Quantification of Methotrexate in Human Serum Using Surface-Enhanced Raman Scattering—Toward Therapeutic Drug Monitoring

Abstract

Therapeutic drug monitoring (TDM) can improve clinical care when using drugs with pharmacokinetic variability and a narrow therapeutic window. Rapid, reliable and easy to use detection methods are required in order to decrease the time of analysis, and can enable TDM also in resource-limited settings or even at bedside. Monitoring methotrexate (MTX), an anti-cancer drug, is critical, since it is needed to follow the drug clearance rate and decide how to administer the rescue drug, leucovorin (LV), in order to avoid toxicity and even death. We show that with the optimized nanopillar assisted separation (NPAS) method using surface-enhanced Raman scattering (SERS) we were able to measure MTX in PBS and serum in the linear range of 5 to 150 µM and confirmed that MTX detection can be carried out even in the presence of LV. Additionally, when NPAS was combined with centrifugal filtration a quantification limit of 2.1 µM for MTX in human serum sample was achieved. The developed detection method enables fast detection (10 min) and quantification of MTX from human serum (>90% accuracy). Furthermore, we show the potential of the developed method for TDM, when quantifying MTX from clinical samples, collected from patients, who are undergoing high-dose MTX therapy.

Methotrexate Detection in Serum at Clinically Relevant Levels with Electrochemically Assisted SERS on a Benchtop, Custom Built Raman Spectrometer

“…The studies mentioned above were carried out with nanoparticles in suspension, which have an intrinsic limitation since the reproducibility strongly depends on the synthesis, experience, and training in handling the particles. Previously, we have reported a LoD of 6.3 μM using AgNP by means of a nanopillar-assisted separation method . Thus, the electrochemically assisted SERS technique appears to have comparable sensitivity to the previously reported SERS-based detection approaches using nanoparticles and showed an improved sensitivity with NP SERS substrates.…”

“…60−62 Previously, we showed that organic solvent based protein precipitation and ultracentrifugation improved the SERS assay sensitivity when working with human serum samples. 27 In the current work, organic solvents were also assessed, but mixing the serum sample with a large amount of organic solvent was not compatible with the electrochemically assisted SERS assay. Protein precipitation using acids was evaluated since this combined efficient protein precipitation 62,63 and sample acidification.…”

“…19,29−33 Recently, we have developed an assay for MTX detection in serum, based on ultrafiltration and nanopillar-assisted separation, reaching the limit of quantification of 2.1 μM. 27 When aiming to further improve the label-free detection of MTX, to achieve clinically relevant limits, 5 the goal is to bring the target analyte closer than 0.7 nm to SERS "hotspots" on the surface to maximize the enhancement effect. 34 Electrochemical-SERS (EC-SERS) is a promising technique considering that analytes may lack affinity to the SERS substrate.…”

One-Step Preparation of S, N Co-Doped Carbon Quantum Dots for the Highly Sensitive and Simple Detection of Methotrexate

“…Methotrexate (MTX) is a folic-acid-type antitumor agent that is commonly used in high doses in the treatment of lymphoma, acute lymphoblastic leukemia, chorionic epithelial carcinoma, and other types of malignant tumors, as well as rheumatoid arthritis [ 1 ]. MTX is used to prevent the transferring of folic acid into folinic acid with physiological activity through inhibiting the activity of dihydrofolate, which can reduce the synthesis of DNA and protein, then block the synthesis of tumor cells [ 2 ].…”

“…There are several methods available for determining MTX, such as surface-enhanced Raman scattering (SPRS) [ 1 ], surface plasmon resonance (SPR) [ 2 ], high-performance liquid chromatography (HPLC) [ 7 , 8 , 9 ], electroanalysis [ 10 , 11 , 12 ], and immunoassays [ 13 ]. In comparison to the methods mentioned above, fluorescence has gained attention due to its remarkable sensitivity, quick response, easy operation, good biocompatibility, minimal toxicity, and the ability to detect non-destructively [ 14 , 15 ].…”