Establishment and validation of a LC-MS/MS method for the determination of anlotinib in human plasma: Application to therapeutic drug monitoring.

Anlotinib is a novel small molecule multitarget tyrosine kinase inhibitor for the treatment of several cancers. We developed and validated a highly sensitive, rapid and stable high-performance liquid chromatography-mass spectrometrymethod for the determination of anlotinib in human plasma with anlotinib-d5 as a stable isotopically labeled internal standard (SIL-IS). To explore the feasibility of therapeutic drug monitoring in the treatment of tumors with anlotinib, human plasma samples were prepared by protein precipitation. The mobile phases comprised of (A) 5.0 mm NH4 AC aqueous solution containing 0.1% formic acid and (B) 100% methanol containing 0.1% formic acid. A gradient mobile phase system was adopted for chromatographic separation using a BEH C18 (2.1 × 50 mm, 1.7 µm) column. A positive ion pattern was chosen for quantification under multiple reaction monitoring mode. The ion pairs were detected at m/z 408.2 â†’ 339.1 and m/z 413.4 â†’ 344.3 for anlotinib and anlotinib-d5 (SIL-IS), respectively. The total run time was 5.0 min. The calibration curve was found to be linear within a plasma concentration range of 2-400 ng·ml-1 . The precision and accuracy, matrix effect, extraction recovery and stability were all validated and met the requirements of international guidelines. The proposed methods were successfully applied to support therapeutic drug monitoring in breast and thyroid cancer patients receiving anlotinib for therapy. Clinical data showed that in the 12 mg dose group, the mean plasma concentrations of anlotinib in breast cancer patients and thyroid cancer patients were 87.1 and 118.8 ng·ml-1 , respectively. The data demonstrate that the peak concentration of anlotinib may be related to the different tumor types in patients.

The Role of the Mesencephalic Astrocyte-Derived Neurotrophic Factor in Patients in Intensive Care Units Receiving Voriconazole Therapy.

Recent publications regarding the role of mesencephalic astrocyte-derived neurotrophic factor (MANF) in various metabolic and degenerative disorders suggest that MANF is both a marker of disease and a possible therapeutic agent. We investigate the role of plasma MANF levels in patients in intensive care units (ICUs) receiving voriconazole (VCZ) therapy while also comparing MANF levels in healthy individuals. A single-center prospective study was conducted. The plasma MANF level in patients in ICU was found to have high interindividual variability and was significantly higher than that in healthy controls (P < .01). Compared with patients using VCZ only, patients using both VCZ and amikacin had 3-fold lower MANF concentrations (P < .05). The MANF concentrations also decreased when alkaline phosphatase (ALP) and serum creatinine levels were above the upper limits of the normal range (P < .05) and the estimated glomerular filtration rate (eGFR) was below the lower limit of the normal range (P < .01). Receiver operating characteristic curve analysis indicated that low MANF levels were associated with high ALP levels, high creatinine levels, and low eGFR. The cut-off value of MANF for ALP levels higher than 126 U/L was 0.35 ng/mL (area under curve, AUC = 0.62, 95%CI = 0.50-0.74, P = .044); for serum creatinine levels higher than 104 µmol/L, the cut-off value was 0.41 ng/mL (AUC = 0.74, 95%CI = 0.62-0.87, P = .001); and for eGFR below 80 mL/min, the cut-off value was 0.75 ng/mL (AUC = 0.70, 95%CI = 0.59-0.81, P = .002). Monitoring plasma MANF levels may be of value for clinical decision-making regarding the choice of antibiotics and the prediction of impaired liver function and renal function in patients admitted to an ICU.

Prediction of plasma trough concentration of voriconazole in adult patients using machine learning.

OBJECTIVE: Plasma trough concentration of voriconazole (VCZ) was associated with its toxicity and efficacy. However, the nonlinear pharmacokinetic characteristics of VCZ make it difficult to determine the relationship between clinical characteristics and its concentration. We intended to present a machine learning (ML)-based method to predict toxic plasma trough concentration of VCZ (>5 µg/mL). METHODS: A single center retrospective study was conducted. Three ML algorithms were used to estimate the concentration in adult patients, including random forest (RF), gradient boosting (GB), and extreme gradient boosting (XGBoost). The importance of variables was recognized by the SHapley Additive exPlanations (SHAP) method. In addition, an external validation set was used to validate the robustness of models. RESULTS: A total of 1318 VCZ plasma concentration were included, with 33 variables enrolled in the model. Nine classification models were developed using the RF, GB, and XGBoost algorithms. Most models performed well for both the training set and test set, with an average balanced accuracy (BA) of 0.704 and an average accuracy (ACC) of 0.788. In addition, the average Matthews correlation coefficient value reached 0.484, which indicated the predicted values are meaningful. Based on the average BA and ACC values, the predictive ability of the models can be ranked from best to worst as follows: younger adult models > mixed models > elderly models, and XGBoost models > GBT models > RF models. The SHAP results showed that the top five influencing factors in younger adult patients (<60 years) were albumin, total bile acid (TBA), platelets count, age, and inflammation, while the top five influencing factors in elderly patients were albumin, TBA, aspartate aminotransferase, creatinine, and alanine aminotransferase. Furthermore, the prediction of external validation set for VCZ concentrations verified the high reliability of the models, for the ACC value of 0.822 by the best model. CONCLUSIONS: The ML models can be reliable tools for predicting toxic concentration exposure of VCZ. The SHAP results may provide useful guidelines for dosage adjustment of VCZ.

Simultaneous quantitative analysis and in vitro anti-arthritic effects of five polyphenols from Terminalia chebula.

Background: The fruit of Terminalia chebula has been widely used for a thousand years for treating diarrhea, ulcers, and arthritic diseases in Asian countries. However, the active components of this Traditional Chinese medicine and their mechanisms remain unclear, necessitating further investigation. Objectives: To perform simultaneous quantitative analysis of five polyphenols in T. chebula and evaluate their anti-arthritic effects including antioxidant and anti-inflammatory activity in vitro. Materials and methods: Water, 50% water-ethanol, and pure ethanol were used as extract solvents. Quantitative analysis of gallic acid, corilagin, chebulanin, chebulagic acid, and ellagic acid in the three extracts was performed using high-performance liquid chromatography (HPLC). Antioxidant activity was assessed by the 2,2-diphenylpicrylhydrazyl (DPPH) radical-scavenging assay, and anti-inflammatory activity was evaluated by detecting interleukin (IL)-6 and IL-8 expression in IL-1ß-stimulated MH7A cells. Results: The 50% water-ethanol solvent was the optimal solvent yielding the highest total polyphenol content, and the concentrations of chebulanin and chebulagic acid were much higher than those of gallic acid, corilagin, and ellagic acid in the extracts. The DPPH radical-scavenging assay showed that gallic acid and ellagic acid were the strongest antioxidative components, while the other three components showed comparable antioxidative activity. As for the anti-inflammatory effect, chebulanin and chebulagic acid significantly inhibited IL-6 and IL-8 expression at all three concentrations; corilagin and ellagic acid significantly inhibited IL-6 and IL-8 expression at high concentration; and gallic acid could not inhibit IL-8 expression and showed weak inhibition of IL-6 expression in IL-1ß-stimulated MH7A cells. Principal component analysis indicated that chebulanin and chebulagic acid were the main components responsible for the anti-arthritic effects of T. chebula. Conclusion: Our findings highlight the potential anti-arthritic role of chebulanin and chebulagic acid from T. chebula.

Inflammation Affects Liver Function and the Metabolism of Voriconazole to Voriconazole-N-Oxide in Adult and Elderly Patients.

Background: The inner association of inflammation with voriconazole (VCZ) metabolism has not been fully investigated. We intend to investigate the effects of inflammation on liver function, VCZ trough concentration (C0), C0/dose ratio and the ratio of VCZ to VCZ-N-oxide concentration (C0/CN) in adult and elderly patients. Methods: A single-center retrospective study was conducted among patients who were treated in our hospital between January 2018 and December 2021. For each eligible patient, demographic details, medical history, laboratory parameters, procalcitonin (PCT), C reactive protein (CRP), and interleukin-6 (IL-6) were collected from the medical chart. VCZ CN, TNF-α, IL-1ß, IL-8, and IL-10 concentrations were detected in blood samples. Results: A total of 356 patients were included in our study, with 195 patients in the adult cohort (<60 years) and 161 patients in the elderly cohort (≥60 years). In adult patients, CRP and IL-8 levels showed moderate association with VCZ C0/CN ratio (CRP: r = 0.512, p < 0.001; IL-8: r = 0.476, p = 0.002). IL-6 level shallowly associated with VCZ C0/CN ratio both in adult and elderly patients (r = 0.355, p = 0.003; r = 0.386, p = 0.001). A significantly higher VCZ C0, C0/dose ratio and C0/CN ratio was observed in adult patients with severe inflammation compared with patients with moderate inflammation and no to mild inflammation, as reflected by PCT levels (p < 0.05). However, there was no significant difference observed among different inflammation degrees in elderly patients. Lower albumin (AL) and higher total bilirubin (TBIL) were observed along with the degree of inflammation in both adult and elderly patients, as reflected by CRP and PCT levels (p < 0.05). Conclusion: Inflammation may affect the metabolism of VCZ to VCZ-N-oxide both in adult and elderly patients, and decreased plasma AL levels and increased TBIL levels under inflammatory conditions may also alter VCZ metabolism.

The expression and significance of efferocytosis and immune checkpoint related molecules in pancancer samples and the correlation of their expression with anticancer drug sensitivity.

Background: The efferocytosis-related molecules have been considered to be correlated with the resistance to cancer chemotherapy. The aim of this study was to investigate the expression and significance of efferocytosis-related molecules in cancers and the correlation of their expression with anticancer drug sensitivity, and provide new potential targets and treatment options for cancers. Methods: We investigated the differential expression of 15 efferocytosis-related molecules (Axl, Tyro3, MerTK, CX3CL1, Tim-4, BAI1, Stab2, Gas6, IDO1, Rac1, MFGE8, ICAM-1, CD47, CD31, and PD-L1) and other 12 common immune checkpoint-related molecules in tumor and normal tissues, the correlation between their expression and various clinicopathological features in 16 types of cancers using publicly available pancancer datasets in The Cancer Genome Atlas. We also analyzed the correlation of the expression of efferocytosis and immune checkpoint related molecules with 126 types of anticancer drugs sensitivity using drug-RNA-seq data. Results: There is a panel of circulating molecules among the 27 molecules. Based on the results of differential expression and correlation with various clinicopathological features of efferocytosis-related molecules in cancers, we identified new potential therapeutic targets for anticancer therapy, such as Axl for kidney renal clear cell carcinoma, Tyro3 for liver hepatocellular carcinoma, and IDO1 for renal papillary cell carcinoma. Except for BAI1, CD31, and MerTK, the enhanced expressions of Axl, Tyro3, Gas6, MFGE8, Stab2, Tim-4, CX3CL1, IDO1, Rac1, and PD-L1 were associated with decreased sensitivity of the cancer cells to many anti-cancer drugs; however, for other common immune checkpoint-related molecules, only enhanced expressions of PD-1, CD28, CTLA4, and HVEM were associated with decreased sensitivity of the cancer cells to a few drugs. Conclusion: The efferocytosis-related molecules were significantly associated with clinical outcomes in many types of cancers and played important roles in resistance to chemotherapy. Combination therapy targeting efferocytosis-related molecules and other immune checkpoint-related molecules is necessary to reduce resistance to chemotherapy.

γ-Core Guided Antibiotic Design Based on Human Enteric Defensin 5.

An increase in the number of infections caused by resistant bacteria worldwide necessitates the development of alternatives to antibiotics. Human defensin (HD) 5 is an innate immune peptide with broad-spectrum antibacterial activity, but its complicated structure makes its preparation difficult. Herein, we truncated the HD5 structure by extracting the highly conserved γ-core motif. A structure-activity study showed that this motif was ineffective in killing bacteria in the absence of specific spatial conformation. Notably, after the introduction of two intramolecular disulfide bonds, its antibacterial activity was markedly improved. Glu and Ser residues were then replaced with Arg to create the derivative RC18, which exhibited stronger potency than HD5, particularly against methicillin-resistant S. aureus (MRSA). Mechanistically, RC18 bound to lipid A and lipoteichoic acid at higher affinities than HD5. Furthermore, RC18 was more efficient than HD5 in penetrating the bacterial membranes. Molecular dynamics simulation revealed that five Arg residues, Arg1, Arg7, Arg9, Arg15, and Arg18, mediated most of the polar interactions of RC18 with the phospholipid head groups during membrane penetration. In vivo experiments indicated that RC18 decreased MRSA colonization and dramatically improved the survival of infected mice, thus demonstrating that RC18 is a promising drug candidate to treat MRSA infections.

The Gut Microbiota of Healthy Aged Chinese Is Similar to That of the Healthy Young.

The microbiota of the aged is variously described as being more or less diverse than that of younger cohorts, but the comparison groups used and the definitions of the aged population differ between experiments. The differences are often described by null hypothesis statistical tests, which are notoriously irreproducible when dealing with large multivariate samples. We collected and examined the gut microbiota of a cross-sectional cohort of more than 1,000 very healthy Chinese individuals who spanned ages from 3 to over 100 years. The analysis of 16S rRNA gene sequencing results used a compositional data analysis paradigm coupled with measures of effect size, where ordination, differential abundance, and correlation can be explored and analyzed in a unified and reproducible framework. Our analysis showed several surprising results compared to other cohorts. First, the overall microbiota composition of the healthy aged group was similar to that of people decades younger. Second, the major differences between groups in the gut microbiota profiles were found before age 20. Third, the gut microbiota differed little between individuals from the ages of 30 to >100. Fourth, the gut microbiota of males appeared to be more variable than that of females. Taken together, the present findings suggest that the microbiota of the healthy aged in this cross-sectional study differ little from that of the healthy young in the same population, although the minor variations that do exist depend upon the comparison cohort. IMPORTANCE We report the large-scale use of compositional data analysis to establish a baseline microbiota composition in an extremely healthy cohort of the Chinese population. This baseline will serve for comparison for future cohorts with chronic or acute disease. In addition to the expected difference in the microbiota of children and adults, we found that the microbiota of the elderly in this population was similar in almost all respects to that of healthy people in the same population who are scores of years younger. We speculate that this similarity is a consequence of an active healthy lifestyle and diet, although cause and effect cannot be ascribed in this (or any other) cross-sectional design. One surprising result was that the gut microbiota of persons in their 20s was distinct from those of other age cohorts, and this result was replicated, suggesting that it is a reproducible finding and distinct from those of other populations.