A population pharmacokinetic model using allometric scaling for baricitinib in patients with juvenile idiopathic arthritis.

Baricitinib is approved for the treatment of rheumatoid arthritis (RA) in more than 70 countries, and juvenile idiopathic arthritis (JIA) in the European Union. Population pharmacokinetic (PK) models were developed in a phase 3 trial to characterize PK in pediatric patients with JIA and identify weight-based dosing regimens. The phase 3, randomized, double-blind, placebo-controlled withdrawal, efficacy and safety trial, JUVE-BASIS, enrolled patients (aged 2 to <18 years) with polyarticular course JIA. During a safety/PK period, baricitinib concentration data from age-based dose cohorts were compared to concentrations from adult patients receiving 4-mg QD. PK data were used to develop a population PK model with allometric scaling to determine a weight-based posology in pediatric patients with JIA that matched the adult 4-mg exposure. Baricitinib plasma concentrations from 217 pediatric patients were used to characterize PK. Based on the adult model, pediatric PK was best described using a 2-compartment model with allometric scaling on clearance and volume of distribution and renal function (estimated with glomerular filtration rate [GFR], a known covariate affecting PK of baricitinib) on clearance. The PK modeling suggested the optimal dosing regimen based on weight for pediatric patients as: 2-mg QD for patients 10 to <30 kg and 4-mg QD for patients ≥30 kg. The use of a population PK model of baricitinib treatment in adult patients with RA, with the addition of allometric scaling for weight on clearance and volume terms, was useful to predict exposures and identify weight-based dosing in pediatric patients with JIA.

Simple and rapid quantification of ribociclib in rat plasma by protein precipitation and LC-MS/MS: An application to pharmacokinetics of ribociclib nanoparticles in rats.

Ribociclib is a cyclin-dependent kinase (CDK4/6) inhibitor and is a standard of care for treating metastatic breast cancer. The drug has moderate oral bioavailability and exhibits permeability-controlled absorption. Novel formulations to enhance ribociclib pharmacokinetics are being developed and tested in rats. This requires developing analytical assays for quantifying ribociclib monitoring in rat plasma. We present a fully validated, sensitive, and simple LC-MS/MS method for measuring ribociclib in rat plasma. Ribociclib-D6 was utilized as an internal standard, and a simple protein precipitation procedure with acetonitrile was used in sample preparation. Excellent assay linearity was observed over a standard curve concentration of 1.008-1027.624 ng/mL. Acceptable intra- and inter-day accuracy and reproductivity were demonstrated for ribociclib quality controls (bias and CV% within ±15%). Complete extraction recovery of ribociclib was achieved, and a negligible matrix effect of analyte to internal standard ratio was observed. Ribociclib was stable at various conditions, including bench-top, freeze-thaw, and short-term stability. Overall, the presented method is simple, sensitive, accurate, and precise and was successfully applied to quantify ribociclib in plasma samples from a pharmacokinetic study of ribociclib suspension and nanoparticle formulation in rats.

Phase I or II Study of Ribociclib in Combination With Topotecan-Temozolomide or Everolimus in Children With Advanced Malignancies: Arms A and B of the AcSé-ESMART Trial.

PURPOSE: AcSé-ESMART is a proof-of-concept, phase I or II, platform trial, designed to explore targeted agents in a molecularly enriched cancer population. Arms A and B aimed to define the recommended phase II dose and activity of the CDK4/6 inhibitor ribociclib with topotecan and temozolomide (TOTEM) or everolimus, respectively, in children with recurrent or refractory malignancies. PATIENTS AND METHODS: Ribociclib was administered orally once daily for 16 days after TOTEM for 5 days (arm A) or for 21 days with everolimus orally once daily continuously in a 28-day cycle (arm B). Dose escalation followed the continuous reassessment method, and activity assessment the Ensign design. Arms were enriched on the basis of molecular alterations in the cell cycle or PI3K/AKT/mTOR pathways. RESULTS: Thirty-two patients were included, 14 in arm A and 18 in arm B, and 31 were treated. Fourteen patients had sarcomas (43.8%), and 13 brain tumors (40.6%). Main toxicities were leukopenia, neutropenia, and lymphopenia. The recommended phase II dose was ribociclib 260 mg/m2 once a day, temozolomide 100 mg/m2 once a day, and topotecan 0.5 mg/m2 once a day (arm A) and ribociclib 175 mg/m2 once a day and everolimus 2.5 mg/m2 once a day (arm B). Pharmacokinetic analyses confirmed the drug-drug interaction of ribociclib on everolimus exposure. Two patients (14.3%) had stable disease as best response in arm A, and seven (41.2%) in arm B, including one patient with T-acute lymphoblastic leukemia with significant blast count reduction. Alterations considered for enrichment were present in 25 patients (81%) and in eight of nine patients with stable disease; the leukemia exhibited CDKN2A/B and PTEN deficiency. CONCLUSION: Ribociclib in combination with TOTEM or everolimus was well-tolerated. The observed activity signals initiated a follow-up study of the ribociclib-everolimus combination in a population enriched with molecular alterations within both pathways.

A Highly Sensitive Nonextraction-Assisted HPLC Method with Fluorescence Detection for Quantification of Duvelisib in Plasma Samples and its Application to Pharmacokinetic Study in Rats.

BACKGROUND: Duvelisib (DUV) is a new oral phosphoinositide-3-kinase (PI3K)-δ and PI3K-γ inhibitor. It has been recently granted an accelerated approval for treatment of adult patients with relapsed or refractory chronic lymphocytic leukemia (CLL) and small lymphocytic lymphoma (SLL). It is also effective in therapy of T-cell lymphoma, solid tumors, and non-Hodgkin's lymphoma. In literature, there is no method valid for quantitation of DUV in human plasma for its therapeutic monitoring and pharmacokinetic studies. PURPOSE: The purpose of this study is the establishment of a highly sensitive HPLC method with fluorescence detection for quantitation of DUV in plasma for its therapeutic monitoring and pharmacokinetic studies of DUV. METHODS: The resolution of DUV and the internal standard (IS) olaparib (OLA) was achieved on Nucleosil CN column, with a mobile phase composed of acetonitrile:water (25:75, v/v) at a flow rate of 1.7 mL min-1. The fluorescence of both DUV and OLA was detected at 410 nm after excitation at 280 nm. The method was validated according to the guidelines of bioanalytical method validation. RESULTS: The method was linear in the range of 5-100 ng mL-1, and its limit of detection (LOD) and limit of quantitation (LOQ) were 2.12 ng mL-1 and 7 ng mL-1, respectively. The precisions of the method were ≤ 8.26%, and its accuracies were ≥ 95.32%. All the other validation parameters were satisfactory. The proposed method was successfully employed to the investigation of the pharmacokinetic profile of DUV in rats following a 25 mg/kg single dose of oral administration. CONCLUSION: The method is characterized with high sensitivity, accuracy, simple sample pretreatment, rapidity, eco-friendly as it consumes low volumes of organic solvent in the mobile phase and has high analysis throughput as its run time was short (~ 10 min).

Ribociclib Population Pharmacokinetics and Pharmacokinetic/Pharmacodynamic Analysis of Neutrophils in Cancer Patients.

The population pharmacokinetics (popPK) of ribociclib and population pharmacokinetic/pharmacodynamic (PK/PD) relationship between ribociclib and absolute neutrophil count (ANC) were characterized in patients with cancer. PopPK and ANC PK/PD modeling were both conducted in 2 rounds per data availability. Initial models were developed based on data sets from early-phase trials and qualified using external data from the phase III MONALEESA-2 trial. The second round of analyses was performed using updated data sets that included 2 more phase III trials (MONALEESA-3 and -7). The popPK and ANC PK/PD models adequately described the data and demonstrated reasonable predictive ability. Covariate analysis showed that ribociclib PK were not affected by age, sex, race, baseline Eastern Cooperative Oncology Group (ECOG) status (grade 1), mild/moderate renal impairment, mild hepatic impairment, or concomitant use of combination partners, including aromatase inhibitors (letrozole, anastrozole) or fulvestrant, proton-pump inhibitors, or weak cytochrome P450 3A4/5 inhibitors. Body weight had no impact on ribociclib clearance to warrant dose adjustment. The ANC PK/PD relationship was not affected by age, weight, sex, race, baseline ECOG status (grade 1), or concomitant use of letrozole, anastrozole, or fulvestrant. The PK/PD analysis confirmed reversibility of ribociclib's effect on ANC; it also suggested that lowering the dose of ribociclib would mitigate ANC decrease and neutropenia risk. The popPK and ANC PK/PD analyses support the use of ribociclib in combination with an aromatase inhibitor or fulvestrant in patients with hormone receptor-positive, human epidermal growth factor receptor-2-negative advanced or metastatic breast cancer without dose adjustment in subpopulations, and the use of dose interruption/reduction to mitigate potential treatment-emergent neutropenia.

Pharmacokinetics of Ribociclib in Subjects With Hepatic Impairment.

Ribociclib is an orally bioavailable, highly selective small-molecule inhibitor of cyclin-dependent kinases 4 and 6. It is currently approved for the treatment of hormone receptor-positive, human epidermal growth factor receptor 2-negative breast cancer with a starting dose of 600 mg. Both in vitro and in vivo studies indicate that ribociclib is primarily metabolized in the liver via cytochrome P450 3A4. A phase 1, open-label, multicenter, parallel cohort, single-oral-dose study was conducted to characterize the pharmacokinetics of ribociclib in subjects with varying degrees of hepatic impairment as measured by the Child-Pugh classification. Subjects were divided into 4 cohorts determined by their degree of hepatic impairment: normal, mild, moderate, or severe. Thirty subjects were enrolled and received a single 400 mg dose of ribociclib. Ribociclib exposure was similar in subjects with mild hepatic impairment compared with subjects with normal hepatic function, but was increased by approximately 30% in subjects with moderate and severe hepatic impairment. At a dose of 400 mg, ribociclib was generally well tolerated in all subjects regardless of the level of hepatic impairment. These results indicate that no dose adjustment (recommended dose of ribociclib is 600 mg daily, 3 weeks on and 1 week off) is necessary for patients with mild hepatic impairment but that a reduced dose of 400 mg daily, 3 weeks on and 1 week off in patients with moderate or severe hepatic impairment is recommended.

Pharmacogenetic studies of thiopurine methyltransferase genotype-phenotype concordance and effect of methotrexate on thiopurine metabolism.

The discovery and implementation of thiopurine methyltransferase (TPMT) pharmacogenetics has been a success story and has reduced the suffering from serious adverse reactions during thiopurine treatment of childhood leukaemia and inflammatory bowel disease. This MiniReview summarizes four studies included in Dr Zimdahl Kahlin's doctoral thesis as well as the current knowledge on this field of research. The genotype-phenotype concordance of TPMT in a cohort of 12 663 individuals with clinically analysed TPMT status is described. Notwithstanding the high concordance, the benefits of combined genotyping and phenotyping for TPMT status determination are discussed. The results from the large cohort also demonstrate that the factors of gender and age affect TPMT enzyme activity. In addition, characterization of four previously undescribed TPMT alleles (TPMT*41, TPMT*42, TPMT*43 and TPMT*44) shows that a defective TPMT enzyme could be caused by several different mechanisms. Moreover, the folate analogue methotrexate (MTX), used in combination with thiopurines during maintenance therapy of childhood leukaemia, affects the metabolism of thiopurines and interacts with TPMT, not only by binding and inhibiting the enzyme activity but also by regulation of its gene expression.

Development of Sustained Release Baricitinib Loaded Lipid-Polymer Hybrid Nanoparticles with Improved Oral Bioavailability.

Baricitinib (BTB) is an orally administered Janus kinase inhibitor, therapeutically used for the treatment of rheumatoid arthritis. Recently it has also been approved for the treatment of COVID-19 infection. In this study, four different BTB-loaded lipids (stearin)-polymer (Poly(d,l-lactide-co-glycolide)) hybrid nanoparticles (B-PLN1 to B-PLN4) were prepared by the single-step nanoprecipitation method. Next, they were characterised in terms of physicochemical properties such as particle size, zeta potential (ζP), polydispersity index (PDI), entrapment efficiency (EE) and drug loading (DL). Based on preliminary evaluation, the B-PLN4 was regarded as the optimised formulation with particle size (272 ± 7.6 nm), PDI (0.225), ζP (-36.5 ± 3.1 mV), %EE (71.6 ± 1.5%) and %DL (2.87 ± 0.42%). This formulation (B-PLN4) was further assessed concerning morphology, in vitro release, and in vivo pharmacokinetic studies in rats. The in vitro release profile exhibited a sustained release pattern well-fitted by the Korsmeyer-Peppas kinetic model (R2 = 0.879). The in vivo pharmacokinetic data showed an enhancement (2.92 times more) in bioavailability in comparison to the normal suspension of pure BTB. These data concluded that the formulated lipid-polymer hybrid nanoparticles could be a promising drug delivery option to enhance the bioavailability of BTB. Overall, this study provides a scientific basis for future studies on the entrapment efficiency of lipid-polymer hybrid systems as promising carriers for overcoming pharmacokinetic limitations.

First orally bioavailable prodrug of proteolysis targeting chimera (PROTAC) degrades cyclin-dependent kinases 2/4/6 in vivo.

A growing number of reports suggested that the inhibitor targeting cyclin-dependent kinases (CDK) 2/4/6 can act as a more feasible chemotherapy strategy. In the present paper, a novel PROTAC molecule was developed based on the structure of Ribociclib's derivative. In malignant melanoma cells, the degrader can not only degrade CDK 2/4/6 simultaneously and effectively, but also remarkably induce cell cycle arrest and apoptosis of melanoma cells. Moreover, PROTAC molecules with CRBN ligands always have poor oral bioavailability. We developed the orally bioavailable prodrug for the first time. It would provide general solution for oral administration of the PROTAC molecules, derived from CRBN ligands, for animal test conveniently.

Regulation of Anxiety and Depression by Mitochondrial Translocator Protein-Mediated Steroidogenesis: the Role of Neurons.

Pharmacological studies have implicated the translocator protein (TSPO) in the regulation of complex behaviors including anxiety and depression, effects thought to be mediated by increased synthesis of neuroactive steroid hormones. However, TSPO function in the brain remains to be corroborated in vivo via genetic studies. To address this, we developed global TSPO knockout (TSPO-KO) and neuronal TSPO transgenic (TSPO-Tg) mouse models to investigate TSPO function in the regulation of anxiety- and depression-related behaviors using elevated plus maze and forced swim test paradigms. Neuroactive steroid hormones were measured in the brain by mass spectrometry. In vivo TSPO ligand pharmacokinetics was investigated using competitive PET with 18F-FE-DAA1106. Genetic TSPO deficiency increased anxiety-related behavior and impaired brain steroidogenesis but did not affect depressive behaviors. Using the TSPO-KO model, we then demonstrated the specificity of Ac-5216, also known as XBD-173 or Emapunil, as an anxiolytic targeting TSPO at doses optimized by competitive PET for high cortical occupancy. Neuronal TSPO overexpression decreased depressive behaviors, an effect that was dependent on steroidogenesis, and partially reversed anxiogenic behavior in TSPO-KO mice. These findings demonstrate that TSPO is critical for brain steroidogenesis and modulates anxiety- and depression-related behaviors. However, we demonstrate that key differences in the contribution of neuronal TSPO to the modulation of these complex behaviors, illustrating the tissue- and cell-specific importance of TSPO. The TSPO-KO and TSPO-Tg mice provide the tools and rationale for the development of therapeutic approaches targeting TSPO in the brain for treatment of neuropsychiatric conditions.

DBS Assay with LC-MS/MS for the Determination of Idelalisib, A Selective PI3K-δ Inhibitor in Mice Blood and Its Application to a Pharmacokinetic Study.

Idelalisib is a selective and second-generation PI3K-δ inhibitor, approved for the treatment of non-Hodgkin lymphoma and chronic lymphocytic leukemia. In this paper, we present a fully validated dried blood spot (DBS) method for the quantitation of idelalisib from mice blood using an LC-MS/MS, which was operated under multiple reaction monitoring mode. To the punched DBS discs, acidified methanol enriched with internal standard (IS; larotrectinib) was added and extracted using tert-butyl methyl ether as an extraction solvent with sonication. Chromatographic separation of idelalisib and the IS was achieved on an Atlantis dC18 column using a mixture of 10 mM ammonium formate:acetonitrile (25:75, v/v). The flow-rate and injection volume were 0.80 mL/min and 2.0 µL, respectively. Idelalisib and the IS were eluted at ~0.98 and 0.93 min, respectively and the total run time was 2.00 min. Idelalisib and the IS were analyzed using positive ion scan mode and parent-daughter mass to charge ion (m/z) transition of 416.1→176.1 and 429.1→342.1, respectively was used for the quantitation. The calibration range was 1.01-4 797 ng/mL. No matrix effect and carry over were observed. Haematocrit did not influence DBS idelalisib concentrations. All the validation parameters met the acceptance criteria. The applicability of the validated method was shown in a mice pharmacokinetic study.

Potent and orally active purine-based fetal hemoglobin inducers for treating ß-thalassemia and sickle cell disease.

Reactivation of fetal hemoglobin (HbF) expression by therapeutic agents has been suggested as an alternative treatment to modulate anemia and the related symptoms of severe ß-thalassemia and sickle cell disease (SCD). Hydroxyurea (HU) is the first US FDA-approved HbF inducer for treating SCD. However, approximately 25% of the patients with SCD do not respond to HU. A previous study identified TN1 (1) as a small-molecule HbF inducer. However, this study found that the poor potency and oral bioavailability of compound 1 limits the development of this inducer for clinical use. To develop drug-like compounds, further structure-activity relationship studies on the purine-based structure of 1 were conducted. Herein, we report our discovery of a more potent inducer, compound 13a, that can efficiently induce γ-globin gene expression at non-cytotoxic concentrations. The molecular mechanism of 13a, for the regulation HbF expression, was also investigated. In addition, we demonstrated that oral administration of 13a can ameliorate anemia and the related symptoms in SCD mice. The results of this study suggest that 13a can be further developed as a novel agent for treating hemoglobinopathies, such as ß-thalassemia and SCD.

Validated HPLC-UV method for simultaneous quantification of phosphatidylinositol 3-kinase inhibitors, copanlisib, duvelisib and idelalisib, in rat plasma: Application to a pharmacokinetic study in rats.

Phosphatidylinositol 3-kinase (PI3K) inhibitors are a novel class of anticancer drugs that are approved to treat various malignancies. We report the development and validation of a HPLC method for the simultaneous quantitation of three PI3K inhibitors, namely copanlisib, duvelisib and idelalisib, in rat plasma as per the regulatory guidelines of the United States Food and Drug Administration. The method involves extraction of copanlisib, duvelisib and idelalisib along with an internal standard (IS; filgotinib) from rat plasma (100 µL) using a liquid-liquid extraction process. The chromatographic separation of the analytes was achieved using step-wise gradient elution on a Hypersil Gold C18 column. The UV detection wavelength was set at λmax = 280 nm. Copanlisib, duvelisib, idelalisib and the IS eluted at 7.16, 12.6, 11.9 and 9.86 min, respectively, with a total run time of 15 min. The calibration curve ranged from 50 to 5000 ng/mL for all the analytes. Inter- and intra-day precision and accuracy, stability studies, dilution integrity and incurred sample reanalysis were investigated for all three analytes, and the results met the acceptance criteria. The validated HPLC method was successfully applied to a pharmacokinetic study in rats.

Physiologically Based Pharmacokinetic Modeling of Central Nervous System Pharmacokinetics of CDK4/6 Inhibitors to Guide Selection of Drug and Dosing Regimen for Brain Cancer Treatment.

A better understanding of the human central nervous system (CNS) pharmacokinetics is critical to the selection of the right drug and refinement of dosing regimen for more effective treatment of primary and metastatic brain cancer. Using the physiologically-based pharmacokinetic (PBPK) modeling approach, we systematically compared the CNS pharmacokinetics of three cyclin D-cyclin dependent kinase 4 and 6 (CDK4/6) inhibitors (ribociclib, palbociclib, and abemaciclib) in patients with cancer. A PBPK model platform was developed and verified for predicting plasma and CNS pharmacokinetics. Target engagement ratio (TER), defined as the ratio of the average steady-state unbound drug brain concentration to the in vitro half-maximal inhibitory concentration (IC50 ) for CDK4/6 inhibition, was used as a crude predictor of efficacy. As compared with ribociclib and palbociclib, abemaciclib penetrated into the human brain to a larger extent, but at a slower rate, and was retained in the brain longer. Following the standard dosing regimens, the predicted CDK4/6 TERs were 26/5.2 for abemaciclib, 2.4/0.62 for ribociclib, and 0.36/0.27 for palbociclib. Simulations suggested that abemaciclib achieved comparable TERs following twice daily or daily dosing; ribociclib may sufficiently inhibit both CDK4 and CDK6 at the maximum tolerated dose; whereas, palbociclib achieved TERs < 0.5 even at a dose 50% higher than the standard dose. In conclusion, the PBPK modeling, supported by available preclinical and clinical evidence, suggests that abemaciclib is the best CDK4/6 inhibitor for brain cancer treatment, whereas palbociclib is not recommended. The model refined dosing regimen is 300 mg daily on a 4-weeks-on schedule for abemaciclib, and 900 mg daily on a 3-weeks-on/1-week-off schedule for ribociclib.

Clinical Pharmacokinetics and Pharmacodynamics of the Cyclin-Dependent Kinase 4 and 6 Inhibitors Palbociclib, Ribociclib, and Abemaciclib.

Palbociclib, ribociclib, and abemaciclib are inhibitors of the cyclin-dependent kinases 4 and 6 approved for the treatment of locally advanced or metastatic breast cancer. In this review, we provide an overview of the available clinical pharmacokinetic and pharmacodynamic characteristics of these novel drugs, summarize the results of food-effect and drug-drug interaction studies, and highlight exposure-response and exposure-toxicity relationships. All three drugs exhibit a large inter-individual variability in exposure (coefficient of variation range 40-95% for minimum plasma concentration), are extensively metabolized by cytochrome P450 3A4, and have their brain penetration limited by efflux transporters. Abemaciclib has three active metabolites with similar potency that are clinically relevant (i.e., M2, M20, M18), whereas the metabolites of palbociclib and ribociclib are not of clinical significance. Pharmacokinetic exposure increases in a dose-proportional manner for palbociclib, whereas exposure increases under- and over-proportionally with an increasing dose for abemaciclib and ribociclib, respectively. High exposure is associated with an increased risk of neutropenia, and for ribociclib also to corrected QT prolongation. For abemaciclib, a clear exposure-efficacy relationship has been described, while for palbociclib and ribociclib exposure-response analyses remain inconclusive. Future studies are needed to address exposure-efficacy relationships to further improve dosing.

A phase I/II study of ribociclib following radiation therapy in children with newly diagnosed diffuse intrinsic pontine glioma (DIPG).

PURPOSE: Cyclin-dependent kinase-retinoblastoma (CDK-RB) pathway is dysregulated in some diffuse intrinsic pontine gliomas (DIPG). We evaluated safety, feasibility, and early efficacy of the CDK4/6-inhibitor ribociclib, administered following radiotherapy in newly-diagnosed DIPG patients. METHODS: Following radiotherapy, eligible patients received ribociclib in 28-day cycles (350 mg/m2; 21 days on/7 days off). Feasibility endpoints included tolerability for at least 6 courses, and a less than 2-week delay in restarting therapy after 1 dose reduction. Early efficacy was measured by 1-year and median overall survival (OS). Patient/parent-by-proxy reported outcomes measurement information system (PROMIS) assessments were completed prospectively. RESULTS: The study included 10 evaluable patients, 9 DIPG and 1 diffuse midline glioma (DMG)-all 3.7 to 19.8 years of age. The median number of courses was 8 (range 3-14). Three patients required dose reduction for grade-4 neutropenia, and 1 discontinued therapy for hematological toxicity following course 4. The most common grade-3/4 toxicity was myelosuppression. After 2 courses, MRI evaluations in 4 patients revealed increased necrotic volume, associated with new neurological symptoms in 3 patients. The 1-year and median OS for DIPG was 89% and 16.1 months (range 10-30), respectively; the DMG patient died at 6 months post-diagnosis. Five patients donated brain tissue and tumor; 3 were RB+ . CONCLUSIONS: Ribociclib administered following radiotherapy is feasible in DIPG and DMG. Increased tumor necrosis may represent a treatment effect. These data warrant further prospective volumetric analyses of tumors with necrosis. Feasibility and stabilization findings support further investigation of ribociclib in combination therapies. TRIAL REGISTRATION: NCT02607124.

Tofacitinib and Baricitinib Are Taken up by Different Uptake Mechanisms Determining the Efficacy of Both Drugs in RA.

BACKGROUND: Rheumatoid arthritis (RA) is a systemic autoimmune disease in which synovial fibroblasts (SF) play a key role. Baricitinib and Tofacitinib both act intracellularly, blocking the ATP-binding side of JAK proteins and thereby the downstream signalling pathway via STAT-3. Therefore, we investigated the role of organic cation transporters (OCTs) in Baricitinib and Tofacitinib cellular transport. METHODS: OCT expression was analysed in SF isolated from RA and osteoarthritis (OA) patients, as well as peripheral blood mononuclear cells. The interaction of Baricitinib and Tofacitinib with OCTs was investigated using quenching experiments. The intracellular accumulation of both drugs was quantified using LC/MS. Target inhibition for both drugs was tested using Western blot for phosphorylated JAK1 and STAT3 upon stimulation with IL-6. RESULTS: MATE-1 expression increased in OASF compared to RASF. The other OCTs were not differentially expressed. The transport of Baricitinib was not OCT dependent. Tofacitinib; however, was exported from RASF in a MATE-1 dependent way. Tofacitinib and Baricitinib showed comparable inhibition of downstream signalling pathways. CONCLUSION: We observed different cellular uptake strategies for Baricitinib and Tofacitinib. Tofacitinib was exported out of healthy cells due to the increased expression of MATE1. This might make Tofacitinib the favourable drug.

Pharmacokinetics, Safety, and Tolerability of Single- and Multiple-Dose Once-Daily Baricitinib in Healthy Chinese Subjects: A Randomized Placebo-Controlled Study.

The objective of this phase 1 study was to evaluate the pharmacokinetics, safety, and tolerability of baricitinib after single and multiple doses in healthy Chinese adults. Eligible subjects received a once-daily dose of baricitinib 2, 4, or 10 mg or placebo on day 1 (single dose) and days 4 through 10 for 7 consecutive days (multiple doses). Plasma pharmacokinetic samples were collected up to 48 hours after dosing on days 1 and 10, with predose samples collected before dosing on day 1 and days 4 through 10. Safety and tolerability were also assessed. Baricitinib was rapidly absorbed, reaching peak plasma concentrations within 0.5 to 1 hour (median). Plasma concentrations declined rapidly following the attainment of peak concentrations, with a mean terminal half-life of 5.7 to 7.3 hours. Steady-state plasma concentrations of baricitinib were achieved after the second day of once-daily dosing, with minimal accumulation of baricitinib in plasma (up to 10% increase in area under the plasma concentration-time curve). Single- and multiple-dose mean values for area under the plasma concentration-time curve from time zero to infinity and maximum plasma concentration appeared to increase in an approximately dose-proportional manner across the dose range. Single and multiple oral doses of once-daily baricitinib up to 10 mg were well tolerated by healthy Chinese subjects.

Current Understanding of the Intestinal Absorption of Nucleobases and Analogs.

It has long been suggested that a Na+-dependent carrier-mediated transport system is involved in the absorption of nucleobases and analogs, including some drugs currently in therapeutic use, for their uptake at the brush border membrane of epithelial cells in the small intestine, mainly based on studies in non-primate experimental animals. The presence of this transport system was indeed proved by the recent identification of sodium-dependent nucleobase transporter 1 (SNBT1/Slc23a4) as its molecular entity in rats. However, this transporter has been found to be genetically deficient in humans and higher primates. Aware of this deficiency, we need to revisit the issue of the absorption of these compounds in the human small intestine so that we can understand the mechanisms and gain information to assure the more rational use and development of drugs analogous to nucleobases. Here, we review the current understanding of the intestinal absorption of nucleobases and analogs. This includes recent knowledge about the efflux transport of those compounds across the basolateral membrane when exiting epithelial cells, following brush border uptake, in order to complete the overall absorption process; the facilitative transporters of equilibrative nucleoside transporter 1 (ENT1/SLC29A1) and equilibrative nucleobase transporter 1 (ENBT1/SLC43A3) may be involved in that in many animal species, including human and rat, without any major species differences.

Development of a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for characterizing caffeine, methylliberine, and theacrine pharmacokinetics in humans.

Coffea liberica possesses stimulant properties without accumulating the methylxanthine caffeine. The basis for this peculiar observation is that methylurates (e.g., theacrine and methylliberine) have replaced caffeine. The stimulant properties of methylurates, alone and in combination with caffeine, have recently been investigated. However, human pharmacokinetics and LC-MS/MS methods for simultaneous measurement of methylxanthines and methylurates are lacking. To address this deficiency, we conducted a pharmacokinetic study in which subjects (n = 12) were orally administered caffeine (150 mg), methylliberine (Dynamine™, 100 mg), and theacrine (TeaCrine®, 50 mg) followed by blood sampling over 24 h. Liquid-liquid extraction of plasma samples containing purine alkaloids and internal standard (13C-Caffeine) were analyzed using a C18 reversed-phase column and gradient elution (acetonitrile and water, both containing 0.1% formic acid). A Waters Xevo TQ-S tandem mass spectrometer (positive mode) was used to detect caffeine, methylliberine, theacrine, and IS transitions of m/z 195.11 â†’ 138.01, 225.12 â†’ 168.02, 225.12 â†’ 167.95, and 198.1 â†’ 140.07, respectively. The method was validated for precision, accuracy, selectivity, and linearity and was successfully applied to characterize the oral pharmacokinetics of caffeine, methylliberine, and theacrine in human plasma. Successful development and application of LC-MS/MS-based methods such as ours for the simultaneous measurement of methylxanthines and methylurates are essential for the characterization of potential pharmacokinetic and pharmacodynamic interactions.