The development of oral delivery systems for protein and peptide drugs has long been the goal of the pharmaceutical field. However, extremely low bioavailability has been an unsolved problem in this research. This is related to the physical and chemical properties of the protein and peptides, as well as the three physiological barriers: the complex pH and enzyme environment after oral administration, the mucus barrier and the intestinal epithelial tissue barrier. With its porous rigid structure and high drug loading and strong stability, inorganic particle carrier has the ability to penetrate mucus and transport intestinal epithelia after surface modification, thus overcoming the three physiological barriers. This study introduces the current clinical application of protein and peptide drugs for oral administration, and the mechanism and coping strategies of physiological barrier that limit their oral absorption. Additionally, the application of inorganic materials as carriers in the oral delivery system of protein and peptide drugs are summarized.

Recently, chimeric antigen receptor (CAR)-T cell therapy has made remarkable success in hematological malignancies but faces a series of challenges in solid tumors. One of the major problems is that CAR-T cells overexpress programmed death-1 (PD-1) in tumor microenvironment. Therefore, blocking PD-1 can rescue the effector functions of CAR-T cells and reduce tumor burden significantly. Herein, it is aimed to summarize the progress in preclinical and clinical research on immunotherapy of combining CAR-T cells with PD-1 blockade for solid tumors.

Circular RNAs (circRNAs) are a class of single-stranded RNA molecules with covalently closed continuous loops which are involved in the occurrence and development of diseases. Neurovascular units(NVU) refer to the cellular and molecular interfaces between the circulatory system and central nervous system(CNS), mainly composed of microvascular cells, glial cells, neurons, and extracellular matrix, which are of great significance for maintaining brain homeostasis. However, the role of circRNA on NVU remains unknown. This article summarizes the latest research progress, with the aim of providing new targets and strategies for the treatment of related diseases.

OBJECTIVE To design and synthesize trifluoromethoxy chalcone derivatives based on the natural licorice chalcone parent as the lead compound backbone, and to study their anti-cervical cancer activity in vitro. METHODS New trifluoromethoxy chalcone derivatives were synthesized by Claisen-Schmidt aldol condensation. The structures were confirmed by ¹H-NMR, ¹³C-NMR and HR-ESI-MS. The cytotoxic activity of the target compounds on cervical cancer cell lines HeLa, SiHa, C-33A and normal cervical epithelial immortalized H8 cells were determined by MTT assay, and the structure activity relationship (SAR) was analyzed and the candidate compounds were selected. The effects of candidate compound 3o on invasion, migration, apoptosis and cell cycle of HeLa cells were determined by Transwell and flow cytometry. The candidate compound 3o was docked to the MDM2 and protein target by molecular docking method, and the binding ability and binding characteristics of the compound to the target protein molecules were determined. The regulatory effects of the candidate compounds on MDM2 and p53 proteins were assessed using Western Blot analysis. RESULTS Twenty new trifluoromethoxy chalcones were synthesized. Candidate compound 3o showed the strongest inhibitory activity against cervical cancer cells (IC₅₀=4.60±0.40 μmol·L^-1), which was significantly better than that of positive drug cisplatin (IC₅₀=17.16±0.93 μmol·L^-1). The candidate compound 3o could effectively inhibit the invasion and migration of HeLa cells, induce apoptosis and arrest the cell cycle at G₀/G₁ phase. Candidate compound 3o binds to key amino acids in p53 binding pocket of MDM2 protein (binding energy -37.62 kcal·mol^-1). The compounds significantly downregulated MDM2 protein expression while upregulating p53 protein levels. CONCLUSION The research results provide experimental evidence for screening new chalcone derivatives as targeted, effective, and low-toxicity anti-tumor candidates against cervical cancer.

OBJECTIVE To establish the cell model of reporter gene for detecting histamine phosphate. METHODS The plasmids of human histamine phosphate H1 receptor (H1R), H2 receptor (H2R) and the response elements of G protein-coupled receptor alpha subunits, including Gs, Gi or Gq, cloned with reporter gene were co-transfected into HEK293 cells. The transfected ratio of the plasmid was optimized. Subsequently, the expression and function of H1R and H2R were verified by detecting the change of cAMP content and cellular [Ca²⁺] after the effect of agonists. The chemiluminescence activities of HEK293 cells transfected with H1R and/or H2R under different concentrations of histamine were compared. Finally, the cell model was verified by adding histamine phosphate into compound amino acid injection, succinyl gelatin injection, and enoxaparin sodium solution to simulate the detection and calculating the recovery rate. RESULTS When the amount of three plasmids was 1∶1∶10, the response value of cells to histamine phosphate was higher, this ratio was used for subsequent transfection. The changes of cAMP and Ca²⁺contents in cells verified the overexpression of H1R and H2R and the function of reporter gene response element. When the concentration of histamine phosphate was higher than 0.64 nmol·L^-1, the chemiluminescence value of cells overexpressing H1R and H2R reporter genes (H1R/H2R-Luc cells), was higher than that of other groups (P<0.05). This cell model was used to detect the histamine phosphate added in amino acid injection, succinyl gelatin injection, and enoxaparin sodium solution. The recovery rates were between 88%-121% when the concentration of histamine phosphate was between 3.2-400 nmol·L^-1. CONCLUSION The H1R/H2R-Luc cells constructed successfully in the present study would be used for the detection of histamine phosphate.

OBJECTIVE To explore the characteristic and microscopic features of Perillae Folium and Perillae Folium sum Cacumen and provide reference for the identification of Perillae Folium and Perillae Folium sum Cacumen. METHODS Characteristic identification and microscopic identification of 6 batches of Perillae Folium and Perillae Folium sum Cacumen were carried out. Samples were collected at different growing stages, including seeding stage, vegetative growth stage and reproductive stage. RESULTS For Perillae Folium collected at different growthing stages, the transverse sections of leaf veins were compared. As plant grows into older stages, more areas of the leaf veins appeared purple color. Additionally, the color of excited luminescence varied between Perillae Folium collected at different growing stages. These phenomena could be utilized to differentiate between Perillae Folium collected at different growing stages. Next, this experiment explored the microscopic difference of Perillae Folium and its counterfeit, Perillae Folium sum Cacumen. The leaf surface characters of Perillae Folium and Perillae Folium sum Cacumen were found to appear in different shapes. The density of peltare glandular hair and capitate glandular hair, as well as the numerical value of stomatal index ratio (lower/upper surface), was observed to be different between two kinds of leaves. CONCLUSION Through this experiment, the characteristic and microscopic features of Perillae Folium and Perillae Folium sum Cacumen are summarized, offering a reference for the supervision inspection, clinical application and standard revision of Perillae Follium.

OBJECTIVE To explore the mechanism of Wendan Decoction in the treatment of chronic obstructive pulmonary disease (COPD). METHODS Firstly, based on the network pharmacology method, the potential mechanism of Wendan Decoction in the treatment of COPD was revealed by using the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP), Swiss Target Prediction and Metascape databases. Secondly, the intervention effect of Wendan Decoction on endogenous metabolite disorder induced by COPD was studied by metabolomics method. Finally, the results of network pharmacology and metabolomics were integrated through MetScape algorithm, and molecular simulation docking and molecular biology methods were used to study the mechanism of Wendan Decoction in the treatment of COPD. RESULTS The results of network pharmacology revealed 102 active components of Wendan Decoction and 98 potential targets for the treatment of COPD. Further analysis showed that 11 potential pharmacodynamic substances of Wendan Decoction, including 3-tert-butyladipic acid, 6-methylgingediacetate and licoisoflavanone may play a role in regulating lipid and atherosclerosis, CAMP signaling pathway, PI3K-Akt signaling pathway and IL-17 signaling pathway. On the other hand, metabolomics studies have shown that 9 endogenous substances, such as 2-propanol, N-acetylneuraminic acid, threonine, protocholic acid, 2-oxoisovalerate, acetylphosphate, citrate and isobutyric acid, may be the key metabolites of Wendan Decoction in the treatment of COPD. Finally, the integrated analysis showed that lipid metabolism and amino acid metabolism might be the key processes of Wendan Decoction in the treatment of COPD. XDH, KDR, PDE5 A, CYP2C9 and CYP2C19, which were related to oxidation and inflammation, were the potential key targets of Wendan Decoction in the treatment of COPD. The results showed that Wendan Decoction could effectively improve the pathological state of pulmonary congestion, edema and inflammatory cell infiltration in COPD rats, significantly reduce the content of IL-6, TNF-α and MDA activity in lung tissue of COPD rats, and increase SOD activity. Moreover, the molecular docking results also revealed the correlation between potential pharmacodynamic substances and core target genes. CONCLUSION The integrated pharmacological studies based on metabolomics, network pharmacology and molecular docking showed that Wendan Decoction might achieve the effect of treating COPD by regulating inflammatory response, oxidative stress, lipid metabolism and amino acid metabolism.

OBJECTIVE To design and optimize the enzymatic preparation process of 3-hydroxyphloridzin and investigate the in vivo anti-vitiligo activity of 3-hydroxyphloridzin, thus to provide scientific basis for achieving the mass production of 3-hydroxyphloridzin and vitiligo prevention and treatment. METHODS 3-hydroxyphloridzin was prepared in a system containing L-ascorbic acid and crude polyphenol oxidase (PPO) solution, with phloridzin from the aqueous extract of young leaves of Malus hupehensis being the substrate. The molar yield of 3-hydroxyphloridzin was taken as the index, and factors including phloridzin concentration, pH, temperature, reaction time, L-ascorbic acid concentration, enzyme concentration, and stirring speed were selected to optimize the preparation process conditions of 3-hydroxyphloridzin by a complete randomized design, Placket-Burman (PB) test, path of steepest ascent method, and central composite design (CCD)-effect surface method. To verify the feasibility of the preparation process, the reaction system was scaled up to 250 L. Besides, Vitiligo model was constructed by applying hydroquinone cream to the back skin of C57BL/6 mice for 75 consecutive days, and the mice were randomly divided into blank group, model group, positive group (8-methoxypsoralen (8-mop), 4.25 mg·kg^-1), and 3-hydroxyphloridzin low, medium, and high dosage group (10, 20, and 40 mg·kg^-1) on the 20th day of modeling, and were administered with 55 days of gavage. The mice in each group were depilated every 5 days, while the modeling area was videotaped and the hair decoloration was scored. At the end of the administration, blood was taken by removing the mice eyeballs, and the serum tyrosinase (TYR), cholinesterase (CHE) activity, and malondialdehyde (MDA) content of mice were detected using the kit. In addition, skin was taken from the dorsal lesions of mice and sections were stained with hematoxylin-eosin (HE), then under a light microscope, 50 hair follicles were observed to count those containing melanin RESULTS The optimal preparation process of 3-hydroxyphloridzin was as follows: phloridzin concentration of 1.70 mmol·L^-1, reaction time of 6.0 h, and PPO concentration of 25.0 U·mL^-1. In the amplified experiment, the reaction solution was decontaminated by a macroporous resin, concentrated, and dried to obtain 334.80 g of crude 3-hydroxyphloridzin with a purity of 70.32%, and the extraction rate of 8.37%, and the purity of 98.10% of the 3-hydroxyphloridzin was obtained by further isolation and purification, which yielded a pure product of 3-hydroxyphloridzin with a purity of 98.10% and the yield of 68.50%. In vivo activity experiments showed that compared with the model group, the hair decoloration of mice in the 3-hydroxyphloridzin low, medium and high dose groups was significantly improved, with a significant decrease in hair decoloration scores, serum MDA content and CHE activity (P<0.05) and a significant increase in the number of melanin-containing hair follicles and serum TYR activity (P<0.05). CONCLUSION 3-hydroxyphloridzin can be prepared by enzymatic extraction of the aqueous extract of young leaves of Malus hupehensis, and the method has the feasibility of quantitative production. 3-hydroxyphloridzin has therapeutic activity for vitiligo, and the mechanism of action is related to enhancement of tyrosinase activity to increase melanin synthesis and improvement of oxidative stress damage to slow down the development of vitiligo.

OBJECTIVE To prepare and characterize transferrin (Tf) modified liposomes containing photosensitizing agent indocyanine green (IR820), and to provide a new idea for the clinical treatment of breast cancer by combining photodynamic therapy (PDT) in 4T1 breast cancer cells and the mouse model of breast cancer in situ. METHODS The liposome loaded with IR820 (IR820@Lipo) and the Tf modified liposome loaded with IR820 (Tf-IR820@Lipo) were prepared by the film dispersion method, respectively. The encapsulation efficiency was detected by UV method, and the physical and chemical properties were investigated by transmission electron microscope and particle size meter. CCK8 assay was used to evaluate the inhibitory effect of the two drugs on the proliferation of 4TI cells. The intracellular fluorescence intensity of the two drugs under laser irradiation was detected by reactive oxygen species (ROS). In vivo pharmacodynamic study was performed to analyze the anticancer effect and toxic side effects of drugs. RESULTS The average particle size of IR820@Lipo and Tf-IR820@Lipo were (84.30±15.66 ) and(116.20±14.68) nm, respectively, and the Zeta potential were (-8.21±2.06) and (-5.23±1.19) mV, respectively. Transmission electron microscopy showed that the liposome was spherical and uniformly distributed. The encapsulation efficiency of IR820@Lipo was (94.61±0.67)%, and the drug loading was (8.82±0.92)%. The encapsulation efficiency of Tf-IR820@Lipo was (95.55±0.83)%, and the drug loading was (8.92±1.01)%. Tf-IR820@Lipo could significantly inhibit the proliferation and promote the apoptosis of 4T1 cells under laser irradiation. ROS detection results showed that Tf-IR820@Lipo could significantly enhance the fluorescence intensity of 4T1 cells under laser irradiation. The results of in vivo experiments in tumor-bearing mice showed that the Tf-IR820@Lipo light groups had higher tumor targeting in mice. The pharmacodynamic study in vivo showed that Tf-IR820@Lipo had the strongest inhibitory effect on breast cancer, and did not cause liver and kidney function damage in mice, and had no obvious toxic side effects. CONCLUSION Tf-IR820@Lipo is expected to become a new agent for the treatment of breast cancer due to its high targeting, high efficacy and low toxicity.

OBJECTIVE To optimize the formulation process of liposomes, construct a complex adjuvant liposome peptide vaccine, and evaluate its immune effect in mice. METHODS Taking particle size distribution and polydispersity index as evaluation indicators, the formulation process was optimized through single factor experiments. The physical properties were investigated using a laser particle size analyzer, and the microstructure was observed using transmission electron microscopy. The encapsulation efficiency was calculated using BCA protein concentration assay, and the stability was investigated by storing at 4 and 25 ℃ for 0, 90, and 180 d. Three composite adjuvant peptide vaccines were constructed by combining it with squalene, monophosphoryl lipid A(MPLA), and QS-21. Then, BALB/c mice were randomly divided into a blank control group and an experimental group. The experimental group was subcutaneously injected with free peptide vaccine and three kinds of liposomal peptide vaccine, respectively. The samples were taken 14 d after the second immunization. ELISA was used to detect the specific IgG antibody and its subtype titer in mouse serum, and flow cytometry was used to detect the spleen lymphocyte typing, to evaluate the immune effects of different peptide vaccine formulations. RESULTS The optimal preparation conditions obtained from single factor experiments were as follows: membrane material ratio of 5∶1, ultrasonic power of 30 W, ultrasonic frequency of 40 times, and high-pressure homogenization time of 6 min. The average particle size of the liposomes was reduced from 329.7 nm to 132.0 nm, with a polydispersity index of 17.8% and an encapsulation efficiency of 76.9%. The morphology under transmission electron microscopy was regular and approximately spherical, and the stability was good after storage at 4 ℃ for 180 d. The results of the in vivo immune experiment in mice showed that the liposomal peptide vaccine could produce high titer IgG and IgG2a antibodies, up to 6.4×10⁴ and 3.2×10⁴, respectively; upregulate the proportion of CD4⁺T and CD8⁺T cells in lymphocytes (P<0.05), and enhance cellular immunity in mice. CONCLUSION The physicochemical properties of the optimized formulation process are stable and good, and the polypeptide vaccine formulated with adjuvant can enhance humoral and cellular immunity in mice。 It is a potential carrier for the delivery of polypeptide antigen immunization.

OBJECTIVE To investigate metagenomic sequencing in the identification and tracing of bacteria contaminating drugs, and evaluate its application value. METHODS Both metagenomic sequencing and isolation culturing were employed to identify and type bacteria in two batches of Chinese patent medicines and environmental samples from the testing laboratory. The metagenomic sequencing results were analyzed at the strain level using MetaPhlan and StrainSifter software. RESULTS The analysis revealed a substantial contamination of Bacillus cereus-like organisms in both medicines and environmental samples, with the presence of the same B. cereus-like strain in medicine B18 and environmental sample HJ2. Through isolation culturing, five B. cereus-like strains were obtained from the medicines and environment. A phylogenetic tree was constructed based on the gyrA and panC gene sequences amplified by B. cereus-specific PCR, and multilocus sequence typing (MLST) results, all of which demonstrated that the B. cereus-like strain in medicine B18 belonged to the same group as one strain from the environment. CONCLUSION These findings indicate that metagenomic sequencing technology is reliable and has advantage over isolation culturing in comprehensively detecting potentially hazardous bacterial species and accurately performing strain-level traceability analysis. It holds significant application value in identification and tracing of pharmaceutical contamination.

OBJECTIVE To establish a self-contrast HPLC method with correction factor for determination of the related substances in tamsulosin hydrochloride, and validate the limits of impurities based on the prediction of genotoxicity using Nexus 2.6 software (with Derek and Sarah). METHODS ZORBAX SB-C₁₈(4.6 mm×150 mm,3.5 μm)column was used for the determination of correction factors of the eight known impurities in tamsulosin hydrochloride with mobile phase consisting of perchlorate buffer solution-acetonitrile by gradient elution under the detection wavelength of 225 nm, the related substances in tamsulosin hydrochloride were determine by self-contrast HPLC method with the correction factor, and the genotoxicity of the impurities was predicted by using Nexus 2.6 software. RESULTS There was no significant difference (the deviation is within ±0.02%) between the results by relative correction factors and external standards. The validation test showed that the proposed method met the requirements for the intended analytical applications, and the predicted result of impurity I by Nexus 2.6 software was positive (ICH M7 class 3), and all the others were negative (ICH M7 class 5). CONCLUSION The established method is rapid, efficient, accurate and sensitive, and a limit of 0.1% is established for each known impurity according to the predicted genotoxicity. This study provides a basis for revising pharmacopoeia standards.

OBJECTIVE To evaluate the economic value of denosumab compared with alendronate and zoledronate in the treatment of postmenopausal women and men with osteoporosis (OP) from the perspective of Chinese health system. METHODS Markov model was built and the treatment period was set to 5 years. The clinical efficacy, health utilities and cost data were obtained from published literature. Lifetime health outcomes and costs were calculated for each intervention with the consideration of adherence and treatment residual effect. With the incremental cost-effectiveness ratio as the index, the threshold of a willingness-to-pay was set as 3 times GDP per capita. Scenario analysis and sensitivity analysis were conducted to test the result's robustness. RESULTS Denosumab was dominant compared with alendronate or zoledronate in postmenopausal women with OP, associated with higher QALY by 0.06 and 0.03 and lower costs by CNY1 471 and CNY3 327 respectively. Denosumab was also dominant compared with alendronate or zoledronate in men with OP, associated with higher QALY by 0.07 and 0.03 and lower CNY2 costs by 158 and CNY3 526 respectively. Sensitivity analyses verified the robustness of study results. CONCLUSION In the treatment of postmenopausal women and men with OP, denosumab has absolute advantages over alendronate and zoledronate, and should be a preferred or long-term treatment in clinical practice.