2024 Vol. 14, No. 6

News
High-strength antiswelling adhesive achieves both hemostasis and wound healing
Xin Zhao, Jinlong Luo, Ying Huang, Lei Mu, Jueying Chen, Zhen Liang, Zhanhai Yin, Dake Chu, Yong Han, Baolin Guo
2024, 14(6): 100955. doi: 10.1016/j.jpha.2024.02.009
Abstract:
Review papers
Basic regulatory science behind drug substance and drug product specifications of monoclonal antibodies and other protein therapeutics
Patanachai K. Limpikirati, Sorrayut Mongkoltipparat, Thinnaphat Denchaipradit, Nathathai Siwasophonpong, Wudthipong Pornnopparat, Parawan Ramanandana, Phumrapee Pianpaktr, Songsak Tongchusak, Maoxin Tim Tian, Trairak Pisitkun
2024, 14(6): 100916. doi: 10.1016/j.jpha.2023.12.006
Abstract:

In this review, we focus on providing basics and examples for each component of the protein therapeutic specifications to interested pharmacists and biopharmaceutical scientists with a goal to strengthen understanding in regulatory science and compliance. Pharmaceutical specifications comprise a list of important quality attributes for testing, references to use for test procedures, and appropriate acceptance criteria for the tests, and they are set up to ensure that when a drug product is administered to a patient, its intended therapeutic benefits and safety can be rendered appropriately. Conformance of drug substance or drug product to the specifications is achieved by testing an article according to the listed tests and analytical methods and obtaining test results that meet the acceptance criteria. Quality attributes are chosen to be tested based on their quality risk, and consideration should be given to the merit of the analytical methods which are associated with the acceptance criteria of the specifications. Acceptance criteria are set forth primarily based on efficacy and safety profiles, with an increasing attention noted for patient-centric specifications. Discussed in this work are related guidelines that support the biopharmaceutical specification setting, how to set the acceptance criteria, and examples of the quality attributes and the analytical methods from 60 articles and 23 pharmacopeial monographs. Outlooks are also explored on process analytical technologies and other orthogonal tools which are on-trend in biopharmaceutical characterization and quality control.

New perspectives on the therapeutic potential of quercetin in non-communicable diseases: Targeting Nrf2 to counteract oxidative stress and inflammation
Li Zhang, Li-Yue Xu, Fei Tang, Dong Liu, Xiao-Lan Zhao, Jing-Nan Zhang, Jia Xia, Jiao-Jiao Wu, Yu Yang, Cheng Peng, Hui Ao
2024, 14(6): 100930. doi: 10.1016/j.jpha.2023.12.020
Abstract:

Non-communicable diseases (NCDs), including cardiovascular diseases, cancer, metabolic diseases, and skeletal diseases, pose significant challenges to public health worldwide. The complex pathogenesis of these diseases is closely linked to oxidative stress and inflammatory damage. Nuclear factor erythroid 2-related factor 2 (Nrf2), a critical transcription factor, plays an important role in regulating antioxidant and anti-inflammatory responses to protect the cells from oxidative damage and inflammation-mediated injury. Therefore, Nrf2-targeting therapies hold promise for preventing and treating NCDs. Quercetin (Que) is a widely available flavonoid that has significant antioxidant and anti-inflammatory properties. It modulates the Nrf2 signaling pathway to ameliorate oxidative stress and inflammation. Que modulates mitochondrial function, apoptosis, autophagy, and cell damage biomarkers to regulate oxidative stress and inflammation, highlighting its efficacy as a therapeutic agent against NCDs. Here, we discussed, for the first time, the close association between NCD pathogenesis and the Nrf2 signaling pathway, involved in neurodegenerative diseases (NDDs), cardiovascular disease, cancers, organ damage, and bone damage. Furthermore, we reviewed the availability, pharmacokinetics, pharmaceutics, and therapeutic applications of Que in treating NCDs. In addition, we focused on the challenges and prospects for its clinical use. Que represents a promising candidate for the treatment of NCDs due to its Nrf2-targeting properties.

Bio-soft matter derived from traditional Chinese medicine: Characterizations of hierarchical structure, assembly mechanism, and beyond
Guiya Yang, Yue Liu, Yuying Hu, Yue Yuan, Yunan Qin, Quan Li, Shuangcheng Ma
2024, 14(6): 100943. doi: 10.1016/j.jpha.2024.01.011
Abstract:

Structural and functional explorations on bio-soft matter such as micelles, vesicles, nanoparticles, aggregates or polymers derived from traditional Chinese medicine (TCM) has emerged as a new topic in the field of TCM. The discovery of such cross-scaled bio-soft matter may provide a unique perspective for unraveling the new effective material basis of TCM as well as developing innovative medicine and biomaterials. Despite the rapid rise of TCM-derived bio-soft matter, their hierarchical structure and assembly mechanism must be unambiguously probed for a further in-depth understanding of their pharmacological activity. In this review, the current emerged TCM-derived bio-soft matter assembled from either small molecules or macromolecules is introduced, and particularly the unambiguous elucidation of their hierarchical structure and assembly mechanism with combined electron microscopic and spectroscopic techniques is depicted. The pros and cons of each technique are also discussed. The future challenges and perspective of TCM-derived bio-soft matter are outlined, particularly the requirement for their precise in situ structural determination is highlighted.

Original articles
In situ visualization of the cellular uptake and sub-cellular distribution of mussel oligosaccharides
Zhenjie Yu, Huarong Shao, Xintian Shao, Linyan Yu, Yanan Gao, Youxiao Ren, Fei Liu, Caicai Meng, Peixue Ling, Qixin Chen
2024, 14(6): 100932. doi: 10.1016/j.jpha.2023.12.022
Abstract:

Unlike chemosynthetic drugs designed for specific molecular and disease targets, active small-molecule natural products typically have a wide range of bioactivities and multiple targets, necessitating extensive screening and development. To address this issue, we propose a strategy for the direct in situ microdynamic examination of potential drug candidates to rapidly identify their effects and mechanisms of action. As a proof-of-concept, we investigated the behavior of mussel oligosaccharide (MOS-1) by tracking the subcellular dynamics of fluorescently labeled MOS-1 in cultured cells. We recorded the entire dynamic process of the localization of fluorescein isothiocyanate (FITC)-MOS-1 to the lysosomes and visualized the distribution of the drug within the cell. Remarkably, lysosomes containing FITC-MOS-1 actively recruited lipid droplets, leading to fusion events and increased cellular lipid consumption. These drug behaviors confirmed MOS-1 is a candidate for the treatment of lipid-related diseases. Furthermore, in a high-fat HepG2 cell model and in high-fat diet-fed apolipoprotein E (ApoE) mice, MOS-1 significantly promoted triglyceride degradation, reduced lipid droplet accumulation, lowered serum triglyceride levels, and mitigated liver damage and steatosis. Overall, our work supports the prioritization of in situ visual monitoring of drug location and distribution in subcellular compartments during the drug development phase, as this methodology contributes to the rapid identification of drug indications. Collectively, this methodology is significant for the screening and development of selective small-molecule drugs, and is expected to expedite the identification of candidate molecules with medicinal effects.

Oxalate regulates crystal-cell adhesion and macrophage metabolism via JPT2/PI3K/AKT signaling to promote the progression of kidney stones
Qianlin Song, Chao Song, Xin Chen, Yunhe Xiong, Ziqi He, Xiaozhe Su, Jiawei Zhou, Hu Ke, Caitao Dong, Wenbiao Liao, Sixing Yang
2024, 14(6): 100956. doi: 10.1016/j.jpha.2024.02.010
Abstract:

Oxalate is an organic dicarboxylic acid that is a common component of plant foods. The kidneys are essential organs for oxalate excretion, but excessive oxalates may induce kidney stones. Jupiter microtubule associated homolog 2 (JPT2) is a critical molecule in Ca2+ mobilization, and its intrinsic mechanism in oxalate exposure and kidney stones remains unclear. This study aimed to reveal the mechanism of JPT2 in oxalate exposure and kidney stones. Genetic approaches were used to control JPT2 expression in cells and mice, and the JPT2 mechanism of action was analyzed using transcriptomics and untargeted metabolomics. The results showed that oxalate exposure triggered the upregulation of JPT2, which is involved in nicotinic acid adenine dinucleotide phosphate (NAADP)-mediated Ca2+ mobilization. Transcriptomic analysis revealed that cell adhesion and macrophage inflammatory polarization were inhibited by JPT2 knockdown, and these were dominated by phosphatidylinositol 3-kinase (PI3K)/AKT signaling, respectively. Untargeted metabolomics indicated that JPT2 knockdown inhibited the production of succinic acid semialdehyde (SSA) in macrophages. Furthermore, JPT2 deficiency in mice inhibited kidney stones mineralization. In conclusion, this study demonstrates that oxalate exposure facilitates kidney stones by promoting crystal-cell adhesion, and modulating macrophage metabolism and inflammatory polarization via JPT2/PI3K/AKT signaling.

MGMT activated by Wnt pathway promotes cisplatin tolerance through inducing slow-cycling cells and nonhomologous end joining in colorectal cancer
Haowei Zhang, Qixin Li, Xiaolong Guo, Hong Wu, Chenhao Hu, Gaixia Liu, Tianyu Yu, Xiake Hu, Quanpeng Qiu, Gang Guo, Junjun She, Yinnan Chen
2024, 14(6): 100950. doi: 10.1016/j.jpha.2024.02.004
Abstract:

Chemotherapy resistance plays a pivotal role in the prognosis and therapeutic failure of patients with colorectal cancer (CRC). Cisplatin (DDP)-resistant cells exhibit an inherent ability to evade the toxic chemotherapeutic drug effects which are characterized by the activation of slow-cycle programs and DNA repair. Among the elements that lead to DDP resistance, O6-methylguanine (O6-MG)-DNA-methyltransferase (MGMT), a DNA-repair enzyme, performs a quintessential role. In this study, we clarify the significant involvement of MGMT in conferring DDP resistance in CRC, elucidating the underlying mechanism of the regulatory actions of MGMT. A notable upregulation of MGMT in DDP-resistant cancer cells was found in our study, and MGMT repression amplifies the sensitivity of these cells to DDP treatment in vitro and in vivo. Conversely, in cancer cells, MGMT overexpression abolishes their sensitivity to DDP treatment. Mechanistically, the interaction between MGMT and cyclin dependent kinase 1 (CDK1) inducing slow-cycling cells is attainted via the promotion of ubiquitination degradation of CDK1. Meanwhile, to achieve nonhomologous end joining, MGMT interacts with XRCC6 to resist chemotherapy drugs. Our transcriptome data from samples of 88 patients with CRC suggest that MGMT expression is co-related with the Wnt signaling pathway activation, and several Wnt inhibitors can repress drug-resistant cells. In summary, our results point out that MGMT is a potential therapeutic target and predictive marker of chemoresistance in CRC.

CA IX-targeted Ag2S quantum dots bioprobe for NIR-II imaging-guided hypoxia tumor chemo-photothermal therapy
Xinyue Cui, Zhuang Hu, Ruihan Li, Peng Jiang, Yongchang Wei, Zilin Chen
2024, 14(6): 100969. doi: 10.1016/j.jpha.2024.100969
Abstract:

Hypoxia is the common characteristic of almost all solid tumors, which prevents therapeutic drugs from reaching the tumors. Therefore, the development of new targeted agents for the accurate diagnosis of hypoxia tumors is widely concerned. As carbonic anhydrase IX (CA IX) is abundantly distributed on the hypoxia tumor cells, it is considered as a potential tumor biomarker. 4-(2-Aminoethyl) benzenesulfonamide (ABS) as a CA IX inhibitor has inherent inhibitory activity and good targeting effect. In this study, Ag2S quantum dots (QDs) were used as the carrier to prepare a novel diagnostic and therapeutic bioprobe (Ag2S@polyethylene glycol (PEG)-ABS) through ligand exchange and amide condensation reaction. Ag2S@PEG-ABS can selectively target tumors by surface-modified ABS and achieve accurate tumor imaging by the near infrared-II (NIR-II) fluorescence characteristics of Ag2S QDs. PEG modification of Ag2S QDs greatly improves its water solubility and stability, and therefore achieves high photothermal stability and high photothermal conversion efficiency (PCE) of 45.17%. Under laser irradiation, Ag2S@PEG-ABS has powerful photothermal and inherent antitumor combinations on colon cancer cells (CT-26) in vitro. It also has been proved that Ag2S@PEG-ABS can realize the effective treatment of hypoxia tumors in vivo and show good biocompatibility. Therefore, it is a new efficient integrated platform for the diagnosis and treatment of hypoxia tumors.

Paeoniflorin ameliorates chronic colitis via the DR3 signaling pathway in group 3 innate lymphoid cells
Shaowei Huang, Xueqian Xie, Bo Xu, Zengfeng Pan, Junjie Liang, Meiling Zhang, Simin Pan, Xiaojing Wang, Meng Zhao, Qing Wang, Jinyan Chen, Yanyang Li, Lian Zhou, Xia Luo
2024, 14(6): 100940. doi: 10.1016/j.jpha.2024.01.008
Abstract:

Inhibiting the death receptor 3 (DR3) signaling pathway in group 3 innate lymphoid cells (ILC3s) presents a promising approach for promoting mucosal repair in individuals with ulcerative colitis (UC). Paeoniflorin, a prominent component of Paeonia lactiflora Pall., has demonstrated the ability to restore barrier function in UC mice, but the precise mechanism remains unclear. In this study, we aimed to delve into whether paeoniflorin may promote intestinal mucosal repair in chronic colitis by inhibiting DR3 signaling in ILC3s. C57BL/6 mice were subjected to random allocation into 7 distinct groups, namely the control group, the 2% dextran sodium sulfate (DSS) group, the paeoniflorin groups (25, 50, and 100 mg/kg), the anti-tumor necrosis factor-like ligand 1A (anti-TL1A) antibody group, and the IgG group. We detected the expression of DR3 signaling pathway proteins and the proportion of ILC3s in the mouse colon using Western blot and flow cytometry, respectively. Meanwhile, DR3-overexpressing MNK-3 cells and 2% DSS-induced Rag1-/- mice were used for verification. The results showed that paeoniflorin alleviated DSS-induced chronic colitis and repaired the intestinal mucosal barrier. Simultaneously, paeoniflorin inhibited the DR3 signaling pathway in ILC3s and regulated the content of cytokines (interleukin-17A, granulocyte-macrophage colony stimulating factor, and interleukin-22). Alternatively, paeoniflorin directly inhibited the DR3 signaling pathway in ILC3s to repair mucosal damage independently of the adaptive immune system. We additionally confirmed that paeoniflorin-conditioned medium (CM) restored the expression of tight junctions in Caco-2 cells via coculture. In conclusion, paeoniflorin ameliorates chronic colitis by enhancing the intestinal barrier in an ILC3-dependent manner, and its mechanism is associated with the inhibition of the DR3 signaling pathway.

Fangchinoline induces antiviral response by suppressing STING degradation
Jinyong Wang, Fang Xie, Xin Jia, Xuejiao Wang, Lingdong Kong, Yiying Li, Xue Liang, Meiqi Zhang, Yuting He, Wandi Feng, Tong Luo, Yao Wang, Anlong Xu
2024, 14(6): 100972. doi: 10.1016/j.jpha.2024.100972
Abstract:

The stimulator of interferon genes (STING), an integral adaptor protein in the DNA-sensing pathway, plays a pivotal role in the innate immune response against infections. Additionally, it presents a valuable therapeutic target for infectious diseases and cancer. We observed that fangchinoline (Fan), a bis-benzylisoquinoline alkaloid (BBA), effectively impedes the replication of vesicular stomatitis virus (VSV), encephalomyocarditis virus (EMCV), influenza A virus (H1N1), and herpes simplex virus-1 (HSV-1) in vitro. Fan treatment significantly reduced the viral load, attenuated tissue inflammation, and improved survival in a viral sepsis mouse model. Mechanistically, Fan activates the antiviral response in a STING-dependent manner, leading to increased expression of interferon (IFN) and interferon-stimulated genes (ISGs) for potent antiviral effects in vivo and in vitro. Notably, Fan interacts with STING, preventing its degradation and thereby extending the activation of IFN-based antiviral responses. Collectively, our findings highlight the potential of Fan, which elicits antiviral immunity by suppressing STING degradation, as a promising candidate for antiviral therapy.

Secondary metabolites of mulberry leaves exert anti-lung cancer activity through regulating the PD-L1/PD-1 signaling pathway
Guiqin Ye, Xin Sun, Jiuzhou Li, Yuanyuan Mai, Ruilan Gao, Jianbin Zhang
2024, 14(6): 100926. doi: 10.1016/j.jpha.2023.12.016
Abstract:

Lung cancer ranks the top of malignancies that cause cancer-related deaths worldwide. The leaves of Morus alba L are traditional Chinese medicine widely applied in respiratory diseases. Our previous work has demonstrated the anti-lung cancer effect of secondary metabolites of mulberry leaf, but their mechanism of action has still not fully elucidated. We synthesized Moracin N (MAN)-Probe conjugated with alkyne to label lung cancer cells and identified protein targets by chemical proteomic analysis. MAN and its probe exerted similar growth-inhibitory effect on human lung cancer cells. Chemical proteomic results showed that MAN targeted the programmed death ligand 1 (PD-L1) checkpoint pathway and T cell receptor (TCR) signaling pathway, indicating its immune-regulatory function. Cell-free surface plasmon resonance (SPR) results showed the direct interaction of MAN with PD-L1 protein. Molecular docking analysis demonstrated that MAN bound to E158 residue of PD-L1 protein. MAN downregulated the expression levels of PD-L1 in a time- and dose-dependent manner and disrupted the PD-L1/programmed death 1 (PD-1) binding, including other secondary metabolites of mulberry leaves Guangsangon E (GSE) and Chalcomoracin (CMR). Human peripheral blood mononuclear cells (PBMCs) co-cultured with MAN-treated A549 cells, resulting in the increase of CD8+ GZMB+ T cells and the decrease of CD8+ PD-1+ T cells. It suggested that MAN exerts anti-cancer effect through blocking the PD-L1/PD-1 signaling. In vivo, MAN combined with anti-PD-1 antibody significantly inhibited lung cancer development and metastasis, indicating their synergistic effect. Taken together, secondary metabolites of mulberry leaves target the PD-L1/PD-1 signaling, enhance T cell-mediated immunity and inhibit the tumorigenesis of lung cancer. Their modulatory effect on tumor microenvironment makes them able to enhance the therapeutic efficacy of immune checkpoint inhibitors in lung cancer.

Short communications
Establishment of microneurovascular units in alginate hydrogel microspheres to reveal the anti-hypoxic effect of salidroside
Shiyu Chen, Yuxuan Li, Yingrui Zhang, Hongren Yao, Tong Xu, Yi Zhang, Jin-Ming Lin, Xian-Li Meng
2024, 14(6): 100967. doi: 10.1016/j.jpha.2024.100967
Abstract:
Medcheck: A novel software for automated de-formulation of traditional Chinese medicine (TCM) prescriptions by liquid chromatography-mass spectrometry
Xiao-lan Li, Jian-qing Zhang, Yun Li, Xuan-jing Shen, Huan-ya Yang, Lin Yang, Meng Xu, Qi-rui Bi, Chang-liang Yao, De-an Guo
2024, 14(6): 100958. doi: 10.1016/j.jpha.2024.02.012
Abstract:
Corrigenda
Corrigendum to “Active components of Dengzhan Shengmai ameliorate cognitive impairment by facilitating hippocampal long-term potentiation via the NMDA receptor-mediated Ca2+/CaMKII pathway” [J. Pharm. Anal. 14 (2024) 435–438]
Congyu Ma, Ning Sheng, Jun Wu, Lulu Wang, Jiandong Jiang, Haibo Yu, Jinlan Zhang
2024, 14(6): 101025. doi: 10.1016/j.jpha.2024.101025
Abstract:
Corrigendum to “Nitrosamines crisis in pharmaceuticals - Insights on toxicological implications, root causes and risk assessment: A systematic review” [J. Pharm. Anal. 14 (2024) 100919]
Hemanth P. R. Vikram, Tegginamath Pramod Kumar, Gunjan Kumar, Narasimha M. Beeraka, Rajashree Deka, Sheik Mohammed Suhail, Sandeep Jat, Namitha Bannimath, Gayatiri Padmanabhan, Ravandur S. Chandan, Pramod Kumar, Bannimath Gurupadayya
2024, 14(6): 101033. doi: 10.1016/j.jpha.2024.101033
Abstract: