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关键词： Endostar   HIF-1   MHC-I   beta 2m   lung cancer  

摘要： Endostar is a human recombinant endostatin which is an attractive anti-angiogenesis protein. Because inefficient antigen presenting MHC class I expression (which can be downregulated by HIF-1) is an important strategy for cancer immune evasion, besides its anti-angiogenesis effect, it remains unclear whether Endostar has an inhibitory effect on HIF-1 expression by upregulating MHC class I expression in cancer cells to facilitate immunotherapies, including PD-1/PD-L1 inhibitors. In this study, A549 and NCI-H1299 lung cancer cells were treated with Endostar (6.25 mu g/ml, 12.5 mu g/ml, and 25 mu g/ml, respectively). HIF-1 expression was detected by Immunocytochemistry and Western blot. Proteins of the MHC class I alpha-heavy chain and beta 2 m light chain, STAT3 and pSTAT3 were detected by Western blot. The mRNAs of MHC class I alpha-heavy chain and beta 2 m light chain were detected by RT-qPCR. It was shown that decreased expression of HIF-1 and promotion of beta 2-microglobulin were observed after Endostar treatment. In addition, elevated levels of MHC class I alpha-heavy chain mRNA and protein, as well as downregulation of STAT3 and pSTAT3, were also observed following Endostar treatment. Endostar inhibited HIF-1 expression in A549 and NCI-H1299 lung cancer cells, upregulated expression of MHC class I alpha-heavy chain and beta 2 m light chain, with the upregulation of STAT3 and pSTAT3, suggesting involvement of STAT3 pathway. It is important because only in combination with MHC class I on target cells can tumor antigenic peptides be recognized by CD8+ CTLs which destroy target cells. However, MHC class I is frequently deficient in cancer cells.

关键词： Autophagy   IL-10   macrophage   Mycobacterium abscessus subsp. abscessus   RUBCN  

摘要： Mycobacterium abscessus subsp. abscessus (***) is a nontuberculous mycobacterium that can infect human lung macrophages, which poses a public health concern. Understanding its mechanism is crucial for developing strategies to combat *** infections. *** survives within host cells by inhibiting autophagy, a defense mechanism used against intracellular pathogens;therefore, we investigated the mechanism underlying autophagy inhibition and human lung macrophage infection by ***. This study focuses on the *** UC22 strain, which exhibits stronger inhibition of autophagic flux compared to the *** ATCC 19,977 strain. Central to this study is MAB_0676c, a protein secreted by *** UC22, and its effects on autophagic flux and the innate immune response, particularly its role in enhancing IL-10 production, a known autophagy regulator. Experiments showed that MAB_0676c expression stabilizes autophagy-related proteins while reducing LC3-LAMP2 co-localization in macrophages, thereby inhibiting autophagy and promoting bacterial growth. Furthermore, blocking IL-10 reduced both autophagy-related protein levels and the intracellular growth of MAB_0676c-expressing bacteria. Therefore, *** UC22 mediates intracellular survival by inhibiting autophagy through IL-10 production. Our study reveals bacterial immune-evasion tactics and identifies a potential therapeutic target for treating infectious diseases caused by nontuberculous mycobacteria.

关键词： IL-15 transpresentation   mRNA delivery   NK cell immunotherapy   ovarian cancer  

摘要： Ovarian cancer (OC) is the most lethal gynecological malignancy. As high numbers of Natural Killer (NK) cells in ascites associate with improved survival, the adoptive transfer of allogeneic NK cells is an attractive therapeutic strategy. An approach to further improve NK cell expansion and anti-tumor functionality post-infusion includes IL-15 transpresentation (transIL-15), which involves surface expression of the IL-15 cytokine bound to IL-15R alpha. However, others have substantiated that systemic administration of ALT/N-803, a soluble molecule mimicking transIL-15, leads to T cell-mediated rejection of the infused allogeneic NK cell product. In addition, whether transIL-15 induce superior expansion and functionality of our hematopoietic progenitor cell-derived NK cells (HPC-NK) remains understudied. Here, we propose to transfect OC cells with IL-15 and IL-15R alpha mRNA and evaluate HPC-NK cell stimulation in vitro. Co-transfection of both mRNAs resulted in surface co-expression of both components, thus mimicking the transIL-15. Importantly, co-culture of HPC-NK cells with transIL-15 OC cells resulted in superior proliferation, IFN gamma production, cytotoxicity and granzyme B secretion. Furthermore, we observed uptake of IL-15R alpha by HPC-NK cells when co-cultured with transIL-15 OC cells, which associates with NK cell long-term proliferation and survival. Superior killing and granzyme B secretion were also observed in transIL-15 OC spheroids. Our results demonstrate that local delivery of IL-15 and IL-15R alpha mRNA to OC tumors may be a safer strategy to boost HPC-NK cell therapy of OC through IL-15 transpresentation.

关键词： Calpain 2   hepatocellular carcinoma   IL-1 alpha   MDSCs   tumor microenvironment  

摘要： Tumor-promoting inflammation significantly impacts cancer progression, and targeting inflammatory cytokines has emerged as a promising therapeutic approach in clinical trials. Interleukin (IL)-1 alpha, a member of the IL-1 cytokine family, plays a crucial role in both inflammation and carcinogenesis. How IL-1 alpha is secreted in the tumor microenvironment has been poorly understood, and we previously showed that calpain 1 cleaves pro-IL-1 alpha for mature IL-1 alpha secretion, which exacerbates hepatocellular carcinoma by recruiting myeloid-derived suppressor cells. In this study, we report that calpain 2 also modulates IL-1 alpha secretion. Notably, a deficiency in calpain 2 resulted in enhanced hepatocellular carcinoma development within an IL-1 alpha-enriched tumor microenvironment. Further investigations revealed that calpain 2 deficiency increased calpain 1 expression, implying a compensatory mechanism between the two calpains. Mechanistically, calpain 2 deficiency led to increased expression of FoxO3, which is a forkhead transcription factor that promotes calpain 1 expression. Collectively, these results suggest that calpain 2 modulates calpain 1 expression, and therefore IL-1 alpha secretion through the induction of FoxO3, offering novel potential therapeutic targets for cancer treatment.

关键词： Alzheimer disease   nanobody   single-domain antibody   TNF alpha   TNF alpha inhibition   nanobody humanization  

摘要： Tumor necrosis factor-alpha (TNF alpha) is a key pro-inflammatory cytokine implicated in the pathogenesis of numerous inflammatory and autoimmune diseases, including rheumatoid arthritis, inflammatory bowel disease, and neurodegenerative disorders such as Alzheimer's disease. Effective inhibition of TNF alpha is essential for mitigating disease progression and improving patient outcomes. In this study, we present the development and comprehensive characterization of potent humanized TNF alpha inhibitory nanobodies (TNFI-Nbs) derived from camelid single-domain antibodies. In silico analysis of the original camelid nanobodies revealed low immunogenicity, which was further reduced through machine learning-guided humanization and developability optimization. The two humanized TNFI-Nb variants we developed demonstrated high anti-TNF alpha activity, achieving IC50 values in the picomolar range. Binding assays confirmed their high affinity for TNF alpha, underscoring robust neutralization capabilities. These TNFI-Nbs present valid alternatives to conventional monoclonal antibodies currently used in human therapy, offering potential advantages in potency, specificity, and reduced immunogenicity. Our findings establish a solid foundation for further preclinical development and clinical translation of TNF alpha-targeted nanobody therapies in TNF alpha-mediated diseases.

关键词： Oligomerized polyphenol   bladder cancer   complementary medicine  

摘要： Oligonol (R) (Olg) is an oligomerized polyphenol with a higher bioavailability and exhibits excellent antioxidant and anti-inflammatory properties. However, the effects and underlying mechanisms of Olg on bladder cancer are still largely unknown. The current study found that Olg markedly reduced the viability of bladder cancer cells through the induction of caspase-dependent apoptosis while exhibiting no noticeable toxicity to normal bladder cells. Combining Olg and cisplatin resulted in a synergistic cytotoxic effect on bladder cancer cells. Olg also inhibited cell migration and invasion by affecting the expression of E-cadherin, Snail, N-cadherin, and vimentin. These effects of Olg were significantly weaker in the NF-kappa B silencing cells, revealing that Olg may inhibit cell viability and motility by modulation of the TNF-alpha/NF-kappa B/Snail pathway in human bladder cancer cells. Olg acts against cancer cells and enhances cisplatin chemotherapy, and it holds great potential as an anticancer drug or complementary medicine for bladder cancer.

关键词： Atopic dermatitis   Loranthus tanakae Franch. & Sav   ovalbumin   JAK2   STAT1  

摘要： Loranthus tanakae Franch. and Sav. is a traditional herbal remedy with anti-inflammatory and antioxidative properties, used to treat joint and respiratory inflammation. In this study, we investigated the therapeutic effects of L. tanakae ethanol extract (LTE) on atopic dermatitis (AD). An ovalbumin (OVA)-induced AD animal model and a human keratinocyte cell line, HaCaT, were used to assess LTE treatment effects on AD. An in vitro experiment showed that LTE treatment significantly decreased the production of regulated upon activation, normal T cell expressed and secreted (RANTES) cytokines and macrophage-derived chemokines (MDC) in tumor necrosis factor-alpha (TNF-alpha)/interferon-gamma (IFN-gamma) (TNF-alpha/IFN-gamma)-stimulated HaCaT cells in a concentration-dependent manner. In addition, treatment with LTE markedly reduced the translocation of signal transducer and activator transcription 1 (STAT1) protein to the nucleus and the phosphorylation of Janus kinase 2 (JAK2) in TNF-alpha/IFN-gamma-stimulated HaCaT cells. In the in vivo experiment, administration of LTE significantly decreased the levels of immunoglobulin E (IgE) and interleukin-13 (IL-13) of OVA-induced AD mice, which was supported by histological evidence. Moreover, LTE treatment markedly reduced inflammatory cell infiltration and edema in the OVA-induced AD mice's damaged lesions. In addition, applying LTE notably inhibited the phosphorylation of JAK2 and STAT1 in the OVA-induced AD mice, supported by in vitro results. In conclusion, LTE effectively alleviated the AD-induced skin inflammation in the OVA-induced AD animal model and TNF-alpha/IFN-gamma-stimulated HaCaT cells;this was related to the suppression of JAK2 and STAT1 phosphorylation. These findings suggest that LTE has potential as a therapeutic agent for AD management.

关键词： Panaxatriol saponins   chronic kidney disease   renal fibrosis   TNF-alpha   TGF-beta 1/smad3   UUO model  

摘要： Renal fibrosis is a key pathological process in the progression of chronic kidney disease (CKD). Panaxatriol saponins (PTS), the main bioactive compounds extracted from Panax notoginseng (Burk.) F.H. Chen, have demonstrated antioxidative and anti-inflammatory activities. This study aimed to investigate the potential protective effects of PTS against renal fibrosis and explore the underlying pharmacological mechanisms. A unilateral ureteral obstruction (UUO) model was established in Sprague-Dawley (SD) rats to induce renal fibrosis. Histopathological changes were assessed using hematoxylin and eosin (HE) staining, Masson's trichrome staining, and transmission electron microscopy (TEM). Network pharmacology and molecular docking approaches were employed to identify potential signaling molecules through which PTS may mitigate renal fibrosis. Western blotting, quantitative real-time PCR (qRT-PCR), and immunohistochemistry were utilized to validate the involvement of specific signaling pathways in PTS-mediated anti-fibrotic effects. Our data demonstrated that PTS alleviated renal dysfunction and provided protective effects against renal fibrosis, primarily through the TNF-alpha and TGF-beta 1 signaling pathways. Moreover, PTS treatment significantly downregulated pro-inflammatory cytokines such as TNF-alpha, IL-6, and Smad3 activity. Additionally, PTS inhibited the expression of key fibrosis markers, including alpha-SMA, collagen I, and fibronectin. Our study suggests that PTS exert a prevention effect in renal fibrosis by blocking the TGF-beta 1/Smad3 signaling pathway.

关键词： Tumor-infiltrating lymphocytes   interleukin 2   adoptive cell therapy   checkpoint blockade   anti-PD-1   immune profiling  

摘要： This study evaluated the efficacy and safety of unengineered tumor-infiltrating lymphocytes (TILs) combined with pembrolizumab and either high (HD, Arm-1) or low (LD, Arm-2) doses of IL-2 in patients with metastatic melanoma (MM). Patients were lymphodepleted with cyclophosphamide and fludarabine, followed by TIL infusion and IL-2 (Arm-1: 720,000 IU/kg IV q 8 hrs up to 15 doses;Arm-2: 2 million IU SC for 14 days). Patients received pembrolizumab 200 mg IV starting 21 days post-TIL infusion, and every 3 weeks for up to 2 years. The primary endpoint was overall response rate (ORR) per RECIST 1.1. Blood samples were collected for longitudinal flow cytometry and cytokine analysis. In Arm-1 (n = 7), one patient had a partial response (PR) for 10 months, two had stable disease (SD), three had progressive disease (PD), and one was not evaluable (NE). In Arm-2 (n = 7), one patient had an ongoing PR for over 76 months, one had SD, and five had PD. The toxicity profiles were comparable;however, patients in Arm-2 had lower grade 3 febrile neutropenia (57% vs. 71%) and shorter hospitalization (median 16 days vs. 18 days). No correlation was observed between TIL phenotype and clinical response, although PR patients received high numbers of TIL with a high CD8+/CD4+ T cell ratio. IL-2 dose did not affect the frequency, phenotype, or proliferation of circulating T cell subsets, and anti-PD-1 did not boost T-cell proliferation. No significant differences were observed between IL-2 doses, suggesting low-dose IL-2 as an alternative to high-dose IL-2 after TIL administration.