Plasma Samples Research Articles

Associations between sex, systemic iron and inflammatory status and subcortical brain iron.

Brain iron increases in several neurodegenerative diseases are associated with disease progression. However, the causes of increased brain iron remain unclear. This study investigates relationships between subcortical iron, systemic iron and inflammatory status. Brain magnetic resonance imaging (MRI) scans and blood plasma samples were collected from cognitively healthy females (n = 176, mean age = 61.4 ± 4.5 years, age range = 28-72 years) and males (n = 152, mean age = 62.0 ± 5.1 years, age range = 32-74 years). Regional brain iron was quantified using quantitative susceptibility mapping. To assess systemic iron, haematocrit, ferritin and soluble transferrin receptor were measured, and total body iron index was calculated. To assess systemic inflammation, C-reactive protein (CRP), neutrophil:lymphocyte ratio (NLR), macrophage colony-stimulating factor 1 (MCSF), interleukin 6 (IL6) and interleukin 1β (IL1β) were measured. We demonstrated that iron levels in the right hippocampus were higher in males compared with females, while iron in the right caudate was higher in females compared with males. There were no significant associations observed between subcortical iron levels and blood markers of iron and inflammatory status indicating that such blood measures are not markers of brain iron. These results suggest that brain iron may be regulated independently of blood iron and so directly targeting global iron change in the treatment of neurodegenerative disease may have differential impacts on blood and brain iron.

Pharmacokinetics and Alterations in Glucose and Insulin Levels After a Single Dose of Canagliflozin in Healthy Icelandic Horses.

Canagliflozin (CFZ) is a sodium-glucose cotransporter-2 inhibitor that has shown promising results as a drug for the treatment of insulin dysregulation in horses. Even though CFZ is used clinically, no pharmacokinetic data has previously been published. In this study, the pharmacokinetics of CFZ after administration of a single oral dose of 1.8 mg/kg in eight healthy Icelandic horses was examined. Additionally, the effect of treatment on glucose and insulin levels in response to a graded glucose infusion was investigated. Plasma samples for CFZ quantification were taken at 0, 0.33, 0.66, 1, 1.33, 1.66, 2, 2.33, 2.66, 3, 3.5, 4, 5, 6, 8, 12, 24, 32, and 48 h post administration. CFZ was quantified using UHPLC coupled to tandem quadrupole mass spectrometry (UHPLC-MS/MS). A non-compartmental analysis revealed key pharmacokinetic parameters, including a median Tmax of 7 h, a Cmax of 2350 ng/mL, and a t1/2Z of 28.5 h. CFZ treatment reduced glucose (AUCGLU, p = 0.001) and insulin (AUCINS, p = 0.04) response to a graded glucose infusion administered 5 h after treatment. This indicates a rapid onset of action following a single dose in healthy Icelandic horses. No obvious adverse effects related to the treatment were observed.

Plasma collagen neoepitopes are associated with multiorgan disease in the ACCESS and GRADS sarcoidosis cohorts

IntroductionThe pathogenesis of sarcoidosis involves tissue remodelling mediated by the accumulation of abnormal extracellular matrix, which is partly the result of an imbalance in collagen synthesis, cross-linking and degradation. During...

Colorectal Cancer Recurrence Prediction Using a Tissue-Free Epigenomic Minimal Residual Disease Assay.

Post-treatment detection of circulating tumor DNA (ctDNA) is strongly predictive of recurrence. Most molecular residual disease (MRD) assays require prior tissue testing to guide ctDNA analysis, resulting in lengthy time to initial results and unevaluable patients. We assessed a tissue-free assay (Guardant Reveal) that bioinformatically evaluates >20,000 epigenomic regions for ctDNA detection in 1,977 longitudinally collected post-operative plasma samples from 342 patients with resected colorectal cancer (CRC). We observed sensitive and specific detection of MRD associated with clinically meaningful differences in recurrence-free interval at each timepoint evaluated with a median lead time of 5.3 months. Longitudinal sensitivity in stage II or higher colon cancer was 81%. Sensitivity increased with serial measurement and varied by recurrence site: higher for liver (100%) versus lung (53%) and peritoneal (40%). Sensitivity among rectal cancer patients was 60% owing to a high proportion of lung metastases. Specificity was 98.2% among 1,461 post-treatment samples (99.1% among those with follow-up longer than the upper interquartile range of the lead time observed in this study). Our data demonstrate the potential clinical utility of ctDNA as a tool to improve management of stage II and higher CRC with a methodology that is non-invasive, accessible, and allows for rapid evaluation to inform clinical decisions.

Circulating tumor DNA dynamics reveal KRAS G12C mutation heterogeneity and response to treatment with the KRAS G12C inhibitor divarasib in solid tumors.

To inform prognosis, treatment response, disease biology, and KRAS G12C mutation heterogeneity, we conducted exploratory circulating tumor DNA (ctDNA) profiling on 134 patients with solid tumors harboring a KRAS G12C mutation treated with single-agent divarasib (GDC-6036) in a phase 1 study. Plasma samples were collected for serial ctDNA profiling at baseline (Cycle 1 Day 1 prior to treatment) and multiple on-treatment time points (Cycle 1 Day 15 and Cycle 3 Day 1). KRAS G12C ctDNA was detectable from plasma samples in 72.9% (43/59) and 92.6% (50/54) of patients with non-small cell lung cancer and colorectal cancer, respectively, the majority of whom were eligible for study participation based on a local test detecting the KRAS G12C mutation in tumor tissue. Baseline ctDNA tumor fraction was associated with tumor type, disease burden, and metastatic sites. A decline in ctDNA level was observed as early as Cycle 1 Day 15. Serial assessment showed a decline in ctDNA tumor fraction associated with response and progression-free survival. Except for a few cases of KRAS G12C sub-clonality, on-treatment changes in KRAS G12C variant allele frequency mirrored changes in the overall ctDNA tumor fraction. Across tumor types, the KRAS G12C mutation likely represents a truncal mutation in the majority of patients. Rapid and deep decline in ctDNA tumor fraction was observed in patients responding to divarasib treatment. Early on-treatment dynamics of ctDNA were associated with patient outcomes and tumor response to divarasib treatment.

Circulating Tumor DNA Assessment for Treatment Monitoring Adds Value to PSA in Metastatic Castration Resistant Prostate Cancer.

Enzalutamide after abiraterone progression is commonly used in metastatic castration resistant prostate cancer (mCRPC) despite a low rate of clinical benefit. Analyzing IMbassador250, a phase III trial assessing enzalutamide with or without atezolizumab after abiraterone, we hypothesized that baseline and early changes in circulating tumor DNA (ctDNA) tumor fraction (TF) may identify patients more likely to exhibit survival benefit from enzalutamide. ctDNA was quantified from plasma samples using a tissue-agnostic assay without buffy coat sequencing. Baseline ctDNA TF, changes in ctDNA TF from baseline to cycle 3 day 1 (C3D1), and detection at C3D1 alone, were compared vs overall response rate (ORR), radiographic progression-free survival (rPFS), median OS (mOS), and 50% reduction in PSA. ctDNA TF detection at baseline and/or C3D1 was associated with shorter rPFS and OS in 494 evaluable patients. Detection of ctDNA TF at C3D1, with or without detection at C1D1, was associated with worse rPFS and mOS than lack of detection. When ctDNA TF and PSA response at C3D1 were discordant, patients with [ctDNA TF undetected/PSA not reduced] had more favorable outcomes than [ctDNA TF detected/PSA reduced] (mOS 22.1 months vs. 16 months, p<0.001). In a large cohort of mCRPC patients receiving enzalutamide after abiraterone, we demonstrate the utility of a new tissue-agnostic assay for monitoring molecular response based on ctDNA TF detection and dynamics. CtDNA TF provides a minimally-invasive, complementary biomarker to PSA testing and may refine personalized treatment approaches.

Development and Validation of a Sensitive Liquid Chromatography-Tandem Mass Spectrometry Method for Therapeutic Drug Monitoring of Ceftolozane and Tazobactam in Human Plasma Microsamples.

Ceftolozane/tazobactam (C-T) is a novel beta-lactam/beta-lactamase inhibitor combination approved for the treatment of various infections caused by difficult-to-treat Pseudomonas aeruginosa. In critically ill patients, C-T may exhibit significant pharmacokinetic variability, both between individuals and within individuals, warranting therapeutic drug monitoring for clinical purposes. We aim to develop and validate a novel and sensitive analytical method for concurrently determining C and T in human plasma microsamples (3μL). The method was developed using liquid chromatography-tandem mass spectrometry (LC-MS/MS) with positive electrospray ionization and multiple reaction monitoring (MRM) detection modes, employing specific mass transitions for both drugs. Sample preparation was simple, and the chromatographic run lasted only 4 minutes. Validation was conducted according to European Medicines Agency (EMA) guidelines, encompassing specificity, sensitivity, linearity, precision, accuracy, matrix effect, extraction recovery, limit of quantification, and drug stability. The validated method was applied to measure C and T in 32 plasma samples collected from critically ill patients with multidrug-resistant, gram-negative, bacterial infections. The method ensured accurate (BIAS% 2.1-9.6 for C and -2.2 to 15.2 for T) and precise intraday CV% for C: 6.7-5.5; for T: 1.3-8.9; interday CV% for C 6.0-10.8; for T 4.1-10.2) measurements of C-T over a wide concentration range (0.2-200.0 mg/L for C and 0.1-100.0 mg/L for T). Overall, the recovery at quality control concentration levels was high for both C and T (mean values: 90-91 for C and 89-92 for T). Analyte stability was satisfactory in both human plasma and extracts under various storage conditions. The clinical applicability of the assay was confirmed by the reliably quantifying C and T in clinical plasma samples. The developed and validated LC-MS/MS method is sensitive and suitable for monitoring C and T in human plasma microsamples.

Inverted Sandwich-Type e-LCR Aided by Lambda Exonuclease-RecJf Combination Enables Ultrasensitive Detection of Low-Frequency EGFR-L858R Mutation in NSCLC.

Highly sensitive detection of low-frequency EGFR-L858R mutation is particularly important in guiding targeted therapy of nonsmall-cell lung carcinoma (NSCLC). To this end, a ligase chain reaction (LCR)-based electrochemical biosensor (e-LCR) with an inverted sandwich-type architecture was provided by combining a cooperation of lambda exonuclease-RecJf exonuclease (λ-RecJf exo). In this work, by designing a knife-like DNA substrate (an overhang ssDNA part referred to the "knife arm") and introducing the λ-RecJf exo, the unreacted DNA probes in the LCR were specially degraded while only the ligated products were preserved, after which the ligated knife-like DNA products were hybridized with capture probes on the gold electrode surface through the "knife arms", forming the inverted sandwich-type DNA structure and bringing the methylene blue-label close to the electrode surface to engender the electrical signal. Finally, the sensitivity of the e-LCR could be improved by 3 orders of magnitude with the help of the λ-RecJf exo, and due to the mutation recognizing in the ligation site of the employed ligase, this method could detect EGFR-L858R mutation down to 0.01%, along with a linear range of 1 fM-10 pM and a limit detection of 0.8 fM. Further, the developed method could distinguish between L858R positive and negative mutations in cultured cell samples, tumor tissue samples, and plasma samples, whose accuracy was verified by the droplet digital PCR, holding a huge potential in liquid biopsy for precisely guiding individualized-treatment of NSCLC patients with advantages of high sensitivity, low cost, and adaptability to point-of-care testing.

Identification of Amino Acids and Polyphenolic Metabolites in Human Plasma by UHPLC-ESI-QTOF-MS/MS, after the Chronic Intake of a Functional Meal in an Elderly Population

Novel foods especially formulated and targeted for the elderly population should provide sufficient nutrients and bioactive ingredients to counteract the natural age-related deterioration of various organs and tissues. Dietary protein and phenolic compounds achieve this goal; however, older adults have alterations in their gastrointestinal system that may impact their bioavailability and few studies have been aimed at this population. Since phenolic compounds are the subject of multiple biotransformations by host and microbiome enzymes during the digestion process, identification of their bioavailable forms in human plasma or tissues represents a considerable analytical challenge. In this study, UHPLC-ESI-QTOF/MS-MS, chemometrics, and multivariate statistical methods were used to identify the amino acids and phenolic compounds that were increased in the plasma of elderly adults after a 30-day intervention in which they had consumed an especially formulated muffin and beverage containing Brosimum alicastrum Sw. seed flour. A large interindividual variation was observed regarding the amino acids and phenolic metabolites identified in the plasma samples, before and after the intervention. Three phenolic metabolites were significantly increased in the population after the intervention: protocatechuic acid, 5-(methoxy-4′-hydroxyphenyl) valerolactone, and phloretic acid. These metabolites, as well as others that were not significantly increased (although they did increase in several individuals), are probably the product of the microbiota metabolism of the major phenolic compounds present in the B. alicastrum Sw. seed flour and other food ingredients. A significant decrease in 4-ethyl-phenol, a biomarker of stress, was observed in the samples. Results showed that the incorporation of foods rich in phenolic compounds into the regular diet of older adults contributes to the increase in bioactive compounds in plasma, that could substantially benefit their mental, cardiovascular, and digestive health.

Evaluation of the Fluorion HCV QNP v3.0 real-time PCR assay for quantifying HCV RNA in Moroccan patients: A comparative study with COBAS AmpliPrep/COBAS TaqMan HCV v2.0

PurposeHepatitis C Virus (HCV) RNA quantification is crucial for diagnosing and monitoring chronic HCV treatment. Cost-effective methods are crucial to ensure accessibility. This study evaluated the Fluorion HCV QNP v3.0 Real-Time PCR assay's effectiveness in EDTA-plasma and serum, comparing it with the COBAS AmpliPrep/COBAS TaqMan HCV v2.0 (CAP/CTM HCV v2.0) test. Methods105 matched pairs of EDTA-plasma and serum specimens (91 positive and 14 negative) from HCV infected Moroccan patients were analyzed using the Fluorion HCV QNP v3.0 assay and compared to the CAP/CTM HCV v2.0 test, being the reference method. ResultsThe values obtained by the Fluorion HCV QNP v3.0 in plasma were slightly higher than those in serum (3.94 ± 2.23 log10 IU/mL versus 3.91 ± 2.22 log10 IU/mL) and were both significantly lower than those quantified by the CAP/CTM HCV v2.0 assay (4.34 ± 2.28 log10 IU/mL; p < 0.001). High correlations were observed between the Fluorion HCV QNP v3.0 serum and CAP/CTM HCV v2.0 (R2 = 0.9433), the Fluorion HCV QNP v3.0 plasma and CAP/CTM HCV v2.0 (R2 = 0.949) and the Fluorion HCV QNP v3.0 serum and plasma (R2 = 0.9954). HCV RNA was detected in all tested genotypes by both assays. ConclusionThe Fluorion HCV QNP v3.0 assay demonstrated excellent performance in comparison with the CAP/CTM HCV v2.0 on both plasma and serum samples which can be used interchangeably for HCV quantification. The test was shown to be suitable for disease monitoring including all HCV genotypes.

Determination of ivermectin in plasma and whole blood using LC-MS/MS

Background Ivermectin is a widely used drug for the treatment of helminthiasis and filariasis worldwide, and it has also shown promise for malaria elimination through its potent mosquito-lethal activity. The objective of this study was to develop and validate a high-throughput and sensitive method to quantify ivermectin in plasma and whole blood samples, using automated sample extraction followed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Methods Phospholipids were removed in patient whole blood (100 µl) and plasma (100 µl) samples using a 96-well plate Hybrid-solid phase extraction technique. Ivermectin and its isotope-labelled internal standard (ivermectin-D2) were separated on an Agilent Poroshell 120 EC-C18 50mm × 3.0mm I.D. 2.7µm, using a mobile phase of acetonitrile: ammonium formate 2 mM containing 0.5% formic acid (90: 10, v/v). Detection was performed using a triple quadrupole mass spectrometer in the positive ionization mode. Results The method was validated in the concentration range 0.970 - 384 ng/ml in both plasma and whole blood matrices. Intra- and inter-batch precisions during the validation were below 15%. There was no carryover or matrix effects detected. Ivermectin is a stable compound and results showed no degradation in the different stability tests. Conclusions The validated method proved to have high sensitivity and precision, good selectivity and to be suitable for clinical application or laboratory quantification of ivermectin in plasma or whole blood samples.

43 Epigenomic profiling of translocation renal cell carcinoma via liquid biopsy

Abstract Background Translocation renal cell carcinoma (tRCC) is an aggressive subtype of kidney cancer usually driven by a fusion involving the TFE3 gene. Due to histologic overlap with other subtypes of RCC, tRCC is frequently misclassified. Methods for accurate diagnosis and detection of this molecularly distinct entity are therefore a pressing need. Epigenomic profiling of ctDNA via plasma chromatin immunoprecipitation and sequencing (ChIP-seq) has recently emerged as a powerful tool to detect and molecularly subtype cancers and may offer a more sensitive and specific detection of molecular fusions. We aimed to detect tRCC in plasma and to discriminate tRCC from ccRCC based on epigenomic profiling of cfDNA. Methods We first identified differentially expressed gene, methylated regions (DMRs) and regulatory elements (REs) specific to tRCC vs. ccRCC via RNA-seq, methylated DNA immunoprecipitation sequencing (MeDIP-seq) and ChIP-Seq, of 4 tRCC and 5 ccRCC cell lines. We collected 16 plasma samples from metastatic patients with tRCC, 11 with ccRCC and 9 healthy patients (HP). Ultra low pass whole genome sequencing (ulpWGS) was performed to infer ctDNA fraction (TF), and cfMeDIP-seq, H3K4me3/H3K27ac cfChIP-seq for epigenomic profiling. Signal at tRCC-specific regions derived from cell lines profiling was aggregated for each mark and normalized to common active REs and DMRs, then compared between classes using a Wilcoxon rank-sum test. Classification performance was evaluated using the area under the receiver operating characteristic (AUROC) curve. Results Overall 8/16 tRCC and 5/12 ccRCC samples had &amp;gt;3% TF by ulpWGS. H3K4me3 and H3K27ac cfChIP-seq signal was significantly higher in all tRCC samples compared to healthy patients (p&amp;lt;10-6, AUROC =1), at tRCC-specific promoters and tRCC-specific REs respectively. Furthermore, H3K27ac signal in plasma was also significantly higher at tRCC-specific REs in tRCC samples compared to ccRCC (p&amp;lt;10-4, AUROC=0.95). MeDIP-seq could not discriminate between tRCC and ccRCC. Conclusions Although a majority of tRCC plasma samples profiled had TF &amp;lt; 3%, all could be distinguished from healthy samples on the basis of cfChIP. H3K27ac cfChIP-Seq was also discriminatory for tRCC vs. ccRCC. Epigenomic profiling of cfDNA appears as a powerful tool for both detection of tRCC and discrimination from ccRCC/healthy plasma, with potential implications for diagnosis and guiding therapy.

Plasma extracellular vesicle long RNA profiling identifies a predictive signature for immunochemotherapy efficacy in lung squamous cell carcinoma

IntroductionThe introduction of Immune Checkpoint Inhibitors (ICIs) has marked a paradigm shift in treating Lung Squamous Cell Carcinoma (LUSC), emphasizing the urgent need for precise molecular biomarkers to reliably forecast therapeutic efficacy. This study aims to identify potential biomarkers for immunochemotherapy efficacy by focusing on plasma extracellular vesicle (EV)-derived long RNAs (exLRs).MethodsWe enrolled 78 advanced LUSC patients undergoing first-line immunochemotherapy. Plasma samples were collected, and exLR sequencing was conducted to establish baseline profiles. A retrospective analysis was performed on 42 patients to identify differentially expressed exLRs. Further validation of the top differentially expressed exLRs was conducted using quantitative reverse transcription PCR (qRT-PCR). Univariate Cox analysis was applied to determine the prognostic significance of these exLRs. Based on these findings, we developed a predictive signature (p-Signature).ResultsIn the retrospective analysis of 42 patients, we identified 460 differentially expressed exLRs, with pathways related to leukocyte migration notably enriched among non-responders. Univariate Cox analysis revealed 45 exLRs with prognostic significance. The top 6 protein-coding exLRs were validated using qRT-PCR, identifying CXCL8, SSH3, and SDHAF1 as differentially expressed between responders and non-responders. The p-Signature, comprising these three exLRs, demonstrated high accuracy in distinguishing responders from non-responders, with an Area Under the Curve (AUC) of 0.904 in the retrospective cohort and 0.812 in the prospective cohort.DiscussionThis study highlighted the potential of plasma exLR profiles in predicting LUSC treatment efficacy. Intriguingly, lower p-Signature scores were associated with increased abundance of activated CD4+ and CD8+ T cells, indicating a more robust immune environment. These findings suggest that the p-Signature could serve as a valuable tool in guiding personalized and effective therapeutic strategies for LUSC.

44 Liquid biopsy epigenomic profiling for the detection of sarcomatoid renal cell carcinoma

Abstract Background Sarcomatoid differentiation (SD) in renal cell carcinoma (RCC) is associated with poor survival and heightened response to immune checkpoint blockade. Detection of SD can be challenging due to spatial heterogeneity and sampling error. Herein, we introduce a novel tissue–informed epigenomic approach to noninvasively identify sarcomatoid differentiation in patients with RCC from cell-free DNA (cfDNA) using 1mL of plasma. Methods Chromatin immunoprecipitation and sequencing (ChIP-seq) for H3K27ac – a histone modification associated with active regulatory elements (REs) – was performed on pathologically reviewed clear cell RCC frozen tissue samples with and without SD (sarcomatoid-RCC and epithelioid-RCC, resp.) collected at the Dana-Farber Cancer Institute. Differentially marked REs between sarcomatoid and epithelioid subtypes were identified using DESeq2 (false discovery rate of q &amp;lt; 0.01). After establishing tissue signatures, ChIP-seq was then performed on cell-free chromatin (cfChIP-seq) in plasma from patients with sarc-RCC and epi-RCC. A Sarcomatoid Score was derived for each sample by aggregating the plasma H3K27ac signal at tissue-derived sarcomatoid-specific REs (sarc-REs), while normalizing to signal at epithelioid-specific REs (epi-REs). Scores were compared between the two groups using a Wilcoxon rank-sum test. A classifier was built to distinguish sarc-RCC from epi-RCC based on the Sarcomatoid Score and its performance was evaluated using the area under the receiver operating characteristic (AUROC) curve. Results We identified 25,919 differentially marked REs between 8 sarc-RCC and 8 epi-RCC tissue samples at a false discovery rate of q &amp;lt; 0.01. We selected 12,868 REs that are enriched in sarcomatoid vs. epithelioid. We generated cfChIP-seq profiles from plasma of 29 patients, 17 with sarc-RCC and 12 with epi-RCC. The Sarcomatoid Scores were significantly higher in sarc-RCC vs. epi-RCC plasma samples (p=1.6×10-4; Figure 1A). These scores achieved an AUROC curve of 0.9 for classifying patients with sarc-RCC from patients with epi-RCC (Figure 1B). Conclusions We present a proof-of-concept study in 1 cc of plasma for the detection of sarcomatoid differentiation in RCC based on the assessment of histone modification signals in cfDNA. This approach could help overcome the challenges of spatial heterogeneity and sampling error from tissue that make identification of sarc-RCC difficult. More generally, it establishes a paradigm for identifying histologic subtypes of cancer based on their epigenomic correlates from cfDNA, with possible therapeutic implications in real-time. Figure 1. (A) Sarcomatoid Score in plasma from patients with RCC at sarcomatoid-specific REs, comparing sarcomatoid (orange) and epithelioid RCC (blue). (B) ROC curves for distinguishing sarcomatoid from epithelioid RCC plasma samples using H3K27ac cell-free ChIP-seq signal at sarcomatoid-specific REs. ‘AUC’ indicates area under the ROC curve.

Penicillin concentrations in bone and subcutaneous tissue: A porcine microdialysis study comparing oral and intravenous treatment.

Penicillin is available in both an oral (penicillin V) and intravenous formulation (penicillin G), theoretically allowing for a safe transition between the two. However, the use of oral penicillin remains a topic of debate due to low and variable bioavailability. This study aimed to assess the time for which the free penicillin concentration exceeded targeted minimum inhibitory concentrations for Staphylococcus aureus and Streptococcus species (0.125, 0.25, and 0.5 mg/L) in cancellous bone and subcutaneous tissue after intravenous penicillin and oral penicillin administration. 12 female pigs (68-75 kg) were assigned, according to local standard clinical regimes, to either intravenous penicillin (1.2 g) or oral penicillin (0.8 g) treatment every 6 h over an 18 h period. Microdialysis catheters were placed for sampling in tibial cancellous bone and adjacent subcutaneous tissue. Data was dynamic/continually collected in the first dosing interval (0-6 h), simulating a prophylactic situation, and the third dosing interval (12-18 h), simulating a therapeutic setting. Plasma samples were collected for reference. For all investigated targets, intravenous treatment resulted in a longer mean time above relevant minimum inhibitory concentrations in cancellous bone during the first dosing interval, and in both cancellous bone and subcutaneous tissue during the third dosing interval compared to oral treatment. With clinically relevant dosing, intravenous penicillin provides superior exposure compared to oral penicillin in both a prophylactic and therapeutic setting.

Absorption, Metabolism, and Excretion of [14C]-Labeled Anaprazole: A New Proton Pump Inhibitor, After a Single Oral Administration in Healthy Chinese Male Subjects.

Anaprazole is a proton pump inhibitor. This study aims to elucidate absorption, metabolism, and excretion pathways of anaprazole sodium in the human body. A total of 4 healthy Chinese male subjects were administered a single oral dose of 20mg/100µCi of [14C]-anaprazole sodium enteric-coated capsules. The whole blood, plasma, and excreta were analyzed for a total radioactivity (TRA) and metabolite profile. The cumulative radioactivity excretion rate was 93.2%, with 53.3% and 39.9% of the radioactive dose excreted in urine and feces, respectively, and 91.6% of dose recovered within 96hours after dosing. The parent drug, anaprazole, showed good absorption and was extensively metabolized majorly to thioether M8-1 via nonenzymatic metabolism. Overall, 35 metabolites were identified in plasma, urine, and fecal samples. Anaprazole was the most abundant component in plasma followed by the thioether M8-1, accounting for 28.3% and 16.6%, respectively, of the plasma TRA. Thioether carboxylic acid XZP-3409 (26.3% of urine TRA) and XZP-3409 oxidation and dehydrogenation product M417a (15.1% of fecal TRA) were the major metabolites present in urine and feces, respectively. Anaprazole was undetectable in urine, while fecal samples showed traces (0.07% dose). Blood/plasma ratios of the radioactivity (approximately 0.60) remained consistent over time. Anaprazole showed good absorption and was extensively metabolized majorly to thioether M8-1 via nonenzymatic metabolism, and cytochrome P450 3A4 also contributed to its metabolism in healthy individuals.

Pharmacokinetics of carprofen in lactating dogs after intravenous treatment

Analgesia is an important component of veterinary care with nonsteroidal antiinflammatory drugs, particularly carprofen (Rimadyl®) commonly used for dogs. Pharmacokinetic studies of carprofen in lactating dogs have not been examined fully, questioning the use and safety of carprofen. The extent to which lactation changes the pharmacokinetics is unknown, nor is the extent of transfer into milk of lactating dogs and their neonates or pups. We hypothesized that after a single dose of intravenous (4.4 mg/kg) to dogs, concentrations in milk are low and not high enough to produce harmful exposure in pups. We tested our hypothesis in healthy, adult lactating dogs (n = 4) using pharmacokinetic methods and nonchiral and chiral specific assays measuring total, R-, and S-enantiomer carprofen concentrations with high performance liquid chromatography in maternal plasma, milk, and neonatal plasma samples. The CMAX, elimination half-life, and clearance for maternal plasma were 9.09 and 7.3 µg/ml (R-, S+), 6.82 and 6.22 hours (R-, S+), and a higher clearance rate of 95.81 ml/hr/kg (R-) and 73.87 ml/hr/kg (S+) than reported. None of the neonatal plasma concentrations at any time point were more than 10% of the total maternal plasma concentrations and milk:plasma ratio was &lt; 1. This study confirmed the approved dose is appropriate for perioperative and chronic pain in lactating dogs and is safe with reduced exposure to nursing pups. This study suggested that the pharmacokinetics of carprofen in lactating dogs may be different from their nonlactating counterparts. However, further pharmacokinetic studies with more dogs are needed to confirm these findings.

Clinical Effects and Pharmacokinetic Profile of Intramuscular Dexmedetomidine (10 μg/kg) in Cats

This study investigated the pharmacokinetic profile of and pharmacodynamic response to dexmedetomidine administered intramuscularly (IM) at a dose of 10 μg/kg in healthy cats. Nine adult cats were evaluated before and after administration of the drug, with serial collections of plasma samples. Dexmedetomidine induced deep sedation, with a rapid onset of action and a duration of one hour, reaching a peak between 20 and 30 min after administration. The half-life (T½) was 70.2 ± 48 min, with a maximum concentration (Cmax) of 2.2 ± 1.9 ng/mL and time to reach maximum concentration (Tmax) of 26.4 ± 19.8 min. The area under the curve (AUC) was 167.1 ± 149.1 ng/mL*min, with a volume of distribution (Vd) of 2159.9 ± 3237.8 mL/kg and clearance (Cl) of 25.8 ± 33.0 mL/min/kg. There was a reduction in heart rate (HR) and respiratory rate (RR) in relation to the baseline, with a slight decrease in systolic (SBP), diastolic (DBP), and mean (MAP) blood pressure in the first hour. Blood glucose increased after 60 min. Dexmedetomidine proved to be effective and safe, with rapid absorption, metabolization, and elimination, promoting good sedation with minimal adverse effects after IM administration in healthy cats.

Genetic Factors Contributing to Interindividual Variability of α-Tocopherol Levels in Subcutaneous Adipose Tissue among Healthy Adult Males

In humans, α-tocopherol (α-TOC) is mainly stored in adipose tissue, where it participates in preventing damages induced by inflammation and reactive oxygen species. Factors, including genetic ones, that explain adipose tissue α-TOC concentration remain poorly understood. This study, therefore, aimed to characterize the interindividual variability of adipose tissue α-TOC concentration in healthy individuals and to identify single nucleotide polymorphisms (SNPs) associated with it. The study used a randomized cross-over design with 42 healthy adult males. α-TOC concentration was measured in fasting plasma and periumbilical adipose tissue samples, both at fast and 8 h after consumption of three standard meals. Partial least squares (PLS) regression was performed to identify SNPs associated with the interindividual variability of adipose tissue α-TOC concentration. Adipose tissue α-TOC concentration was not associated with fasting plasma concentration (Pearson’s r = 0.24, 95% CI: [−0.08, 0.51]). There was a high interindividual variability of adipose tissue α-TOC concentration (CV = 61%). A PLS regression model comprising 10 SNPs in five genes (PPARG, ABCA1, BUD13, CD36, and MGLL) explained 60% (adjusted R2) of the variability of this concentration. The interindividual variability of adipose tissue α-TOC concentration in humans is due, at least partly, to SNPs in genes involved in α-TOC and triglyceride metabolism.

Mechanisms of resistance and correlation between pre-treatment co-alterations and p-prognosis to osimertinib in chemo-naïve advanced non-small cell lung cancer

BackgroundSeveral patients treated with osimertinib experience progressive disease. The aim was to clarify the mechanisms underlying resistance to osimertinib. MethodsELUCIDATOR: A multi-centre, prospective, observational study involved chemotherapy-naive patients with advanced non-small cell lung cancer receiving osimertinib. Mutations in cancer-associated genes, detected via ultrasensitive next-generation sequencing of circulating tumour deoxyribonucleic acid samples, were collected at baseline and after progressive disease detection. These paired plasma samples were compared. ResultsOf 188 patients enrolled (May 2019-January 2021), 178 (119 females [67 %]) median age 74 years, were included. Patients, n = 95 (53 %) had epidermal growth factor receptor exon 19 deletion mutations. Among 115 patients with progressive disease, circulating tumour deoxyribonucleic acid levels of 85 patients were analysed. MET amplification (n = 4), TP53 mutations (n = 4), PIK3CA mutations (n = 3), BRINP3 mutation (n = 2), BRAF mutation (n = 2), APC mutation (n = 1), RET mutation (n = 1) and epidermal growth factor receptor (EGFR) resistance mutation, and C797S (n = 1) were detected. Patients with baseline TP53 mutations, with MET or EGFR amplification had shorter progression-free (PFS) and overall survival. Patients with PIK3CA mutations tended to shorter PFS. ConclusionMET amplification and PIK3CA mutation mechanisms underly resistance to osimertinib in patients. Patients with coexisting mutations or amplifications at baseline had shorter PFS and overall survival.