Search

Promising biomaterials should be tested in appropriate large animal models that recapitulate human inflammatory and regenerative responses. Previous studies have shown tyrosine-derived polycarbonates (TyrPC) are versatile biomaterials with a wide range of applications across multiple disciplines. The library of TyrPC has been well studied and consists of thousands of polymer compositions with tunable mechanical characteristics and degradation and resorption rates that are useful for nerve guidance tubes (NGTs). NGTs made of different TyrPCs have been used in segmental nerve defect models in small animals. The current study is an extension of this work and evaluates NGTs made using two different TyrPC compositions in a 1 cm porcine peripheral nerve repair model. We first evaluated a nondegradable TyrPC formulation, demonstrating proof-of-concept chronic regenerative efficacy up to 6 months with similar nerve/muscle electrophysiology and morphometry to the autograft repair control. Next, we characterized the acute regenerative response using a degradable TyrPC formulation. After 2 weeks in vivo, TyrPC NGT promoted greater deposition of pro-regenerative extracellular matrix (ECM) constituents (in particular collagen I, collagen III, collagen IV, laminin, and fibronectin) compared to commercially available collagen-based NGTs. This corresponded with dense Schwann cell infiltration and axon extension across the lumen. These findings confirmed results reported previously in a mouse model and reveal that TyrPC NGTs were well tolerated in swine and facilitated host axon regeneration and Schwann cell infiltration in the acute phase across segmental defects - likely by eliciting a favorable neurotrophic ECM milieu. This regenerative response ultimately can contribute to functional recovery., (© 2020 Wiley Periodicals LLC.)

Background: The mechanism by which immune cells regulate metastasis is unclear. Understanding the role of immune cells in metastasis will guide the development of treatments improving patient survival., Methods: We used syngeneic orthotopic mouse tumour models (wild-type, NOD/scid and Nude), employed knockout (CD8 and CD4) models and administered CXCL4. Tumours and lungs were analysed for cancer cells by bioluminescence, and circulating tumour cells were isolated from blood. Immunohistochemistry on the mouse tumours was performed to confirm cell type, and on a tissue microarray with 180 TNBCs for human relevance. TCGA data from over 10,000 patients were analysed as well., Results: We reveal that intratumoral immune infiltration differs between metastatic and non-metastatic tumours. The non-metastatic tumours harbour high levels of CD8 + T cells and low levels of platelets, which is reverse in metastatic tumours. During tumour progression, platelets and CXCL4 induce differentiation of monocytes into myeloid-derived suppressor cells (MDSCs), which inhibit CD8 + T-cell function. TCGA pan-cancer data confirmed that CD8 low Platelet high patients have a significantly lower survival probability compared to CD8 high Platelet low ., Conclusions: CD8 + T cells inhibit metastasis. When the balance between CD8 + T cells and platelets is disrupted, platelets produce CXCL4, which induces MDSCs thereby inhibiting the CD8 + T-cell function., (© 2021. The Author(s).)

Many oncogenic insults deregulate RNA splicing, often leading to hypersensitivity of tumors to spliceosome-targeted therapies (STTs). However, the mechanisms by which STTs selectively kill cancers remain largely unknown. Herein, we discover that mis-spliced RNA itself is a molecular trigger for tumor killing through viral mimicry. In MYC-driven triple-negative breast cancer, STTs cause widespread cytoplasmic accumulation of mis-spliced mRNAs, many of which form double-stranded structures. Double-stranded RNA (dsRNA)-binding proteins recognize these endogenous dsRNAs, triggering antiviral signaling and extrinsic apoptosis. In immune-competent models of breast cancer, STTs cause tumor cell-intrinsic antiviral signaling, downstream adaptive immune signaling, and tumor cell death. Furthermore, RNA mis-splicing in human breast cancers correlates with innate and adaptive immune signatures, especially in MYC-amplified tumors that are typically immune cold. These findings indicate that dsRNA-sensing pathways respond to global aberrations of RNA splicing in cancer and provoke the hypothesis that STTs may provide unexplored strategies to activate anti-tumor immune pathways., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Abstract: Neurogastroenterology and motility (NGM) disorders are common in childhood and are often very debilitating. Although pediatric gastroenterology fellows are expected to obtain training in the diagnosis and management of patients with these disorders, there is an ongoing concern for unmet needs and lack of exposure and standardized curriculum. In the context of tailoring training components, outcome and expressed needs of pediatric gastroenterology fellows and programs, members of the North American Society for Pediatric Gastroenterology, Hepatology and Nutrition (NASPGHAN) and American Neurogastroenterology and Motility Society (ANMS) developed guidelines for NGM training in North America in line with specific expectations and goals of training as delineated through already established entrustable professional activities (EPAs). Members of the joint task force applied their expertise to identify the components of knowledge, skills, and management, which are expected of NGM consultants. The clinical knowledge, skills and management elements of the NGM curriculum are divided into domains based on anatomic regions including esophagus, stomach, small bowel, colon and anorectum. In addition, dedicated sections on pediatric functional gastrointestinal (GI) disorders, research and collaborative approach, role of behavioral health and surgical approaches to NGM disorders and transition from pediatric to adult neurogastroenterology are included in this document. Members of the NASPGHAN-ANMS task force anticipate that this document will serve as a resource to break existing barriers to pursuing a career in NGM and provide a framework towards uniform training expectations at 3 hierarchical tiers corresponding to EPA levels., Competing Interests: The authors report no conflicts of interest., (Copyright © 2020 by European Society for Pediatric Gastroenterology, Hepatology, and Nutrition and North American Society for Pediatric Gastroenterology, Hepatology, and Nutrition.)

The word myeloid is derived from the Greek word muelós which means "marrow". Therefore, myeloid cells are described as cells that arise in the bone marrow. They can be distinguished from lymphoid cells based on their different differentiation trajectories-Lymphoid cells (B and T cells) are usually born in the bone marrow, but they need to migrate to lymphoid organs to mature and differentiate usually in response to antigens produced due to infections and diseases like cancer. On the other hand, myeloid cells do not follow this differentiation trajectory. They arise from the bone marrow, and do not need an encounter with antigens to gain their functionality. Thus, while lymphoid cells are a part of the adaptive immune system, myeloid cells are a part of the innate immune system. Hematopoiesis gives rise to two progenitor cells-the common myeloid progenitor (CMP) and the common lymphoid progenitor (CLP). The CMP can give rise to megakaryocytes, erythrocytes, mast cells and myeloblasts. Myeloblasts in turn lead to the formation of basophils, neutrophils, eosinophils and monocytes that can further differentiate into macrophages. This review will focus on macrophages as well as the phenotypes they acquire with the tumor immune microenvironment (TIME) in triple-negative breast cancer (TNBC). It will address how cancer cells in the tumor microenvironment (TME) recruit macrophages and may switch to recruiting neutrophils upon depletion of these tumor-associated macrophages (TAMs). Finally, it will also shed light on past and current treatment options that specifically target these cells and how those affect patient outcomes in TNBC.

Peripheral nerve injury (PNI) impacts millions annually, often leaving debilitated patients with minimal repair options to improve functional recovery. Our group has previously developed tissue engineered nerve grafts (TENGs) featuring long, aligned axonal tracts from dorsal root ganglia (DRG) neurons that are fabricated in custom bioreactors using the process of axon "stretch-growth." We have shown that TENGs effectively serve as "living scaffolds" to promote regeneration across segmental nerve defects by exploiting the newfound mechanism of axon-facilitated axon regeneration, or "AFAR," by simultaneously providing haptic and neurotrophic support. To extend this work, the current study investigated the efficacy of living versus nonliving regenerative scaffolds in preserving host sensory and motor neuronal health following nerve repair. Rats were assigned across five groups: naïve, or repair using autograft, nerve guidance tube (NGT) with collagen, NGT + non-aligned DRG populations in collagen, or TENGs. We found that TENG repairs yielded equivalent regenerative capacity as autograft repairs based on preserved health of host spinal cord motor neurons and acute axonal regeneration, whereas NGT repairs or DRG neurons within an NGT exhibited reduced motor neuron preservation and diminished regenerative capacity. These acute regenerative benefits ultimately resulted in enhanced levels of functional recovery in animals receiving TENGs, at levels matching those attained by autografts. Our findings indicate that TENGs may preserve host spinal cord motor neuron health and regenerative capacity without sacrificing an otherwise uninjured nerve (as in the case of the autograft) and therefore represent a promising alternative strategy for neurosurgical repair following PNI., (© 2020 John Wiley & Sons, Ltd.)

The purpose of this review is to highlight several areas of lncRNA biology and cancer that we hope will provide some new insights for future research. These include the relationship of lncRNAs and the epithelial to mesenchymal transition (EMT) with a focus on transcriptional and alternative splicing mechanisms and mRNA stability through miRNAs. In addition, we highlight the potential role of enhancer e-lncRNAs, the importance of transposable elements in lncRNA biology, and finally the emerging area of using antisense oligonucleotides (ASOs) and small molecules to target lncRNAs and their therapeutic implications.

Stem cells need to be protected from genotoxic and proteotoxic stress to maintain a healthy pool throughout life 1-3 . Little is known about the proteostasis mechanism that safeguards stem cells. Here we report endoplasmic reticulum-associated degradation (ERAD) as a protein quality checkpoint that controls the haematopoietic stem cell (HSC)-niche interaction and determines the fate of HSCs. The SEL1L-HRD1 complex, the most conserved branch of ERAD 4 , is highly expressed in HSCs. Deletion of Sel1l led to niche displacement of HSCs and a complete loss of HSC identity, and allowed highly efficient donor-HSC engraftment without irradiation. Mechanistic studies identified MPL, the master regulator of HSC identity 5 , as a bona fide ERAD substrate that became aggregated in the endoplasmic reticulum following ERAD deficiency. Restoration of MPL signalling with an agonist partially rescued the number and reconstitution capacity of Sel1l-deficient HSCs. Our study defines ERAD as an essential proteostasis mechanism to safeguard a healthy stem cell pool by regulating the stem cell-niche interaction.

Background: Millions of Americans experience residual deficits from traumatic peripheral nerve injury (PNI). Despite advancements in surgical technique, repair typically results in poor functional outcomes due to prolonged periods of denervation resulting from long regenerative distances coupled with slow rates of axonal regeneration. Novel surgical solutions require valid preclinical models that adequately replicate the key challenges of clinical PNI., Objective: To develop a preclinical model of PNI in swine that addresses 2 challenging, clinically relevant PNI scenarios: long segmental defects (≥5 cm) and ultra-long regenerative distances (20-27 cm). Thus, we aim to demonstrate that a porcine model of major PNI is suitable as a potential framework to evaluate novel regenerative strategies prior to clinical deployment., Methods: A 5-cm-long common peroneal nerve or deep peroneal nerve injury was repaired using a saphenous nerve or sural nerve autograft, respectively. Histological and electrophysiological assessments were performed at 9 to 12 mo post repair to evaluate nerve regeneration and functional recovery. Relevant anatomy, surgical approach, and functional/histological outcomes were characterized for both repair techniques., Results: Axons regenerated across the repair zone and were identified in the distal stump. Electrophysiological recordings confirmed these findings and suggested regenerating axons reinnervated target muscles., Conclusion: The models presented herein provide opportunities to investigate peripheral nerve regeneration using different nerves tailored for specific mechanisms of interest, such as nerve modality (motor, sensory, and mixed fiber composition), injury length (short/long gap), and total regenerative distance (proximal/distal injury)., (Copyright © 2020 by the Congress of Neurological Surgeons.)

Mechanisms driving tumor progression from less aggressive subtypes to more aggressive states represent key targets for therapy. We identified a subset of luminal A primary breast tumors that give rise to HER2-enriched (HER2E) subtype metastases, but remain clinically HER2 negative (cHER2-). By testing the unique genetic and transcriptomic features of these cases, we developed the hypothesis that FGFR4 likely participates in this subtype switching. To evaluate this, we developed 2 FGFR4 genomic signatures using a patient-derived xenograft (PDX) model treated with an FGFR4 inhibitor, which inhibited PDX growth in vivo. Bulk tumor gene expression analysis and single-cell RNA sequencing demonstrated that the inhibition of FGFR4 signaling caused molecular switching. In the Molecular Taxonomy of Breast Cancer International Consortium (METABRIC) breast cancer cohort, FGFR4-induced and FGFR4-repressed signatures each predicted overall survival. Additionally, the FGFR4-induced signature was an independent prognostic factor beyond subtype and stage. Supervised analysis of 77 primary tumors with paired metastases revealed that the FGFR4-induced signature was significantly higher in luminal/ER+ tumor metastases compared with their primaries. Finally, multivariate analysis demonstrated that the FGFR4-induced signature also predicted site-specific metastasis for lung, liver, and brain, but not for bone or lymph nodes. These data identify a link between FGFR4-regulated genes and metastasis, suggesting treatment options for FGFR4-positive patients, whose high expression is not caused by mutation or amplification.

Background: Entrustable professional activities (EPAs) are critical activities performed by medical professionals, which can be observed and assessed. Adding on to common EPAs for all pediatric subspecialty trainees, specialty-specific EPAs for pediatric gastroenterology, hepatology, and nutritional fellowship were developed by the North American Society for Pediatric Gastroenterology, Hepatology, and Nutrition (NASPGHAN) EPA Task Force., Methods: Having developed specialty-specific EPAs, building EPA assessments is the next logical step, as EPAs are included under a larger umbrella of competency-based assessment. Thus, the NASPGHAN EPA Task Force and Training Committee collaborated on an assessment tool and associated curricular resources to aid in tracking trainees' progression to entrustment within individual EPAs and readiness for independent practice., Results: This manuscript reports the development of an EPA assessment tool, including guiding principles and the theory behind the assessment tool, with a focus on simple, meaningful assessments that can provide crucial performance feedback to trainees. In addition, curricular resources were developed, based on the assessment tool, to support training. Ultimately, it is the hope of the NASPGHAN EPA Task Force and Training Committee that this tool can aid training programs in providing formative feedback for trainees, and can be used by training programs and clinical competency committees for summative evaluation.

Polyclonal metastases frequently arise from clusters of circulating tumor cells (CTCs). CTC clusters metastasize better than single CTCs, but the underlying molecular mechanisms are poorly understood. Here, we show that polyclonal metastatic seeds exhibit higher resistance to natural killer (NK) cell killing. Using breast cancer models, we observed higher proportions of polyclonal lung metastasis in immunocompetent mice compared with mice lacking NK cells. Depleting NK cells selectively increased monoclonal but not polyclonal metastases, suggesting that CTC clusters are less sensitive to NK-mediated suppression. Transcriptional analyses revealed that clusters have elevated expression of cell-cell adhesion and epithelial genes, which is associated with decreased expression of NK cell activating ligands. Furthermore, perturbing tumor cell epithelial status altered NK ligand expression and sensitivity to NK-mediated killing. Collectively, our findings show that NK cells can determine the fate of CTCs of different epithelial and mesenchymal states, and impact metastatic clonal evolution by favoring polyclonal seeding., (© 2020. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Background: The COVID-19 pandemic has drastically changed healthcare systems and training around the world. The Training Committee of the North American Society for Pediatric Gastroenterology, Hepatology and Nutrition sought to understand how COVID-19 has affected pediatric gastroenterology fellowship training., Methods: A 21 question survey was distributed to all 77 pediatric gastroenterology fellowship program directors (PDs) in the North American Society for Pediatric Gastroenterology, Hepatology and Nutrition program director database via email on April 7. Responses collected through April 19, 2020 were analyzed using descriptive statistics., Results: Fifty-one of 77 (66%) PDs from the United States, Canada, and Mexico responded to the survey. Forty-six of 51 (90%) PDs reported that they were under a "stay-at-home" order for a median of 4 weeks at the time of the survey. Two of the 51 (4%) programs had fellows participating in outpatient telehealth before COVID-19 and 39 of 51 (76%) at the time of the survey. Fellows stopped participating in outpatient clinics in 22 of 51 (43%) programs and endoscopy in 26 of 51 (52%) programs. Changes to inpatient care included reduced fellow staffing, limiting who entered patient rooms, and rounding remotely. Fellows in 3 New York programs were deployed to adult medicine units. Didactics were moved to virtual conferences in 47 of 51 (94%) programs, and fellows used various online resources. Clinical research and, disproportionately, bench research were restricted., Conclusions: This report provides early information of the impact of COVID-19 on pediatric fellowship training. Rapid adoption of telehealth and reduced clinical and research experiences were important changes. Survey information may spur communication and innovation to help educators adapt.

Preclinical models that faithfully recapitulate tumor heterogeneity and therapeutic response are critical for translational breast cancer research. Immortalized cell lines are easy to grow and genetically modify to study molecular mechanisms, yet the selective pressure from cell culture often leads to genetic and epigenetic alterations over time. Patient-derived xenograft (PDX) models faithfully recapitulate the heterogeneity and drug response of human breast tumors. PDX models exhibit a relatively short latency after orthotopic transplantation that facilitates the investigation of breast tumor biology and drug response. The transplantable genetically engineered mouse models allow the study of breast tumor immunity. The current protocol describes the method to orthotopically transplant breast tumor fragments into the mammary fat pad followed by drug treatments. These preclinical models provide valuable approaches to investigate breast tumor biology, drug response, biomarker discovery and mechanisms of drug resistance.

Transposable elements (TEs) comprise a large proportion of long non-coding RNAs (lncRNAs). Here, we employed CRISPR to delete a short interspersed nuclear element (SINE) in Malat1, a cancer-associated lncRNA, to investigate its significance in cellular physiology. We show that Malat1 with a SINE deletion forms diffuse nuclear speckles and is frequently translocated to the cytoplasm. SINE-deleted cells exhibit an activated unfolded protein response and PKR and markedly increased DNA damage and apoptosis caused by dysregulation of TDP-43 localization and formation of cytotoxic inclusions. TDP-43 binds stronger to Malat1 without the SINE and is likely 'hijacked' by cytoplasmic Malat1 to the cytoplasm, resulting in the depletion of nuclear TDP-43 and redistribution of TDP-43 binding to repetitive element transcripts and mRNAs encoding mitotic and nuclear-cytoplasmic regulators. The SINE promotes Malat1 nuclear retention by facilitating Malat1 binding to HNRNPK, a protein that drives RNA nuclear retention, potentially through direct interactions of the SINE with KHDRBS1 and TRA2A, which bind to HNRNPK. Losing these RNA-protein interactions due to the SINE deletion likely creates more available TDP-43 binding sites on Malat1 and subsequent TDP-43 aggregation. These results highlight the significance of lncRNA TEs in TDP-43 proteostasis with potential implications in both cancer and neurodegenerative diseases., (© The Author(s) 2019. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Background: Due to scarcity of scientific literature on pediatric gastroparesis, there is a need to summarize current evidence and identify areas requiring further research. The aim of this study was to provide an evidence-based review of the available literature on the prevalence, pathogenesis, clinical presentation, diagnosis, treatment, and outcomes of pediatric gastroparesis., Methods: A search of the literature was performed using the Preferred Reporting Items for Systematic Reviews and Meta-analyses guidelines with the following databases: PubMed, EMBASE, Cochrane Database of Systematic Reviews, Database of Abstracts of Reviews of Effects, Cochrane Central Register of Controlled Trials, and Web of Science. Two independent reviewers screened abstracts for eligibility., Key Results: Our search yielded 1085 original publications, 135 of which met inclusion criteria. Most articles were of retrospective study design. Only 12 randomized controlled trials were identified, all of which were in infants. The prevalence of pediatric gastroparesis is unknown. Gastroparesis may be suspected based on clinical symptoms although these are often non-specific. The 4-hour nuclear scintigraphy scan remains gold standard for diagnosis despite lack of pediatric normative comparison data. Therapeutic approaches include dietary modifications, prokinetic drugs, and postpyloric enteral tube feeds. For refractory cases, intrapyloric botulinum toxin and surgical interventions such as gastric electrical stimulation may be warranted. Most interventions still lack rigorous supportive data., Conclusions: Diagnosis and treatment of pediatric gastroparesis are challenging due to paucity of published evidence. Larger and more rigorous clinical trials are necessary to improve outcomes., (© 2019 John Wiley & Sons Ltd.)

It has been almost 30 years since C/EBPß was discovered. Seminal studies have shown that C/EBPß is a master regulator of mammary gland development and has been shown to control and influence proliferation and differentiation through varying mechanisms. The single-exon C/EBPß mRNA yields at least three different protein isoforms which have diverse, specific, context-dependent, and often non-overlapping roles throughout development and breast cancer progression. These roles are dictated by a number of complex factors including: expression levels of other C/EBP family members and their stoichiometry relative to the isoform in question, binding site affinity, post-translational modifications, co-factor expression, and even hormone levels and lactogenic status. Here we summarize the historical work up to the latest findings in the field on C/EBPß in the mammary gland and in breast cancer. With the current emphasis on improving immunotherapy in breast cancer the role of specific C/EBPß isoforms in regulating specific chemokine and cytokine expression and the immune microenvironment will be of increasing interest.

Cancer-induced immune responses affect tumour progression and therapeutic response. In multiple murine models and clinical datasets, we identified large variations of neutrophils and macrophages that define 'immune subtypes' of triple-negative breast cancer (TNBC), including neutrophil-enriched (NES) and macrophage-enriched subtypes (MES). Different tumour-intrinsic pathways and mutual regulation between macrophages (or monocytes) and neutrophils contribute to the development of a dichotomous myeloid compartment. MES contains predominantly macrophages that are CCR2-dependent and exhibit variable responses to immune checkpoint blockade (ICB). NES exhibits systemic and local accumulation of immunosuppressive neutrophils (or granulocytic myeloid-derived suppressor cells), is resistant to ICB, and contains a minority of macrophages that seem to be unaffected by CCR2 knockout. A MES-to-NES conversion mediated acquired ICB resistance of initially sensitive MES models. Our results demonstrate diverse myeloid cell frequencies, functionality and potential roles in immunotherapies, and highlight the need to better understand the inter-patient heterogeneity of the myeloid compartment.

Metastatic breast cancer is an extremely complex disease with limited treatment options due to the lack of information about the major characteristics of metastatic disease. There is an urgent need, therefore, to understand the changes in cellular complexity and dynamics that occur during metastatic progression. In the current study, we analyzed the cellular and molecular differences between primary tumors and paired lung metastases using a syngeneic p53-null mammary tumor model of basal-like breast cancer. Distinct subpopulations driven by the Wnt- and/or STAT3 signaling pathways were detected in vivo using a lentiviral Wnt- and STAT3 signaling reporter system. A significant increase in the overlapping populations driven by both the Wnt- and STAT3 signaling pathways was observed in the lung metastases as compared to the primary tumors. Furthermore, the overlapping populations showed a higher metastatic potential relative to the other populations and pharmacological inhibition of both signaling pathways was shown to markedly reduce the metastatic lesions in established lung metastases. An analysis of the unique molecular features of the lung metastases revealed a significant association with immune response signatures. Specifically, Foxp3 gene expression was markedly increased and elevated levels of Foxp3 + Treg cells were detected in close proximity to lung metastases. Collectively, these studies illustrate the importance of analyzing intratumoral heterogeneity, changes in population dynamics, and the immune microenvironment during metastatic progression.

The Wnt gene family encodes an evolutionarily conserved group of proteins that regulate cell growth, differentiation and stem cell self-renewal. Aberrant Wnt signaling in human breast tumors has been proposed as a driver of tumorigenesis, especially in the basal-like tumor subtype where canonical Wnt signaling is both enriched and predictive of poor clinical outcomes. The development of effective Wnt-based therapeutics, however, has been slowed in part by a limited understanding of the context-dependent nature with which these aberrations influence breast tumorigenesis. We previously reported that MMTV-Wnt1 mice, an established model for studying Wnt signaling in breast tumors, develop two subtypes of tumors by gene expression classification: Wnt1-Early Ex and Wnt1-Late Ex Here, we extend this initial observation and show that Wnt1-Early Ex tumors exhibit high expression of canonical Wnt, non-canonical Wnt, and EGFR signaling pathway signatures. Therapeutically, Wnt1-Early Ex tumors showed a dynamic reduction in tumor volume when treated with an EGFR inhibitor. Wnt1-Early Ex tumors had primarily Cd49f pos /Epcam neg FACS profiles, but it was not possible to serially transplant these tumors into wild-type FVB female mice. Conversely, Wnt1-Late Ex tumors had a bloody gross pathology, which was highlighted by the presence of 'blood lakes' identified by H&E staining. These tumors had primarily Cd49f pos /Epcam pos FACS profiles, but also contained a secondary Cd49f pos /Epcam neg subpopulation. Wnt1-Late Ex tumors were enriched for activating Hras1 mutations and were capable of reproducing tumors when serially transplanted into wild-type FVB female mice. This study definitively shows that the MMTV-Wnt1 mouse model produces two phenotypically distinct subtypes of mammary tumors that differ in multiple biological aspects including sensitivity to an EGFR inhibitor., Competing Interests: Competing interestsC.M.P. is an equity stock holder, consultant and Board of Director Member of BioClassifier LLC and GeneCentric Therapeutics. C.M.P. is also listed as inventor on patent applications on the Breast PAM50 assay., (© 2019. Published by The Company of Biologists Ltd.)

Over the last decade, both early diagnosis and targeted therapy have improved the survival rates of many cancer patients. Most recently, immunotherapy has revolutionized the treatment options for cancers such as melanoma. Unfortunately, a significant portion of cancers (including lung and breast cancers) do not respond to immunotherapy, and many of them develop resistance to chemotherapy. Molecular characterization of non-responsive cancers suggest that an embryonic program known as epithelial-mesenchymal transition (EMT), which is mostly latent in adults, can be activated under selective pressures, rendering these cancers resistant to chemo- and immunotherapies. EMT can also drive tumor metastases, which in turn also suppress the cancer-fighting activity of cytotoxic T cells that traffic into the tumor, causing immunotherapy to fail. In this review, we compare and contrast immunotherapy treatment options of non-small cell lung cancer (NSCLC) and triple negative breast cancer (TNBC). We discuss why, despite breakthrough progress in immunotherapy, attaining predictable outcomes in the clinic is mostly an unsolved problem for these tumors. Although these two cancer types appear different based upon their tissues of origin and molecular classification, gene expression indicate that they possess many similarities. Patient tumors exhibit activation of EMT, and resulting stem cell properties in both these cancer types associate with metastasis and resistance to existing cancer therapies. In addition, the EMT transition in both these cancers plays a crucial role in immunosuppression, which exacerbates treatment resistance. To improve cancer-related survival we need to understand and circumvent, the mechanisms through which these tumors become therapy resistant. In this review, we discuss new information and complementary perspectives to inform combination treatment strategies to expand and improve the anti-tumor responses of currently available clinical immune checkpoint inhibitors.

Metastasis is the most devastating stage of cancer progression and causes the majority of cancer-related deaths. Clinical observations suggest that most cancers metastasize to specific organs, a process known as "organotropism." Elucidating the underlying mechanisms may help identify targets and treatment strategies to benefit patients. This review summarizes recent findings on tumor-intrinsic properties and their interaction with unique features of host organs, which together determine organ-specific metastatic behaviors. Emerging insights related to the roles of metabolic changes, the immune landscapes of target organs, and variation in epithelial-mesenchymal transitions open avenues for future studies of metastasis organotropism., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Background: Triple-negative breast cancers (TNBCs), which lack receptors for estrogen, progesterone, and amplification of epidermal growth factor receptor 2, are highly aggressive. Consequently, patients diagnosed with TNBCs have reduced overall and disease-free survival rates compared to patients with other subtypes of breast cancer. TNBCs are characterized by the presence of cancer cells with mesenchymal properties, indicating that the epithelial to mesenchymal transition (EMT) plays a major role in the progression of this disease. The EMT program has also been implicated in chemoresistance, tumor recurrence, and induction of cancer stem cell (CSC) properties. Currently, there are no targeted therapies for TNBC, and hence, it is critical to identify the novel targets to treat TNBC., Methods: A library of compounds was screened for their ability to inhibit EMT in cells with mesenchymal phenotype as assessed using the previously described Z-cad reporters. Of the several drugs tested, GSK3β inhibitors were identified as EMT inhibitors. The effects of GSK3β inhibitors on the properties of TNBC cells with a mesenchymal phenotype were assessed using qRT-PCR, flow cytometry, western blot, mammosphere, and migration and cell viability assays. Publicly available datasets also were analyzed to examine if the expression of GSK3β correlates with the overall survival of breast cancer patients., Results: We identified a GSK3β inhibitor, BIO, in a drug screen as one of the most potent inhibitors of EMT. BIO and two other GSK3β inhibitors, TWS119 and LiCl, also decreased the expression of mesenchymal markers in several different cell lines with a mesenchymal phenotype. Further, inhibition of GSK3β reduced EMT-related migratory properties of cells with mesenchymal properties. To determine if GSK3β inhibitors target mesenchymal-like cells by affecting the CSC population, we employed mammosphere assays and profiled the stem cell-related cell surface marker CD44+/24- in cells after exposure to GSK3β inhibitors. We found that GSK3β inhibitors indeed decreased the CSC properties of cell types with mesenchymal properties. We treated cells with epithelial and mesenchymal properties with GSK3β inhibitors and found that GSK3β inhibitors selectively kill cells with mesenchymal attributes while sparing cells with epithelial properties. We analyzed patient data to identify genes predictive of poor clinical outcome that could serve as novel therapeutic targets for TNBC. The Wnt signaling pathway is critical to EMT, but among the various factors known to be involved in Wnt signaling, only the higher expression of GSK3β correlated with poorer overall patient survival., Conclusions: Taken together, our data demonstrate that GSK3β is a potential target for TNBCs and suggest that GSK3β inhibitors could serve as selective inhibitors of EMT and CSC properties for the treatment of a subset of aggressive TNBC. GSK3β inhibitors should be tested for use in combination with standard-of-care drugs in preclinical TNBC models.

Mammary gland development is fueled by stem cell self-renewal and differentiation. External cues from the microenvironment coupled with internal cues such as post-transcriptional regulation exerted by microRNAs regulate stem cell behavior and fate. Here, we have identified a miR-205 regulatory network required for mammary gland ductal development and stem cell regeneration following transplantation into the cleared mammary fat pad. In the postnatal mammary gland, miR-205 is predominantly expressed in the basal/stem cell enriched population. Conditional deletion of miR-205 in mammary epithelial cells impairs stem cell self-renewal and mammary regenerative potential in the in vitro mammosphere formation assay and in vivo mammary reconstitution. miR-205 null transplants display significant changes in basal cells, basement membrane, and stroma. NKD1 and PTPA, which inhibit the Wnt signaling pathway, and AMOT, which causes YAP cytoplasmic retention and inactivation were identified as miR-205 downstream mediators. These studies also confirmed that miR-205 is a direct ΔNp63 target gene that is critical for the regulation of basal cell identity. Stem Cells 2018;36:1875-15., (© AlphaMed Press 2018.)

Cooperativity between WNT and FGF signaling is well documented in embryonic development and cancer progression, but the molecular mechanisms underlying this cross-talk remain elusive. In this study, we interrogated the dynamics of RNA levels, ribosome occupancy, and protein expression as a function of inducible FGF signaling in mouse mammary glands with constitutive WNT hyperactivation. Multiomics correlation analysis revealed a substantial discrepancy between RNA and ribosome occupancy levels versus protein levels. However, this discrepancy decreased as cells became premalignant and dynamically responded to FGF signaling, implicating the importance of stringent gene regulation in nontransformed cells. Analysis of individual genes demonstrated that acute FGF hyperactivation increased translation of many stem cell self-renewal regulators, including WNT signaling components, and decreased translation of genes regulating cellular senescence. WNT pathway components translationally upregulated by FGF signaling had long and structured 5' UTRs with a high frequency of polypurine sequences, several of which harbored (CGG) 4 motifs that can fold into either stable G-quadruplexes or other stable secondary structures. The FGF-mediated increase in translation of WNT pathway components was compromised by silvestrol, an inhibitor of EIF4A that clamps EIF4A to polypurine sequences to block 43S scanning and inhibits its RNA-unwinding activity important for translation initiation. Moreover, silvestrol treatment significantly delayed FGF-WNT-driven tumorigenesis. Taken together, these results suggest that FGF signaling selectively enhances translation of structured mRNAs, particularly WNT signaling components, and highlight their vulnerability to inhibitors that target the RNA helicase EIF4A. Significance: The RNA helicase EIF4A may serve as a therapeutic target for breast cancers that require FGF and WNT signaling. Cancer Res; 78(15); 4229-40. ©2018 AACR ., (©2018 American Association for Cancer Research.)

The Notch signaling pathway regulates important cellular processes involved in stem cell maintenance, proliferation, development, survival, and inflammation. These responses to Notch signaling involving both canonical and non-canonical pathways can be spatially and temporally variable and are highly cell-type dependent. Notch signaling can elicit opposite effects in regulating tumorigenicity (tumor-promoting versus tumor-suppressing function) as well as controlling immune cell responses. In various cancer types, Notch signaling elicits a "cancer stem cell (CSC)" phenotype that results in decreased proliferation, but resistance to various therapies, hence potentially contributing to cell dormancy and relapse. CSCs can reshape their niche by releasing paracrine factors and inflammatory cytokines, and the niche in return can support their quiescence and resistance to therapies as well as the immune response. Moreover, Notch signaling is one of the key regulators of hematopoiesis, immune cell differentiation, and inflammation and is implicated in various autoimmune diseases, carcinogenesis (leukemia), and tumor-induced immunosuppression. Notch can control the fate of various T cell types, including Th1, Th2, and the regulatory T cells (Tregs), and myeloid cells including macrophages, dendritic cells, and myeloid-derived suppressor cells (MDSCs). Both MDSCs and Tregs play an important role in supporting tumor cells (and CSCs) and in evading the immune response. In this review, we will discuss how Notch signaling regulates multiple aspects of the tumor-promoting environment by elucidating its role in CSCs, hematopoiesis, normal immune cell differentiation, and subsequently in tumor-supporting immunogenicity.

Functional abdominal pain disorders are highly prevalent in children. These disorders can be present in isolation or combined with organic diseases, such as celiac disease and inflammatory bowel diseases. Intestinal inflammation (infectious and non-infectious) predisposes children to the development of visceral hypersensitivity that can manifest as functional abdominal pain disorders, including irritable bowel syndrome. The new onset of irritable bowel syndrome symptoms in a patient with an underlying organic disease, such as inflammatory bowel disease, is clinically challenging, given that the same symptomatology may represent a flare-up of the inflammatory bowel disease or an overlapping functional abdominal pain disorder. Similarly, irritable bowel syndrome symptoms in a child previously diagnosed with celiac disease may occur due to poorly controlled celiac disease or the overlap with a functional abdominal pain disorder. There is little research on the overlap of functional abdominal disorders with organic diseases in children. Studies suggest that the overlap between functional abdominal pain disorders and inflammatory bowel disease is more common in adults than in children. The causes for these differences in prevalence are unknown. Only a handful of studies have been published on the overlap between celiac disease and functional abdominal pain disorders in children. The present article provides a review of the literature on the overlap between celiac disease, inflammatory bowel disease, and functional abdominal pain disorders in children and establish comparisons with studies conducted on adults., (Copyright © 2018 Asociación Mexicana de Gastroenterología. Publicado por Masson Doyma México S.A. All rights reserved.)

The etiology of chronic pelvic pain syndromes remains unknown. In a murine urinary tract infection (UTI) model, lipopolysaccharide of uropathogenic E. coli and its receptor TLR4 are required for post-UTI chronic pain development. However, downstream mechanisms of post-UTI chronic pelvic pain remain unclear. Because the TRPV1 and MCP-1/CCR2 pathways are implicated in chronic neuropathic pain, we explored their role in post-UTI chronic pain. Mice were infected with the E. coli strain SΦ874, known to produce chronic allodynia, and treated with the TRPV1 antagonist capsazepine. Mice treated with capsazepine at the time of SΦ874 infection failed to develop chronic allodynia, whereas capsazepine treatment of mice at two weeks following SΦ874 infection did not reduce chronic allodynia. TRPV1-deficient mice did not develop chronic allodynia either. Similar results were found using novelty-suppressed feeding (NSF) to assess depressive behavior associated with neuropathic pain. Imaging of reporter mice also revealed induction of MCP-1 and CCR2 expression in sacral dorsal root ganglia following SΦ874 infection. Treatment with a CCR2 receptor antagonist at two weeks post-infection reduced chronic allodynia. Taken together, these results suggest that TRPV1 has a role in the establishment of post-UTI chronic pain, and CCR2 has a role in maintenance of post-UTI chronic pain.

Peripheral nerve injuries impose significant health and economic consequences, yet no surgical repair can deliver a complete recovery of sensory or motor function. Traditional methods of repair are less than ideal: direct coaptation can only be performed when tension-free repair is possible, and transplantation of nerve autograft can cause donor-site morbidity and neuroma formation. Cell-based therapy delivered via nerve conduits has thus been explored as an alternative method of nerve repair in recent years. Stem cells are promising sources of the regenerative core material in a nerve conduit because stem cells are multipotent in function, abundant in supply, and more accessible than the myelinating Schwann cells. Among different types of stem cells, undifferentiated adipose-derived stem cell (uASC), which can be processed from adipose tissue in less than two hours, is a promising yet underexplored cell type. Studies of uASC have emerged in the past decade and have shown that autologous uASCs are non-immunogenic, easy to access, abundant in supply, and efficacious at promoting nerve regeneration. Two theories have been proposed as the primary regenerative mechanisms of uASC: in situ trans-differentiation towards Schwann cells, and secretion of trophic and anti-inflammatory factors. Future studies need to fully elucidate the mechanisms, side effects, and efficacy of uASC-based nerve regeneration so that uASCs can be utilized in clinical settings., Competing Interests: JMR is a board-certified plastic surgeon who specializes in reconstructive surgery, peripheral nerve surgery, microsurgery, and many other areas of plastic surgery. He has published and presented extensively on peripheral nerve repair, regenerative medicine, and telemedicine and cybercare. JMR holds a United States patent on a microelectronic axon processor that restores nerve function after severance

The unfolded protein response (UPR) is a cellular homeostatic mechanism that is activated in many human cancers and plays pivotal roles in tumor progression and therapy resistance. However, the molecular mechanisms for UPR activation and regulation in cancer cells remain elusive. Here, we show that oncogenic MYC regulates the inositol-requiring enzyme 1 (IRE1)/X-box binding protein 1 (XBP1) branch of the UPR in breast cancer via multiple mechanisms. We found that MYC directly controls IRE1 transcription by binding to its promoter and enhancer. Furthermore, MYC forms a transcriptional complex with XBP1, a target of IRE1, and enhances its transcriptional activity. Importantly, we demonstrate that XBP1 is a synthetic lethal partner of MYC. Silencing of XBP1 selectively blocked the growth of MYC-hyperactivated cells. Pharmacological inhibition of IRE1 RNase activity with small molecule inhibitor 8866 selectively restrained the MYC-overexpressing tumor growth in vivo in a cohort of preclinical patient-derived xenograft models and genetically engineered mouse models. Strikingly, 8866 substantially enhanced the efficacy of docetaxel chemotherapy, resulting in rapid regression of MYC-overexpressing tumors. Collectively, these data establish the synthetic lethal interaction of the IRE1/XBP1 pathway with MYC hyperactivation and provide a potential therapy for MYC-driven human breast cancers.

Chronic pelvic pain causes significant patient morbidity and is a challenge to clinicians. Using a murine neurogenic cystitis model that recapitulates key aspects of interstitial cystitis/bladder pain syndrome (IC), we recently showed that pseudorabies virus (PRV) induces severe pelvic allodynia in BALB/c mice relative to C57BL/6 mice. Here, we report that a quantitative trait locus (QTL) analysis of PRV-induced allodynia in F2 CxB progeny identified a polymorphism on chromosome 13, rs6314295 , significantly associated with allodynia (logarithm of odds = 3.11). The nearby gene encoding acyloxyacyl hydrolase ( Aoah) was induced in the sacral spinal cord of PRV-infected mice. AOAH-deficient mice exhibited increased vesicomotor reflex in response to bladder distension, consistent with spontaneous bladder hypersensitivity, and increased pelvic allodynia in neurogenic cystitis and postbacterial chronic pain models. AOAH deficiency resulted in greater bladder pathology and tumor necrosis factor production in PRV neurogenic cystitis, markers of increased bladder mast cell activation. AOAH immunoreactivity was detectable along the bladder-brain axis, including in brain sites previously correlated with human chronic pelvic pain. Finally, AOAH-deficient mice had significantly higher levels of bladder vascular endothelial growth factor, an emerging marker of chronic pelvic pain in humans. These findings indicate that AOAH modulates pelvic pain severity, suggesting that allelic variation in Aoah influences pelvic pain in IC.

A novel freeze-cast porous chitosan conduit for peripheral nerve repair with highly-aligned, double layered porosity, which provides the ideal mechanical and chemical properties was designed, manufactured, and assessed in vivo. Efficacies of the conduit and the control inverted nerve autograft were evaluated in bridging 10-mm Lewis rat sciatic nerve gap at 12 weeks post-implantation. Biocompatibility and regenerative efficacy of the porous chitosan conduit were evaluated through the histomorphometric analysis of longitudinal and transverse sections. The porous chitosan conduit was found to have promising regenerative characteristics, promoting the desired neovascularization, and axonal ingrowth and alignment through a combination of structural, mechanical and chemical cues.

Translation is principally regulated at the initiation stage. The development of the translation initiation (TI) sequencing (TI-seq) technique has enabled the global mapping of TIs and revealed unanticipated complex translational landscapes in metazoans. Despite the wide adoption of TI-seq, there is no computational tool currently available for analyzing TI-seq data. To fill this gap, we develop a comprehensive toolkit named Ribo-TISH, which allows for detecting and quantitatively comparing TIs across conditions from TI-seq data. Ribo-TISH can also predict novel open reading frames (ORFs) from regular ribosome profiling (rRibo-seq) data and outperform several established methods in both computational efficiency and prediction accuracy. Applied to published TI-seq/rRibo-seq data sets, Ribo-TISH uncovers a novel signature of elevated mitochondrial translation during amino-acid deprivation and predicts novel ORFs in 5'UTRs, long noncoding RNAs, and introns. These successful applications demonstrate the power of Ribo-TISH in extracting biological insights from TI-seq/rRibo-seq data.

Puberty is characterized by dynamic tissue remodeling in the mammary gland involving ductal elongation, resolution into the mature epithelial bilayer, and lumen formation. To decipher the cellular mechanisms underlying these processes, we studied the fate of putative stem cells, termed cap cells, present in terminal end buds of pubertal mice. Employing a p63 CreERT2 -based lineage-tracing strategy, we identified a unipotent fate for proliferative cap cells that only generated cells with basal features. Furthermore, we observed that dislocated "cap-in-body" cells underwent apoptosis, which aided lumen formation during ductal development. Basal lineage-specific profiling and genetic loss-of-function experiments revealed a critical role for FOXO transcription factors in mediating these proliferative versus apoptotic fates. Importantly, these studies revealed a mode of WNT signaling-mediated FOXO1 inhibition, potentially mediated through AKT. Together, these data suggest that the WNT pathway confers proliferative and survival advantages on cap cells via regulation of FOXO1 localization., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Introduction: Functional gastrointestinal disorders (FGIDs) are highly prevalent worldwide and are thought to result from the interplay of multiple factors that can vary from region to region. Nationwide studies can help understand the regional epidemiology and the pathogenesis of FGIDs. The objectives of this study were to determine the prevalence of FGIDs in school-children of Colombia and assess associated factors., Methods: A cross-sectional study was carried out at twelve private and public schools in ten cities distributed through the four main geographic regions of Colombia. School-children completed a validated questionnaire to assess functional gastrointestinal disorders according to Rome III criteria. Demographic information and past medical and family history was obtained from the parents., Results: A total of 4394/5062 (86.8%) children participated in the study, with ages ranging from 8-18 years (mean = 11.9, SD = 2.3). The percentage of children with al least on FGID was 23.7%. Disorders of defecation were the most common category FGID (11.7%) followed by abdominal pain related-functional gastrointestinal disorders (10.4%). Children have increased odds of FGIDs if they have separated parents (OR 1.22, P=0.007), attend private school (OR 1.54, P<0.001), or have nausea (OR 3.16, P<0.001)., Conclusions: This large epidemiological study of pediatric FGIDs is the first to evaluate a broad cross-section of children throughout a nation in the Americas. High prevalence of FGIDs and identified associations with their likelihood are relevant when providing medical care and when planning public health efforts., (Copyright © 2017 Hospital Infantil de México Federico Gómez. Publicado por Masson Doyma México S.A. All rights reserved.)

Cellular heterogeneity is a common feature in breast cancer, yet an understanding of the coexistence and regulation of various tumor cell subpopulations remains a significant challenge in cancer biology. In the current study, we approached tumor cell heterogeneity from the perspective of Wnt pathway biology to address how different modes of Wnt signaling shape the behaviors of diverse cell populations within a heterogeneous tumor landscape. Using a syngeneic TP53-null mouse model of breast cancer, we identified distinctions in the topology of canonical Wnt β-catenin-dependent signaling activity and non-canonical β-catenin-independent Ror2-mediated Wnt signaling across subtypes and within tumor cell subpopulations in vivo. We further discovered an antagonistic role for Ror2 in regulating canonical Wnt/β-catenin activity in vivo, where lentiviral shRNA depletion of Ror2 expression augmented canonical Wnt/β-catenin signaling activity across multiple basal-like models. Depletion of Ror2 expression yielded distinct phenotypic outcomes and divergent alterations in gene expression programs among different tumors, despite all sharing basal-like features. Notably, we uncovered cell state plasticity and adhesion dynamics regulated by Ror2, which influenced Ras Homology Family Member A (RhoA) and Rho-Associated Coiled-Coil Kinase 1 (ROCK1) activity downstream of Dishevelled-2 (Dvl2). Collectively, these studies illustrate the integration and collaboration of Wnt pathways in basal-like breast cancer, where Ror2 provides a spatiotemporal function to regulate the balance of Wnt signaling and cellular heterogeneity during tumor progression.

Objectives: Eosinophilic esophagitis (EoE) is a chronic antigen-mediated immune disorder of the esophagus. Consensus guidelines recommend obtaining esophageal, gastric, and duodenal biopsies at diagnostic endoscopy when EoE is suspected. The utility of repeated gastric and duodenal biopsies during follow-up endoscopy in patients previously diagnosed with EoE is not established. The aim of the present study was to explore the role of gastric and duodenal biopsies in children with an established diagnosis of EoE undergoing repeat endoscopy to assess histological response to treatment., Methods: Retrospective chart review of children diagnosed with EoE at a tertiary care center was conducted. A total of 160 patients with EoE with demographic clinical, endoscopic, and histological data at diagnosis and follow-up endoscopy were included. The frequency of gastric and duodenal biopsies at follow-up endoscopy with abnormal histology and their correlation to endoscopic findings was determined., Results: At follow-up endoscopy, 83% (132/160) of patients had gastric and 74% (118/160) had duodenal biopsies. Histology was normal in 81% of gastric and 92% of duodenal biopsies. The most frequent gastric abnormalities were chemical and inactive chronic gastritis. The most frequent duodenal abnormality was villous blunting with increased intraepithelial lymphocytes. Two patients with normal gastric and duodenal histology progressed to eosinophilic gastroenteritis at follow-up endoscopy., Conclusions: Gastric and duodenal biopsies obtained in EoE patients during follow-up endoscopy show pathology in a minority of patients, increase costs, and may add potential risk of adverse events. Large multicenter, prospective studies of endoscopic practice during follow-up of EoE are warranted to provide evidence supporting best practices.

Mechanisms regulating the transition of mammary epithelial cells (MECs) to mammary stem cells (MaSCs) and to tumor-initiating cells (TICs) have not been entirely elucidated. The p53 family member, p63, is critical for mammary gland development and contains transactivation domain isoforms, which have tumor-suppressive activities, and the ΔN isoforms, which act as oncogenes. In the clinic, p63 is often used as a diagnostic marker, and further analysis of the function of TAp63 in the mammary gland is critical for improved diagnosis and patient care. Loss of TAp63 in mice leads to the formation of aggressive metastatic mammary adenocarcinoma at 9-16 months of age. Here we show that TAp63 is crucial for the transition of mammary cancer cells to TICs. When TAp63 is lost, MECs express embryonic and MaSC signatures and activate the Hippo pathway. These data indicate a crucial role for TAp63 in mammary TICs and provide a mechanism for its role as a tumor- and metastasis-suppressor in breast cancer.

In this review, we will discuss how the cell of origin may modulate breast cancer intratumoral heterogeneity (ITH) as well as the role of ITH in the evolution of cancer. The clonal evolution and the cancer stem cell (CSC) models, as well as a model that integrates clonal evolution with a CSC hierarchy, have all been proposed to explain the development of ITH. The extent of ITH correlates with clinical outcome and reflects the cellular complexity and dynamics within a tumor. A unique subtype of breast cancer, the claudin-low subtype that is highly resistant to chemotherapy and most closely resembles mammary epithelial stem cells, will be discussed. Furthermore, we will review how the interactions among various tumor cells, some with distinct mutations, may impact breast cancer treatment. Finally, novel technologies that may help advance our understanding of ITH and lead to improvements in the design of new treatments also will be discussed., (Copyright © 2017 Cold Spring Harbor Laboratory Press; all rights reserved.)

Svoronos and colleagues observed estrogen receptor alpha-positive cells in the tumor stroma of patients with ovarian cancer that appeared to be independent of both the tumor's estrogen receptor status and tumor type. These cells were identified as immunosuppressive myeloid-derived suppressor cells (MDSC) and could be targeted by antiestrogen therapy, thereby leading to the hypothesis that endocrine therapy when combined with immunotherapy may provide a potential therapeutic benefit by helping to reduce immunosuppressive MDSCs. Cancer Discov; 7(1); 17-9. ©2017 AACRSee related article by Svoronos et al., 72., Competing Interests: The authors disclose no potential conflicts of interest., (©2017 American Association for Cancer Research.)

Metastatic competence is contingent upon the aberrant activation of a latent embryonic program, known as the epithelial-mesenchymal transition (EMT), which bestows stem cell properties as well as migratory and invasive capabilities upon differentiated tumor cells. We recently identified the transcription factor FOXC2 as a downstream effector of multiple EMT programs, independent of the EMT-inducing stimulus, and as a key player linking EMT, stem cell traits and metastatic competence in breast cancer. As such, FOXC2 could serve as a potential therapeutic target to attenuate metastasis. However, as FOXC2 is a transcription factor, it is difficult to target by conventional means such as small-molecule inhibitors. Herein, we identify the serine/threonine-specific kinase p38 as a druggable upstream regulator of FOXC2 stability and function that elicits phosphorylation of FOXC2 at serine 367 (S367). Using an orthotopic syngeneic mouse tumor model, we make the striking observation that inhibition of p38-FOXC2 signaling selectively attenuates metastasis without impacting primary tumor growth. In this model, circulating tumor cell numbers are significantly reduced in mice treated with the p38 inhibitor SB203580, relative to vehicle-treated counterparts. Accordingly, genetic or pharmacological inhibition of p38 decreases FOXC2 protein levels, reverts the EMT phenotype and compromises stem cell attributes in vitro. We also identify the EMT-regulator ZEB1-known to directly repress E-cadherin/CDH1-as a downstream target of FOXC2, critically dependent on its activation by p38. Consistent with the notion that activation of the p38-FOXC2 signaling axis represents a critical juncture in the acquisition of metastatic competence, the phosphomimetic FOXC2(S367E) mutant is refractory to p38 inhibition both in vitro and in vivo, whereas the non-phosphorylatable FOXC2(S367A) mutant fails to elicit EMT and upregulate ZEB1. Collectively, our data demonstrate that FOXC2 regulates EMT, stem cell traits, ZEB1 expression and metastasis in a p38-dependent manner, and attest to the potential utility of p38 inhibitors as antimetastatic agents., Competing Interests: SJW, NS, ANP, RS and SAM are inventors of a patent application based on the work described here. The remaining authors declare no conflict of interest.

Background: The purpose was to evaluate the overlap frequency of irritable bowel syndrome (IBS), gastroesophageal reflux disease (GERD), and overactive bladder syndrome (OBS), as well as other gastrointestinal and systemic symptoms, in functional dyspepsia (FD). Additionally, we sought to determine whether adult Rome III FD subtypes were uniquely related to overlap syndromes or symptoms., Methods: The study was a retrospective review of 100 consecutive pediatric patients, age 8-17 years, diagnosed with FD. All had completed a standardized medical history including gastrointestinal and systemic symptoms as well as specific symptoms related to GERD and OBS. The frequency of overlap with IBS, GERD, and OBS were determined for the whole group and for those fulfilling adult FD subtype criteria. Individual symptoms were also compared by FD subtype., Results: Overlap IBS was present in 33 % of the FD patients. At least one GERD symptom was present in 74 % of patients with 41 % reporting heartburn. At least one OBS symptom was present in 44 % of patients with 29 % reporting urinary urgency. Other than pain, the most common reported gastrointestinal symptom was nausea (86 %). Systemic symptoms were common. Overlap syndromes/symptoms did not vary by FD subtype. Postprandial distress syndrome was associated with pain with eating, weight loss, and waking at night to have a stool., Conclusions: FD is a heterogeneous condition in children and adolescents with significant variability in the presence of gastrointestinal and non-gastrointestinal symptoms and overlap syndromes. Varying symptom profiles need to be accounted for and analyzed in studies involving subjects with FD.

Targeted therapies against basal-like breast tumors, which are typically 'triple-negative breast cancers (TNBCs)', remain an important unmet clinical need. Somatic TP53 mutations are the most common genetic event in basal-like breast tumors and TNBC. To identify additional drivers and possible drug targets of this subtype, a comparative study between human and murine tumors was performed by utilizing a murine Trp53-null mammary transplant tumor model. We show that two subsets of murine Trp53-null mammary transplant tumors resemble aspects of the human basal-like subtype. DNA-microarray, whole-genome and exome-based sequencing approaches were used to interrogate the secondary genetic aberrations of these tumors, which were then compared to human basal-like tumors to identify conserved somatic genetic features. DNA copy-number variation produced the largest number of conserved candidate personalized drug targets. These candidates were filtered using a DNA-RNA Pearson correlation cut-off and a requirement that the gene was deemed essential in at least 5% of human breast cancer cell lines from an RNA-mediated interference screen database. Five potential personalized drug target genes, which were spontaneously amplified loci in both murine and human basal-like tumors, were identified: Cul4a, Lamp1, Met, Pnpla6 and Tubgcp3 As a proof of concept, inhibition of Met using crizotinib caused Met-amplified murine tumors to initially undergo complete regression. This study identifies Met as a promising drug target in a subset of murine Trp53-null tumors, thus identifying a potential shared driver with a subset of human basal-like breast cancers. Our results also highlight the importance of comparative genomic studies for discovering personalized drug targets and for providing a preclinical model for further investigations of key tumor signaling pathways., (© 2016. Published by The Company of Biologists Ltd.)

Background: The epithelial to mesenchymal transition (EMT) has been implicated in metastasis and therapy resistance of carcinomas and can endow cancer cells with cancer stem cell (CSC) properties. The ability to detect cancer cells that are undergoing or have completed EMT has typically relied on the expression of cell surface antigens that correlate with an EMT/CSC phenotype. Alternatively these cells may be permanently marked through Cre-mediated recombination or through immunostaining of fixed cells. The EMT process is dynamic, and these existing methods cannot reveal such changes within live cells. The development of fluorescent sensors that mirror the dynamic EMT state by following the expression of bona fide EMT regulators in live cells would provide a valuable new tool for characterizing EMT. In addition, these sensors will allow direct observation of cellular plasticity with respect to the epithelial/mesenchymal state to enable more effective studies of EMT in cancer and development., Results: We generated a lentiviral-based, dual fluorescent reporter system, designated as the Z-cad dual sensor, comprising destabilized green fluorescent protein containing the ZEB1 3' UTR and red fluorescent protein driven by the E-cadherin (CDH1) promoter. Using this sensor, we robustly detected EMT and mesenchymal to epithelial transition (MET) in breast cancer cells by flow cytometry and fluorescence microscopy. Importantly, we observed dynamic changes in cellular populations undergoing MET. Additionally, we used the Z-cad sensor to identify and isolate minor subpopulations of cells displaying mesenchymal properties within a population comprising predominately epithelial-like cells. The Z-cad dual sensor identified cells with CSC-like properties more effectively than either the ZEB1 3' UTR or E-cadherin sensor alone., Conclusions: The Z-cad dual sensor effectively reports the activities of two factors critical in determining the epithelial/mesenchymal state of carcinoma cells. The ability of this stably integrating dual sensor system to detect dynamic fluctuations between these two states through live cell imaging offers a significant improvement over existing methods and helps facilitate the study of EMT/MET plasticity in response to different stimuli and in cancer pathogenesis. Finally, the versatile Z-cad sensor can be adapted to a variety of in vitro or in vivo systems to elucidate whether EMT/MET contributes to normal and disease phenotypes.

Myeloid-derived suppressor cells (MDSCs) play critical roles in primary and metastatic cancer progression. MDSC regulation is widely variable even among patients harbouring the same type of malignancy, and the mechanisms governing such heterogeneity are largely unknown. Here, integrating human tumour genomics and syngeneic mammary tumour models, we demonstrate that mTOR signalling in cancer cells dictates a mammary tumour's ability to stimulate MDSC accumulation through regulating G-CSF. Inhibiting this pathway or its activators (for example, FGFR) impairs tumour progression, which is partially rescued by restoring MDSCs or G-CSF. Tumour-initiating cells (TICs) exhibit elevated G-CSF. MDSCs reciprocally increase TIC frequency through activating Notch in tumour cells, forming a feedforward loop. Analyses of primary breast cancers and patient-derived xenografts corroborate these mechanisms in patients. These findings establish a non-canonical oncogenic role of mTOR signalling in recruiting pro-tumorigenic MDSCs and show how defined cancer subsets may evolve to promote and depend on a distinct immune microenvironment.