1104. Comparison of Antibiotic Sampling Techniques: Predicting Plasma Vancomycin Concentrations Using Volumetric Absorptive Microsampling (VAMS) from Capillary and Venous/Arterial Whole Blood

Background Therapeutic drug monitoring (TDM) is paramount to optimize the safety and efficacy of vancomycin (VAN). In children, TDM is challenged by difficulty in obtaining venous samples, impeding timely sampling. We assessed the ability of volumetric absorptive microsampling (VAMS) as a novel, whole blood sampling technique to predict plasma VAN concentrations in plasma. Methods We conducted a prospective pilot study among critically ill children prescribed VAN for clinical care. Coincident with VAN TDM in plasma (P), we collected 20 µL of capillary whole blood (C) and venous/arterial whole blood (V) using VAMS. Paired VAMS-P samples drawn >5 mins apart and VAMS samples with over- or under-loaded filter tip on visual inspection were excluded. Plasma concentrations were measured via chemiluminescent immunoassay in the Chemistry Laboratory. VAMS C and V concentrations were measured using LC/MS in the Bioanalytic Core Laboratory. Plasma concentrations were predicted from whole blood VAMS with Passing-Bablok regression using 3 methods: 1) uncorrected VAMS measures, 2) hematocrit-corrected VAMS, and 3) lab-corrected VAMS (Figure 1). We then assessed bias, imprecision, and accuracy of plasma predictions from VAMS (C and V) as compared to coincident P concentrations for each technique (Figure 1). Figure 1. Methods for relating whole blood vancomycin concentrations collected via VAMS to plasma concentrations and measure to evaluate predictive performance. Results Paired samples were collected from 31 enrolled subjects (Figure 2), with a median age of 3.3 years (range 0.1-17.9). Measured P concentrations ranged from 4.6 - 54.9 mg/L. 11 C samples (29%) and 3 V samples (10%) were excluded due to collection issues. Prediction results are shown in Figure 3. The 3 prediction techniques had similar performance characteristics, with each method displaying minimal bias (-0.4-2.0%) and reasonable imprecision (13.7-20.2%). The accuracy of prediction of P concentrations using VAMS was better for V than C samples. Figure 2. Flow diagram from sample collection to evaluation. Abbreviations: C-P, capillary VAMS-plasma; V-P, venous/arterial VAMS-plasma; VAMS, volumetric absorptive microsampling. Figure 3. Performance of 3 techniques to predict plasma vancomycin concentrations using whole blood collected via VAMS. Conclusion Our pilot highlights the challenges of using VAMS for TDM. Sample collection issues were common. When VAMS is used, education on collection techniques is imperative. The predictive performance of VAMS was modest and V sampling had higher accuracy than C, although our sample size was small. Larger studies will be needed to further evaluate the predictive performance of the regression equations derived by our study. Disclosures Kevin J. Downes, MD, Merck, Inc. (Grant/Research Support)

ANALYTICAL METHOD VALIDATION OF DOXORUBICIN AND DOXORUBICINOL IN VOLUMETRIC ABSORPTIVE MICROSAMPLING BY LIQUID CHROMATOGRAPHY-TANDEM MASS SPECTROMETRY

Doxorubicin is a broad-spectrum anthracycline antibiotic which has antineoplastic activity. Doxorubicin can cause cardiotoxic effects due to the formation of doxorubicinol as its main metabolite. One of the latest bio sampling methods, Volumetric Absorptive Microsampling (VAMS), has various advantages which include blood sampling by finger-prick, hematocrit does not influence it, and it can be stored at room temperature. This study aims to determine the optimum analysis conditions and sample preparation method in VAMS with daunorubicin as an internal standard, and to develop sensitive, specific measurements as well as validate the analytical method using Ultra High Performance Liquid Chromatography-Tandem Mass Spectrometry (UPLC-MS/MS). Analytical quantification analysis using mass spectrometry with a mass analyzer type is a triple quadrupole with a positive type of electrospray ionization (ESI). The separation was carried out with the Acquity® UPLC BEH C18 column (2.1 x 100 mm; 1.7 μm), with a flow rate of 0.2 mL/min, and gradient elution using 0.1% formic acid in water and 0.1% formic acid in acetonitrile for 7 minutes. Multiple reaction monitoring (MRM) values were set at m/z 544.22 > 396.9 for doxorubicin; m/z 546.22 > 398.9 for doxorubicinol; and m/z 528.5 > 362.95 for daunorubicin. Sample preparation used protein precipitation with methanol as the extracting solution, the drying time of the VAMS was 2 hours, and the sonication time was 30 minutes. LLOQ values obtained were 8 ng/mL for doxorubicin and 3 ng/mL for doxorubicinol with the linearity of 0.9904 for doxorubicin and 0.9902 for doxorubicinol.

Development and Validation of the Quantification Method for Hydroxychloroquine in Volumetric Absorptive Microsampling (VAMS) Using High-Performance Liquid Chromatography-Photodiode Array

Hydroxychloroquine is an antimalarial drug used for systemic lupus erythematosus, rheumatoid arthritis, and malaria treatment. However, hydroxychloroquine has several side effects such as ocular toxicity, neurotoxicity, gastrointestinal disorder, and also severe toxicity such as cardiotoxicity. Therefore, therapeutic drug monitoring of high dose or long-term use of hydroxychloroquine is needed. This study aims to obtain an optimum and validated analysis and preparation method for hydroxychloroquine in volumetric absorptive microsampling (VAMS) using the high-performance liquid chromatography–photodiode array detector based on the Food and Drug Administration guidelines (2018). Hydroxychloroquine quantification was performed using HPLC-PDA with Waters Sunfire™ C18 (5 µm; 250 × 4,6 mm) column. Mobile phase consists of acetonitrile-diethylamine 1% (65 : 35, v/v) (isocratic elution) and delivered at a flow rate of 0.8 mL/min throughout the 12 minutes run. Sample in VAMS is extracted by liquid-liquid extraction with ammonia 1% and n-hexane-ethyl acetate (50 : 50 v/v) as a extraction solvent. This method has successfully qualified the Food and Drug Administration (2018) parameters, with 2 ng/mL of LLOQ, range of calibration curve 2–6500 ng/mL, and coefficient of correlation 0.9993–0.9997.

Volumetric absorptive microsampling: its use in COVID-19 research and testing

COVID-19 led to changes in the way blood samples are collected. As societies were isolated to control viral spread, access to facilities became limited. Remote sample collection with a volumetric microsampling approach, using Mitra® devices based on VAMS® technology, proved to be highly effective. It allowed people to collect high-quality samples at home and post them to a laboratory. This enabled scientists to conduct large serosurveillance studies, with results showing that seroprevalence of COVID-19 was higher than initially expected. Furthermore, remote microsampling studies by several institutions were conducted to measure the relationship between antigen levels and antibody response and duration. VAMS technology was also used in COVID-19 clinical trials. In summary, the independent research reviewed in this paper proved that VAMS is an effective sample collection alternative.

Antibody Mediated Immunity to SARS-CoV-2 and Human Coronaviruses: Multiplex Beads Assay and Volumetric Absorptive Microsampling to Generate Immune Repertoire Cartography

The COVID-19 pandemic is caused by SARS-CoV-2, a novel zoonotic coronavirus. Emerging evidence indicates that preexisting humoral immunity against other seasonal human coronaviruses (HCoVs) plays a critical role in the specific antibody response to SARS-CoV-2. However, current work to assess the effects of preexisting and cross-reactive anti-HCoVs antibodies has been limited. To address this issue, we have adapted our previously reported multiplex assay to simultaneously and quantitatively measure anti-HCoV antibodies. The full mPlex-CoV panel covers the spike (S) and nucleocapsid (N) proteins of three highly pathogenic HCoVs (SARS-CoV-1, SARS-CoV-2, MERS) and four human seasonal strains (OC43, HKU1, NL63, 229E). Combining this assay with volumetric absorptive microsampling (VAMS), we measured the anti-HCoV IgG, IgA, and IgM antibodies in fingerstick blood samples. The results demonstrate that the mPlex-CoV assay has high specificity and sensitivity. It can detect strain-specific anti-HCoV antibodies down to 0.1 ng/ml with 4 log assay range and with low intra- and inter-assay coefficients of variation (%CV). We also estimate multiple strain HCoVs IgG, IgA and IgM concentration in VAMS samples in three categories of subjects: pre-COVID-19 (n=21), post-COVID-19 convalescents (n=19), and COVID-19 vaccine recipients (n=14). Using metric multidimensional scaling (MDS) analysis, HCoVs IgG concentrations in fingerstick blood samples were well separated between the pre-COVID-19, post-COVID-19 convalescents, and COVID-19 vaccine recipients. In addition, we demonstrate how multi-dimensional scaling analysis can be used to visualize IgG mediated antibody immunity against multiple human coronaviruses. We conclude that the combination of VAMS and the mPlex-Cov assay is well suited to performing remote study sample collection under pandemic conditions to monitor HCoVs antibody responses in population studies.

Therapeutic Drug Monitoring of Antiseizure Medications Using Volumetric Absorptive Microsampling: Where Are We?

Therapeutic drug monitoring (TDM) of antiseizure medications (ASMs) represents a valuable tool to establish an appropriate patient therapy, to collect important information about drugs’ interactions and to evaluate patient’s metabolic capabilities. In recent years, a new volumetric absorptive microsampling technique using VAMS® technology and Mitra® devices, consisting of a sampling technique for the collection of fixed-volume capillary blood, was developed. These new devices provide a new home-sampling technique for whole blood that has been spread out to simplify sample collection from finger-pricks. This review is aimed to compare published articles concerning the application of VAMS® in epilepsy and to identify the strengths and improvement points for the TDM of antiseizure medications. VAMS® allowed a minimally invasive blood sampling even in the absence of trained personnel. Good stability data have indicated that storage and delivery can be facilitated only for specific ASMs. Trueness and precision parameters have been evaluated, and the hematocrit (HCT) effect was minimized.