant; RIF-S, rifampicin susceptible.
d-Cycloserine Concentration Effect Against Extracellular and IntracellularMtb
Following 7 days of coincubation with extracellular log-phase growthMtb,dcycloserine achieved a maximal kill (Emax) of 5.13 ± 0.28 log10CFU/mL and a concentration mediating 50% of maximal kill (EC50) of 5.44 ± 0.54 mg/L (r2= 0.97);最大杀死下面瘀(瘀= 0bacterial burden) was 4.61 log10CFU/mL. After 7 days of coincubation with intracellularMtb, the Emaxwas 2.55 ± 0.06 log10CFU/mL and the EC50was 6.87 ± 0.29 mg/L (r2= 0.99); maximal kill below stasis was only 1.59 log10CFU/mL. Thus,dcycloserine maximal kill of intracellularMtbwas >1000-fold (ie, 3.02 log10) less than for extracellularMtb, and was also less potent (EC50is higher).dcycloserine had no effect on the viability of adherent THP-1 cells.
PK/PD ofd-Cycloserine Microbial Kill in the HFS-TB
A 1-compartment model with first-order input and elimination best described the HFS-TB pharmacokinetic parameters, based on Akaike information criteria (AIC). The observed vs model-predicted concentrations are shown inSupplementary Figure 1A.Supplementary Figure 1B and 1Cshows the modeled and observed concentration-time profiles, which demonstrate that our dose fractionation exercise was successful.
dcycloserine microbial kill in the hollow fiber system model of tuberculosis.A, Time to positivity (TTP) as a measure of bacterial burden whendcycloserine doses were administered daily; the doses are shown as the 0- to 24-hour area under the concentration–time curve (AUC0–24) values. TTP increases as bacterial burden decreases. The highest 2 doses with an area under the concentration–time curve (AUC) of 182 and 273 mg × h/L achieved TTP >56 days, and thus negative culture. The TTP >56 days is a more stringent cutoff point for negative cultures compared to the 42 days used in the clinic, though this varies between clinical laboratories.B, The once-weekly regimens did worse, with TTPs only increasing after 3 weeks; doses are shown as AUC0–24values.C, When microbial kill was measured using colony-forming units (CFU)/mL, the 2 highest doses achieved negative cultures by day 7, unlike what was seen with TTP. The CFU/mL assay is less sensitive at lower bacterial burdens.D, Based on CFU/mL, the once-weekly dosing schedules demonstrated microbial kill during the first 10 days, then failed, with regrowth after 21 days. The kill slopes were less steep compared to the daily dosing schedule. Doses are shown as AUC0–24values. AUC, area under the concentration–time curve; CFU, colony-forming units; MGIT, mycobacterial growth indicator tube; TTP, time to positivity.
Akaike Information Criteria for Pharmacokinetic/Pharmacodynamic Indices on Different Sampling Days in the Hollow Fiber System Model of Tuberculosis
| PK/PD Index | Day 7 | Day 14 | Day 21 | Day 28 |
|---|---|---|---|---|
| Microbial kill | ||||
| AUC0–24/MIC | 20.22 | 25.82 | 29.99 | 30.85 |
| Peak/MIC | 35.03 | 36.00 | 36.39 | 37.69 |
| %TMIC | 29.96 | 23.53 | 18.47 | 20.69 |
| Resistance log10CFU/mL | ||||
| AUC0–24/MIC | –8.37 | –0.35 | 9.30 | 13.44 |
| Peak/MIC | 7.42 | 12.35 | 14.81 | 16.89 |
| %TMIC | 11.62 | 5.69 | 8.94 | –0.23 |
| PK/PD Index | Day 7 | Day 14 | Day 21 | Day 28 |
|---|---|---|---|---|
| Microbial kill | ||||
| AUC0–24/MIC | 20.22 | 25.82 | 29.99 | 30.85 |
| Peak/MIC | 35.03 | 36.00 | 36.39 | 37.69 |
| %TMIC | 29.96 | 23.53 | 18.47 | 20.69 |
| Resistance log10CFU/mL | ||||
| AUC0–24/MIC | –8.37 | –0.35 | 9.30 | 13.44 |
| Peak/MIC | 7.42 | 12.35 | 14.81 | 16.89 |
| %TMIC | 11.62 | 5.69 | 8.94 | –0.23 |
Values in bold indicate the PK/PD parameter with the lowest AIC scores for microbial kill and ADR on the different sampling days.
Abbreviations: %TMIC, percentage of time concentration persisting above the minimum inhibitory concentration; AUC0–24, 0- to 24-hour area under the concentration–time curve; CFU, colony-forming units; MIC, minimum inhibitory concentration; PK/PD, pharmacokinetic/pharmacodynamic.
Akaike Information Criteria for Pharmacokinetic/Pharmacodynamic Indices on Different Sampling Days in the Hollow Fiber System Model of Tuberculosis
| PK/PD Index | Day 7 | Day 14 | Day 21 | Day 28 |
|---|---|---|---|---|
| Microbial kill | ||||
| AUC0–24/MIC | 20.22 | 25.82 | 29.99 | 30.85 |
| Peak/MIC | 35.03 | 36.00 | 36.39 | 37.69 |
| %TMIC | 29.96 | 23.53 | 18.47 | 20.69 |
| Resistance log10CFU/mL | ||||
| AUC0–24/MIC | –8.37 | –0.35 | 9.30 | 13.44 |
| Peak/MIC | 7.42 | 12.35 | 14.81 | 16.89 |
| %TMIC | 11.62 | 5.69 | 8.94 | –0.23 |
| PK/PD Index | Day 7 | Day 14 | Day 21 | Day 28 |
|---|---|---|---|---|
| Microbial kill | ||||
| AUC0–24/MIC | 20.22 | 25.82 | 29.99 | 30.85 |
| Peak/MIC | 35.03 | 36.00 | 36.39 | 37.69 |
| %TMIC | 29.96 | 23.53 | 18.47 | 20.69 |
| Resistance log10CFU/mL | ||||
| AUC0–24/MIC | –8.37 | –0.35 | 9.30 | 13.44 |
| Peak/MIC | 7.42 | 12.35 | 14.81 | 16.89 |
| %TMIC | 11.62 | 5.69 | 8.94 | –0.23 |
Values in bold indicate the PK/PD parameter with the lowest AIC scores for microbial kill and ADR on the different sampling days.
Abbreviations: %TMIC, percentage of time concentration persisting above the minimum inhibitory concentration; AUC0–24, 0- to 24-hour area under the concentration–time curve; CFU, colony-forming units; MIC, minimum inhibitory concentration; PK/PD, pharmacokinetic/pharmacodynamic.
where the EC50is %TMICof 40.25. From equation 1, we calculated the %TMICassociated with stasis as 20%; that mediating 1.0 log10CFU/mL kill (cidal) was 30%, while EC80was a %TMICof 64%.
Evolution of Resistance in the HFS-TB
From equation 2, we calculated the %TMIC与完整的ADR相关抑制100%。
Monte Carlo Experiments to Identifyd-Cycloserine Clinical Doses
First we examined how well doses would achieve the exposure that suppresses ADR (%TMIC= 100%); we abandoned the exercise as even doses of 3000 mg a day performed poorly.Table 1compares the MCE-derived PK parameters to those in patients, an internal validation step.Figure 4shows the performance of several different doses and schedules at achieving (1) %TMICassociated with stasis, (2) %TMICassociated with cidal effect (1.0 log10CFU/mL kill), and (3) the EC80, in pulmonary tuberculosis cavities.Figure 4A–Cshows that all doses performed poorly, reflecting the uniformly poor penetration ofdcycloserine into the pulmonary cavity. Based on the poorer efficacy and lower potency against intracellularMtbdemonstrated earlier, performance of doses would even be worse against intracellular bacilli. The performance of all doses fell when examined for the ability to achieve %TMICassociated with 1.0 log10CFU/mL drop (cidal effect) inFigure 4Band fell even more in achieving the EC80target. In the highest dose tested, of 750 mg twice a day, the target attainment probability (TAP) fell below 90% at an MIC of 64 mg/L for cidal effect and 32 mg/L for EC80target.Figure 4Dshows that the dose best able to achieve cidal effect was 750 mg twice a day. Based on this, the dose of 750 mg twice a day was chosen as at least being able to achieve cidal effect inside most patients’ pulmonary cavities.
Performance of differentdcycloserine doses for pulmonary disease.dcycloserine minimum inhibitory concentrations (MICs) based on the Sensititre assay were used.A, For the stasis target (ie, exposure at which there is no kill or growth compared to day 0), the target attainment probabilities (TAPs) are shown for doses of 250–750 mg at 2 dosing schedules. There is clear separation of performance by the 750-mg doses (shown in black) from the rest of the doses. At 750 mg daily, TAP falls at MICs of ≥32 mg/L.B, For cidal activity, the same pattern is shown, except that now there is also separation of TAPs between 750 mg once daily and 750 mg twice daily. At the latter dose and schedule, the MIC above which TAP falls below 90% is 32 mg/L.C, For the target of 80% of maximal kill (EC80), even the dose if 750 mg twice a day would fall below 90% at around 32 mg/L.D, When an expectation was taken over the entire MIC range, the dose of 750 mg twice daily was able to achieve the exposure target of stasis, cidal effect, and EC80,in 93%, 92%, and 81% of 10000 patients, respectively. Abbreviations: bid, twice daily; EC80, 80% of maximal kill; MIC, minimum inhibitory concentration; OD, once daily.
If we bargained to get good microbial kill in the meninges, based on the better penetration ofdcycloserine into subarachnoid space, in exchange for possible increased neurotoxicity, then target attainment in tuberculous meningitis was as shown inFigure 5. A 500 mg twice a day achieved a cumulative fraction of response of 88.2% for the stasis target, 84.7% for cidal effect, and 69.8% for the EC80target. While the target is still shy of the 90% target attainment, it is still acceptable performance when balanced against possible increase neurotoxicity at higher doses.
目标成就(TAP)各种概率dcycloserine doses in meningitis. Shown are TAP values for 10000 patients with tuberculous meningitis (TBM) and extracavitary compartments of tuberculosis.dcycloserine has better penetration into cerebrospinal fluid (CSF) than lung cavities.A, The TAP for stasis target exposure demonstrated good performance for the dose of 500 mg twice daily, and only fell below 90% at a minimum inhibitory concentration (MIC) of 32 mg/L, and for the highest dose at an MIC of 64 mg/L.B, For the cidal exposure target, 500 mg twice daily and 750 mg twice daily achieved good TAP up to an MIC of 32 mg/L, such that the majority of patients with meningitis would be expected to achieve exposures that have cidal effect in CSF at these doses. This means that as long as MICs were <64 mg/L, cidal effect was achieved. Given the paucibacillary nature of TBM, this is thought to be the best pharmacokinetic/pharmacodynamic exposure target for meningitis.C, Even with good penetration into the subarachnoid space, the 80% of maximal kill was more difficult to achieve at all doses for MICs ≥16 mg/L.D, Cumulative fraction of patients who achieve responses for each dose. The dose of 500 mg twice daily falls just short of 90% for the cidal effect, but is proposed for use in TBM. Abbreviations: bid, twice daily; EC80, 80% of maximal kill; MIC, minimum inhibitory concentration; OD, once daily.
DISCUSSION
First, we found a tentative ECOFF value of 64 mg/L based on the Sensititre MYCOTB assay (Figure 2). In MCEs, at the proposed dose of 750 mg twice a day for pulmonary tuberculosis, the TAP falls below 90% at an MIC of 64 mg/L for the cidal effect target (Figure 4). This means that fordcycloserine both the PK/PD-based approach and the tentative ECOFF are in agreement. Based on both, we propose a susceptibility breakpoint of 64 mg/L.
Second,dcycloserine had impressive kill rates againstMtbin the HFS-TB, which rivaled some of the first-line compounds and fluoroquinolones [18,36,51]. Thus the drug is not “static,” as has been traditionally thought. Instead, the limitation of this drug stems from its poor lung cavitary penetration in patients. Another problem is the poor kill of intracellularMtb, which constitutes about 20% of all bacteria in lung cavities [52]. As a result, we propose 750 mg twice a day for pulmonary tuberculosis. While our dose findings are MCE-based, and thus require clinical confirmation, it is interesting that we found that 500 to 750 mg a day would achieve the target of EC80 in 1%–56% of patient’s in lung cavities. In 1962, Rivera and Browning treated 90 patients with 500 mgdcycloserine plus 3 g pyrazinamide each day; sputum conversion plus disappearance of cavity was achieved in only 11% of patients [53]. Similarly, Epstein and colleagues treated patients with pulmonary disease with 1000–1500 mg/day ofdcycloserine [54]. In patients without prior treatment, the culture conversion occurred in 11% on isoniazid-cycloserine combination compared with 13% ondcycloserine monotherapy; in drug-resistant tuberculosis, 33% achieved negative cultures on solid agar. Thus, at high doses, 1000–1500 mg/day cure was achieved in 10%–30% of patients with pulmonary tuberculosis, which is consistent with our MCE.
Third,dcycloserine has good CSF penetration, which likely explains the high rates of neurotoxicity. If we made the Faustian bargain to treat tuberculous meningitis with the potentially neurotoxicdcycloserine, at the high doses of 500 mg twice a day that we propose, kill rates would be equivalent to those of fluoroquinolones. However, it could be that the neuropsychiatric problems would be worse during treatment for a disease that is by definition neurotoxic. Vitamin B6 could be prescribed concurrently with thedcycloserine to minimize toxicity, with some experts recommending 50 mg of pyridoxine for every 250 mg ofdcycloserine [53]. However, human dosing trials ofdcycloserine and pyridoxine in combination with standard tuberculous meningitis regimens have not been performed, and the effectiveness of this approach as yet unproven.
There are some limitations to our study. First, we did not examinedcycloserine in combination with other drugs for synergistic effects, which could potentially lower the exposures ofdcycloserine needed. Second, in contrast to our work with gatifloxacin, levofloxacin, and ethionamide, we had no clinical data to validate the doses or susceptibility breakpoints we identified [18,51,55]. Third, we relied on the Sensititre assay, which is not endorsed by WHO and could differ from conventional media, to define a tentative ECOFF. As an example, the MGIT-derived MIC was systematically 1-tube dilution lower than MYCOTB-derived for our quality control isolate, which could affect the PK/PD target exposures, and hence optimal dose. These limitations mean that the accuracy of ourdcycloserine dose choices remains to be prospectively confirmed in a clinical study.
In summary,dcycloserine has cidal effects againstMtb, provided optimal exposures are achieved. In practice, given the poor penetration into human tuberculous cavities, the drug could be effective to some extent in pulmonary disease but at high doses. The drug should be given at least twice daily to optimize exposure and should preferably be used in the intensive phase of treatment due to its poor intracellular and thus sterilizing efficacy. On the other hand,dcycloserine likely could add to effectiveness of regimens to treat tuberculous meningitis and pulmonary tuberculosis outside cavities, at doses of about 500 mg twice a day.
Supplementary Data
Supplementary materials are available atClinical Infectious Diseasesonline. Consisting of data provided by the authors to benefit the reader, the posted materials are not copyedited and are the sole responsibility of the authors, so questions or comments should be addressed to the corresponding author.
Notes
Author contributions. T. G. and D. D designed the laboratory studies. J.-W. C. A., N. S., and M. G. G. S. designed and performed the systematic analysis and literature review. T. G., D. D., and M. L. C. performed the hollow fiber studies. C. U. K. and T. S. provided critical feedback and editing on MIC distributions, ECOFF, clinical breakpoints, and MIC methods. H. M. provided information on pharmacokinetics of cycloserine, especially as terizidone. D. D wrote the first draft of the manuscript. P. S. L. performed drug concentration assays. T. K. performed DNA extraction. T. G. performed PK/PD modeling and MCEs. K. D. and T. G. performed the clinical study that identifieddcycloserine penetration into lung cavities. S. G. M., S. B., S. F., O. O., S. P., E. R. H., and S. K. H. identified MICs in the MDR-TB clinical studies and took part in thedcycloserine population pharmacokinetics study. D. D., S. K. H., and T. G. wrote the manuscript. All authors edited and contributed to the final version of the manuscript.
Acknowledgments. We would like to thank Dr Charles Peloquin (University of Florida) for allowing us use his group’s ongoing population PK study. We acknowledge the work of Dr Onno W. Akkerman, Dr Mathieu S. Bolhuis, and Samiksha Ghimire (University of Groningen, University Medical Center Groningen) on the systematic review.
Financial support. This work was supported by the Baylor Research Institute, Dallas, Texas (to T. G.) and the National Institute of Allergy and Infectious Diseases of the National Institutes of Health (grant numbers R01AI116155 to H. M. and T. G. and U01AI115594 to S. K. H.). C. U. K. is a research associate at Wolfson College, University of Cambridge.
Supplement sponsorship. This supplement is sponsored by the Baylor Institute of Immunology Research of the Baylor Research Institute.
Potential conflicts of interest. C. U. K. is a consultant for the Foundation for Innovative New Diagnostics which involves work for the Cepheid Inc., Hain Lifescience and the WHO. C. U. K. is an advisor to GenoScreen and QuantuMDx Group Ltd. The Bill & Melinda Gates Foundation, Janssen Pharmaceutica, and PerkinElmer covered C. U. K.’s travel and accommodation to present at meetings. The European Society of Mycobacteriology awarded C. U. K. the Gertrud Meissner Award, which is sponsored by Hain Lifescience. The Global Alliance for TB Drug Development Inc. and Otsuka Novel Products GmbH have supplied C.U.K. with antibiotics forin vitroresearch. All other authors report no potential conflicts. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed.
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