Miniature mass spectrometer-based point-of-care assay for cabotegravir and rilpivirine in whole blood

Abstract

HIV prevention and treatment with injectable cabotegravir and/or rilpivirine administered once every 4 to 8 weeks is an attractive alternative to daily therapy. Prescribed dosage and drug concentrations in plasma are based on patient data collected in clinical trials, but actual patients are expected to exhibit more variability in drug concentrations, which is important to quantify. Here, we demonstrate the first quantitative point-of-care assay with a miniature mass spectrometer to assess these drug concentrations in whole blood. Quantitative performance is obtained using paper spray ionization in combination with tandem mass spectrometry (MS/MS) in the clinically relevant concentration range of both drugs. Limits of quantitation (LoQs) of cabotegravir and rilpivirine are measured to be 750 ng/mL and 20 ng/mL, respectively. The assay turnaround time is < 4 min, and strong linear relationships are established between MS/MS responses and concentration, with percentage of relative standard deviations (RSDs) that are <15% at concentrations above the LoQs. The speed, portability, low power consumption, and specificity offered by the miniature instrument should make it an appropriate platform for measuring drug concentrations in a walk-in clinic using small volumes of patient blood.

Keywords: Ambient ionization; Antiretrovirals; Drug quantitation; HIV treatment; Paper spray.

Figure 1.

Point-of-care analysis of CAB and RPV in whole blood with paper spray ionization and a miniature mass spectrometer. (a) Add 100 μL of solvent with internal standard and shake for one minute. (b) Deposit the supernatant layer onto a triangular piece of Q5 filter paper. (c) Apply solvent and high voltage and acquire MS/MS spectra in the positive mode using the Mini-12.

Figure 2.

MS/MS spectra of the protonated forms of (a) 1000 ng/mL CAB (b) 1000 ng/mL CAB-d5 (c) 100 ng/mL RPV (d) 100 ng/mL RPV-d6 recorded using the Mini-12 with nanoESI in the positive ion mode. Spectra (a) and (b) were recorded in the mass range m/z 205 - 445, whereas (c) and (d) were recorded in the mass range m/z 205 - 400. Each spectrum is an average of six scans recorded at a scan speed of 3500 m/z per second.

Figure 3.

Calibration curves of (a) CAB and (b) RPV with the Mini-12. Red and green lines correspond to 25^th and 75^th percentile concentrations observed in the registrational clinical trials. n=4 for each concentration.

Figure 4.

Comparison of quantitative performance over three different days for (a) CAB (b) RPV using the Mini-12; n = 4 for each data point.