The treatment of
drug-resistant Mycobacterium tuberculosis relies on complex
antibiotic therapy. Inadequate
antibiotic exposure can lead to treatment failure, acquired drug resistance, and an increased risk of adverse events. Therapeutic
drug monitoring (TDM) can be used to optimize the
antibiotic exposure. Therefore, we aimed to develop a single-run multiplex assay using high-performance liquid chromatography-mass spectrometry (HPLC-MS) for TDM of patients with multidrug-resistant, pre-extensively
drug-resistant and
extensively drug-resistant tuberculosis. A target profile for sufficient performance, based on the intended clinical application, was established and the assay was developed accordingly.
Antibiotics were analyzed on a zwitterionic hydrophilic interaction liquid chromatography column and a triple quadrupole mass spectrometer using stable
isotope-labeled internal standards. The assay was sufficiently sensitive to monitor
drug concentrations over five half-lives for
rifampicin,
rifabutin,
levofloxacin,
moxifloxacin,
bedaquiline,
linezolid,
clofazimine,
terizidone/
cycloserine,
ethambutol,
delamanid,
pyrazinamide,
meropenem,
prothionamide, and para-amino
salicylic acid (PAS). Accuracy and precision were sufficient to support clinical decision making (≤±15% in clinical samples and ±20-25% in spiked samples, with 80% of future measured concentrations predicted to fall within ±40% of nominal concentrations). The method was applied in the TDM of two patients with complex
drug-resistant tuberculosis. All relevant
antibiotics from their regimens could be quantified and high-dose
therapy was initiated, followed by microbiological conversion. In conclusion, we developed a multiplex assay that enables TDM of the relevant first- and second-line anti-
tuberculosis medicines in a single run and was able to show its applicability in TDM of two
drug-resistant tuberculosis patients.