Oral mitochondrial transplantation using nanomotors to treat ischaemic heart disease.

Mitochondrial transplantation is an important therapeutic strategy for restoring energy supply in patients with ischaemic heart disease (IHD); however, it is limited by the invasiveness of the transplantation method and loss of mitochondrial activity. Here we report successful mitochondrial transplantation by oral administration for IHD therapy. A nitric-oxide-releasing nanomotor is modified on the mitochondria surface to obtain nanomotorized mitochondria with chemotactic targeting ability towards damaged heart tissue due to nanomotor action. The nanomotorized mitochondria are packaged in enteric capsules to protect them from gastric acid erosion. After oral delivery the mitochondria are released in the intestine, where they are quickly absorbed by intestinal cells and secreted into the bloodstream, allowing delivery to the damaged heart tissue. The regulation of disease microenvironment by the nanomotorized mitochondria can not only achieve rapid uptake and high retention of mitochondria by damaged cardiomyocytes but also maintains high activity of the transplanted mitochondria. Furthermore, results from animal models of IHD indicate that the accumulated nanomotorized mitochondria in the damaged heart tissue can regulate cardiac metabolism at the transcriptional level, thus preventing IHD progression. This strategy has the potential to change the therapeutic strategy used to treat IHD.

Carbon monoxide-releasing nanomotors based on endogenous biochemical reactions for tumor therapy.

The lack of selective release ability in the tumor microenvironment and the limited efficacy of monotherapy are important factors that limit the current use of carbon monoxide (CO) donors for tumor therapy. Herein, inspired by endogenous biochemical reactions in vivo, one kind of CO-releasing nanomotor was designed for the multimodal synergistic treatment of tumor. Specifically, glucose oxidase (GOx) and 5-aminolevulinic acid (5-ALA) were co-modified onto metal-organic framework material (MIL-101) to obtain MIL-GOx-ALA nanomotors (M-G-A NMs), which exhibit excellent biocompatibility and degradation ability in tumor microenvironment. Subsequently, the released 5-ALA generates CO in the tumor microenvironment through an endogenous reaction and further acts on mitochondria to release large amounts of reactive oxygen species (ROS), which directly kill tumor cells. Furthermore, the produced ROS and the degradation products of M-G-A NMs can also provide the reaction substrate for the Fenton reaction, thereby enhancing chemodynamic therapy (CDT) and inducing apoptosis of tumor cells. Both in vitro and in vivo experimental data confirm the successful occurrence of the above process, and the combination of CO gas therapy/enhanced CDT can effectively inhibit tumor growth. This CDT-enhancing agent designed based on endogenous biochemical reactions has good prospects for tumor treatment application.

Liver fibrosis: pathological features, clinical treatment and application of therapeutic nanoagents.

Liver fibrosis is a reversible damage-repair response, the pathological features of which mainly include damage to hepatocytes, sinusoid capillarization, hepatic stellate cells activation, excessive accumulation of extracellular matrix and inflammatory response. Although some treatments (including drugs and stem cell therapy) for these pathological features have been shown to be effective, more clinical trials are needed to confirm their effectiveness. In recent years, nanomaterials-based therapies have emerged as an innovative and promising alternative to traditional drugs, being explored for the treatment of liver fibrosis diseases. Natural nanomaterials (including extracellular vesicles) and synthetic nanomaterials (including inorganic nanomaterials and organic nanomaterials) are developed to facilitate drug targeting delivery and combination therapy. In this review, the pathological features of liver fibrosis and the current anti-fibrosis drugs in clinical trials are briefly introduced, followed by a detailed introduction of the therapeutic nanoagents for the precise delivery of anti-fibrosis drugs. Finally, the future development trend in this field is discussed.

Flexible MXene/Nanocellulose Composite Aerogel Film with Cellular Structure for Electromagnetic Interference Shielding and Photothermal Conversion.

With the rapid development of wearable devices and integrated systems, protection against electromagnetic waves is an issue. For solving the problems of poor flexibility and a tendency to corrode traditional electromagnetic interference (EMI) shielding materials, two-dimensional (2D) nanomaterial MXene was employed to manufacture next-generation EMI shielding materials. Vacuum-assisted filtration combined with the liquid nitrogen prefreezing strategy was adopted to prepare flexible MXene/cellulose nanofibers (CNFs) composite aerogel film with unique cellular structure. Here, CNFs were employed as the reinforcement, and such a cellular structure design can effectively improve the shielding effectiveness (SE). In particular, the composite shows an outstanding EMI SE of 54 dB. Furthermore, the MXene/CNFs composite aerogel film exhibited prominent and steady photothermal conversion ability, which could obtain the maximum equilibrium temperature of 89.4 °C under an 808 nm NIR laser. Thus, our flexible composite aerogel film with appealing cellular construction holds great promise for wearable EMI shielding materials and heating applications in a cold and complex practical environment.

Improved analysis ZIC-HILIC-HCD-Orbitrap method for mapping the glycopeptide by mass spectrometry.

Glycosylation is one of the most common post-translational modifications (PTMs). Protein glycosylation analysis is the bottleneck to deeply understand their functions. At present, the LC-MS analysis of glycosylated post-translational modification is mainly focused on the analysis of glycopeptides. However, the factors affecting the identification of glycopeptides were not fully elucidated. In the paper, we have carefully studied the factors, e.g., HILIC materials, search engines, protein amount, gradient duration, extraction solution, etc. According to the results, HILIC materials were the most important factors affecting the glycopeptides identification, and the amphoteric sulfoalkyl betaine stationary phase enriched glycopeptides 6-fold more compared to the amphiphilic ion-bonded fully porous spherical silica stationary phase. We explored the influence of the extraction solutions on glycan identification. Comparing sodium dodecyl sulfate (SDS) and urea (UA), the results showed that N-glycolylneuraminic acid (NeuGc) type of glycan content was found to be increased 1.4-fold in the SDS compared to UA. Besides, we explored the influence of the search engine on glycopeptide identification. Comparing pGlyco3.0 and MSFragger-Glyco, it was observed that pGlyco3.0 outperformed MSFragger-Glyco in identifying glycopeptides. Then, using our optimized method we found that there was a significant difference in the distribution of monosaccharide types in plasma and brain tissue, e.g., the content of NeuAc in brain was 5-fold higher than that in plasma. To importantly, two glycoproteins (Neurexin-2 and SUN domain-containing protein 2) were also found for the first time by our method. In summary, we have comprehensively studied the factors influencing glycopeptide identification than any previous research, and the optimized method could be widely used for identifying the glycoproteins or glycolpeptides biomarkers for disease detection and therapeutic targets.

Improved Environmental Stimulus and Biological Competition Tactics Interactive Artificial Ecological Optimization Algorithm for Clustering.

An interactive artificial ecological optimization algorithm (SIAEO) based on environmental stimulus and a competition mechanism was devised to find the solution to a complex calculation, which can often become bogged down in local optimum because of the sequential execution of consumption and decomposition stages in the artificial ecological optimization algorithm. Firstly, the environmental stimulus defined by population diversity makes the population interactively execute the consumption operator and decomposition operator to abate the inhomogeneity of the algorithm. Secondly, the three different types of predation modes in the consumption stage were regarded as three different tasks, and the task execution mode was determined by the maximum cumulative success rate of each individual task execution. Furthermore, the biological competition operator is recommended to modify the regeneration strategy so that the SIAEO algorithm can provide consideration to the exploitation in the exploration stage, break the equal probability execution mode of the AEO, and promote the competition among operators. Finally, the stochastic mean suppression alternation exploitation problem is introduced in the later exploitation process of the algorithm, which can tremendously heighten the SIAEO algorithm to run away the local optimum. A comparison between SIAEO and other improved algorithms is performed on the CEC2017 and CEC2019 test set.

New insight into the microbiome, resistome, and mobilome on the dental waste water in the context of heavy metal environment.

Object: Hospital sewage have been associated with incorporation of antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs) into microbes, which is considered as a key indicator for the spread of antimicrobial resistance (AMR). The compositions of dental waste water (DWW) contain heavy metals, the evolution of AMR and its effects on the water environment in the context of heavy metal environment have not been seriously investigated. Thus, our major aims were to elucidate the evolution of AMR in DWW. Methods: DWW samples were collected from a major dental department. The presence of microbial communities, ARGs, and MGEs in untreated and treated (by filter membrane and ozone) samples were analyzed using metagenomics and bioinformatic methods. Results: DWW-associated resistomes included 1,208 types of ARGs, belonging to 29 antibiotic types/subtypes. The most abundant types/subtypes were ARGs of multidrug resistance and of antibiotics that were frequently used in the clinical practice. Pseudomonas putida, Pseudomonas aeruginosa, Chryseobacterium indologenes, Sphingomonas laterariae were the main bacteria which hosted these ARGs. Mobilomes in DWW consisted of 93 MGE subtypes which belonged to 8 MGE types. Transposases were the most frequently detected MGEs which formed networks of communications. For example, ISCrsp1 and tnpA.5/4/11 were the main transposases located in the central hubs of a network. These significant associations between ARGs and MGEs revealed the strong potential of ARGs transmission towards development of antimicrobial-resistant (AMR) bacteria. On the other hand, treatment of DWW using membranes and ozone was only effective in removing minor species of bacteria and types of ARGs and MGEs. Conclusion: DWW contained abundant ARGs, and MGEs, which contributed to the occurrence and spread of AMR bacteria. Consequently, DWW would seriously increase environmental health concerns which may be different but have been well-documented from hospital waste waters.

Highly Stereodivergent Synthesis of Chiral C4-Ester-Quaternary Pyrrolidines: A Strategy for the Total Synthesis of Spirotryprostatin A.

In this work, we disclose two sets of highly diastereo- and enantioselective [3 + 2] cycloadditions of iminoesters with various α-substituted acrylates, especially for sterically hindered and weakly activated α-aryl or alkyl-substituted acrylates and alkenal, alkynal, or unstable aliphatic aldehyde-derived iminoesters, catalyzed by the AgHMDS/DTBM-Segphos or Ag2O/CA-AA-Amidphos catalytic system, achieving the stereodivergent synthesis of chiral C4-ester-quaternary exo- or endo-pyrrolidines with high yields and excellent diastereo- and enantioselectivities (up to >99:1 dr and >99% ee). More importantly, the gram-scale synthetic exo-adduct displays significant applications in the aspect of realizing the total synthesis of the spirotryprostatin A alkaloid via nine steps in a 36% overall yield.

Multifunctional MXene Conductive Zwitterionic Hydrogel for Flexible Wearable Sensors and Arrays.

Conductive hydrogels have good prospects in the fields of flexible electronic devices and artificial intelligence due to their biocompatibility, durability, and functional diversity. However, the process of hydrogel polymerization is time-consuming and energy-consuming, and freezing at zero temperature is inevitable, which seriously hinders its applications and working life. Herein, zwitterionic conductive hydrogels with self-adhesive and antifreeze properties were prepared in one minute by introducing two-dimensional (2D) MXene nanosheets into the autocatalytically enhanced system composed of tannic acid-modified cellulose nanofibers and zinc chloride. The system has strong environmental applicability (-60 to 40 °C), good stretchability (ductility ≈ 980%), durable adhesion (even after 30 days of exposure to air), and strong electrical conductivity (20 °C, 30 mS cm-1). By virtue of these advantages, the prepared zwitterionic hydrogels can be developed into flexible strain sensors to monitor large human movements and subtle physiological signals over a wide temperature range and to capture signals from handwriting and voice recognition. In addition, multiple flexible sensors can be assembled into a three-dimensional (3D) array, which can detect the magnitude and spatial distribution of strain or force. These results demonstrate that the prepared zwitterionic hydrogels have promising applications in the fields of medical monitoring and artificial intelligence.

Optimized approach for active peptides identification in Cerebrolysin by nanoLC-MS.

Cerebrolysin (CBL) is a peptide-rich preparation made by hydrolysis and purified extraction of porcine brain. CBL contains various neuroprotective peptides, such as neurotrophic factor, nerve growth factor and ciliary neurotrophic factor, which can be used to treat neurodegenerative diseases. However, the active peptides in CBL had not been studied in depth. In this study, the following was carried out in order to investigate the active peptides in CBL. First, CBL samples were treated using organic reagents (acetonitrile and acetone) to precipitate the proteins and different solid phase extraction methods (MCX mixed-mode cartridges, C18 SPE cartridge columns and HILIC sorbent). Then the samples were analyzed using nanoLC-MS, followed by the identification of peptides using different sequence analysis software (PEAKS, pNovo and novor). Finally, bioinformatics analysis was performed to predict peptides with potential neuroprotective functions in CBL, such as anti-inflammatory and antioxidant peptides. Results showed that the number of peptides obtained by the MCX method coupled with PEAKS was the highest and the method was the most stable. Bioinformatic analysis of the detected peptides showed that two anti-inflammatory peptides (LLNLQPPPR and LSPSLRLP) and an antioxidant peptide (WPFPR) might be neuroprotective peptides in CBL. In addition, this study found that some peptides in CBL were present in myelin basic protein and tubulin beta chain. The results of this study for the detection of active peptides in CBL laid the foundation for the subsequent study of its active ingredients.

Efficacy and safety of PLDR-IMRT for the re-irradiation of recurrent NPC: A prospective, single-arm, multicenter trial.

Salvage treatment of locoregionally recurrent nasopharyngeal carcinoma (NPC) requires weighing the benefits of re-irradiation against increased risks of toxicity. Here, we evaluated the outcomes of patients treated with intensity-modulated-based pulsed low-dose-rate radiotherapy (PLDR-IMRT) to enhance the curative effect of salvage treatment and reduce RT-related SAEs. A prospective clinical trial was conducted from March 2018 to March 2020 at multiple institutions. NPC patients who experienced relapse after radical therapy were re-irradiated with a median dose of 60 Gy (50.4-70 Gy)/30 f (28-35 f) using PLDR-IMRT. Thirty-six NPC patients who underwent PLDR-IMRT for locoregional recurrence were identified. With a median follow-up of 26.2 months, the objective response rate (ORR) of the entire cohort was 91.6%. The estimated mPFS duration was 28 months (95% CI: 24.9-31.1), and the estimated mLRFS duration was 30.4 months (95% CI: 25.2-35.5). The overall survival (OS) rate for all patients was 80.6%, the progression-free survival (PFS) rate was 75% and the cancer-specific survival (CSS) rate was 88.9% at 1 year. The LRFS and DMFS rates were 88.9% and 91.7%, respectively, at 1 year. A combination of systematic therapies could provide survival benefits to patients who experience NPC relapse (p < 0.05), and a Karnofsky performance status (KPS) score of ≥90 was a favorable factor for local control (p < 0.05). The incidence of acute SAEs (grade 3+) from PLDR was 22.2%, and the incidence of chronic SAEs was 19.4% among all patients. PLDR-IMRT combined with systematic therapy can effectively treat patients with locoregionally recurrent nasopharyngeal carcinoma and causes fewer adverse events than the rates expected with IMRT.

Movement disorder caused by FRRS1L deficiency may be associated with morphological and functional disorders in Purkinje cells.

Ferric Chelate Reductase 1 Like (FRRS1L) protein has been identified as an auxiliary regulatory protein for the α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor (AMPAR). FRRS1L is highly expressed in the cerebellum and other brain regions associated with the control of motor function. Loss of FRRS1L has been shown to lead to impaired synaptic transmission via AMPARs and to movement disorders. We found that deletion of the FRRS1L gene causes hyperactivity, reduced muscle strength, impaired coordination, and ataxia in mice. Deletion also impairs Purkinje cell dendritic spine formation and AMPAR expression in the cerebellum and damages the electrophysiological discharge rhythm of Purkinje cells. Cerebrospinal fluid examination and oleic acid (OA)-induced lipid accumulation monitoring in FRRS1L-knockdown SH-SY5Y cells indicated that FRRS1L deficiency could lead to aberrant metabolism of amino acids, glucose, and lipids. In summary, we found that the deletion of FRRS1L leads to impaired motor coordination and cerebellar ataxia in mice, which might be related to the reduced expression of AMPARs, metabolic deviations, and dysplastic functional defects in Purkinje cells.

Effect of thymoquinone on sepsis-induced cardiac damage via anti-inflammatory and anti-apoptotic mechanisms.

OBJECTIVE: Sepsis is a systemic and deleterious host reaction to severe infection. Cardiac dysfunction is an established serious outcome of multiorgan failure associated with this condition. Therefore, it is important to develop drugs targeting sepsis-induced cardiac damage and inflammation. Thymoquinone (TQ) has anti-inflammatory, anti-oxidant, anti-fibrotic, anti-tumor, and anti-apoptotic effects. This study examined the effects of thymoquinone on sepsis-induced cardiac damage. METHODS: Male BALB/c mice were randomly segregated into four groups: control, TQ, cecal ligation and puncture (CLP), and CLP + TQ groups. CLP was performed after gavaging the mice with TQ for 2 weeks. After 48 hours, we estimated the histopathological changes in the cardiac tissue and the serum levels of cardiac troponin-T. We evaluated the expression of factors associated with inflammation, apoptosis, oxidative stress, and the PI3K/AKT pathway. RESULTS: TQ significantly reduced intestinal histological alterations and inhibited the upregulation of interleukin-6, tumor necrosis factor-α, Bax, NOX4, p-PI3K, and p-AKT. TQ also increased Bcl-2, HO-1, and NRF2 expression. CONCLUSION: These results suggest that TQ effectively modulates pro-inflammatory, apoptotic, oxidative stress, and PI3K/AKT pathways, making it indispensable in the treatment of sepsis-induced cardiac damage.

Cell-cycle and apoptosis related and proteomics-based signaling pathways of human hepatoma Huh-7 cells treated by three currently used multi-RTK inhibitors.

Sorafenib, lenvatinib and regorafenib, the multi-RTK inhibitors with potent anti-angiogenesis effects, are currently therapeutic drugs generally recommended for the patients with advanced hepatocellular carcinoma (HCC). To date, however, there have been no published studies on the mechanism underling differential effects of the three drugs on HCC cell proliferation, and the proteomic analysis in HCC cell lines treated by regorafenib or lenvatinib. The present study for the first time performed a direct comparison of the cell cycle arrest and apoptosis induction in the Huh-7 cells caused by sorafenib, regorafenib and lenvatinib at respective IC50 using flow cytometry technique, as well as their pharmacological interventions for influencing whole cell proteomics using tandem mass tag-based peptide-labeling coupled with the nLC-HRMS technique. Sorafenib, regorafenib and lenvatinib at respective IC50 drove the remaining surviving Huh-7 cells into a G0/G1 arrest, but lenvatinib and regorafenib were much more effective than sorafenib. Lenvatinib produced a much stronger induction of Huh-7 cells into early apoptosis than sorafenib and regorafenib, while necrotic cell proportion induced by regorafenib was 2.4 times as large as that by lenvatinib. The proteomic study revealed 419 proteins downregulated commonly by the three drugs at respective IC50. KEGG pathway analysis of the downregulated proteins indicated the ranking of top six signaling pathways including the spliceosome, DNA replication, cell cycle, mRNA surveillance, P53 and nucleotide excision repair involved in 33 proteins, all of which were directly related to their pharmacological effects on cell cycle and cell apoptosis. Notably, lenvatinib and regorafenib downregulated the proteins of PCNA, Cyclin B1, BCL-xL, TSP1, BUD31, SF3A1 and Mad2 much more strongly than sorafenib. Moreover, most of the proteins in the P53 signaling pathway were downregulated with lenvatinib and regorafenib by more than 36% at least. In conclusion, lenvatinib and regorafenib have much stronger potency against Huh-7 cell proliferation than sorafenib because of their more potent effects on cell cycle arrest and apoptosis induction. The underling mechanism may be at least due to the 33 downregulated proteins centralizing the signal pathways of cell cycle, p53 and DNA synthesis based on the present proteomics study.

Salvia chinensia Benth induces autophagy in esophageal cancer cells via AMPK/ULK1 signaling pathway.

Salvia chinensia Benth (Shijianchuan in Chinese, SJC) has been used as a traditional anti-cancer herb. SJC showed good anti-esophageal cancer efficacy based on our clinical application. However, the current research on SJC is minimal, and its anti-cancer effect lacks scientific certification. This study aims to clarify the inhibitory effect of SJC on esophageal cancer and explore its underlying mechanism. Q-Orbitrap high-resolution LC/MS was used to identify the primary chemical constituents in SJC. Cell proliferation and colony formation assays showed that SJC could effectively inhibit the growth of esophageal tumor cells in vitro. To clarify its mechanism of action, proteomic and bioinformatic analyses were carried out by combining tandem mass labeling and two-dimensional liquid chromatography-mass spectrometry (LC-MS). Data are available via ProteomeXchange with identifier PXD035823. The results indicated that SJC could activate AMPK signaling pathway and effectively promote autophagy in esophageal cancer cells. Therefore, we further used western blotting to confirm that SJC activated autophagy in esophageal cancer cells through the AMPK/ULK1 signaling pathway. The results showed that P-AMPK and P-ULK1 were significantly up-regulated after the treatment with SJC. The ratio of autophagosomes marker proteins LC3II/I was significantly increased. In addition, the expression of the autophagy substrate protein P62 decreased with the degradation of autophagosomes. Using lentiviral transfection of fluorescent label SensGFP-StubRFP-LC3 protein and revalidation of LC3 expression before and after administration by laser confocal microscopy. Compared with the control group, the fluorescence expression of the SJC group was significantly enhanced, indicating that it promoted autophagy in esophageal cancer cells. Cell morphology and the formation of autophagosomes were observed by transmission electron microscopy. Our study shows that the tumor suppressor effect of SJC is related to promoting autophagy in esophageal tumor cells via the AMPK/ULK1 signaling pathway.

Influence of thermal processing on the quality of hawthorn: Quality markers of heat-processed hawthorn.

Hawthorn and its derived products are used worldwide as foods as well as complementary medicine. During the preparation of hawthorn, heating and thermal processing are frequently reported. The thermal processing will change the medicinal purposes and modify the efficacy of hawthorn. However, details including the chemical profile shifting and quality markers of heat-processed hawthorn have not been well understood. In this study, we analyzed the hawthorn samples processed at different temperatures and different times by ultraviolet visible absorption spectrum and liquid-mass spectrometry technologies combined with multivariate statistical analysis. It was revealed for the first time that thermal processing could greatly change the ultraviolet-visible absorption spectra and chemical profiles of hawthorn even with heat treatment at 130°C for 10 min. And the ultraviolet visible absorption spectrum, especially the ratio value (RA500 nm/400 nm ), was a descriptive and qualitative indicator of heating degree for the thermal processing at the macroscopic level. Several components, such as hyperoside, chlorogenic acid, quercetin, and apigenin, decreased or increased in content during the processing, and they could be utilized as the chemical quality markers. The proposed quality markers for heat-processed hawthorn will be helpful for further optimizing the processing conditions of hawthorn.

Deficiency of transmembrane AMPA receptor regulatory protein γ-8 leads to attention-deficit hyperactivity disorder-like behavior in mice.

Attention-deficit hyperactivity disorder (ADHD) is a neurodevelopmental disorder prevalent in school-age children. At present, however, its etiologies and risk factors are unknown. Transmembrane α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor regulatory protein γ-8 (TARP γ-8, also known as calcium voltage-gated channel auxiliary subunit gamma 8 (CACNG8)) is an auxiliary AMPA receptor (AMPAR) subunit. Here, we report an association between TARP γ-8 and ADHD, whereby adolescent TARP γ-8 knockout (KO) mice exhibited ADHD-like behaviors, including hyperactivity, impulsivity, anxiety, impaired cognition, and memory deficits. Human single-nucleotide polymorphism (SNP) analysis also revealed strong associations between intronic alleles in CACNG8 genes and ADHD susceptibility. In addition, synaptosomal proteomic analysis revealed dysfunction of the AMPA glutamate receptor complex in the hippocampi of TARP γ-8 KO mice. Proteomic analysis also revealed dysregulation of dopaminergic and glutamatergic transmissions in the prefrontal cortices of TARP γ-8 KO mice. Methylphenidate (MPH), which is commonly used to treat ADHD, significantly rescued the major behavioral deficits and abnormal synaptosomal proteins in TARP γ-8 KO mice. Notably, MPH significantly reversed the up-regulation of Grik2 and Slc6a3 in the prefrontal cortex. MPH also significantly improved synaptic AMPAR complex function by up-regulating other AMPAR auxiliary proteins in hippocampal synaptosomes. Taken together, our results suggest that TARP γ-8 is involved in the development of ADHD in humans. This study provides a useful alternative animal model with ADHD-like phenotypes related to TARP γ-8 deficiency, which has great potential for the development of new therapies.

Elucidating a Complicated Enantioselective Metabolic Profile: A Study From Rats to Humans Using Optically Pure Doxazosin.

Doxazosin (DOX) is prescribed as a racemic drug for the clinical treatment of benign prostatic hyperplasia and hypertension. Recent studies found that the two enantiomers of DOX exhibit differences in blood concentration and pharmacological effects. However, the stereoselective metabolic characteristics and mechanisms for DOX are not yet clear. Herein, we identified 34 metabolites of DOX in rats based on our comprehensive and effective strategy. The relationship among the metabolites and the most discriminative metabolites between (-)-DOX and (+)-DOX administration was analyzed according to the kinetic parameters using state-of-the-art multivariate statistical methods. To elucidate the enantioselective metabolic profile in vivo and in vitro, we carefully investigated the metabolic characteristics of metabolites after optically pure isomers administration in rat plasma, rat liver microsomes (RLMs) or human liver microsomes (HLMs), and recombinant human cytochrome P450 (CYP) enzymes. As a result, the differences of these metabolites were found based on their exposure and elimination rate, and the metabolic profile of (±)-DOX was more similar to that of (+)-DOX. Though the metabolites identified in RLMs and HLMs were the same, the metabolic profiles of the metabolites from (-)-DOX and (+)-DOX were greatly different. Furthermore, four human CYP enzymes could catalyze DOX to produce metabolites, but their preferences seemed different. For example, CYP3A4 highly specifically and selectively catalyzed the formation of the specific metabolite (M22) from (-)-DOX. In conclusion, we established a comprehensive metabolic system using pure optical isomers from in vivo to in vitro, and the complicated enantioselectivity of the metabolites of DOX was clearly shown. More importantly, the comprehensive metabolic system is also suitable to investigate other chiral drugs.