A quantitative mass spectrometry imaging (QMSI) technique using infrared matrix-assisted laser desorption electrospray ionization (IR-MALDESI) is demonstrated for the antiretroviral (ARV) drug emtricitabine in incubated human cervical tissue. Method development of the QMSI technique leads to a gain in sensitivity and removal of interferences for several ARV drugs. Analyte response was significantly improved by a detailed evaluation of several cationization agents. Increased sensitivity and removal of an isobaric interference was demonstrated with sodium chloride in the electrospray solvent. Voxel-to-voxel variability was improved for the MSI experiments by normalizing analyte abundance to a uniformly applied compound with similar characteristics to the drug of interest. Finally, emtricitabine was quantified in tissue with a calibration curve generated from the stable isotope-labeled analog of emtricitabine followed by cross-validation using liquid chromatography tandem mass spectrometry (LC-MS/MS). The quantitative IR-MALDESI analysis proved to be reproducible with an emtricitabine concentration of 17.2±1.8 μg/gtissue. This amount corresponds to the detection of 7 fmol/voxel in the IR-MALDESI QMSI experiment. Adjacent tissue slices were analyzed using LC-MS/MS which resulted in an emtricitabine concentration of 28.4±2.8 μg/gtissue.
dPersistent HIV replication within active viral reservoirs may be caused by inadequate antiretroviral penetration. Here, we used mass spectrometry imaging with infrared matrix-assisted laser desorption-electrospray ionization to quantify the distribution of efavirenz within tissues from a macaque dosed orally to a steady state. Intratissue efavirenz distribution was heterogeneous, with the drug concentrating in the lamina propria of the colon, the primary follicles of lymph nodes, and the brain gray matter. These are the first imaging data of an antiretroviral drug in active viral reservoirs.
Strategies to prevent HIV infection using pre-exposure prophylaxis (PrEP) are required to curtail the HIV pandemic. The mucosal tissues of the genital and rectal tracts play a critical role in HIV acquisition, but antiretroviral (ARV) disposition and correlates of efficacy within these tissues are not well understood. Pre-clinical and clinical strategies to describe ARV pharmacokinetic-pharmacodynamic relationships (PK/PD) within mucosal tissues are currently being investigated. In this review, we summarize the physiochemical and biologic factors influencing ARV tissue exposure. Further, we discuss the necessary steps to generate relevant PK/PD data and the challenges associated with this process. Finally, we suggest how pre-clinical and clinical data might be practically translated into optimal PrEP dosing strategies for clinical trials testing using mathematical modeling and simulation.
This work describes the coupling of the IR-MALDESI imaging source with the Q Exactive mass spectrometer. IR-MALDESI MSI was used to elucidate the spatial distribution of several HIV drugs in cervical tissues that had been incubated in either a low or high concentration. Serial sections to those analyzed by IR-MALDESI MSI were homogenized and analyzed by LC-MS/MS to quantify the amount of each drug present in the tissue. By comparing the two techniques, an agreement between the average intensities from the imaging experiment with the absolute quantities for each drug was observed. This correlation between these two techniques serves as a prerequisite to quantitative IR-MALDESI MSI. In addition, a targeted MS2 imaging experiment was also conducted to demonstrate the capabilities of the Q Exactive and to highlight the added selectivity that can be obtained with SRM or MRM imaging experiments.
Adherence to a drug regimen can be a strong predictor of health outcomes, and validated measures of adherence are necessary at all stages of therapy from drug development to prescription. Many of the existing metrics of drug adherence (e.g., self-report, pill counts, blood monitoring) have limitations, and analysis of hair strands has recently emerged as an objective alternative. Traditional methods of hair analysis based on LC–MS/MS (segmenting strands at ≥1 cm length) are not capable of preserving a temporal record of drug intake at higher resolution than approximately 1 month. Here, we evaluated the detectability of HIV antiretrovirals (ARVs) in hair from a range of drug classes using infrared matrix-assisted laser desorption electrospray ionization (IR-MALDESI) mass spectrometry imaging (MSI) with 100 µm resolution. Infrared laser desorption of hair strands was shown to penetrate into the strand cortex, allowing direct measurement by MSI without analyte extraction. Using optimized desorption conditions, a linear correlation between IR-MALDESI ion abundance and LC–MS/MS response was observed for six common ARVs with estimated limits of detection less than or equal to 1.6 ng/mg hair. The distribution of efavirenz (EFV) was then monitored in a series of hair strands collected from HIV infected, virologically suppressed patients. Because of the role hair melanin plays in accumulation of basic drugs (like most ARVs), an MSI method to quantify the melanin biomarker pyrrole-2,3,5-tricarboxylic acid (PTCA) was evaluated as a means of normalizing drug response between patients to develop broadly applicable adherence criteria.
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