Autocrine Signaling Loop Research Articles

ATRT-12. EXPLORING FGFR INHIBITION AND COMBINATION THERAPIES FOR ATYPICAL TERATOID/RHABDOID TUMORS

Abstract BACKGROUND Atypical teratoid/rhabdoid tumors (ATRTs) are aggressive pediatric brain neoplasms characterized by an inactivation of the SWI/SNF chromatin remodeling complex members SMARCB1 or SMARCA4. Based on epigenetic profiling ATRTs can be divided into four different molecular subgroups. Despite improvements in treatment strategies, the prognosis for ATRT patients remains dismal. The FGF/FGFR signaling pathway plays a crucial role in many physiological processes, including embryonic development, tissue repair, angiogenesis. Dysregulation of the signaling axis is driving tumorigenesis. Targeted therapeutic approaches, particularly FGFR inhibitors, have demonstrated potential in enhancing treatment outcomes for various tumors, including pediatric brain tumors. METHODS We used bulk and single-cell RNA sequencing (scRNA-seq) data to examine the expression patterns of FGF family members in ATRTs. Subsequently, we conducted in vitro analysis on a panel of FGFR inhibitors and combination treatments. RESULTS Transcriptomic analysis of more than 130 ATRT tissues samples, revealed a widespread expression of FGFR1 and FGFR2 across all ATRT subtypes. Notably, FGFR1 displayed distinctly elevated expression in ATRT-SHH, surpassing levels seen in other ATRT subtypes and brain tumor types. Utilizing scRNA-seq data of murine ATRT models, we elucidated an autocrine FGFR signaling loop. To assess the therapeutic feasibility of targeting FGFR, we evaluated a panel of FGFR inhibitors, currently in clinical development, across 10 in house established patient-derived cell models (6 ATRT-MYC, 3 ATRT-SHH, 1 ATRT-TYR), revealing sensitivity in a nanomolar to a low-micromolar range. We further explored combination therapies, demonstrating that Erdafitinib, the most effective inhibitor when combined with Abemaciclib and radiation, exhibited enhanced synergistic effects in 2D as well as in 3D tumoroid models. Investigations towards testing these combinations in an organoid co-culture system, along with in vivo models are ongoing. CONCLUSIONS This study emphasizes the prospect of combining FGFR inhibition with radiation and CDK4/6 inhibition as a therapeutic target for ATRTs.

Novel VEGFR-2 inhibitors as antiangiogenic and apoptotic agents via paracrine and autocrine cascades: Design, synthesis, and biological evaluation

Appertaining to its paracrine and autocrine signaling loops, VEGFR-2 succeeded in grabbing attention as one of the leading targets in cancer treatment. Based on the foregoing and our comprehensive studies regarding pharmacophoric features and activity of sorafenib, novel phenylpyridazinone based VEGFR-2 inhibitors 4, 6a-e, 7a,b, 9a,b, 12a-c, 13a,b, 14a,b, 15a,b, and 17a-d were optimized. An assortment of biological assays was conducted to assess the antiangiogenic and apoptotic activities of the synthesized derivatives. In vitro VEGFR-2 kinase assay verified the inhibitory activity of the synthesized derivatives with IC50 values from 49.1 to 418.0 nM relative to the reference drug sorafenib (IC50 = 81.8 nM). Antiproliferative activity against HUVECs revealed that compounds 2-{2-[2-(6-oxo-3-phenylpyridazin-1(6H)-yl)acetyl]hydrazineyl}-N-(p-tolyl)acetamide (12c) and 2-[(5-mercapto-4-methyl-4H-1,2,4-triazol-3-yl)methyl]-6-phenylpyridazin-3(2H)-one (13a) possessed superior activity (IC50 values = 11.5 and 12.3 nM, respectively) in comparison to sorafenib (IC50 = 23.2 nM). For the purpose of appraising their antiproliferative effect, derivatives 12c and 13a were exposed to cell cycle analysis, apoptotic, cell invasion and migration assays in addition to determination of VEGFR-2 in protein level. Moreover, cytotoxicity as well as selectivity index against WI-38 cell line was measured to examine safety of derivatives 12c and 13a. After that, molecular docking study was executed on the top five compounds in the in vitro VEGFR-2 kinase assay 6d, 12c, 13a, 14a and 17c to get a deep perception on binding mode of the synthesized compounds and correlate the design strategy with biological results. Finally, physicochemical, pharmacokinetic properties, and drug-likeness studies were performed on the top five derivative in in vitro VEGFR-2 kinase assay.

SLAMF7 engagement superactivates macrophages in acute and chronic inflammation.

Macrophages regulate protective immune responses to infectious microbes, but aberrant macrophage activation frequently drives pathological inflammation. To identify regulators of vigorous macrophage activation, we analyzed RNA-seq data from synovial macrophages and identified SLAMF7 as a receptor associated with a superactivated macrophage state in rheumatoid arthritis. We implicated IFN-γ as a key regulator of SLAMF7 expression and engaging SLAMF7 drove a strong wave of inflammatory cytokine expression. Induction of TNF-α after SLAMF7 engagement amplified inflammation through an autocrine signaling loop. We observed SLAMF7-induced gene programs not only in macrophages from rheumatoid arthritis patients but also in gut macrophages from patients with active Crohn's disease and in lung macrophages from patients with severe COVID-19. This suggests a central role for SLAMF7 in macrophage superactivation with broad implications in human disease pathology.

The Role of c-Jun and Autocrine Signaling Loops in the Control of Repair Schwann Cells and Regeneration.

After nerve injury, both Schwann cells and neurons switch to pro-regenerative states. For Schwann cells, this involves reprogramming of myelin and Remak cells to repair Schwann cells that provide the signals and mechanisms needed for the survival of injured neurons, myelin clearance, axonal regeneration and target reinnervation. Because functional repair cells are essential for regeneration, it is unfortunate that their phenotype is not robust. Repair cell activation falters as animals get older and the repair phenotype fades during chronic denervation. These malfunctions are important reasons for the poor outcomes after nerve damage in humans. This review will discuss injury-induced Schwann cell reprogramming and the concept of the repair Schwann cell, and consider the molecular control of these cells with emphasis on c-Jun. This transcription factor is required for the generation of functional repair cells, and failure of c-Jun expression is implicated in repair cell failures in older animals and during chronic denervation. Elevating c-Jun expression in repair cells promotes regeneration, showing in principle that targeting repair cells is an effective way of improving nerve repair. In this context, we will outline the emerging evidence that repair cells are sustained by autocrine signaling loops, attractive targets for interventions aimed at promoting regeneration.

Biology of the dopamine transporter on human macrophages

Neuromodulators, such as norepinephrine and dopamine, regulate immune responses. In addition to receptor competency, select immune cells express machinery to engage in active neurotransmitter transmission. Specifically, macrophages express the norepinephrine (NET) and dopamine transporters (DAT). However, how these transporters - classically studied in central neurons - affect immunity remains poorly understood. To this end, we investigated the expression and function of NET and DAT in human macrophages. We found that monocyte-derived macrophages express functional NET and DAT in vitro. A subset of human intestinal macrophages were further confirmed to express DAT in situ. Interestingly, we discovered that inhibiting DAT, but not NET, enhanced the pro-inflammatory macrophage program in response to an immune challenge. We attribute this modulation to an observed LPS-induced efflux of dopamine through DAT that engages an autocrine dopamine signaling loop. Collectively, we propose that DAT actively regulates dopamine signaling on immune cells, and, in this context, DAT on macrophages is a novel, dynamic immunomodulator.

PMN-derived netrin-1 attenuates cardiac ischemia-reperfusion injury via myeloid ADORA2B signaling.

Previous studies implicated the neuronal guidance molecule netrin-1 in attenuating myocardial ischemia-reperfusion injury. However, the tissue-specific sources and receptor signaling events remain elusive. Neutrophils are among the first cells responding to an ischemic insult and can be associated with tissue injury or rescue. We found netrin-1 levels were elevated in the blood of patients with myocardial infarction, as well as in mice exposed to myocardial ischemia-reperfusion. Selectively increased infarct sizes and troponin levels were found in Ntn1loxP/loxP Lyz2 Cre+ mice, but not in mice with conditional netrin-1 deletion in other tissue compartments. In vivo studies using neutrophil depletion identified neutrophils as the main source for elevated blood netrin-1 during myocardial injury. Finally, pharmacologic studies using treatment with recombinant netrin-1 revealed a functional role for purinergic signaling events through the myeloid adenosine A2b receptor in mediating netrin-1-elicited cardioprotection. These findings suggest an autocrine signaling loop with a functional role for neutrophil-derived netrin-1 in attenuating myocardial ischemia-reperfusion injury through myeloid adenosine A2b signaling.

Inhibition of relaxin autocrine signaling confers therapeutic vulnerability in ovarian cancer.

Ovarian cancer (OC) is the most deadly gynecological malignancy, with unmet clinical need for new therapeutic approaches. The relaxin peptide is a pleiotropic hormone with reproductive functions in the ovary. Relaxin induces cell growth in several types of cancer, but the role of relaxin in OC is poorly understood. Here, using cell lines and xenograft models, we demonstrate that relaxin and its associated GPCR RXFP1 form an autocrine signaling loop essential for OC in vivo tumorigenesis, cell proliferation, and viability. We determined that relaxin signaling activates expression of prooncogenic pathways, including RHO, MAPK, Wnt, and Notch. We found that relaxin is detectable in patient-derived OC tumors, ascites, and serum. Further, inflammatory cytokines IL-6 and TNF-α activated transcription of relaxin via recruitment of STAT3 and NF-κB to the proximal promoter, initiating an autocrine feedback loop that potentiated expression. Inhibition of RXFP1 or relaxin increased cisplatin sensitivity of OC cell lines and abrogated in vivo tumor formation. Finally, we demonstrate that a relaxin-neutralizing antibody reduced OC cell viability and sensitized cells to cisplatin. Collectively, these data identify the relaxin/RXFP1 autocrine loop as a therapeutic vulnerability in OC.

Game of Bones: How Myeloma Manipulates Its Microenvironment.

Multiple myeloma is a clonal disease of long-lived plasma cells and is the second most common hematological cancer behind Non-Hodgkin’s Lymphoma. Malignant transformation of plasma cells imparts the ability to proliferate, causing harmful lesions in patients. In advanced stages myeloma cells become independent of their bone marrow microenvironment and form extramedullary disease. Plasma cells depend on a rich array of signals from neighboring cells within the bone marrow for survival which myeloma cells exploit for growth and proliferation. Recent evidence suggests, however, that both the myeloma cells and the microenvironment have undergone alterations as early as during precursor stages of the disease. There are no current therapies routinely used for treating myeloma in early stages, and while recent therapeutic efforts have improved patients’ median survival, most will eventually relapse. This is due to mutations in myeloma cells that not only allow them to utilize its bone marrow niche but also facilitate autocrine pro-survival signaling loops for further progression. This review will discuss the stages of myeloma cell progression and how myeloma cells progress within and outside of the bone marrow microenvironment.

Autocrine TGF-β in Cancer: Review of the Literature and Caveats in Experimental Analysis.

Autocrine signaling is defined as the production and secretion of an extracellular mediator by a cell followed by the binding of that mediator to receptors on the same cell to initiate signaling. Autocrine stimulation often operates in autocrine loops, a type of interaction, in which a cell produces a mediator, for which it has receptors, that upon activation promotes expression of the same mediator, allowing the cell to repeatedly autostimulate itself (positive feedback) or balance its expression via regulation of a second factor that provides negative feedback. Autocrine signaling loops with positive or negative feedback are an important feature in cancer, where they enable context-dependent cell signaling in the regulation of growth, survival, and cell motility. A growth factor that is intimately involved in tumor development and progression and often produced by the cancer cells in an autocrine manner is transforming growth factor-β (TGF-β). This review surveys the many observations of autocrine TGF-β signaling in tumor biology, including data from cell culture and animal models as well as from patients. We also provide the reader with a critical discussion on the various experimental approaches employed to identify and prove the involvement of autocrine TGF-β in a given cellular response.

TPL2 enforces RAS-induced inflammatory signaling and is activated by point mutations.

NF-κB transcription factors, driven by the IRAK/IKK cascade, confer treatment resistance in pancreatic ductal adenocarcinoma (PDAC), a cancer characterized by near-universal KRAS mutation. Through reverse-phase protein array and RNA sequencing we discovered that IRAK4 also contributes substantially to MAPK activation in KRAS-mutant PDAC. IRAK4 ablation completely blocked RAS-induced transformation of human and murine cells. Mechanistically, expression of mutant KRAS stimulated an inflammatory, autocrine IL-1β signaling loop that activated IRAK4 and the MAPK pathway. Downstream of IRAK4, we uncovered TPL2 (also known as MAP3K8 or COT) as the essential kinase that propels both MAPK and NF-κB cascades. Inhibition of TPL2 blocked both MAPK and NF-κB signaling, and suppressed KRAS-mutant cell growth. To counter chemotherapy-induced genotoxic stress, PDAC cells upregulated TLR9, which activated prosurvival IRAK4/TPL2 signaling. Accordingly, a TPL2 inhibitor synergized with chemotherapy to curb PDAC growth in vivo. Finally, from TCGA we characterized 2 MAP3K8 point mutations that hyperactivate MAPK and NF-κB cascades by impeding TPL2 protein degradation. Cancer cell lines naturally harboring these MAP3K8 mutations are strikingly sensitive to TPL2 inhibition, underscoring the need to identify these potentially targetable mutations in patients. Overall, our study establishes TPL2 as a promising therapeutic target in RAS- and MAP3K8-mutant cancers and strongly prompts development of TPL2 inhibitors for preclinical and clinical studies.

Abstract B25: An actionable AXL-ABL2-TAZ signaling axis promotes lung adenocarcinoma metastasis to the brain

Abstract Despite recent work to uncover the biologic mechanisms governing tumor cell metastasis to the brain, new therapeutic approaches for treating patients with brain-metastatic disease have remained elusive. Through the use of in vivo models of brain metastasis and relevant in vitro molecular biology techniques, we identify a novel and targetable autocrine signaling axis required for lung adenocarcinoma metastasis to the brain centered upon the transcriptional coactivator TAZ. Nuclear localization of TAZ drives transcription of target genes AXL and ABL2, which in turn engage in bidirectional tyrosine kinase signaling at the protein level. Activation of ABL2 in turn increases TAZ stability and promotes its nuclear localization, establishing an autocrine feed-forward signaling loop and revealing a previously undescribed mechanism of TAZ nuclear localization dependent on tyrosine kinase activity of ABL2. We further reveal that the Neural Adhesion Molecule L1 (L1CAM), previously identified as a critical mediator of brain metastasis, is a direct target gene of TAZ whose expression is regulated by activation of AXL-ABL2-TAZ signaling. Our findings provide evidence for AXL and ABL2 kinase inhibitors as potential therapeutic strategies for the treatment of brain metastases. Citation Format: Jacob P. Hoj, Benjamin J. Mayro, Ann Marie Pendergast. An actionable AXL-ABL2-TAZ signaling axis promotes lung adenocarcinoma metastasis to the brain [abstract]. In: Proceedings of the AACR Special Conference on the Hippo Pathway: Signaling, Cancer, and Beyond; 2019 May 8-11; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Res 2020;18(8_Suppl):Abstract nr B25.

ADAMDEC1 and FGF2/FGFR1 signaling constitute a positive feedback loop to maintain GBM cancer stem cells

ABSTRACTIdentification of targetable mechanisms that maintain glioblastoma cancer stem cells (CSCs) remain a priority. Our study reveals a new mechanism by which a disintegrin and metalloproteinase domain-like protein decysin 1 promotes CSC maintenance through the activation of a fibroblast growth factor autocrine signaling loop, which can be blocked pharmacologically.

CD40L reverse signaling suppresses prevertebral sympathetic axon growth and tissue innervation.

CD40‐activated CD40L reverse signaling is a major physiological regulator of the growth of neural processes in the developing nervous system. Previous work on superior cervical ganglion (SCG) neurons of the paravertebral sympathetic chain has shown that CD40L reverse signaling enhances NGF‐promoted axon growth and tissue innervation. Here we show that CD40L reverse signaling has the opposite function in prevertebral ganglion (PVG) sympathetic neurons. During a circumscribed perinatal window of development, PVG neurons cultured from Cd40 –/– mice had substantially larger, more exuberant axon arbors in the presence of NGF than PVG neurons cultured from wild‐type mice. Tissues that receive their sympathetic innervation from PVG neurons were markedly hyperinnervated in Cd40 –/– mice compared with wild‐type mice. The exuberant axonal growth phenotype of cultured CD40‐deficient perinatal PVG neurons was pared back to wild‐type levels by activating CD40L reverse signaling with a CD40‐Fc chimeric protein, but not by activating CD40 forward signaling with CD40L. The co‐expression of CD40 and CD40L in PVG neurons suggests that these proteins engage in an autocrine signaling loop in these neurons. Our work shows that CD40L reverse signaling is a physiological regulator of NGF‐promoted sympathetic axon growth and tissue innervation with opposite effects in paravertebral and prevertebral neurons.

069 Generation and utilization of a stable IFN response reporter line in keratinocytes

We have recently identified a critical role for IFN-kappa (IFNK), an epidermally derived type I IFN, in various inflammatory skin diseases including autoimmune connective tissue diseases. Through an autocrine signaling loop, that is unique to keratinocytes (KCs), IFN-k maintains basal interferon responses in KCs that are heightened in diseases such as lupus. The evolutionary basis of this may be explained by the role of KCs as a critical barrier against infections, particularly viral. With a series of amino acids and signal peptide sequences of IFN-k being conserved in all the type I IFNs, the KCs might have evolved with distinct function for IFNK expression in the epidermis and for subsequent activation of interferon responses. Hence, we designed a stable reporter line in KCs, using the IFN response gene MX1 to dissect the role of autocrine IFN responses and identify key molecules/pathways involved in this process. The MX1 reporter cell line was established by stably infecting KCs with a lentiviral construct encoding the luciferase gene under the control of the MX1 promoter. The stable MX1 reporter line in KCs is currently being used to identify the critical regulators of IFN responsive elements in KCs for which studies have been limited, and in particular to identify genes which positively or negatively regulate type I IFN responses using small molecule chemical screens and CRISPR/Cas9 based genome-wide screens.

OVA66 promotes tumour angiogenesis and progression through enhancing autocrine VEGF-VEGFR2 signalling.

BackgroundActivation of autocrine VEGF-VEGFR2 signalling in tumour cells activates cell proliferation, survival, and angiogenesis, all of which are crucial for tumour progression. Ovarian cancer-associated antigen 66 (OVA66) is now known to be overexpressed in multiple tumours and plays a role in tumour development, but the underlying mechanisms has not been fully investigated.MethodsWe employed ovarian and cervical cancer cells and mouse models to detect the role of OVA66 in angiogenesis, growth, and metastasis of cancer cells. Immunofluorescence and western blot were used to determine the function of OVA66 in regulating autocrine VEGF-VEGFR2 signalling. Immunohistochemistry and bioinformatics analysis were used to detect the correlation of OVA66 and VEGF expression.FindingsOVA66 overexpression in the cancer cell lines promoted VEGF secretion, tumour growth and angiogenesis in vitro and in vivo. Conversely, shRNA-mediated OVA66 knockdown had the opposite effects. Mechanistically, OVA66 overexpression was found to boost an autocrine VEGF–VEGFR2 positive-feedback signalling loop in the tumour cells, leading to amplified effect of VEGF on tumour angiogenesis and proliferation and increased migration in vitro and in vivo, respectively. Finally, we identified a significant positive correlation between the expression levels of OVA66 and VEGF in ovarian and cervical cancer specimens, and found that OVA66 was significantly associated with advanced ovarian cancer.InterpretationWe identify a novel function for OVA66 in regulating an autocrine VEGF–VEGFR2 feed-forward signalling loop that promotes tumour progression and angiogenesis.FundThis work was supported by the National Natural Science Foundation of China (81602262); and Excellent Youth Scholar Program of Tongji University (2015KJ062).

Targeting an Autocrine Regulatory Loop in Cancer Stem-like Cells Impairs the Progression and Chemotherapy Resistance of Bladder Cancer.

Cancer stem-like cells (CSCs) contribute to bladder cancer chemotherapy resistance and progression, but the associated mechanisms have not been elucidated. This study determined whether blocking an autocrine signaling loop in CSCs improves the therapeutic effects of cis-platinum on bladder cancer. The expression of the epithelial marker OV6 and other markers in human bladder cancer specimens was examined by IHC. The CSC properties of magnetic-activated cell sorting (MACS)-isolated OV6+ and OV6- bladder cancer cells were examined. Molecular mechanisms were assessed through RNA-Seq, cytokine antibody arrays, co-immunoprecipitation (co-IP), chromatin immunoprecipitation (ChIP) and other assays. An orthotopic bladder cancer mouse model was established to evaluate the in vivo effects of a YAP inhibitor (verteporfin) and a PDGFR inhibitor (CP-673451) on the cis-platinum resistance of OV6+ CSCs in bladder cancer. Upregulated OV6 expression positively associated with disease progression and poor prognosis for bladder cancer patients. Compared with OV6- cells, OV6+ bladder cancer cells exhibited strong CSC characteristics, including self-renewal, tumor initiation in NOD/SCID mice, and chemotherapy resistance. YAP, which maintains the stemness of OV6+ CSCs, triggered PDGFB transcription by recruiting TEAD1. Autocrine PDGF-BB signaling through its receptor PDGFR stabilized YAP and facilitated YAP nuclear translocation. Furthermore, blocking the YAP/TEAD1/PDGF-BB/PDGFR loop with verteporfin or CP-673451 inhibited the cis-platinum resistance of OV6+ bladder cancer CSCs in an orthotopic bladder cancer model. OV6 could be a helpful indicator of disease progression and prognosis for patients with bladder cancer, and targeting the autocrine YAP/TEAD1/PDGF-BB/PDGFR loop might serve as a remedy for cis-platinum resistance in patients with advanced bladder cancer.

DDR1 regulates thyroid cancer cell differentiation via IGF-2/IR-A autocrine signaling loop.

Patients with thyroid cancers refractory to radioiodine (RAI) treatment show a limited response to various therapeutic options and a low survival rate. The recent use of multikinase inhibitors has also met limited success. An alternative approach relies on drugs that induce cell differentiation, as the ensuing increased expression of the cotransporter for sodium and iodine (NIS) may partially restore sensitivity to radioiodine. The inhibition of the ERK1/2 pathway has shown some efficacy in this context. Aggressive thyroid tumors overexpress the isoform-A of the insulin receptor (IR-A) and its ligand IGF-2; this IGF-2/IR-A loop is associated with de-differentiation and stem-like phenotype, resembling RAI-refractory tumors. Importantly, IR-A has been shown to be positively modulated by the non-integrin collagen receptor DDR1 in human breast cancer. Using undifferentiated human thyroid cancer cells, we now evaluated the effects of DDR1 on IGF-2/IR-A loop and on markers of cell differentiation and stemness. DDR1 silencing or downregulation caused significant reduction of IR-A and IGF-2 expression, and concomitant increased levels of differentiation markers (NIS, Tg, TSH, TPO). Conversely, markers of epithelial-to-mesenchymal transition (Vimentin, Snail-2, Zeb1, Zeb2 and N-Cadherin) and stemness (OCT-4, SOX-2, ABCG2 and Nanog) decreased. These effects were collagen independent. In contrast, overexpression of either DDR1 or its kinase-inactive variant K618A DDR1-induced changes suggestive of less differentiated and stem-like phenotype. Collagen stimulation was uneffective. In conclusion, in poorly differentiated thyroid cancer, DDR1 silencing or downregulation blocks the IGF-2/IR-A autocrine loop and induces cellular differentiation. These results may open novel therapeutic approaches for thyroid cancer.

IL-17A Attenuates IFN-λ Expression by Inducing Suppressor of Cytokine Signaling Expression in Airway Epithelium.

IFN-λ is a cytokine expressed in epithelial tissues and plays a central role in antiviral mucosal immune response. The expression of IFN-λ in the airway is impaired in chronic airway diseases (e.g., asthma, chronic obstructive pulmonary disease), which renders patients susceptible to viral infection. IL-17A is associated with asthma and chronic obstructive pulmonary disease pathogenesis; however, IL-17A regulation of IFN-λ expression remains unclear. The aim of the current study is to clarify IL-17A-mediated regulatory mechanisms of IFN-λ expression in human airway epithelial cells. In this study, we have shown that polyinosinic:polycytidylic acid (polyI:C) and influenza A virus (IAV) infection increased IFN-λ expression at mRNA and protein levels in primary cultures of normal human bronchial epithelial cells, whereas IL-17A attenuated polyI:C- or IAV-induced IFN-λ expression. IFN-λ receptor 1 knockdown and a JAK inhibitor, ruxolitinib, attenuated polyI:C-induced IFN-λ expression, confirming that a positive autocrine feedback loop, the IFN-λ receptor-JAK-STAT pathway, was involved in IFN-λ expression. In Western blotting analysis, we demonstrated that polyI:C and IAV infection induced STAT1 phosphorylation in normal human bronchial epithelial cells, whereas IL-17A suppressed polyI:C- or IAV-mediated STAT1 phosphorylation. Furthermore, we found that cotreatment with IL-17A and polyI:C or IAV infection synergistically increased suppressor of cytokine signaling (SOCS)1 and SOCS3 expression. SOCS1 small interfering RNA and SOCS3 small interfering RNA negated the inhibitory effect of IL-17A in polyI:C-induced IFN-λ expression by restoring attenuated STAT1 phosphorylation. Taken together, these findings indicate that IL-17A attenuates virus-induced IFN-λ expression by enhancing SOCS1 and SOCS3 expression to inhibit autocrine signaling loops in human airway epithelial cells.

Abstract 4423: MiR551b-mediated Oncostatin M signaling in breast cancer

Abstract We have previously demonstrated that an oncogenic microRNA miR551b-3p (also known as miR551b) regulates STAT3 transcription thereby enhancing tumor growth and metastasis in ovarian cancer. Herein, we have identified that Oncostatin M (OSM) is upregulated through STAT3-driven transcription, which plays an important role in promoting migration and invasion of breast cancer cells. The purpose of this study was to evaluate the role of miR551b mediated STAT3 activation and the precise mechanism that regulates OSM-Receptor (OSMR) signaling for breast cancer growth and metastasis. MDA-MB231 cells were either overexpressed or silenced with miR551b and anti-miR551b respectively and the effects on proliferation, migration/invasion (Matrigel transwell assay), wound healing and expression of OSMR and its downstream targets (Western blotting, Quantitative RT-PCR) were assessed. Our data demonstrate that miR551b promoted survival, proliferation, invasion and migration and epithelial to mesenchymal transition (EMT) and spheroid formation of breast cancer cells. Our results also show that miR551b-mediated STAT3 activation upregulated the components of OSMR signaling through elevated expression of OSMR, Interleukin 31 receptor-alpha (IL31RA) and their ligands Oncostatin (OSM) and IL31 thereby inducing phosphorylation of STAT3 for tumor cell proliferation, migration and invasion. Conversely, anti-miR551b reduced the expression of OSMR, IL31R and their ligands OSM and IL31 as well as the phosphorylation of STAT3. Briefly, our results provide the first report indicating that miR551b regulates OSMR expression by activating STAT3 and thereby promoting tumor cell growth and metastasis in breast cancer cells through autocrine feed-forward signaling loop. A.G, D.P., contributed equally. Citation Format: Anjali Geethadevi, Deepak Parashar, Miriam Ragle-Aure, Bindu Nair, Yiling Lu, Vessela N. Kristensen, Ming You, Anil K. Sood, Gordon B. Mills, Sunila Pradeep, Pradeep Chaluvally Raghavan. MiR551b-mediated Oncostatin M signaling in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4423.

Autocrine activation of platelet-derived growth factor receptor α in metastatic papillary thyroid cancer

Metastatic dissemination of papillary thyroid cancer has been reported to be strongly associated with expression of platelet-derived growth factor (PDGFR) α and altered TTF1 function. However, the status of PDGF ligands in papillary thyroid cancer and the potential role of these ligands in metastatic disease are obscure. We assessed the prevalence of PDGF ligands in benign and malignant thyroid tumors to determine if ligand upregulation is associated with α-isoform (PDGF-AA or PDGF-BB) or the β-isoform (PDGF-BB or PDGF-DD) of PDGFR in individual tumors. The immunohistochemical expression of PDGFRα, PDGF-AA, PDGF-BB, and PDGF-DD was surveyed in follicular adenomas (n=55), papillary thyroid carcinomas (103 with and 59 without nodal metastases), and lymph node metastasis (n=12). There is an augmented tendency for PDGF-AA expression in node-positive papillary thyroid cancer metastases (P<.0001). Although PDGF-BB and -DD were commonly identified, there was no relationship between the presence of these cytokines and malignant disease or metastases. Logistic regression demonstrated that PDGF-AA expression was significantly associated with the presence of PDGFRα (odds ratio=4.6, P=.004) and recurrent disease. When either PDGFRα or PDGF-AA was used to predict the presence of metastases, the sensitivity achieved was 86% and 88%, respectively, whereas specificities were lower at 71% and 61%, respectively. The augmented coexpression of PDGF-AA and PDGFRα in metastatic papillary thyroid cancers suggests that an autocrine signaling loop may contribute to nodal infiltration. Combined testing for the expression of PDGF-AA and PDGFRα may identify those patients with papillary thyroid cancer at risk of metastatic disease and resistance to therapy.