Improvement of Docetaxel Efficacy through Simultaneous Blockade of Transcription Factors NF-κB and STAT-3 Using Pentoxifylline and Stattic in Prostate Cancer Cells.

Prostate cancer (PCa) is a common and deadly disease in men. It is often diagnosed at advanced stages, at which point patients are treated mainly with docetaxel (DTX), which is effective but limited by resistance and side effects. Overactivation of the transcription factors NF-κB and STAT-3 plays a critical role in the development, progression, and chemoresistance of PCa. In this regard, the blockade of NF-κB with pentoxifylline (PTX) or STAT-3 with Stattic (STT) is known to increase the sensitivity of tumor cells to chemotherapy in both in vitro and in vivo models. We investigated whether simultaneous blockade with PTX and STT increases the efficacy of the DTX treatment in inducing apoptosis in metastatic castration-resistant PCa DU-145 cells. Our results showed that the combination of PTX + STT led to higher levels of apoptosis, regardless of whether or not DTX was present in the treatment. Determining caspases and ΔΨm indicates that the intrinsic caspase pathway of apoptosis is principally favored. In addition, this combination inhibited proliferation and colony formation and arrested the cell cycle in the G1 phase. These results indicate that the combination of the PTX + STAT-3 inhibitor could potentiate DTX effectively, opening the possibility of effective treatments in PCa.

Identification of benzo[b]thiophene-1,1-dioxide derivatives as novel PHGDH covalent inhibitors.

The increased de novo serine biosynthesis confers many advantages for tumorigenesis and metastasis. Phosphoglycerate dehydrogenase (PHGDH), a rate-limiting enzyme in serine biogenesis, exhibits hyperactivity across multiple tumors and emerges as a promising target for cancer treatment. Through screening our in-house compound library, we identified compound Stattic as a potent PHGDH inhibitor (IC50 = 1.98 ± 0.66 µM). Subsequent exploration in structural activity relationships led to the discovery of compound B12 that demonstrated the increased enzymatic inhibitory activity (IC50 = 0.29 ± 0.02 µM). Furthermore, B12 exhibited robust inhibitory effects on the proliferation of MDA-MB-468, NCI-H1975, HT1080 and PC9 cells that overexpress PHGDH. Additionally, using a [U-13C6]-glucose tracing assay, B12 was found to reduce the production of glucose-derived serine in MDA-MB-468 cells. Finally, mass spectrometry-based peptide profiling, mutagenesis experiment and molecular docking study collectively suggested that B12 formed a covalent bond with Cys421 of PHGDH.

Chemokine CCL2 promotes cardiac regeneration and repair in myocardial infarction mice via activation of the JNK/STAT3 axis.

Stimulation of adult cardiomyocyte proliferation is a promising strategy for treating myocardial infarction (MI). Earlier studies have shown increased CCL2 levels in plasma and cardiac tissue both in MI patients and mouse models. In present study we investigated the role of CCL2 in cardiac regeneration and the underlying mechanisms. MI was induced in adult mice by permanent ligation of the left anterior descending artery, we showed that the serum and cardiac CCL2 levels were significantly increased in MI mice. Intramyocardial injection of recombinant CCL2 (rCCL2, 1 µg) immediately after the surgery significantly promoted cardiomyocyte proliferation, improved survival rate and cardiac function, and diminished scar sizes in post-MI mice. Alongside these beneficial effects, we observed an increased angiogenesis and decreased cardiomyocyte apoptosis in post-MI mice. Conversely, treatment with a selective CCL2 synthesis inhibitor Bindarit (30 µM) suppressed both CCL2 expression and cardiomyocyte proliferation in P1 neonatal rat ventricle myocytes (NRVMs). We demonstrated in NRVMs that the CCL2 stimulated cardiomyocyte proliferation through STAT3 signaling: treatment with rCCL2 (100 ng/mL) significantly increased the phosphorylation levels of STAT3, whereas a STAT3 phosphorylation inhibitor Stattic (30 µM) suppressed rCCL2-induced cardiomyocyte proliferation. In conclusion, this study suggests that CCL2 promotes cardiac regeneration via activation of STAT3 signaling, underscoring its potential as a therapeutic agent for managing MI and associated heart failure.

Drug-Loaded Mesoporous Silica Nanoparticles Enhance Antitumor Immunotherapy by Regulating MDSCs.

Myeloid-derived suppressor cells (MDSCs) are recognized as major immune suppressor cells in the tumor microenvironment that may inhibit immune checkpoint blockade (ICB) therapy. Here, we developed a Stattic-loaded mesoporous silica nanoparticle (PEG-MSN-Stattic) delivery system to tumor sites to reduce the number of MDSCs in tumors. This approach is able to significantly deplete intratumoral MSDCs and thereby increase the infiltration of T lymphocytes in tumors to enhance ICB therapy. Our approach may provide a drug delivery strategy for regulating the tumor microenvironment and enhancing cancer immunotherapy efficacy.

Anti-virulence and bactericidal activities of Stattic against Shigella sonnei.

IMPORTANCE: Shigella sonnei is a major human enteric pathogen that causes bacillary dysentery. The increasing spread of drug-resistant S. sonnei strains has caused an emergent need for the development of new antimicrobial agents against this pathogenic bacterium. In this study, we demonstrate that Stattic employs two antibacterial mechanisms against S. sonnei. It exerted both anti-virulence activity and bactericidal activity against S. sonnei, suggesting that it shows advantages over traditional antibiotics. Moreover, Stattic showed excellent synergistic effects with kanamycin, ampicillin, chloramphenicol, and gentamicin against S. sonnei. Our findings suggest that Stattic has promising potential for development as a new antibiotic or as an adjuvant to antibiotics for infections caused by S. sonnei.

Signal transducer and activator of transcription 3 inhibition alleviates resistance to BRAF inhibition in anaplastic thyroid cancer.

Anaplastic thyroid cancer (ATC) is a rare type of thyroid cancer (TC) with no effective therapeutic strategy. Although surgery, chemotherapy and radiation are all available for ATC treatment, the median survival for ATC patients is less than 6 months. In this study, we aimed to study on resistant mechanisms to B-Raf proto-oncogene serine/threonine kinase (BRAF) inhibitor and identify effective combinational therapy for ATC patients. TC cells were treated with Vemurafenib and cell apoptosis and viability were analyzed by flow cytometry and MTT assay. Monolayer and sphere cells were isolated from ATC cells to detect the mRNA level of stem cell markers and differentiation markers by RT-PCR. Phosphor-STAT3 level in sphere and monolayer cells was tested by Western blotting. The xenotransplantation animal model has established to analyze the anti-tumor effect of Vemurafenib and Stattic combinational therapy. Undifferentiated TC cells were resistant to Vemurafenib treatment. Sphere cells isolated from ATC showed no significant change in cell viability and apoptosis upon Vemurafenib treatment, and expressed a high level of stem cell marker and phosphor-STAT3. STAT3 inhibition enhanced the tumorigenic capacity and increased Vemurafenib sensitivity in ATC cell lines. Stattic significantly enhanced anti-tumor effect of Vemurafenib in mouse model. Our findings demonstrate that the combinational therapy of Vemurafenib and Stattic is an effective therapeutic treatment for ATC patients.

Deletion of Rap1 protects against myocardial ischemia/reperfusion injury through suppressing cell apoptosis via activation of STAT3 signaling.

Ischemic heart disease is a leading cause of morbidity and mortality. Repressor activator protein 1 (Rap1), an established telomere-associated protein, is a novel modulator of hypoxia-induced apoptosis. This study aimed to explore the potential direct role of Rap1 in myocardial ischemia/reperfusion injury (I/RI) and to determine the underlying molecular mechanism. In a mouse model of myocardial I/RI (30-min of left descending coronary artery ligation followed by 2-h reperfusion), Rap1 deficiency significantly reduced myocardial infarct size (IS) and improved cardiac systolic/diastolic function. This was associated with a reduction in apoptosis in the post-ischemic myocardium. In H9C2 and primary cardiomyocytes, Rap1 knockdown or knockout significantly suppressed hypoxia/reoxygenation (H/R)-induced cell injury and apoptosis through increasing the phosphorylation/activation of STAT3 at site Ser727 and translocation of STAT3 to the nucleus. We surmise this since Stattic (selective STAT3 inhibitor) pretreatment canceled the abovementioned protective effect. Furthermore, co-immunoprecipitation assay revealed a direct interaction between Rap1 and STAT3, but not JAK2, suggesting that the association of Rap1 with STAT3 may contribute to the reduced activity of STAT3 (Ser727 ) upon H/R stimulation. In conclusion, Rap1 deficiency protects the heart from ischemic damage through STAT3-dependent reduction of cardiomyocyte apoptosis, which may yield viable target for pharmacological intervention in ischemic heart disease.

Stattic sensitizes osteosarcoma cells to epidermal growth factor receptor inhibitors via blocking the interleukin 6-induced STAT3 pathway.

Osteosarcoma (OS), the most common malignant bone tumor with high metastatic potential, frequently affects children and adolescents. Epidermal growth factor receptor (EGFR)-targeted tyrosine kinase inhibitors exhibit encouraging anti-tumor activity for patients with solid tumors, whereas their effects on OS remain controversial. In the present study, we aimed to elucidate the anti-tumor activity of gefitinib for OS, as well as to explore the underlying mechanisms. Gefitinib inhibits cell viability, tumor growth, cell migration, and invasion and promotes cell apoptosis and G1 cycle arrest in OS at a relatively high concentration via suppressing the PI3K/Akt and ERK pathways. However, gefitinib treatment results in the feedback activation of signal transducer and activator of transcription 3 (STAT3) induced by interleukin 6 (IL-6) secretion. Combined treatment with gefitinib and stattic, an inhibitor for STAT3 phosphorylation, engenders more evident inhibitory effects on cell proliferation, migration, and invasion and promotive effects on cell apoptosis and G1 phase arrest in OS, compared with the single exposure to gefitinib or stattic. Western blot analysis demonstrates that stattic treatment in gefitinib-treated OS abrogates the IL-6-induced STAT3 activation and subsequently further restrains the activities of EGFR, Akt, and ERK pathways in tumor cells. This study confirms that the EGFR inhibitor of gefitinib has moderate anti-tumor effects on OS through IL-6 secretion-mediated STAT3 activation. Additional administration of stattic in EGFR-targeted therapies may contribute to improve the efficacy for OS.

Hiltonol Cocktail Kills Lung Cancer Cells by Activating Cancer-Suppressors, PKR/OAS, and Restraining the Tumor Microenvironment.

NSCLC (non-small cell lung cancer) is a leading cause of cancer-related deaths worldwide. Clinical trials showed that Hiltonol, a stable dsRNA representing an advanced form of polyI:C (polyinosinic-polycytidilic acid), is an adjuvant cancer-immunomodulator. However, its mechanisms of action and effect on lung cancer have not been explored pre-clinically. Here, we examined, for the first time, how a novel Hiltonol cocktail kills NSCLC cells. By retrospective analysis of NSCLC patient tissues obtained from the tumor biobank; pre-clinical studies with Hiltonol alone or Hiltonol+++ cocktail [Hiltonol+anti-IL6+AG490 (JAK2 inhibitor)+Stattic (STAT3 inhibitor)]; cytokine analysis; gene knockdown and gain/loss-of-function studies, we uncovered the mechanisms of action of Hiltonol+++. We demonstrated that Hiltonol+++ kills the cancer cells and suppresses the metastatic potential of NSCLC through: (i) upregulation of pro-apoptotic Caspase-9 and Caspase-3, (ii) induction of cytosolic cytochrome c, (iii) modulation of pro-inflammatory cytokines (GRO, MCP-1, IL-8, and IL-6) and anticancer IL-24 in NSCLC subtypes, and (iv) upregulation of tumor suppressors, PKR (protein kinase R) and OAS (2'5' oligoadenylate synthetase). In silico analysis showed that Lys296 of PKR and Lys66 of OAS interact with Hiltonol. These Lys residues are purportedly involved in the catalytic/signaling activity of the tumor suppressors. Furthermore, knockdown of PKR/OAS abrogated the anticancer action of Hiltonol, provoking survival of cancer cells. Ex vivo analysis of NSCLC patient tissues corroborated that loss of PKR and OAS is associated with cancer advancement. Altogether, our findings unraveled the significance of studying tumor biobank tissues, which suggests PKR and OAS as precision oncological suppressor candidates to be targeted by this novel Hiltonol+++ cocktail which represents a prospective drug for development into a potent and tailored therapy for NSCLC subtypes.

LYW-6, a novel cryptotanshinone derived STAT3 targeting inhibitor, suppresses colorectal cancer growth and metastasis.

The constitutive activation of signal transducer and activator of transcription 3(STAT3) is associated with aggressive development and metastasis in colorectal cancer (CRC), but STAT3-targeting drugs remain elusive in clinic. Here, structure-based strategy was used to remodel the natural compound cryptotanshinone into a more effective STAT3 inhibitor LYW-6. Using the Biolayer Interferometry assay, we observed that LYW-6 exhibited specific interactions with STAT3(KD = 6.6 ± 0.7 µM). Western blot analysis and electrophoretic mobility shift assays (EMSA) showed that LYW-6 inhibited the phosphorylation of STAT3 tyrosine 705 (Tyr-705) and had slight effects on STAT1 and STAT5 phosphorylation. Western blot analysis on the upstream kinases of STAT3 confirmed that the inhibitory mechanism on p-STAT3 was independent of upstream kinases. Further investigation demonstrated that LYW-6 downregulated the expression of downstream oncogenes to inhibit cell viability, cell cycle development, and potently increased cell apoptosis in human CRC cells. The invasion and metastasis linked signaling was also blocked by LYW-6 treatment. LYW-6 was found to reduce the metastasis foci in lung on tail-lung metastasis models. In addition, it was observed that LYW-6 markedly diminished STAT3 phosphorylation in tumor tissue and significantly inhibited tumor growth on xenograft models. Tumor development on chemically-induced colorectal cancer model also significantly inhibited by LYW-6 treatment. These findings provided adequate evidence that STAT3 inhibitor LYW-6 might be a potential candidate agent for CRC treatment.

Identification of Nrf2/STAT3 axis in induction of apoptosis through sub-G 1 cell cycle arrest mechanism in HT-29 colon cancer cells.

We investigated the role of stattic as an adjuvant molecule to increase the cytotoxicity of 5-fluorouracil (5-FU) through specific inhibition of molecular targets, signal transducer and activator of transcription 3 (STAT3) and nuclear factor erythroid 2-related factor 2 (Nrf2) in HT-29 colon cancer cells. Cytotoxicity and apoptotic effects were investigated by methylthiazolyldiphenyl-​tetrazolium bromide assay and flow cytometry analysis, respectively. Real-time polymerase chain reaction was applied to assess the messenger RNA (mRNA) level of STAT3, Nrf2, and apoptotic genes including Bax, Bcl-xl, and Bcl-2. The antitumor effect of 5-FU in combination with stattic induced synergistic effect in HT-29 cells with combination indexes (CIs) 0.49. Flow cytometric results related to apoptotic confirmed that there was up to 40% increase in the population of apoptotic cells in HT-29 colon cancer cells incubated with 5-FU and stattic compared with control groups. Our data from gene expression determined a substantial diminish in the mRNA levels of the Nrf2 and antiapoptotic gene Bcl-2 along with a noticeable increase in the level of the proapoptotic Bax in HT-29 colon cells that underwent cotreatment with 5-FU and stattic (P < 0.05). Moreover, the results exhibited that stattic can be used as adjuvant chemotherapy besides the 5-FU. This therapeutic approach in colon cancer could mediate 5-FU chemoresistance via modulating therapeutic targets (ie, STAT3 and Nrf2 pathways) and decreased 5-FU-related adverse effects.

Cryptotanshinone protects against pulmonary fibrosis through inhibiting Smad and STAT3 signaling pathways.

Cryptotanshinone (CTS), a lipophilic compound extracted from root of Salvia miltiorrhiza (Danshen), has demonstrated multiple pharmacological activities, including anti-inflammation, anti-proliferation and anti-infection. However, the effect of CTS on pulmonary fibrosis is unknown. This study aims to investigate the effects of CTS treatment on pulmonary fibrosis and its underlying mechanism. The pulmonary fibrosis model was established by intratracheal instillation of bleomycin (5 mg/kg) in Sprague-Dawley rats (in vivo) and stimulating human fetal lung fibroblasts (HLFs) with transforming growth factor-beta 1 (TGF-ß1) (in vitro). CTS (7.5, 15, 30, 60 mg/kg/day) and pirfenidone (150 mg/kg/day, positive control) were administered by oral gavage for 28 days. In this study, we found CTS treatment improved pulmonary function, relieved pathological changes and attenuated the accumulation of extracellular matrix in pulmonary fibrosis rat model induced by bleomycin. Mechanistically, CTS suppressed phosphorylation of Smad2/3 and STAT3 induced by TGF-ß1 in HLFs. Stattic, a 1-benzothiophene based small-molecule STAT3 inhibitor, resulted in a significant down-regulation of fibrosis biomarkers including fibronectin, collagen type I and alpha smooth muscle actin (α-SMA). Overexpression of STAT3 promoted expression of fibrosis biomarkers in HLFs cell model induced by TGF-ß1 and partially blocked the inhibitory effect of CTS on TGF-ß1-induced fibrosis response. Taken together, these results suggested that CTS protects against pulmonary fibrosis via inhibition of Smad and STAT3 signaling pathways. Thus, CTS may represent a promising drug candidate for treating pulmonary fibrosis.

Activated STAT3 may participate in tumor progression through increasing CD133/survivin expression in early stage of colon cancer.

The activation of signal transducer and activator of transcription 3 (STAT3) by elevated interleukin (IL) levels has been reported to regulate tumorigenesis both in vitro and in vivo. However, the clinical implication of p-STAT3 expression in colon cancer is still controversial. In this study, we evaluated the effect of STAT3 inactivation on biologic behavior of primary (Caco-2) and metastatic colon cancer cells (LoVo and SNU407) and the relation of p-STAT3 expression with the invasion of colon tumor. In vitro study, the STAT3 inhibition by siRNA and stattic treatment significantly reduced colony formation and cell migration and decreased CD133 and survivin the expression compared with a control in all three cell lines. Furthermore, primary cancer cells exhibited a marked decrease in CD133 expression and increased apoptosis compared to metastatic cells after stattic treatment. The immunohistochemical assay using clinical samples of colonic tumors with various invasion depth showed that p-STAT3 expression was inversely associated with tumor invasion (p = 0.001, hazard ratio (HR) = 0.328, 95% confidence interval (95%CI): 0.170-0.632). In conclusion, p-STAT3 may participate in the progression of the early stage of colon cancer through the up-regulation of CD133, which in turn induces survivin expression. However, the regulatory mechanism of these molecules in tumor progression in vivo is need to be more verified.

EGFR-mediated apoptosis via STAT3.

The Epidermal Growth Factor Receptor (EGFR) is a cell surface receptor with primary implications in cell growth in both normal and malignant tissue. Paradoxically, cell lines that hyperexpress the EGFR have been documented to undergo receptor-mediated apoptosis. The underlying mechanism by which EGF-induced apoptosis occurs however remains inexplicit. In an attempt to identify this mechanism, we assessed downstream effectors of EGFR in MDA-MB-468 cells during conditions of EGF-induced apoptosis. The effector assessment revealed STAT3 as a potential mediator of EGF-induced apoptosis. Alternative strategies for activating STAT3, independent of EGFR stimulation, resulted in the induction of the apoptotic pathways. A reduction in STAT3 expression via RNAi resulted in a significant attenuation of EGF-induced PARP cleavage. Our findings support STAT3 as a positive mediator of EGF-induced apoptosis in MDA-MB-468 cells.

Subcutaneous administration of Stattic alleviates neuropathic pain by relieving inflammation in a mouse model of postherpetic neuralgia.

Stattic, a commercial inhibitor of STAT3, can drive the development of neuropathic pain. Exploring the connection between Stattic and JAK1/STAT3 signaling may facilitate the understanding of neuropathic pain caused by postherpetic neuralgia (PHN). In the current study, as crucial regulators of inflammation, STAT3 and its associated JAK1/STAT3 pathway were found to be upregulated and activated in the L4-L6 dorsal root ganglion (DRG) of mice in response to resiniferatoxin (RTX)-induced PHN, while subcutaneous administration of Stattic was found to downregulate STAT3 expression and phosphorylation in a PHN model. Stattic administration further attenuated hypersensitivity to mechanical and thermal stimuli in PHN mice, and alleviated inflammation and cell death in the L4-L6 DRG of mice. Overexpression of STAT3 via microinjection of a lentiviral-STAT3 overexpression vector reversed the abnormal decrease of STAT3 at both the mRNA and protein levels in the L4-6 DRGs of PHN mice and significantly promoted hypersensitivity to mechanical stimuli in the mice. Collectively, we found that subcutaneous static administration alleviated RTX-induced neuropathic pain by deactivating JAK1/STAT3 in mice.

Intratumoral injection of mRNA encoding survivin in combination with STAT3 inhibitor stattic enhances antitumor effects.

Intratumoral delivery of mRNA encoding immunostimulatory molecules can initiate a robust, global antitumor response with little side effects by enhancing local antigen presentation in the tumor and the tumor draining lymph node. Neoantigen-based mRNA nanovaccine can inhibit melanoma growth in mice by intratumoral injection. Myeloid-derived suppressor cells (MDSCs) suppress antitumor immune responses by secreting immunosuppressive agents, such as reactive oxygen species (ROS). Suppression of STAT3 activity by stattic may reduce MDSC-mediated immunosuppression in the TME and promote the antitumor immune responses. In this study, in vitro transcribed mRNA encoding tumor antigen survivin was prepared and injected intratumorally in BALB/c mice bearing subcutaneous colon cancer tumors. In vivo studies demonstrated that intratumoral survivin mRNA therapy could induce antitumor T cell response and inhibit tumor growth of colon cancer. Depletion of CD8+ T cells could significantly inhibit survivin mRNA-induced antitumor effects. RT-qPCR and ELISA analysis indicated that survivin mRNA treatment led to increased expression of receptor activator nuclear factor-κB ligand (RANKL). In vitro experiment showed that MDSCs could be induced from mouse bone marrow cells by RANKL and RANKL-induced MDSCs could produce high level of ROS. STAT3 inhibitor stattic suppressed activation of STAT3 and NF-κB signals, thereby inhibiting expansion of RANKL-induced MDSCs. Combination therapy of survivin mRNA and stattic could significantly enhance antitumor T cell response, improve long-term survival and reduce immunosuppressive tumor microenvironment compared to each monotherapy. In addition, combined therapy resulted in a significantly reduced level of tumor cell proliferation and an obviously increased level of tumor cell apoptosis in CT26 colon cancer-bearing mice, which could be conducive to inhibit the tumor growth and lead to immune responses to released tumor-associated antigens. These studies explored intratumoral mRNA therapy and mRNA-based combined therapy to treat colon cancer and provide a new idea for cancer therapy.

STAT3 inhibitor Stattic Exhibits the Synergistic Effect with FGFRs Inhibitor Erdafitinib in FGFR1-positive Lung Squamous Cell Carcinoma.

Lung squamous cell carcinoma (LUSC), a subset of non-small cell lung cancer (NSCLC), accounts for about 30% of all lung cancers (LC) and exhibits a dismal response to current therapeutic protocols. Existed studies have indicated that aberrations in fibroblast growth factor receptors (FGFRs) play a pivotal role in the progression of LUSC, rendering them as attractive targets for therapeutic intervention in this cancer type. This study found that Erdafitinib (Erda), a novel pan-FGF receptor tyrosine kinase inhibitor (TKI), exerted a cytotoxic effect on LUSC cells. However, STAT3, the downstream target of FGFRs, remained still activated despite Erdafitinib treatment. Then, a STAT3 inhibitor, Stattic (Sta), was concurrently used with Erdafitinib, and the combined treatment demonstrated a synergistic efficacy in both in vitro and in vivo models of LUSC when compared to that of the treatment of the Erdafitinib or Stattic alone. Further molecular studies showed that such an effect of Erdafitinib and Stattic was associated with their concurrently inhibitory effect on FGFR1 and STAT3 signaling in LUSC cells. Therefore, the findings of this study indicated that the concurrent use of Erdafitinib and Stattic is a promising therapeutic approach for the treatment of FGFR1-positive LUSC.

The Combination of Afatinib With Dasatinib or Miransertib Results in Synergistic Growth Inhibition of Stomach Cancer Cells.

Background: Of various human epidermal growth factor receptor (HER) inhibitors, only the anti-HER2 monoclonal antibody (mAb) Herceptin/trastuzumab and the antibody-drug conjugate trastuzumab deruxtecan (T-Dxd) has been approved for the treatment of patients with stomach cancer. However, the duration of response may be short in many patients, with tumor heterogeneity being one contributing factor. Methods: We investigated the effect of various types of targeted agents on growth in vitro and migration of a panel of human stomach cancer cells (HSCCLs) and the impact of cell proliferation rate on the anti-tumor activities of these agents. We also investigated the association between the cell surface expression of the HER family members, hepatocyte growth factor receptor (c-Met), anaplastic lymphoma kinase (ALK)7 and cancer stem cell markers CD44 and CD133, and the response to the targeted agents. Results: Of the 18 agents examined, the cyclin dependent kinase (CDK) 1/2/5/9 inhibitor dinaciclib was the most effective and inhibited the growth of all human HSCCLs at 50% inhibitory concentration (IC50) values between 9 nM to 23 nM. Of various HER inhibitors, the irreversible pan-HER family inhibitors (e.g., afatinib) were more effective than the reversible dual epidermal growth factor receptor (EGFR)/HER2 tyrosine kinase inhibitor (TKI) lapatinib and the EGFR-specific TKI erlotinib in inhibiting the growth of HSCCLs. Of agents targeting different downstream cell signaling molecules, dasatinib targeting Ab1/Src/C-Kit, trametinib targeting MERK1/2 and miransertib targeting AKT1/2/3 inhibited growth of majority of HSCCLs, with the IC50 values ranging from 2 nM to 7 µM. Many of these agents were more effective in inhibiting the growth of HSCCLs when they were proliferating at a slower rate. Treatment with neratinib, afatinib, dinaciclib, dasatinib, stattic, miransertib and paclitaxel significantly inhibited migration of stomach cancer cells. Interestingly, treatment with a combination of afatinib and dasatinib or afatinib and miransertib resulted in synergistic and additive growth inhibition of stomach cancer cells. Conclusions: These results suggest that treatment with a combination of these agents may be of therapeutic value in stomach cancer and warrants further investigations.

Stattic ameliorates the cecal ligation and puncture-induced cardiac injury in septic mice via IL-6-gp130-STAT3 signaling pathway.

AIM: Sepsis-induced cardiac dysfunction is the leading cause of higher morbidity and mortality with poor prognosis in septic patients. Our recent previous investigation provides evidence of the hallmarks of signal transducer and activator of transcription3 (STAT3) activation in sepsis and targeting of STAT3 with Stattic, a small-molecule inhibitor of STAT3, has beneficial effects in various septic tissues. We investigated the possible cardioprotective effects of Stattic on cardiac inflammation and dysfunction in mice with cecal ligation and puncture (CLP)-induced sepsis. MAIN METHODS: A polymicrobial sepsis model was induced by CLP in mice and Stattic (25 mg/kg) was intraperitoneally given at one and twelve hours after CLP operation. The cecum was exposed in sham-control mice without CLP. After 18 h of surgery, electrocardiogram (ECG) for anaesthized mice was registered followed by collecting of samples of blood and tissues for bimolecular and histopathological assessments. Myeloperoxidase, a marker of neutrophil infiltration, was assessed immunohistochemically. KEY FINDINGS: CLP profoundly impaired cardiac functions as evidenced by ECG changes in septic mice as well as elevation of cardiac enzymes, and inflammatory markers with myocardial histopathological and immunohistochemical alterations. While, Stattic markedly reversed the CLP-induced cardiac abnormalities and restored the cardiac function by its anti-inflammatory activities. SIGNIFICANCE: Stattic treatment had potential beneficial effects against sepsis-induced cardiac inflammation, dysfunction and damage. Its cardioprotective effects were possibly attributed to its anti-inflammatory activities by targeting STAT3 and downregulation of IL-6 and gp130. Our investigations suggest that Stattic could be a promising target for management of cardiac sepsis and inflammation-related cardiac damage.

Stattic alleviates pulmonary fibrosis in a mouse model of rheumatoid arthritis-relevant interstitial lung disease.

Approximately 20% of rheumatoid arthritis (RA) patients have RA-related interstitial lung disease (RA-ILD). Stattic, an STAT3 inhibitor, has been confirmed to be relevant to both RA and ILD. Therefore, this study explored the effect of Stattic on the progression of joint disease and pulmonary fibrosis in zymosan-treated female SKG mice, an established model for autoimmune arthritis. The experimental mice developed pulmonary interstitial pneumonia, which is similar to human cellular and fibrotic nonspecific interstitial pneumonia. Oral gavage of Stattic (60 mg/kg/d) was initiated 10 weeks after zymosan injection. Arthritis and lung fibrosis outcome scores decreased significantly following Stattic treatment. An obvious decrease in lung collagen levels, measured using hydroxyproline level determination and collagen staining, was detected after 6 weeks in Stattic-exposed mice with established disease. Stattic also dramatically restricted arthritis progression, based on joint evaluation. Transforming growth factor beta 1 (TGF-ß1) is a pivotal fibrosis-causing cytokine, used here to treat myofibroblasts, thereby establishing a lung fibrosis cell model. Stattic treatment can mitigate the TGF-ß1-triggered inflammatory response, myofibroblast activation, oxidative stress, and hyperproliferation by modulating the JAK1/STAT3 pathway. Our observations support a direct role of Stattic-inhibited STAT3 activation in lung fibrosis, which may be particularly relevant in the RA-ILD context.