The expression of Catsup in the hindgut is essential for zinc homeostasis in Drosophila melanogaster.

Zinc excretion is crucial for zinc homeostasis. However, the mechanism of zinc excretion has not been well characterized. Zinc homeostasis in Drosophila seems well conserved to mammals. In this study, we screened all members of the zinc transporters ZnT (SLC30) and Zip (SLC39) for their potential roles in Drosophila hindgut, an insect organ that belongs to the excretory system. The results indicated that Catecholamines up (Catsup, CG10449), a ZIP member localized to the Golgi, is responsible for zinc homeostasis in the hindgut of Drosophila hindgut-specific knockdown of Catsup leads to a developmental arrest in the larval stage, which could be rescued well by human ZIP7. Further study suggested that Catsup RNAi in the hindgut reduced zinc levels in the excretory system (containing the Malpighian tubule and hindgut) but exhibited systemic zinc overload. Besides, more calculi were observed in the Malpighian tubules of Catsup RNAi flies. The developmental arrest and calculi in the Malpighian tubules of hindgut-specific Catsup RNAi flies could be rescued by dietary zinc restriction but hypersensitivity to zinc. These results will help us understand the fundamental process of zinc excretion in higher eukaryotes.

Gene expression profiling of peripheral blood in patients with steroid-induced osteonecrosis of the femoral head.

Aim: Steroid-induced osteonecrosis of the femoral head (SONFH) is a severe complication following glucocorticoid therapy. This study aimed to identify the differential mRNA expression and investigate the molecular mechanisms of SONFH. Materials & methods: RNA sequencing was performed in eight SONFH patients, five non-SONFH patients and five healthy individuals. Results: A total of 1555, 3997 and 5276 differentially expressed mRNAs existed between the following combinations: SONFH versus non-SONFH, SONFH versus healthy subjects and non-SONFH versus healthy subjects. Increased ISM1 expression might contribute to a high risk of SONFH through antiangiogenesis. Decreased FOLR3 expression might affect the metabolism of homocysteine, leading to avascular necrosis of the femoral head. KCNJ2, which plays a pivotal role in regulating bone development, was also deregulated. Conclusion: ISM1, FOLR3 and KCNJ2 might be related to the occurrence of SONFH.

A Prediction Model for Tacrolimus Daily Dose in Kidney Transplant Recipients With Machine Learning and Deep Learning Techniques.

Tacrolimus is a major immunosuppressor against post-transplant rejection in kidney transplant recipients. However, the narrow therapeutic index of tacrolimus and considerable variability among individuals are challenges for therapeutic outcomes. The aim of this study was to compare different machine learning and deep learning algorithms and establish individualized dose prediction models by using the best performing algorithm. Therefore, among the 10 commonly used algorithms we compared, the TabNet algorithm outperformed other algorithms with the highest R2 (0.824), the lowest prediction error [mean absolute error (MAE) 0.468, mean square error (MSE) 0.558, and root mean square error (RMSE) 0.745], and good performance of overestimated (5.29%) or underestimated dose percentage (8.52%). In the final prediction model, the last tacrolimus daily dose, the last tacrolimus therapeutic drug monitoring value, time after transplantation, hematocrit, serum creatinine, aspartate aminotransferase, weight, CYP3A5, body mass index, and uric acid were the most influential variables on tacrolimus daily dose. Our study provides a reference for the application of deep learning technique in tacrolimus dose estimation, and the TabNet model with desirable predictive performance is expected to be expanded and applied in future clinical practice.

Population Pharmacokinetics of Mycophenolic Acid in Renal Transplant Patients: A Comparison of the Early and Stable Posttransplant Stages.

Mycophenolic acid (MPA) is an antimetabolic immunosuppressive drug widely used in solid organ transplantation and autoimmune diseases. Pharmacokinetics (PK) of MPA demonstrates high inter- and intra-variability. The aim of this study was to compare the population PK properties of MPA in adult renal transplant patients in the early and stable post-transplant stages and to simulate an optimal dosing regimen for patients at different stages. A total of 51 patients in the early post-transplant period (1 week after surgery) and 48 patients in the stable state (5.5-10 years after surgery) were included in the study. In the two-compartment population PK model, CL/F (23.36 L/h vs. 10.25 L/h) and V/F (78.07 vs. 16.24 L) were significantly different between the two stages. The dose-adjusted area under the concentration time curve (AUCss,12h/dose) for patients in the early stage were significantly lower than those for patients in the stable state (40.83 ± 22.26 mg h/L vs. 77.86 ± 21.34 mg h/L; p < 0.001). According to Monte Carlo simulations, patients with 1.0-1.5 g of mycophenolate mofetil twice daily in the early phase and 0.50-0.75 g twice daily in the stable phase had a high probability of achieving an AUCss,12h of 30-60 mg h/L. In addition, limited sampling strategies showed that two 4-point models (C0-C1-C2-C4 and C1-C2-C3-C6) performed well in predicting MPA exposure by both Bayesian estimate and regression equation and could be applied in clinical practice to assist therapeutic drug monitoring of MPA.

The Heterogeneity of Immune Cell Infiltration Landscape and Its Immunotherapeutic Implications in Hepatocellular Carcinoma.

Immunotherapy, closely associated with immune infiltration and tumor mutation burden (TMB), is emerging as a promising strategy for treating tumors, but its low response rate in hepatocellular carcinoma (HCC) remains a major challenge. Herein, we applied two algorithms to uncover the immune infiltration landscape of the immune microenvironment in 491 HCC patients. Three immune infiltration patterns were defined using the CIBERSORT method, and the immune cell infiltration (ICI) scores were established using principal component analysis. In the high ICI score group, the activation of the Wnt/ß-catenin pathway was significantly enriched and expressions of immune checkpoint genes increased, which showed a pessimistic outcome. The low ICI score group was characterized by increased TMB and enrichment of metabolism-related pathways. Further analysis found that the ICI score exhibited a significant difference in age ≥65/age <65, grade I/grade II-IV, and response to immunotherapy. Moreover, the CTNNB1 mutation status was found to be closely associated with prognosis and immunotherapeutic efficiency, significantly affecting the ICI score and TMB, which might be regarded as a potential marker for the treatment of HCC. The evaluation of immune infiltration patterns can improve the understanding of the tumor immune microenvironment and provide new directions for the study of individualized immunotherapy strategies for HCC.

UPLC-MS/MS Method for Simultaneous Determination of 14 Antimicrobials in Human Plasma and Cerebrospinal Fluid: Application to Therapeutic Drug Monitoring.

Pharmacokinetics/pharmacodynamics is the foundation for guiding the rational application of antibiotics in clinical practice, so it is necessary to establish quantitative methods for accurate drug concentration determination. This study aimed to develop a rapid and simple ultrahigh-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for simultaneous quantification of 14 antibiotics (amikacin, etimicin, ceftazidime, cefepime, cefoperazone, ceftriaxone, daptomycin, latamoxef, linezolid, meropenem, biapenem, ampicillin, norvancomycin, and vancomycin) in human plasma and cerebrospinal fluid. Antibiotics were chromatographically separated on a Waters ACQUITY UPLC BEH C18 column (2.1 mm × 50 mm, 1.7 µm) via gradient elution within 3 minutes and were monitored using positive ion fitted with multiple reaction monitoring. The lower limit of quantification was 0.05-2.0 µg·mL-1. The method was verified according to the FDA bioanalysis method validation guidelines, which showed excellent accuracy (from 86.75% to 110.85%) and precision (from 0.46% to 10.97%). At last, this method was successfully applied to therapeutic drug monitoring in 113 patients under antibiotics treatment.

Impact of CYP2C19 Phenotype and Drug-Drug Interactions on Voriconazole Concentration in Pediatric Patients.

Voriconazole (VRC), a first-line agent for the treatment of invasive fungal infections, is mainly metabolized by human cytochrome P450 (CYP) 2C19. In this study, a retrospective analysis was performed to investigate the key factors that influence the plasma trough concentration (Cmin) of VRC, and an appropriate dosing regimen for pediatric patients was drafted subsequently. Overall, factors such as age, CYP2C19 phenotype, and combination medication with proton pump inhibitors accounted for 23.4% of variability in dose-normalized Cmin values of VRC by a multiple linear regression analysis. Dose-normalized Cmin values in the poor metabolizers (PMs) and intermediate metabolizers (IMs) were significantly higher than those in extensive metabolizers (EMs) (P < 0.001). To achieve therapeutic Cmin for CYP2C19 ultrarapid metabolizers (UMs) or EMs, patients aged no more than 12 and more than 12 years required doses of 6.53 ± 2.08 and 3.95 ± 0.85 mg/kg of body weight twice daily (P = 0.007). For CYP2C19 PMs or IMs, patients aged under 12 and over 12 years required doses of 5.75 ± 1.73 and 4.23 ± 0.76 mg/kg twice daily, respectively (P = 0.019). Furthermore, coadministration of rifamycin sodium or omeprazole exhibited significant effects on VRC Cmin. Taken together, it is necessary to pay attention to the impact of CYP2C19 phenotype and drug-drug interactions to achieve optimal therapy.

Deciphering the Relationship Between the Trough Concentration of Posaconazole and Its Efficacy and Safety in Chinese Patients With Hematological Disorders.

Posaconazole (PCZ) is effective in preventing and salvage treatment invasive fungal infections in patients with hematologic disorders. However, PCZ displays highly variable individual pharmacokinetics affecting its efficacy and safety. To investigate the correlation between PCZ concentration and efficacy and safety, the following key influencing factors were explored. A total of 285 trough plasma concentrations (Cmin) of 81 Chinese patients receiving PCZ oral suspension for prophylaxis or treatment of invasive fungal infections were collected in this study. The relationships between Cmin values and clinical response and hepatotoxicity were investigated as well as the incidence of clinical response under different Cmin values of PCZ with a logistic regression model. The concentration of PCZ showed remarkable differences among patients with haematologic disorders. PCZ Cmin values of 0.76 and 1.0 µg/mL were both associated with an over 80% probability of successful response to prophylaxis and treatment of fungal infections, respectively. No association between Cmin values and hepatotoxicity was noted (P > 0.05). Gender, albumin, and co-administration of proton pump inhibitor (PPI) were identified as independent factors influencing PCZ Cmin by multiple linear regression analysis. Furthermore, patients' C-reactive protein (CRP), albumin, and co-administration of PPI exhibited significant effects on the therapeutic window of patients receiving PCZ for prophylaxis. The plasma concentration is closely associated with therapeutic efficacy of PCZ. It is necessary to adjust the dosing regimens based on PCZ Cmin to obtain an optimal therapeutic response.

Tacrolimus Starting Dose Prediction Based on Genetic Polymorphisms and Clinical Factors in Chinese Renal Transplant Recipients.

Aims: Tacrolimus has extensive pharmacokinetic variability among patients and a narrow therapeutic window. The U.S. Clinical Pharmacogenetics Implementation Consortium recommends a starting dose for tacrolimus of 0.15-0.3 mg/kg/day, which is much higher compared with 0.05-0.15 mg/kg/day used in China. The purpose of this study was to investigate the influence of clinical factors and single nucleotide polymorphisms (SNPs) on tacrolimus concentrations in Chinese renal transplant recipients. Methods: This study enrolled 406 tacrolimus-treated patients. After renal transplantation, the first tacrolimus trough concentration and corresponding clinical information were collected from all patients. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry was used to genotype 15 SNPs. The relationship between the genetic and clinical factors and dose-adjusted tacrolimus trough concentration was examined. The tacrolimus starting dose was predicted using a classification and regression tree analysis. Results: Examination of the 15 SNPs and several clinical factors identified the CYP3A5 genotype (p = 5.6 × 10-11) and hemoglobin (p = 8.4 × 10-10) as the most significant determinants of tacrolimus C0/D. Accordingly, a concise tacrolimus recommendation dosage model, a classification scheme, and a regression tree were developed. Conclusion: A new classification and regression tree model was developed for establishing the starting dose of tacrolimus based on the CYP3A5 genotype and hemoglobin values. This result may help clinicians prescribe an appropriate initial tacrolimus dose. ClinicalTrials.gov ID: 2020-KY-147.

NUDT15 and TPMT Genetic Polymorphisms Are Related to Azathioprine Intolerance in Chinese Patients with Rheumatic Diseases.

Aims: Azathioprine (AZA) is commonly used to treat autoimmune diseases, but its applications have been limited due to significant adverse effects, particularly leukopenia. The aim of this study was to investigate the associations of NUDT15, TPMT, and inosine triphosphatase (ITPA) polymorphisms with AZA-induced toxicity. Materials and Methods: A total of 86 Chinese patients with autoimmune diseases were recruited, and the NUDT15*2-*6, TPMT*3C, and ITPA rs7270101 genotypes of these patients were characterized by Sanger sequencing. Sociodemographic data and clinical records over a period of 6 months were also collected. Results: The TPMT*3C and NUDT15*3 genotypes were significantly associated with AZA-induced leukopenia (p = 0.007 and 4.475 × 10-6, respectively). The p-value for the correlation between ITPA rs7270101 and leukopenia was 0.059. In addition, NUDT15*3 was significantly associated with gastrointestinal effects, erythropenia, hypochromia, and thrombocytopenia [p = 0.002, 1.109 × 10-5, 1.653 × 10-7, and 9.110 × 10-6, respectively; allelic odds ratio (95% confidence interval): 5.714 (1.56-20.95), 9.333 (2.96-29.47), 13.18 (4.15-41.87), and 20.13 (3.40-119.18), respectively]. The TPMT*3C genotypes were also significantly associated with gastrointestinal discomfort [p = 0.028, 12.08 (0.71-204.49)], alopecia [p = 2.864 × 10-4, 33 (1.80-606.47)], and hypochromia [p = 0.045, 10.33 (0.61-173.66)]. Conclusion: This study demonstrated that NUDT15*3 and TPMT*3C are both highly predictive genetic markers for AZA-induced toxicity in Chinese populations with rheumatic diseases.

Research advances in the detection of miRNA.

MicroRNAs (miRNAs) are a family of endogenous, small (approximately 22 nucleotides in length), noncoding, functional RNAs. With the development of molecular biology, the research of miRNA biological function has attracted significant interest, as abnormal miRNA expression is identified to contribute to serious human diseases such as cancers. Traditional methods for miRNA detection do not meet current demands. In particular, nanomaterial-based methods, nucleic acid amplification-based methods such as rolling circle amplification (RCA), loop-mediated isothermal amplification (LAMP), strand-displacement amplification (SDA) and some enzyme-free amplifications have been employed widely for the highly sensitive detection of miRNA. MiRNA functional research and clinical diagnostics have been accelerated by these new techniques. Herein, we summarize and discuss the recent progress in the development of miRNA detection methods and new applications. This review will provide guidelines for the development of follow-up miRNA detection methods with high sensitivity and specificity, and applicability to disease diagnosis and therapy.

A novel human lncRNA SANT1 cis-regulates the expression of SLC47A2 by altering SFPQ/E2F1/HDAC1 binding to the promoter region in renal cell carcinoma.

SLC47A2 encodes MATE 2-K in the kidney, which mediates the secretion of certain endogenous and exogenous compounds. SLC47A2 was dramatically repressed in patients with renal cell carcinoma (RCC), and a lower level of SLC47A2 might act as a negative prognostic marker, although the mechanism is not well understood. In this study, we aimed to investigate the mechanism via which SLC47A2 is downregulated in RCC. Based on the annotation information of the SLC47A2 locus available in the UCSC genome browser database, we identified a novel lncRNA, which is transcribed from the SLC47A2 locus and named it SANT1. Overexpression and knock-down assays were performed to investigate the effects of SANT1 on cis-regulation of SLC47A2. We verified the direct binding between SANT1 and SFPQ/E2F1/HDAC1 using the cross-linking and immunoprecipitation (CLIP) assay. Chromatin immunoprecipitation was performed to confirm the molecular mechanism via which SANT1 activates the transcription of the SLC47A2 coding region. We observed that SANT1 can cis-regulate its own genetic locus. In tumour-adjacent tissues, the SLC47A2 locus highly expresses SANT1, which can remove the regulatory SFPQ/E2F1/HDAC1 suppressor complex from the promoter region, thereby significantly increasing the levels of the H3K27ac modification and RNAPII binding. Owing to a low SANT1 level, the binding of this inhibitory complex in the promoter region is upregulated in RCC, which results in silencing of the SLC47A2 coding region. In conclusion, we identified a novel lncRNA and elucidated the mechanism via which it regulates SLC47A2 expression in RCC.

A novel helper qPCR system for platinum detection via Pt-DNA coordination.

A novel real-time polymerase chain reaction (qPCR) platform for the simple and robust detection of platinum is described for the first time. Compared with conventional qPCR, a helper template, which is related to the active template for performing qPCR, was introduced in our helper qPCR system. Several guanine (G) bases were introduced in the helper template to obtain a platinum-responsive on/off switch based on G-Pt-G coordination. Because of the helper template, a slight change in platinum concentration would significantly change the signal in the qPCR. This novel helper qPCR technique easily detects platinum with high sensitivity (1 ng/mL) and selectivity over other metal ions. Therefore, it will be a promising technique for the detection of platinum in biomedical and environmental samples.

Platinum(II)-Oligonucleotide Coordination Based Aptasensor for Simple and Selective Detection of Platinum Compounds.

Wide use of platinum-based chemotherapeutic regimens for the treatment for carcinoma calls for a simple and selective detection of platinum compound in biological samples. On the basis of the platinum(II)-base pair coordination, a novel type of aptameric platform for platinum detection has been introduced. This chemiluminescence (CL) aptasensor consists of a designed streptavidin (SA) aptamer sequence in which several base pairs were replaced by G-G mismatches. Only in the presence of platinum, coordination occurs between the platinum and G-G base pairs as opposed to the hydrogen-bonded G-C base pairs, which leads to SA aptamer sequence activation, resulting in their binding to SA coated magnetic beads. These Pt-DNA coordination events were monitored by a simple and direct luminol-peroxide CL reaction through horseradish peroxidase (HRP) catalysis with a strong chemiluminescence emission. The validated ranges of quantification were 0.12-240 µM with a limit of detection of 60 nM and selectivity over other metal ions. This assay was also successfully used in urine sample determination. It will be a promising candidate for the detection of platinum in biomedical and environmental samples.