Signal Transducer And Activator 3 Research Articles

E-cadherin maintains the undifferentiated state of mouse spermatogonial progenitor cells via β-catenin

BackgroundCadherins play a pivotal role in facilitating intercellular interactions between spermatogonial progenitor cells (SPCs) and their surrounding microenvironment. Specifically, E-cadherin serves as a cellular marker of SPCs in many species. Depletion of E-cadherin in mouse SPCs showed no obvious effect on SPCs homing and spermatogenesis.ResultsHere, we investigated the regulatory role of E-cadherin in regulating SPCs fate. Specific deletion of E-cadherin in germ cells was shown to promote SPCs differentiation, evidencing by reduced PLZF+ population and increased c-Kit+ population in mouse testes. E-cadherin loss down-regulated the expression level of β-catenin, leading to the reduced β-catenin in nuclear localization for transcriptional activity. Remarkably, increasing expression level of Cadherin-22 (CDH22) appeared specifically after E-cadherin deletion, indicating CDH22 played a synergistic effect with E-cadherin in SPCs. By searching for the binding partners of β-catenin, Lymphoid enhancer-binding factor 1 (LEF1), T-cell factor (TCF3), histone deacetylase 4 (HDAC4) and signal transducer and activator 3 (STAT3) were identified as suppressors of SPCs differentiation by regulating acetylation of differentiation genes with PLZF.ConclusionsTwo surface markers of SPCs, E-cadherin and Cadherin-22, synergically maintain the undifferentiation of SPCs via the pivotal intermediate molecule β-catenin. LEF1, TCF3, STAT3 and HDAC4 were identified as co-regulatory factors of β-catenin in regulation of SPC fate. These observations revealed a novel regulatory pattern of cadherins on SPCs fate.

Dihydroartemisinin attenuates pulmonary inflammation and fibrosis in rats by suppressing JAK2/STAT3 signaling.

Coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2, has induced a worldwide pandemic since early 2020. COVID-19 causes pulmonary inflammation, secondary pulmonary fibrosis (PF); however, there are still no effective treatments for PF. The present study aimed to explore the inhibitory effect of dihydroartemisinin (DHA) on pulmonary inflammation and PF, and its molecular mechanism. Morphological changes and collagen deposition were analyzed using hematoxylin-eosin staining, Masson staining, and the hydroxyproline content. DHA attenuated early alveolar inflammation and later PF in a bleomycin-induced rat PF model, and inhibited the expression of interleukin (IL)-1β, IL-6, tumor necrosis factor α (TNFα), and chemokine (C-C Motif) Ligand 3 (CCL3) in model rat serum. Further molecular analysis revealed that both pulmonary inflammation and PF were associated with increased transforming growth factor-β1 (TGF-β1), Janus activated kinase 2 (JAK2), and signal transducer and activator 3(STAT3) expression in the lung tissues of model rats. DHA reduced the inflammatory response and PF in the lungs by suppressing TGF-β1, JAK2, phosphorylated (p)-JAK2, STAT3, and p-STAT3. Thus, DHA exerts therapeutic effects against bleomycin-induced pulmonary inflammation and PF by inhibiting JAK2-STAT3 activation. DHA inhibits alveolar inflammation, and attenuates lung injury and fibrosis, possibly representing a therapeutic candidate to treat PF associated with COVID-19.

Cathelicidin Modulates Vascular Smooth Muscle Cell Phenotypic Switching through ROS/IL-6 Pathway.

Vascular smooth muscle cells (VSMC) are stromal cells of the blood vessels and their differentiation is thought to be essential during atherosclerosis. Cathelicidin-related antimicrobial peptides (CRAMP) are suggested to play a role in the development of atherosclerosis. Even so, the relationship of CRAMP and VSMC remains unclear. The present study was to determine whether CRAMP regulates VSMC phenotypic transformation and underlying mechanisms. We demonstrated that CRAMP could reverse platelet-derived growth factor-BB (PDGF-BB)-induced VSMC phenotypic transformation, evidencing by increasing α-smooth muscle actin (α-SMA), smooth muscle 22α (SM22α) and decreasing of proliferation and migration. Further studies showed that CRAMP inhibited nuclear factor κB (NF-κB)-induced autocrine of interleukin-6 (IL-6), which further activated of janus kinase 2 (JAK2)/signal transducer and activator 3 (STAT3). Meanwhile, our data showed that CRAMP can significantly inhibit PDGF-BB enhanced intracellular reactive oxygen species (ROS) level which further affected the NF-κB signaling pathway, indicating that CRAMP can regulate the phenotypic transformation of VSMC by regulating oxidative stress. These results indicated that CRAMP regulated the differentiation of VSMC by inhibiting ROS-mediated IL-6 autocrine, suggesting that targeting CRAMP is a potential avenue for regulating the differentiation of VSMC and treatment of atherosclerosis.

The EGFR Liver Regeneration Pathway is Downregulated in Alcoholic Hepatitis

BackgroundAlcoholic liver disease (ALD) is a spectrum of progressive alcohol‐induced liver disorders including steatosis, steatohepatitis, and fibrosis. Alcoholic Hepatitis (AH), a clinical manifestation of severe liver damage, is associated with a hyper‐activated inflammatory immune response. The liver has an innate ability to regenerate after injury and inflammation. In severe liver diseases, including AH, this regenerative process becomes impaired. Not all the mechanisms are fully understood in the development/progression of AH, including the molecular events involved in alterations of liver tissue repair and regeneration. Recently, the epidermal growth factor receptor (EGFR) signaling pathway has been implicated in liver regeneration. The downstream effector, the E3 ubiquitin protein ligase, NEDD4, was demonstrated to be essential for EGFR internalization and signaling in vivo. NEDD4 facilitates EGFR internalization and downstream extracellular regulated kinase 1/2(ERK1/2) and signal transducer and activator 3 (STAT3) activity. The role of this pathway in liver regeneration in AH is not well defined.ObjectiveTo identify alterations of EGFR signaling in the liver of AH patients by proteomic and phosphoproteomic approach.MethodsLiquid chromatography and tandem mass spectrometry analysis were performed to measure proteomic and phosphoproteomic changes in liver tissue samples from AH patients (n=6, 44.3 ± 11.1 year old with MELD 37.2 ± 4.3, ABIC 9.61 ± 2.2, and Lille Score 0.57 ± 0.3), and age‐ and sex‐matched controls (individuals without ALD, n=12). A Student’s two‐tailed unpaired t‐test was used to determine statistical significance between the two groups (p<0.05). Linear regression analysis was performed between proteomic/phosphoproteomic data and markers of AH severity/prognosis (MELD, ABIC, and Lille Scores).ResultsEGFR protein was downregulated in AH relative to controls (1.2‐fold, p=0.02). Phosphorylation of EGFR at Y1069 was reduced 1.7‐fold, p=0.05 demonstrating decreased receptor activity. Hepatic EGFR was negatively associated with MELD (R2=0.2646), ABIC (R2=0.6719), and Lille Score (R2=0.4768). Hepatic NEDD4 was trending downward in AH relative to controls, and was negatively correlated with MELD (R2=0.8472), ABIC (R2=0.2982) and Lille Score (R2=0.52). Phosphorylated T202/Y204 relative to total ERK1 was decreased 1.5‐fold, p=0.0007 in AH relative to controls. Phospho‐ERK1 was also negatively associated with ABIC (R2=0.4523) and Lille Score (R2=0.4446). STAT3, the transcription factor that transcriptionally regulates proliferation genes in hepatocytes was decreased in AH relative to controls (1.2‐fold, p=0.01). Activation of STAT3 via phosphorylation of S714 was negatively correlated with ABIC (R2=0.8327, p=0.01) and Lille Score (R2=0.94, p<0.002).ConclusionCollectively, the data demonstrated that EGFR signaling was downregulated in AH patients which was negatively associated with disease severity. Downregulation of the EGFR pathway may potentially contribute to compromised liver regeneration in AH patients.Support or Funding InformationNIH NIAAA

Dietary Fat-Accelerating Leptin Signaling Promotes Protumorigenic Gastric Environment in Mice.

Excess of fat intake leads to obesity and causes a variety of metabolic diseases and cancer. We previously demonstrated that high-lard diet induces intestinal metaplasia, a precancerous lesion of the stomach mediated by leptin signaling. This study aims to investigate which kinds of dietary fat cause the intestinal metaplasia onset. We fed eight kinds of high-fat diets (HFDs) of animal or plant origin to mice evaluated their effect on gastric pathogenesis. Five types of dietary fat were divided according to their observed effects: Obese with high metaplasia (group I; beef tallow, lard, and hydrogenated coconut oil), non-obese with high metaplasia (group II; linseed oil), obese without metaplasia (group III; corn oil and olive oil), non-obese without metaplasia (group IV, soybean oil) and lean without metaplasia (group V; cocoa butter). The group I and II diets induced leptin, phosphorylated leptin receptor (ObR), signal transducer and activator 3 (STAT3), and increased intracellular β-catenin accumulation in the stomach. Moreover, mice fed these HFDs with 1-methyl-3-nitro-1-nitrosoguanidine (MNNG), a gastric carcinogen, and further accelerated dysplasia in the stomach. Lactobacillus occupancy in the stomach increased in all HFDs except hydrogenated coconut oil. Our findings suggest that HFDs inducing leptin signaling accelerate the enhancement of protumorigenic gastric microenvironment independent of body mass gain or microbiome changes.

Expression of biologically active bovine interleukin 7 and evaluating the activity in vitro.

Aim:Interleukin 7 (IL-7) is a ϒc family cytokine involved in the homeostatic proliferation and maintenance of immune cells. In the present study, we report the expression of bovine IL-7 (bIL-7) in Escherichia coli and evaluated for its biological activity.Materials and Methods:The sequence coding for bIL-7 (mature protein) was amplified from primary bovine kidney cell culture and cloned into pET28-a vector and expressed in E. coli (BL 21 DE3). The expressed protein was purified by nickel-nitrilotriacetatechromatography, and the reactivity of the protein was confirmed by western blotting using monoclonal antibodies raised against human IL-7. The biological activity of expressed bIL-7 was evaluated by analyzing its effect on the expression of anuclear factor for activated T-cells c1 (NFATc1), B-cell lymphoma 2 (Bcl2), suppressor of cytokine signaling 3 (SOCS3) molecules in bovine peripheral blood mononuclear cells (PBMCs) by quantitative polymerase chain reaction. Ability of the expressed protein was also analyzed by its effect on phosphorylating signal transducer and activator 3 (STAT3) molecule by immunostaining in human embryonic kidney cells 293 (HEK293) cells.Results:The bIL-7 was able to induce the expression of Bcl2 and NFATc1expression in bovine PBMCs by 7 and 5-folds, respectively, whereas a 2-fold decrease was observed in the case of SOCS3 expression. Immunostaining studies in HEK293 cells using antihuman phospho-STAT3 showed activation and nuclear translocation of STAT3 molecule on bIL-7 treatment.Conclusion:bIL-7 gene was successfully amplified, cloned, and expressed in a prokaryotic expression system. The biological activity study showed that the E. coli expressed bIL-7 protein is biologically active. Considering the role of IL-7 in T-cell homeostasis and memory cell generation, this molecule can be used for enhancing the vaccine response and that has to be proved by further experiments.

Genome-wide screen identified let-7c/miR-99a/miR-125b regulating tumor progression and stem-like properties in cholangiocarcinoma.

Cholangiocarcinoma (CCA), which is a poor prognosis malignancy that arises from the malignant transformation of cholangiocytes, is associated with chronic inflammation of the biliary epithelium. Thus far, the molecular mechanisms of the origin and neoplastic processes of CCA that are promoted by inflammation are still unclear and need to be fully elucidated. Here using small RNA sequencing to determine the microRNA (miRNA) expression profiles in CCA, we found that let-7c, miR-99a and miR-125b, which are three miRNAs of the same cluster, were downregulated in CCA and targeted interleukin 6 (IL-6), IL-6R and type 1 insulin-like growth factor, which are important cytokines and receptors of the IL-6/signal transducer and activator 3 (STAT3) pathway and have key roles in inflammation and CCA initiation. We also found that enforced expression of let-7c, miR-99a or miR-125b could reduce the activity of STAT3 and further suppress CCA tumorigenicity in vivo and inhibit the migration and invasion of CCA cells in vitro. Surprisingly, let-7c/miR-99a/miR-125b cluster also significantly decreased the ability of CCA cells for cancer stem cell-like mammosphere generation by downregulating CD133 and CD44, which suggests the pivotal roles of let-7c, miR-99a and miR-125b in CCA by regulating both inflammation and stem-like properties. Our findings showed potential links between miRNAs and inflammation, and provide a potential treatment strategy for developing an miRNA-based therapy via IL-6/STAT3 targeting for CCA.

Abstract B15: Pediatric glioblastoma cell lines maintain response to developmental growth factors

Abstract Glioblastoma (GBM) is a highly invasive brain tumor arising from an astrocytic cell lineage. In the United States, nearly 500 children each year develop GBMs. Pediatric GBM has a different mutational, methylation, and gene expression spectra than adult GBM, implicating a strong role for developmental processes in disease pathogenesis. The developmental growth factors that define the astrocytic lineage include epidermal growth factor (EGF) and platelet-derived growth factor (PDGF). Expression of these growth factors also distinguishes pediatric versus adult GBM. Here, we use EGF or PDGF as a stimulus for malignant behavior in GBM cell lines isolated from either adult or pediatric patients to understand whether the cell line models parallel growth factor receptor signaling preferences observed clinically. In response to these stimuli, pediatric cell lines demonstrate distinct protein signaling and invasion phenotypes from adult cell lines in vitro, and generate invasive, multifocal lesions in an orthotopic xenograft model. Thus, distinguishing features of adult and pediatric GBM are maintained in the cell line models that will be used to test therapeutic regiments preclinically. Both of the receptors activated by EGF and PDGF are known to interact with a promising therapeutic target in adult glioblastoma: signal transducer and activator 3 (STAT3). STAT3 acts as either an oncogene or tumor suppressor depending on cellular context such as the activating receptor complex and phosphatase and tensin homolog (PTEN) mutational status. Our studies will therefore address the differential implications of STAT3 therapy for pediatric versus adult GBM. Citation Format: Marie Renee Mooney. Pediatric glioblastoma cell lines maintain response to developmental growth factors. [abstract]. In: Proceedings of the AACR Special Conference on Pediatric Cancer at the Crossroads: Translating Discovery into Improved Outcomes; Nov 3-6, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;74(20 Suppl):Abstract nr B15.

Abstract C229: Targeting STAT3 in prostate cancer: Identification of STAT3 as a direct target of the fungal metabolite galiellalactone.

Abstract Introduction: The transcription factor Signal Transducer and Activator 3 (STAT3) is known to affect tumor growth in several malignancies. STAT3 is frequently found to be activated by phosphorylation (p-STAT3) in castration-resistant prostate cancer and constitutes a promising therapeutic target. The fungal metabolite galiellalactone (GL), a potent STAT3-signaling inhibitor, inhibits the growth, both in vitro and in vivo, of prostate cancer cells expressing activated STAT3 and inhibits growth and induces apoptosis of prostate cancer stem cell-like cells expressing p-STAT3. GL inhibits STAT3 signaling without blocking phosphorylation by a mechanism that we here aim to elucidate. Materials and Methods: A biotinylated analogue of galiellalactone (GL-biot) was synthesized for identification of GL target proteins. The p-STAT3 expressing human prostate cancer cell line DU145 was incubated with GL-biot, followed by streptavidin beads to isolate proteins bound to GL-biot. Bound proteins were identified using Western blot analysis. Competition assays were performed by pre-incubating DU145 cell lysates with GL prior to GL-biot treatment. Localization of GL-biot and co-localization of GL-biot with STAT3 was assessed using confocal microscopy. Electrophoretic mobility shift assay (EMSA) was performed on GL treated DU145 cell lysates for analysis of STAT3 binding to DNA. Results: The inhibitory activity of GL-biot on proliferation and inhibition of STAT3-signaling was similar to that of GL. By adding streptavidin beads to GL-biot-treated DU145 cell lysates, STAT3 was isolated and identified as a target protein. Pretreatment with GL prior to addition of GL-biot prevented the binding of GL-biot to STAT3 in a dose dependent manner. GL-biot was observed to interact also with STAT5 but not STAT1. Confocal microscopy revealed GL-biot in both the cytoplasm and nucleus of DU145 cells treated with GL-biot, appearing to co-localize with STAT3 in the nucleus. DU145 cell lysates subjected to EMSA with STAT3 probes showed a dose dependent decrease in STAT3-DNA binding when treated with GL. Conclusions: Our results show that GL binds directly to STAT3 and inhibits DNA binding thus preventing the transcriptional activity of STAT3. This further validates GL as a promising STAT3-inhibitor for the treatment of castration-resistant prostate cancer. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):C229. Citation Format: Nicholas Don-Doncow, Zilma Escobar, Martin Johansson, Eduardo Muñoz, Olov Sterner, Anders Bjartell, Rebecka Hellsten. Targeting STAT3 in prostate cancer: Identification of STAT3 as a direct target of the fungal metabolite galiellalactone. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr C229.

Importance of performance status, corrected calcium, and microvascular invasion in predicting overall survival in patients with metastatic renal cell carcinoma at the era of molecular targeted therapy.

e15588 Background: We have recently reported that the single nucleotide polymorphisms (SNPs) in signal transducer and activator 3 (STAT3) gene are significantly associated with better response to interferon (IFN)-α in patients with metastatic renal cell carcinoma (mRCC) in our prospective trail (Eur Urol 2012, Epub ahead). In the present study, we further evaluated which clinical and pathological factors correlated with overall survival (OS) in the trial. Methods: In the multicenter, prospective study, patients with histologically confirmed RCC that was metastatic, measurable disease, age > 20 years, ECOG performance status (PS) 0-1 and adequate organ function received 3 dosages of 5 million U per week of IFN-α treatment. After the failure of IFN-α, molecular targeted drugs were used. We followed the patients for 2 years to define the date of death or loss to follow-up to calculate OS. Association between OS and clinical, pathological factors, such as age, PS, TNM classification, grade, infiltrating type, sarcomatoid component, microvascular invasion, lactate dehydrogenase (LDH), corrected Calcium (Ca), hemoglobin (Hb), C-reactive protein (CRP), platelet (Plt), etc, was analyzed using the Log-rank test. For multivariate analysis, the Stepwise test was used. Results: Two hundreds three eligible pts were enrolled between December 2006 and October 2009. All pts had prior nephrectomy, and 88.7% had ECOG PS 0. Ninety-four percent of patients had clear cell RCC, and 5% had papillary RCC. The response rate of IFN-α was 13.8% (28/203, 9 CR, 19 PR). Median OS was 42.8 months in mRCC patients initially treated with IFN-α. In univariate analysis, PS, TNM classification, grade, infiltrating type, microvascular invasion, corrected Ca, Hb, CRP, and Plt were significantly associated with OS. Among them, PS, microvascular invasion, and corrected Ca were significantly correlated with OS in multivariate analysis. Conclusions: This prospective study demonstrates that PS, corrected Ca, and microvascular invasion are important in predicting OS in mRCC patients initially treated with IFN-α at the era of molecular targeted therapy. Clinical trial information: UMIN000000731.

YB-1 suppression induces STAT3 proteolysis and sensitizes renal cancer to interferon-α.

Renal cell carcinoma (RCC) accounts for 80-95% of kidney tumors, and approximately 30% of RCC patients have metastatic disease at diagnosis. Conventional chemotherapy is not effective in patients with metastatic RCC (MRCC); therefore, immunotherapy with interferon-α (IFN-α) has been employed to improve survival. However, the response rate of MRCC to IFN-α therapy is low. We previously reported that a signal transducer and activator 3 (STAT3) polymorphism was a useful diagnostic marker to predict the response to IFN-α therapy in patients with MRCC. Therefore, we hypothesized the inhibition of STAT3 in the addition of IFN-α therapy might be useful. Moreover, the blockage of STAT3 itself has been reported to enhance the antitumor effects. However, because IFN-α is thought to elicit its therapeutic effect via enhancement of an antitumor immune response mediated by lymphocytes that can be activated by IFN-α administrations, it is probable that the suppression of STAT3 in vivo relates to autoimmune disorders. In the present study, we found Y-box binding protein-1 (YB-1) was poorly expressed in T lymphocytes, as compared with cancer tissues. YB-1 was reported to have an important effect on the STAT3 pathway. Suppression of STAT3 by YB-1 inhibition did not seem to enhance the potential risk for autoimmune disorders. Moreover, we found sensitivity to IFN-α was increased by YB-1 suppression, and this suppression did not down-regulate IFN-α activation of T lymphocytes.

Use of STAT3 polymorphisms to predict overall survival in patients treated with interferon-alpha for metastatic renal cell carcinoma.

e15048 Background: We previously reported that the single nucleotide polymorphisms (SNPs) in signal transducer and activator 3 (STAT3) gene were most significantly associated with better response to interferon (IFN)-α in patients with metastatic renal cell carcinoma (mRCC) in our retrospective analysis (JCO 25:2785, 2007). We conducted this trial to prospectively confirm the results, and reported the significant association between the SNPs of STAT3-2 and clinical benefits (CR, PR, and SD more than 24 weeks) of IFN-α (p = 0.039) (ASCO 2011, abstract No. 4590). In this study, we have further analyzed the correlation between overall survival (OS), progression-free survival (PFS) and the 11 SNPs reported previously. Methods: In this multicenter, prospective study, patients with histologically confirmed RCC that was metastatic, measurable disease, age > 20 years, ECOG PS 0-1 and adequate organ function received 3 dosages of 5 million U per week of IFN-α treatment. We evaluated the correlation between OS, PFS and SNP allele frequencies of STAT3-2, STAT3-0, SOCS3-1, IL4R-34, PTGS1-3, PTGS1-4, PTGS1-5, PTGS2-12, IRF2-67, ICSBP-38, and TAP2-5 (JCO 25:2785, 2007) in mRCC patients treated with IFN-α. Association between OS, PFS and genetic polymorphisms was analyzed using Logrank test for trend. Results: Two hundreds three eligible patients were enrolled between December 2006 and October 2009. All patients had prior nephrectomy, and 88.7% had ECOG PS 0. Ninety-four percent of patients had clear cell RCC, and 5% had papillary RCC. At the time of this analysis the central review assessed response rate was 13.8% (28/203) (9 CR, 19 PR). CR rate of 4.4% (9/203) was more than we expected. PFS was not associated with any of the 11 SNPs examined. However, 2 SNPs of STAT3 gene, STAT3-2 and STAT3-0, were significantly associated with OS, and the correlation of STAT3-2 (p = 0.0347) was stronger than that of STAT3-0. Namely, C/C genotype of STAT3-2 was significantly associated with prolonged OS compared with the other genotypes (C/T and T/T). Conclusions: This is the first prospective study demonstrating that STAT3 polymorphisms can predict OS in mRCC patients treated with IFN-α.

An analysis of STAT3 polymorphism on outcomes of interferon-alpha treatment in patients with metastatic renal cell carcinoma.

4590 Background: We previously reported that the single nucleotide polymorphisms (SNPs) in signal transducer and activator 3 (STAT3) gene were most significantly associated with better response to interferon (IFN)-α in pts with metastatic renal cell carcinoma (mRCC) in our retrospective analysis (JCO 25:2785, 2007). Japan Immunotherapy SNPs-Study Group for Kidney Cancer (JISG-KC) conducted this trial to prospectively confirm the results. Methods: In this multicenter, prospective study, pts with histologically confirmed RCC that was metastatic, measurable disease, age > 20 years, ECOG PS 0-1 and adequate organ function received 3 dosages of 5 million U per week of IFN-α treatment. The primary end point of the study was to evaluate the correlation between the antitumor effects of IFN-α and SNP allele frequencies of STAT3-2. The secondary end point was to evaluate correlation between the antitumor effects of IFN-α and SNP allele frequencies of STAT3-0, SOCS3-1, IL4R-34, PTGS1-3, PTGS1-4, PTGS1-5, PTGS2-12, IRF2-67, ICSBP-38, and TAP2-5 (JCO 25:2785, 2007). Association between response to IFN-α and genetic polymorphism was analyzed using the logistic model. All pts consented to provide blood samples that would be used for SNPs analysis. Results: Two hundreds four eligible pts were enrolled between December 2006 and October 2009. All pts had prior nephrectomy, and 88.7% had ECOG PS 0. Ninety-four percent of patients had clear cell RCC, and 5% had papillary RCC. At the time of this analysis the central review assessed response rate was 13.7% (28/204) (7 CR, 21 PR). CR rate of 3.4% (7/204) was more than we expected. Response (CR, PR) to IFN-α was not associated with any of the 11 SNPs examined. However, when we assessed pts with CR, PR, and SD more than 24 wks as those with clinical benefits and reevaluated the correlation to the 11 SNPs, the significant association between STAT3-2 and clinical benefits of IFN-α (p = 0.039) was observed. Namely, C/C genotype of STAT3-2 was significantly associated with clinical benefits of IFN-α. Conclusions: This is the first prospective study demonstrating that STAT3 polymorphism can predict clinical benefits of IFN-α in pts with mRCC.

STAT3 Polymorphism Predicts Interferon-Alfa Response in Patients With Metastatic Renal Cell Carcinoma

To clarify the effect of genetic polymorphisms on the response to interferon alfa (IFN-alpha) for metastatic renal cell carcinoma (MRCC), and to find a reliable molecular marker to select those patients with MRCC who would benefit from IFN-alpha immunotherapy. We carried out an association study in which 463 single nucleotide polymorphisms (SNPs) in 33 candidate genes were genotyped in 75 Japanese patients who had received IFN-alpha for MRCC. After adjusting for lung metastasis, stepwise logistic regression analysis revealed that the SNPs in signal transducer and activator 3 (STAT3) were most significantly associated with better response to IFN-alpha. Linkage disequilibrium mapping revealed that the SNP in the 5' region of STAT3, rs4796793, was the most significant predictor of IFN-alpha response (odds ratio [OR] = 2.73; 95% CI, 1.38 to 5.78). The highest OR was shown in the CC genotype at rs4796793 compared to the GG + GC genotypes (OR = 8.38, 95% CI, 1.63 to 42.96). Genotype-dependent expressions of STAT3 in B lymphocyte cell lines and the enhanced growth inhibitory effects of IFN- by STAT3 suppression in an RCC cell line supported the results of the present association study. The present study suggested that the STAT3 polymorphism is a useful diagnostic marker to predict the response to IFN-alpha therapy in patients with MRCC. An efficient response marker for IFN-alpha needs to be utilized to establish individual optimal treatment strategies, even when newer drug therapies are used as first line treatments for MRCC.

Reciprocal Signaling Roles for STAT3 and SHP2

STKE One subunit of the receptor for the cytokine interleukin-6 (IL-6) is gp130. Tyrosine phosphorylation of gp130 recruits both protein phosphatase 2 (SHP2) and signal transducer and activator 3 (STAT3). Ohtani et al. utilized knock-in technology to create mice deficient in only the gp130-mediated SHP2 pathway or the gp130-mediated STAT3 pathway to identify the functions of these two pathways. STAT3 signaling was required for embryonic development, whereas SHP2 signaling-deficient mice were viable and fertile. Reciprocal roles for signaling via STAT3 (positive) and SHP2 (negative) were found for cytokine production in T cells and immunoglobulin responses in B cells, consistent with the observation that eliminating the SHP2 pathway resulted in prolonged IL-6-induced STAT3 phosphorylation.— NG Immunity 12 , 95 (2000).

Reciprocal Signaling Roles for STAT3 and SHP2

One subunit of the interleukin-6 (IL-6) receptor, which activates Janus kinases upon ligand binding, is gp130. Tyrosine phosphorylation of gp130 recruits protein phosphatase 2 (SHP2) and signal transducer and activator 3 (STAT3). Ohtani et al . utilized knock-in technology to create mice deficient in only the gp130-mediated SHP2 pathway or the gp130-mediated STAT3 pathway to dissect the roles these two pathways play in IL-6 signaling. Eliminating the SHP2 response resulted in prolonged IL-6-induced STAT3 phosphorylation. STAT3 signaling was required for embryonic development, where as SHP2 signaling-deficient mice were viable and fertile. STAT3 signaling was required for induction of astrocytes by IL-6 cytokines. Reciprocal roles for STAT3 signaling and SHP2 signaling (STAT3 providing a positive signal and SHP2 providing a negative signal) were found for cytokine and immunoglobulin responses and lymphoid organ homeostasis. Thus, mice deficient in SHP2 signaling developed splenomegaly and lymphadenopathy, and demonstrated increased induction of acute phase proteins and increased antibody production by B cells in response to thymus dependent antigens. Ohtani, T., Ishahara, K., Atsumi, T., Nishida, K., Kaneko, Y., Miyata, T., Itoh, S., Narimatsu, M., Maeda, H., Fukada, T., Itoh, M., Okano, H., Hibi, M., and Hirano, T. (2000) Dissection of signaling cascades through gp130 in vivo: Reciprocal roles for STAT3- and SHP2-mediated signals in immune responses. Immunity 12: 95-105. [Online Journal]