Search

Chimeric antigen receptor T cells (CAR-T) targeting CD19 have achieved significant success in patients with B cell malignancies. To date, implementation of CAR-T in other indications remains challenging due to the lack of truly tumor-specific antigens as well as control of CAR-T activity in patients. CD123 is highly expressed in acute myeloid leukemia (AML) blasts including leukemia-initiating cells making it an attractive immunotherapeutic target. However, CD123 expression in normal hematopoietic progenitor cells and endothelia bears the risk of severe toxicities and may limit CAR-T applications lacking fine-tuned control mechanisms. Therefore, we recently developed a rapidly switchable universal CAR-T platform (UniCAR), in which CAR-T activity depends on the presence of a soluble adapter called targeting module (TM), and confirmed clinical proof-of-concept for targeting CD123 in AML with improved safety. As costimulation via 4-1BB ligand (4-1BBL) can enhance CAR-T expansion, persistence, and effector functions, a novel CD123-specific TM variant (TM123-4-1BBL) comprising trimeric single-chain 4-1BBL was developed for transient costimulation of UniCAR-T cells (UniCAR-T) at the leukemic site in trans . TM123-4-1BBL-directed UniCAR-T efficiently eradicated CD123-positive AML cells in vitro and in a CDX in vivo model. Moreover, additional costimulation via TM123-4-1BBL enabled enhanced expansion and persistence with a modulated UniCAR-T phenotype. In addition, the increased hydrodynamic volume of TM123-4-1BBL prolonged terminal plasma half-life and ensured a high total drug exposure in vivo . In conclusion, expanding the soluble adapter optionality for CD123-directed UniCAR-T maintains the platforms high anti-leukemic efficacy and immediate control mechanism for a flexible, safe, and individualized CAR-T therapy of AML patients., Competing Interests: Conflict-of-interest-disclosure: GE and AE are shareholders of GEMoaB GmbH and CPT Cellex Patient Treatment GmbH. JEM, SL, JD, JS, GJJ, MC, and AE are employees at GEMoaB GmbH and/or its subsidiary company CPT Cellex Patient Treatment GmbH., (© 2021 The Author(s). Published with license by Taylor & Francis Group, LLC.)

Aim: Cinitapride (CIN) is a benzamide-derived molecule used for the treatment of gastroesophageal reflux and dyspepsia. Its pharmacokinetics are controversial due to the use of supratherapeutic doses and the lack of sensitive methodology. Therefore, a sensitive and accurate micromethod was developed for its quantitation in human plasma., Results: CIN was extracted from 300 µl of heparinized plasma by liquid-liquid extraction using cisapride as internal standard, and analyzed with an ultra performance liquid chromatograph employing positive multiple-reaction monitoring-MS., Conclusion: The method proved to be rapid, accurate and stable within a range between 50 and 2000 pg/ml and was successfully validated and applied in a pharmacokinetic interaction trial, where it was demonstrated that oral co-administration of simethicone does not modify the bioavailability of CIN.

Background: Sildenafil is used for the treatment of pediatric pulmonary hypertension. A dried blood spot (DBS)-based LC-MS method for sildenafil quantitation was developed and applied to a group of patients., Results: DBS showed high portability and stability of samples, and the method was selective and linear for quantitation of sildenafil (5-3,000 ng/ml) and N-desmethyl-sildenafil (3-1,500 ng/ml). After a single oral dose of sildenafil (1 mg/kg), method evidenced poor metabolism in these patients., Conclusion: The method was successfully applied in peripheral blood and can be used for both pharmacokinetics and therapeutic drug monitoring. DBS proved to have advantages during sample translation and preservation of analytes. Data suggest that hazardous blood sildenafil levels may be reached in this population.

Background: Nitazoxanide (NTZ) is used for the treatment of gastrointestinal tract colonization by anaerobic bacteria, viruses and other pathogens that represent a major cause of morbidity in Latin America. The aim of the present work was to develop and validate a UPLC-MS/MS method for the selective quantification of tizoxanide (TZN, the major metabolite of NTZ) in human plasma using niclosamide as internal standard; and examine its pharmacokinetic application in healthy volunteers. Nine male subjects received a single oral dose of a NTZ 500-mg tablet under fasting conditions., Results: The method was linear between 0.1 and 10 µg/ml and capable of separating signals from free-TZN and those delivered by in-source collision-induced dissociation of TZN-glucuronide, quantifying it with accuracy and precision. Mean maximum plasma concentration was 6.79 µg/ml and was reached at 2.4 h post-dose., Conclusion: The method was validated, fulfilling regulatory guidelines. Results suggest low pharmacokinetic variability in the assayed population.

Background: A growing number of boron-containing compounds exhibit many important biological activities; of particular interest are the α-amino acid borinic derivatives with activity in the CNS. A validated, sensitive and specific UPLC-MS/MS technique for quantification of the diphenylboroxazolidones of glycine (DBPX-gly), L-aspartate (DPBX-L-asp) and L-glutamate (DPBX-L-glu) in dried blood spots (DBSs) is presented., Results: The most intense signal corresponds to compounds with (11)B. The extraction procedure was liquid elution of 3.2-mm punched DBSs with acetonitrile:aqueous formic acid 0.1% (80:20 v/v). Assays proved to be linear, falling accurately and precisely within the range of 0.3-50 µg/ml for DPBX-L-asp and DPBX-L-glu and 0.1-5 µg/ml for DBPX-gly. Chromatograms exhibit clean 2.0-min running time peaks and S/N ratios for the LLOQ were approximately 15:1. The technique was used to evaluate the pharmacokinetics of the molecules and to correlate these with timecourse toxic effects., Conclusion: DBSs represent an advantage for the collection of small volumes of samples, and also in terms of processing and storage. UPLC-MS/MS allow us not only to identify the isotopic pattern of boron in DBPX, but also to quantify them with accuracy and specificity. Pharmacokinetics of these molecules exhibit a high apparent volume of distribution; it suggests a preference of DPBX-amino acids for fatty tissues such as the CNS.

Background: The aim of the present work was to develop a chromatographic technique coupled with mass spectrometry for the measurement of lamotrigine in plasma. Lamotrigine and guanabenz (internal standard) were measured by selected reaction monitoring. The method was validated and applied in a bioequivalence trial on 26 female volunteers. Lamotrigine chewable tablets (100 mg) were administered and monitored for up to 96 h., Results: The method was linear between 0.05 and 5.0 µg/ml, with acceptable stability, accuracy and precision. Mean maximum plasma concentration was 1.37 µg/ml and was reached at 1.6 h postdose. Elimination half-life was 32.7 h., Conclusion: Lamotrigine tablets were bioequivalent. Ultra-performance liquid chromatography with tandem mass spectrometry represents a powerful tool in terms of sensitivity, specificity and high-throughput analysis.