Synthesis, Antiproliferative Activity and Molecular Docking Analysis of Both Enantiomerically Pure Decursin Derivatives as Anticancer Agents.

Using (S)-decursinol isolated from root of Angelica gigas Nakai (AGN), we semi-synthesized and evaluated a series of both enantiomerically pure decursin derivatives for their antiproliferative activities against A549 human lung cancer cells. All synthesized compounds showed a broad spectrum of inhibitory activities against the growth of A549 cells. Especially, compound (S)-2d with (E)-(furan-3-yl)acryloyl group showed the most potent activity (IC50: 14.03 µM) against A549 cancer cells as compared with the reference compound, decursin (IC50: 43.55 µM) and its enantiomer, (R)-2d (IC50: 151.59 µM). Western blotting assays indicated that (S)-2d more strongly inhibited Janus kinase 1 (JAK1) and signal transducer and activator of transcription activation 3 (STAT3) phosphorylation than decursin in a dose-dependent manner, while having no effect on CXCR7 overexpression and total STAT3 level. In addition, (S)-2d induced cell cycle arrest at G1 phase and subsequent apoptotic cell death in A549 cancer cells. Our combined analysis of molecular docking studies and biological data suggests that the inhibition of JAK1 with (S)-2d resulted in loss of STAT3 phosphorylation and inhibition of cell growth in A549 cancer cells. These overall results strongly suggest that (S)-2d (MRC-D-004) as a novel JAK1 inhibitor may have therapeutic potential in the treatment of A549 human lung cancers by targeting the JAK1/STAT3 signaling pathway.

JAK2 inhibitors improve RA combined with pulmonary fibrosis in rats by downregulating SMAD3 phosphorylation.

BACKGROUND: JAK inhibitors are well known for the treatment of rheumatoid arthritis (RA), but whether they can be used to treat pulmonary fibrosis, a common extra-articular disease of RA, remains to be clarified. METHODS: A jak2 inhibitor, CEP33779 (CEP), was administered to a rat model of RA-associated interstitial lung disease to observe the degree of improvement in both joint swelling and pulmonary fibrosis. HFL1 cells were stimulated with TGF-ß1 to observe the expression of p-JAK2. Then, different concentrations of related gene inhibitors (JAK2, TGFß-R1/2, and p-STAT3) or silencers (STAT3, JAK2) were administered to HFL1 cells, and the expression levels of related proteins were detected to explore the underlying mechanisms of action. RESULTS: CEP not only reduced the degree of joint swelling and inflammation in rats but also improved lung function, inhibited the pro-inflammatory factors IL-1ß and IL-6, reduced lung inflammation and collagen deposition, and alleviated lung fibrosis. CEP decreased the expression levels of TGFß-R2, p-SMAD, p-STAT3, and ECM proteins in rat lung tissues. TGF-ß1 induced HFL1 cells to highly express p-JAK2, with the most pronounced expression at 48 h. The levels of p-STAT3, p-SMAD3, and ECM-related proteins were significantly reduced after inhibition of either JAK2 or STAT3. CONCLUSION: JAK2 inhibitors may be an important and novel immunotherapeutic drug that can improve RA symptoms while also delaying or blocking the development of associated pulmonary fibrotic disease. The mechanism may be related to the downregulation of p-STAT3 protein via inhibition of the JAK2/STAT signaling pathway, which affects the phosphorylation of SMAD3.

The STAT3 inhibitor B9 alleviates lipopolysaccharide-induced acute lung injury through its anti-inflammatory effects.

The development of acute lung injury (ALI), a common respiratory condition with multiple causes, is significantly influenced by the pro-inflammatory environment of signal transducer and activator of transcription 3 (STAT3) in macrophages. Our study aimed to evaluate the anti-inflammatory effects of B9 (N-(4-hydroxyphenyl)-9, 10-dioxo-9, 10-dihydroanthracene-2-sulfonamide), a novel inhibitor targeting the STAT3 SH2 domain, in macrophages and to assess its therapeutic potential for ALI using a mouse model of lipopolysaccharide (LPS)-induced ALI. We found that B9 (30 mg/kg) significantly reduced lung pathological damage and neutrophil infiltration caused by the intratracheal administration of LPS. Additionally, the high expression of pro-inflammatory cytokines (TNF-α, IL-1ß, and IL-6) in alveolar lavage fluid was also inhibited by B9 treatment. The decreased expression of CD86 and increased CD206 in lung tissue demonstrated the anti-inflammatory effect of B9, which was due to its inhibition of the STAT3 signaling pathway in macrophages of ALI mice. Furthermore, B9 suppressed the activation of RAW264.7 cells induced by LPS, characterized by its ability to inhibit the activation of iNOS and STAT3 in a dose-dependent manner, as well as reduce the secretion of IL-6 and IL-1ß. The in vivo preliminary safety evaluation indicated that B9 had a favorable safety profile at the administered doses. These results suggest that B9 exerts a therapeutic effect on LPS-induced ALI, potentially by preventing the phosphorylation of STAT3 Y705 and S727 without affecting the STAT3 protein level. Taken together, these findings provide a foundation for developing B9 as a novel anti-inflammatory agent for ameliorating LPS-induced ALI.

STAT3 inhibition ameliorates renal interstitial inflammation in MRL/lpr mice with diffuse proliferative lupus nephritis.

BACKGROUND AND OBJECTIVES: Acute kidney injury (AKI) is one of the most common and severe clinical syndromes of diffuse proliferative lupus nephritis (DPLN), of which poor prognosis is indicated by aggravated renal function deterioration. However, the specific therapy and mechanisms of AKI in DPLN remain to be explored. METHODS: The correlation between AKI and clinical pathological changes in DPLN patients was analyzed. Expression of STAT3 signaling was detected in MRL/lpr mice with DPLN using immunohistochemical staining and immunoblotting. Inhibition of STAT3 activation by combination therapy was assessed in MRL/lpr mice. RESULTS: Correlation analysis revealed only the interstitial leukocytes were significantly related to AKI in endocapillary DPLN patients. MRL/lpr mice treated with vehicle, which can recapitulate renal damages of DPLN patients, showed upregulation of STAT3, pSTAT3 and caspase-1 in renal cortex. FLLL32 combined with methylprednisolone therapy significantly inhibited the STAT3 activation, improved acute kidney damage, reduced the interstitial infiltration of inflammatory cells and decreased the AKI incidence in MRL/lpr mice. CONCLUSION: STAT3 activation may play an important role in the pathogenesis of DPLN and the development of AKI. Hence, STAT3 inhibition based on the combination of FLLL32 with methylprednisolone may represent a new strategy for treatment of DPLN with AKI.

Exploration of novel isoxazole-fused quinone derivatives as anti-colorectal cancer agents through inhibiting STAT3 and elevating ROS level.

Colorectal cancer (CRC) is trending to be a major health problem throughout the world. Therapeutics with dual modes of action have shown latent capacity to create ideal anti-tumor activity. Signal transducer and activator of transcription 3 (STAT3) has been proved to be a potential target for the development of anti-colon cancer drug. In addition, modulation of tumor redox homeostasis through deploying exogenous reactive oxygen species (ROS)-enhancing agents has been widely applied as anti-tumor strategy. Thus, simultaneously targeting STAT3 and modulation ROS balance would offer a fresh avenue to combat CRC. In this work, we designed and synthesized a novel series of isoxazole-fused quinones, which were evaluated for their preliminary anti-proliferative activity against HCT116 cells. Among these quinones, compound 41 exerted excellent in vitro anti-tumor effect against HCT116 cell line with an IC50 value of 10.18 ± 0.4 nM. Compound 41 was proved to bind to STAT3 by using Bio-Layer Interferometry (BLI) assay, and can significantly inhibit phosphorylation of STAT3. It also elevated ROS of HCT116 cells by acting as a substrate of NQO1. Mitochondrial dysfunction, apoptosis, and cell cycle arrest, which was caused by compound 41, might be partially due to the inhibition of STAT3 phosphorylation and ROS production induced by 41. Moreover, it exhibited ideal anti-tumor activity in human colorectal cancer xenograft model and good safety profiles in vivo. Overall, this study provided a novel quinone derivative 41 with excellent anti-tumor activity by inhibiting STAT3 and elevating ROS level, and gave insights into designing novel anti-tumor therapeutics by simultaneously modulation of STAT3 and ROS.

Network Pharmacology and Molecular Docking Identify the Potential Mechanism and Therapeutic Role of Scutellaria baicalensis in Alzheimer's Disease.

Aim: Scutellaria baicalensis, a traditional Chinese medicinal herb renowned for its anti-inflammatory, antioxidant, and anti-tumor properties, has shown promise in alleviating cognitive impairment associated with Alzheimer's disease. Nonetheless, the exact neuroprotective mechanism of Scutellaria baicalensis against Alzheimer's disease remains unclear. In this study, network pharmacology was employed to explore the possible mechanisms by which Scutellaria baicalensis protects against Alzheimer's disease. Methods: The active compounds of Scutellaria baicalensis were retrieved from the TCMSP database, and their corresponding targets were identified. Alzheimer's disease-related targets were obtained through searches in the GeneCards and OMIM databases. Cytoscape 3.6.0 software was utilized to construct a regulatory network illustrating the "active ingredient-target" relationships. Subsequently, the target genes affected by Scutellaria baicalensis in the context of Alzheimer's disease were input into the String database to establish a PPI network. GO analysis and KEGG analysis were conducted using the DAVID database to predict the potential pathways associated with these key targets. Following this, the capacity of these active ingredients to bind to core targets was confirmed through molecular docking. In vitro experiments were then carried out for further validation. Results: A total of 36 active ingredients from Scutellaria baicalensis were screened out, which corresponded to 365 targets. Molecular docking results demonstrated the robust binding abilities of Baicalein, Wogonin, and 5,2'-Dihydroxy-6,7,8-trimethoxyflavone to key target proteins (SRC, PIK3R1, and STAT3). In vitro experiments showed that the active components of Scutellaria baicalensis can inhibit STAT3 expression by downregulating the PIK3R1/SRC pathway in Neuro 2A cells. Conclusion: In summary, these findings collectively suggest that Scutellaria baicalensis holds promise as a viable treatment option for Alzheimer's disease.

Enhancing cancer therapy: The role of drug delivery systems in STAT3 inhibitor efficacy and safety.

The signal transducer and activator of transcription 3 (STAT3), a member of the STAT family, resides in the nucleus to regulate genes essential for vital cellular functions, including survival, proliferation, self-renewal, angiogenesis, and immune response. However, continuous STAT3 activation in tumor cells promotes their initiation, progression, and metastasis, rendering STAT3 pathway inhibitors a promising avenue for cancer therapy. Nonetheless, these inhibitors frequently encounter challenges such as cytotoxicity and suboptimal biocompatibility in clinical trials. A viable strategy to mitigate these issues involves delivering STAT3 inhibitors via drug delivery systems (DDSs). This review delineates the regulatory mechanisms of the STAT3 signaling pathway and its association with cancer. It offers a comprehensive overview of the current application of DDSs for anti-STAT3 inhibitors and investigates the role of DDSs in cancer treatment. The conclusion posits that DDSs for anti-STAT3 inhibitors exhibit enhanced efficacy and reduced adverse effects in tumor therapy compared to anti-STAT3 inhibitors alone. This paper aims to provide an outline of the ongoing research and future prospects of DDSs for STAT3 inhibitors. Additionally, it presents our insights on the merits and future outlook of DDSs in cancer treatment.

The histamine H4 receptor antagonist 1-[(5-chloro-2,3-dihydro-1-benzofuran-2-yl)methyl]-4-methyl-piperazine(LINS01007) prevents the development of DSS-induced colitis in mice.

Ulcerative colitis (UC) is a chronic inflammatory bowel disease with growing incidence worldwide. Our group reported the compound 5-choro-1-[(2,3-dihydro-1-benzofuran-2-yl)methyl]piperazine (LINS01007) as H4R antagonist (pKi 6.2) and therefore the effects and pharmacological efficacy on a DSS-induced mice model of UC were assessed in this work. Experimental acute colitis was induced in male BALB/c mice (n = 5-10) by administering 3 % DSS in the drinking water for six days. The test compound LINS01007 was administered daily i.p. (5 mg/kg) and compared to control group without treatment. Body weight, water and food consumption, and the presence of fecal blood were monitored during 7-day treatment period. The levels of inflammatory markers (PGE2, COX-2, IL-6, NF-κB and STAT3) were also analyzed. Animals subjected to the acute colitis protocol showed a reduction in water and food intake from the fourth day (p < 0.05) and these events were prevented by LINS01007. Histological signs of edema, hyperplasia and disorganized intestinal crypts, as well as neutrophilic infiltrations, were found in control mice while these findings were significantly reduced in animals treated with LINS01007. Significant reductions in the levels of PGE2, COX-2, IL-6, NF-κB and STAT3 were observed in the serum and tissue of treated animals. The results demonstrated the significant effects of LINS01007 against DSS-induced colitis, highlighting the potential of H4R antagonism as promising treatment for this condition.

10-Secocycloartane (=9,19-cyclo-9,10-secolanostane) triterpenoid saponins: Huangqiyenins M-X from Astragalus membranaceus (Fisch.) Bge.

Phytochemical investigations of the leaves of Astragalus membranaceus (Fisch.) Bge. have led to the isolation of 12 undescribed triterpenoid saponins named huangqiyenins M-X. The structures of the undescribed compounds were determined using NMR and HRESIMS data. The cytotoxicity of these compounds against the RKO and HT-29 colon cancer cell lines was evaluated. Among these compounds, huangqiyenin W exhibited the highest cytotoxic activity against RKO colon cancer cells, whereas huangqiyenin Q and W showed moderate cytotoxic activity against HT-29 colon cancer cells. The network pharmacology results indicated that STAT3, IL-2 and CXCR1 are the correlated targets of huangqiyenin W against colon cancer, with AGE-RAGE and Th17 cell differentiation as the key signaling pathways.

The antiprotozoal drug nitazoxanide improves experimental liver fibrosis in mice.

Nitazoxanide is an FDA-approved antiprotozoal drug. Our previous studies find that nitazoxanide and its metabolite tizoxanide affect AMPK, STAT3, and Smad2/3 signals which are involved in the pathogenesis of liver fibrosis, therefore, in the present study, we examined the effect of nitazoxanide on experimental liver fibrosis and elucidated the potential mechanisms. The in vivo experiment results showed that oral nitazoxanide (75, 100 mg·kg-1) significantly improved CCl4- and bile duct ligation-induced liver fibrosis in mice. Oral nitazoxanide activated the inhibited AMPK and inhibited the activated STAT3 in liver tissues from liver fibrosis mice. The in vitro experiment results showed that nitazoxanide and its metabolite tizoxanide activated AMPK and inhibited STAT3 signals in LX-2 cells (human hepatic stellate cells). Nitazoxanide and tizoxanide inhibited cell proliferation and collagen I expression and secretion of LX-2 cells. Nitazoxanide and tizoxanide inhibited transforming growth factor-ß1 (TGF-ß1)- and IL-6-induced increases of cell proliferation, collagen I expression and secretion, inhibited TGF-ß1- and IL-6-induced STAT3 and Smad2/3 activation in LX-2 cells. In mouse primary hepatic stellate cells, nitazoxanide and tizoxanide also activated AMPK, inhibited STAT3 and Smad2/3 activation, inhibited cell proliferation, collagen I expression and secretion. In conclusion, nitazoxanide inhibits liver fibrosis and the underlying mechanisms involve AMPK activation, and STAT3 and Smad2/3 inhibition.

A novel p55PIK signaling peptide inhibitor alleviates neuroinflammation via the STAT3/NF-kB signaling pathway in experimental stroke.

BACKGROUND: Ischemic stroke remains the predominant contributor to mortality and disability globally. Microglia undergo rapid activation and initiate inflammatory cascade reactions by phenotypic polarization, participating in the regulation of inflammatory injury and tissue repair post-ischemic stroke. Regulating microglia-mediated neuroinflammation is a promising therapeutic strategy for ischemic stroke. Previously, we designed and synthesized a novel p55PIK inhibitor, TAT-N15 polypeptide, which presents inhibitive activity on NF-κB signaling-mediated inflammation in acute conjunctivitis and allergic rhinitis. The present study aimed to explore the therapeutic effect and mechanism of TAT-N15 on ischemia stroke. METHODS: The mouse model of transient cerebral ischemia was made using the intraluminal filament method. After being treated with daily intraperitoneal injections of TAT-N15 (10 mg/kg) for 7 d, the neurological outcomes and the cerebral infarction volume were evaluated. Histopathology of the ischemia cerebral hemisphere was observed by H&E and Nissl staining. Neuronal survival, astrogliosis, and co-labeling of CD86/Iba1 and CD206/Iba1 were detected by immunofluorescence. The cell apoptosis was estimated by TUNEL staining. The expression levels of apoptosis-associated proteins, proinflammatory cytokines, protein markers of M1 and M2 microglia, and the phosphorylation of NF-κB and STAT3 proteins in the ischemic penumbra were detected by Western blot. RESULTS: TAT-N15 treatment significantly decreased the infarct volume and alleviated neurological functional impairment, neuronal injury, and neuron apoptosis. Meanwhile, TAT-N15 treatment restrained the activation of microglia and astrocytes as well as the protein expression of proinflammatory cytokine in ischemic penumbra. Additionally, the administration of TAT-N15 treatment resulted in a significant reduction in the density of M1 phenotype microglia while concurrently increasing the density of M2 phenotype microglia within the ischemic penumbra. Finally, mechanical analysis unveiled that TAT-N15 exerted a substantial inhibitory effect on the protein expression of phosphorylated STAT3 and NF-κB. CONCLUSION: TAT-N15 may inhibit neuroinflammation via regulating microglia activation and polarization through the STAT3/NF-κB pathway, which exhibits the neuroprotection effect in ischemic stroke.

Network pharmacology and experimental insights into STAT3 inhibition by novel isoxazole derivatives of piperic acid in triple negative breast cancer.

STAT3 is a crucial member within a family of seven essential transcription factors. Elevated STAT3 levels have been identified in various cancer types, notably in breast cancer (BC). Consequently, inhibiting STAT3 is recognized as a promising and effective strategy for therapeutic intervention against breast cancer. We herein synthesize a library of isoxazole (PAIs) from piperic acid [2E, 4E)-5-(2H-1,3-Benzodioxol-5-yl) penta-2,4-dienoic acid] on treatment with propargyl bromide followed by oxime under prescribed reaction conditions. Piperic acid was obtained by hydrolysis of piperine extracted from Piper nigrum. First, we checked the binding potential of isoxazole derivatives with breast cancer target proteins by network pharmacology, molecular docking, molecular dynamic (MD) simulation and cytotoxicity analysis as potential anti-breast cancer (BC) agents. The multi-source databases were used to identify possible targets for isoxazole derivatives. A network of protein-protein interactions (PPIs) was generated by obtaining 877 target genes that overlapped gene symbols associated with isoxazole derivatives and BC. Molecular docking and MD modelling demonstrated a strong affinity between isoxazole derivatives and essential target genes. Further, the cell viability studies of isoxazole derivatives on the human breast carcinoma cell lines showed toxicity in all breast cancer cell lines. In summary, our study indicated that the isoxazole derivative showed the significant anticancer activity. The results highlight the prospective utility of isoxazole derivatives as new drug candidates for anticancer chemotherapy, suggesting route for the continued exploration and development of drugs suitable for clinical applications.

STAT3 phosphorylation inhibitor Bt354 exhibits anti-neoplastic activity in glioblastoma multiforme cells.

The high mortality rate of glioblastoma multiforme (GBM), a lethal primary brain tumor, is attributable to postsurgical recurrence. STAT3, an oncogenic protein, is a signal transducer and transcription activator encourages cancer cell migration and proliferation, which results in resistance to therapy. STAT3 inhibition reduces cancer metastasis and improves patient prognosis. Bt354, a small molecule STAT inhibitor, exhibits significant cytotoxic and anti-proliferative activities against certain cancer types. Here, we demonstrated that exposure of GBM cells (U87 MG) to Bt354 had a significant, concentration-dependent growth suppression. Bt354 also induced apoptosis and downregulated the expression of the epithelial-mesenchymal transition genes. Therefore, this study suggests the potential of Bt354 for treating GBM owing to its ability to induce cytotoxicity.

Development of new thieno[2,3-d]pyrimidines as dual EGFR and STAT3 inhibitors endowed with anticancer and pro-apoptotic activities.

In part due to the resilience of cellular feedback pathways that develop therapeutic resistance to targeting the EGFR alone, using EGFR inhibitors alone was demonstrated to be unsuccessful in clinical trials. The over-activation of the signal transducer/activator of transcription 3 (STAT3) during the administration of an EGFR inhibitor is expected to play a substantial part in the failure and resistance of EGFR inhibitor treatment. Therein, we proposed a hypothesis that induced STAT3-mediated resistance to EGFR inhibition therapy could be addressed by a dual inhibition of EGFR and STAT3 method. To this end, we tried to discover new thieno[2,3-d]pyrimidine derivatives "5a-o". Results from the screening on A549 and MCF7 cancer cell lines revealed that compounds 5j and 5k showed two-digit nanomolar with appropriate safety towards the WI-38 cell line. The best molecules, 5j and 5k, were subjected to γ-radiation, and their cytotoxic efficacy didn't change after irradiation, demonstrating that not having to use it avoided its side effects. Compounds 5j and 5k demonstrated the highest inhibition when their potency was tested as dual inhibitors on EGFR 67 and 41 nM, respectively, and STAT3 5.52 and 3.34 nM, respectively, proved with in silico molecular docking and dynamic simulation. In light of the results presented above, the capacity of both powerful compounds to alter the cell cycle and initiate the apoptotic process in breast cancer MCF7 cells was investigated. Caspase-8, Bcl-2, Bax and Caspase-9 apoptotic indicators were studied.

Targeting JAK2/STAT3 for the treatment of cancer: A review on recent advancements in molecular development using structural analysis and SAR investigations.

Cancer is indeed considered a hazardous and potentially life-threatening disorder. The JAK/STAT pathway is an important intracellular signaling cascade essential for many physiological functions, such as immune response, cell proliferation, and differentiation. Dysregulation of this pathway aids in the progression and development of cancer. The downstream JAK2/STAT3 signaling cascades are legitimate targets against which newer anticancer drugs can be developed to prevent and treat cancer. Understanding the mechanisms behind JAK2/STAT3 participation in cancer has paved the way for developing innovative targeted medicines with the potential to improve cancer treatment outcomes. This article provides information on the current scenario and recent advancements in the design and development of anticancer drugs targeting JAK2/STAT3, including structural analysis and SAR investigations of synthesized molecules. Numerous preclinical and clinical trials are ongoing on these inhibitors, which are highlighted to gain more insight into the broader development prospects of inhibitors of JAK2/STAT3.

Repurposing TAK875 as a novel STAT3 inhibitor for treating inflammatory bowel disease.

Inflammatory bowel disease (IBD) is a chronic immune-mediated disease associated with a high recurrence rate and an elevated risk of colon cancer. In this study, we screened a bioactive compound library using a luciferase reporter assay and identified the compound TAK875 as a novel inhibitor of signal transducer and activator of transcription 3 (STAT3). Surface plasmon resonance analysis, differential scanning fluorimetry, and isothermal titration calorimetry demonstrated that TAK875 directly bound to recombinant STAT3. TAK875 suppressed the lipopolysaccharide (LPS)-induced release of nitric oxide, inducible nitric oxide synthase, and inflammatory factors in RAW264.7 cells, likely by inhibiting STAT3 phosphorylation. In addition, TAK875 inhibited the differentiation of CD4+ T cells into T-helper 17 cells, which may partially account for its anti-inflammatory effect. TAK875 also alleviated the LPS-induced accumulation of intracellular reactive oxygen species, thus displaying its antioxidant effects. Finally, we demonstrated its satisfactory anti-inflammatory effect in a dextran sulfate sodium-induced mouse model of ulcerative colitis. In conclusion, this study presented TAK875 as a novel STAT3 inhibitor and demonstrated its anti-inflammatory and antioxidant effects both in vitro and in vivo.

Nifuroxazide repurposing for protection from diabetes-induced retinal injury in rats: Implication of oxidative stress and JAK/STAT3 axis.

The prevalence of diabetes mellitus (DM) is alarmingly increasing worldwide. Diabetic retinopathy (DR) is a prevailing DM microvascular complication, representing the major cause of blindness in working-age population. Inflammation is a crucial player in DR pathogenesis. JAK/STAT3 axis is a pleotropic cascade that modulates diverse inflammatory events. Nifuroxazide (Nifu) is a commonly used oral antibiotic with reported JAK/STAT3 inhibition activity. The present study investigated the potential protective effect of Nifu against diabetes-induced retinal injury. Effect of Nifu on oxidative stress, JAK/STAT3 axis and downstream inflammatory mediators has been also studied. Diabetes was induced in Sprague Dawley rats by single intraperitoneal injection of streptozotocin (50 mg/kg). Animals were assigned into four groups: normal, Nifu control, DM, and DM + Nifu. Nifu was orally administrated at 25 mg/kg/day for 8 weeks. The effects of Nifu on oxidative stress, JAK/STAT3 axis proteins, inflammatory factors, tight junction proteins, histological, and ultrastructural alterations were evaluated using spectrophotometry, gene and protein analyses, and histological studies. Nifu administration to diabetic rats attenuated histopathological and signs of retinal injury. Additionally, Nifu attenuated retinal oxidative stress, inhibited JAK and STAT3 phosphorylation, augmented the expression of STAT3 signaling inhibitor SOCS3, dampened the expression of transcription factor of inflammation NF-κB, and inflammatory cytokine TNF-α. Collectively, the current study indicated that Nifu alleviated DR progression in diabetic rats, suggesting beneficial retino-protective effect. This can be attributed to blocking JAK/STAT3 axis in retinal tissues with subsequent amelioration of oxidative stress and inflammation.

Labdane and ent-halimane diterpenoids with STAT3-inhibitory activity from Leonurus sibiricus.

Nine undescribed labdane diterpenoids (1-9) and one undescribed ent-halimane diterpenoid (10) were isolated from the aerial parts of Leonurus sibiricus, together with four known analogues (11-14) during our searching for naturally occurring antitumor agents. Their structures were established by detailed spectroscopic analyses and electronic circular dichroism analysis. Compound 4 possessed a rare 10-epi labdane scaffold. All compounds except 5 were evaluated for their inhibitory activities against interleukin (IL)-6-stimulated signal transducer and activator of transcription (STAT3) expression using a luciferase reporter assay. Compound 1 showed the most inhibitory effect with the IC50 value 20.31 µM. Compound 1 inhibited the activation of JAK2/STAT3 signal pathway through binding to Gln326 of STAT3 in CNE cells. The antiproliferative evaluation of compound 1 against CNE, CAL-27, A549 and PANC-1 cells demonstrated that CNE cells were the most sensitive to 1. Furthermore, compound 1 showed moderate efficacy in inhibiting cell migration, invasion, and epithelial-mesenchymal transition in CNE cells. In addition, compound 1 also promoted ferroptosis in CNE cells in a dose-dependent manner. These results suggest that compound 1 might be a potential candidate lead for treating nasopharyngeal carcinoma.

STAT3 inhibition enhances gemcitabine sensitivity in pancreatic cancer by suppressing EMT, immune escape and inducing oxidative stress damage.

Pancreatic cancer (PC) is a highly-malignant tumor of the digestive system with a very poor prognosis and high mortality. Chemotherapy and PD-1/PD-L1 immune checkpoint blockade are important treatment strategies for advanced PC. However, chemotherapy resistance and poor therapeutic effect of immune checkpoint inhibitors is are the main clinical problems to be solved urgently at present. The effects of combined application of gemcitabine and STAT3 inhibition on the proliferation, apoptosis, migration, and invasion of PC cells (PCCs) were investigated. In addition, oxidative stress (OS), ferroptosis, immune escape, and the epithelial-mesenchymal transition (EMT) were evaluated. STAT3 inhibition with Stattic enhanced the inhibitory activity of gemcitabine on PCC proliferation by regulating the cell cycle. STAT3 inhibition enhanced mitochondrial-dependent apoptosis in gemcitabine-treated PCCs, but did not induce autophagy and ferroptosis. Further study showed that the anti-proliferative and pro-apoptotic effects may be associated with increased OS damage by inactivating Nrf2-HO-1 signaling, as well as DNA damage by inducing the imbalance between ATM andATR-Chk1 pathway. In addition, STAT3 inhibition strengthened gemcitabine-mediated suppression in PCC invasion and migration by antagonizing Smad2/3-dependent EMT. Moreover, the anti-tumorimmuneresponse of gemcitabine was upregulated by Stattic through reducing the expression of PD-L1 and CD47. Mechanistically, combined application of gemcitabine and Stattic suppressed the phosphorylation and nuclear expression of STAT3. Interestingly, the activities of AKT and ß-catenin signaling were also regulated, suggesting that drug combination has a broad-spectrum signal regulation effect. STAT3 inhibition enhanced the sensitivity of PCCs to the chemotherapy drug gemcitabine by suppressing EMT and immune escape and inducing OS damage.

Pharmacokinetic characteristics of golidocitinib, a highly selective JAK1 inhibitor, in healthy adult participants.

Background: Golidocitinib is an orally available, potent and highly selective JAK (Janus kinase)-1 inhibitor of JAK/STAT3 signaling under clinical development for the treatment of cancer and autoimmune diseases. The objectives of the two reported studies were to investigate the pharmacokinetics (PK), safety, and tolerability of golidocitinib in healthy Chinese participants as compared to those healthy Western participants, as well as the food effect exploration. Methods: Two phase I studies (JACKPOT2 and JACKPOT3) were conducted in USA and China, respectively. In JACKPOT2 study, participants were randomized into placebo or golidocitinib arm in single-ascending dose cohorts (5 - 150 mg) and multiple-ascending dose cohorts (25 - 100 mg, once daily) for 14 days. In the food effect cohort, golidocitinib (50 mg) was administrated shortly after a high-fat meal (fed conditions) as compared to under fasting conditions. In JACKPOT3 study conducted in China, participants were randomized to placebo or golidocitinib arm in single-ascending dose cohorts (25 - 150 mg). Results: Exposure of golidocitinib generally increased in a dose-proportional manner across a dose range of 5 mg to 150 mg (single dose) and 25 mg to 100 mg (once daily). High-fat food did not alter the PK of golidocitinib with statistical significance. Low plasma clearance and extensive volume of distribution characterizes PK of golidoctinib, and long half-life across the dose levels supported once daily dosing. The inter-ethnic difference in primary PK parameters was evaluated. The result suggested slightly higher peak plasma concentrations (Cmax) but comparable area under the plasma concentration-time curve (AUC) was observed in Asian (Chinese) subjects as compared to Caucasian and/or Black subjects, while it was not considered clinically relevant. Golidocitinib was well tolerated without Common Terminology Criteria for Adverse Events (CTCAE) grade 3 or higher drug-related treatment emergent adverse events (TEAE) reported. Conclusion: No noticeable inter-ethnic difference was observed among Asian, Black, and Caucasian healthy subjects in anticipation of the favorable PK properties of golidocitinib. The effect of food on the bioavailability of golidocitinib was minor following a single oral administration of 50 mg. These data guided to use the same dose and regimen for multinational clinical development. Clinical trial registrations: https://clinicaltrials.gov/ct2/show/NCT03728023?term=NCT03728023&draw=2&rank=1, identifier (NCT03728023); http://www.chinadrugtrials.org.cn/clinicaltrials.searchlistdetail.dhtml, identifier (CTR20191011).