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New surgical procedures can expose patients to harm and should be carefully evaluated before widespread use. The InSpace balloon (Stryker, USA) is an innovative surgical device used to treat people with rotator cuff tears that cannot be repaired. We aimed to determine the effectiveness of the InSpace balloon for people with irreparable rotator cuff tears.We conducted a double-blind, group-sequential, adaptive randomised controlled trial in 24 hospitals in the UK, comparing arthroscopic debridement of the subacromial space with biceps tenotomy (debridement only group) with the same procedure but including insertion of the InSpace balloon (debridement with device group). Participants had an irreparable rotator cuff tear, which had not resolved with conservative treatment, and they had symptoms warranting surgery. Eligibility was confirmed intraoperatively before randomly assigning (1:1) participants to a treatment group using a remote computer system. Participants and assessors were masked to group assignment. Masking was achieved by using identical incisions for both procedures, blinding the operation note, and a consistent rehabilitation programme was offered regardless of group allocation. The primary outcome was the Oxford Shoulder Score at 12 months. Pre-trial simulations using data from early and late timepoints informed stopping boundaries for two interim analyses. The primary analysis was on a modified intention-to-treat basis, adjusted for the planned interim analysis. The trial was registered with ISRCTN, ISRCTN17825590.Between June 1, 2018, and July 30, 2020, we assessed 385 people for eligibility, of which 317 were eligible. 249 (79%) people consented for inclusion in the study. 117 participants were randomly allocated to a treatment group, 61 participants to the debridement only group and 56 to the debridement with device group. A predefined stopping boundary was met at the first interim analysis and recruitment stopped with 117 participants randomised. 43% of participants were female, 57% were male. We obtained primary outcome data for 114 (97%) participants. The mean Oxford Shoulder Score at 12 months was 34·3 (SD 11·1) in the debridement only group and 30·3 (10·9) in the debridement with device group (mean difference adjusted for adaptive design -4·2 [95% CI -8·2 to -0·26];p=0·037) favouring control. There was no difference in adverse events between the two groups.In an efficient, adaptive trial design, our results favoured the debridement only group. We do not recommend the InSpace balloon for the treatment of irreparable rotator cuff tears.Efficacy and Mechanism Evaluation Programme, a Medical Research Council and National Institute for Health and Care Research partnership.

Although new drug discoveries are revolutionizing cancer treatments, repurposing existing drugs would accelerate the timeline and lower the cost for bringing treatments to cancer patients. Our goal was to repurpose CPI211, a potent and selective antagonist of the thromboxane A2-prostanoid receptor (TPr), a G-protein–coupled receptor that regulates coagulation, blood pressure, and cardiovascular homeostasis. To identify potential new clinical indications for CPI211, we performed a phenome-wide association study (PheWAS) of the gene encoding TPr, TBXA2R, using robust deidentified health records and matched genomic data from more than 29,000 patients. Specifically, PheWAS was used to identify clinical manifestations correlating with a TBXA2R single-nucleotide polymorphism (rs200445019), which generates a T399A substitution within TPr that enhances TPr signaling. Previous studies have correlated 200445019 with chronic venous hypertension, which was recapitulated by this PheWAS analysis. Unexpectedly, PheWAS uncovered an rs200445019 correlation with cancer metastasis across several cancer types. When tested in several mouse models of metastasis, TPr inhibition using CPI211 potently blocked spontaneous metastasis from primary tumors, without affecting tumor cell proliferation, motility, or tumor growth. Further, metastasis following intravenous tumor cell delivery was blocked in mice treated with CPI211. Interestingly, TPr signaling in vascular endothelial cells induced VE-cadherin internalization, diminished endothelial barrier function, and enhanced transendothelial migration by tumor cells, phenotypes that were decreased by CPI211. These studies provide evidence that TPr signaling promotes cancer metastasis, supporting the study of TPr inhibitors as antimetastatic agents and highlighting the use of PheWAS as an approach to accelerate drug repurposing.

S3. Representative images of cells grown in 3D Matrigel for 14D and treated with ABT-263 (1.0 uM). S4. Whole cell lysates from cells treated with ABT-263 (1.0uM) for 0=24 hours.

An estimated 40,000 deaths will be attributed to breast cancer in 2016, underscoring the need for improved therapies. Evading cell death is a major hallmark of cancer, driving tumor progression and therapeutic resistance. To evade apoptosis, cancers use antiapoptotic Bcl-2 proteins to bind to and neutralize apoptotic activators, such as Bim. Investigation of antiapoptotic Bcl-2 family members in clinical breast cancer datasets revealed greater expression and more frequent gene amplification of MCL1 as compared with BCL2 or BCL2L1 (Bcl-xL) across three major molecular breast cancer subtypes, Luminal (A and B), HER2-enriched, and Basal-like. While Mcl-1 protein expression was elevated in estrogen receptor α (ERα)-positive and ERα-negative tumors as compared with normal breast, Mcl-1 staining was higher in ERα+ tumors. Targeted Mcl-1 blockade using RNAi increased caspase-mediated cell death in ERα+ breast cancer cells, resulting in sustained growth inhibition. In contrast, combined blockade of Bcl-2 and Bcl-xL only transiently induced apoptosis, as cells rapidly acclimated through Mcl-1 upregulation and enhanced Mcl-1 activity, as measured in situ using Mcl-1/Bim proximity ligation assays. Importantly, MCL1 gene expression levels correlated inversely with sensitivity to pharmacologic Bcl-2/Bcl-xL inhibition in luminal breast cancer cells, whereas no relationship was seen between the gene expression of BCL2 or BCL2L1 and sensitivity to Bcl-2/Bcl-xL inhibition. These results demonstrate that breast cancers rapidly deploy Mcl-1 to promote cell survival, particularly when challenged with blockade of other Bcl-2 family members, warranting the continued development of Mcl-1–selective inhibitors for targeted tumor cell killing.Implications: Mcl-1 levels predict breast cancer response to inhibitors targeting other Bcl-2 family members, and demonstrate the key role played by Mcl-1 in resistance to this drug class. Mol Cancer Res; 15(3); 259–68. ©2016 AACR.

Flow chart depicting the decision making steps towards selecting TBXA2R as a potential therapeutic target for drug repurposing of CPI211.

Supplemental Table S1. Kaplan-Meier analysis of clinical cancer expression datasets (RNA-Seq) to assess the relationship between disease free patient survival and TBXA2R expression levels, as determined using Kmplot.com software, using the software-generated auto-select best cutoff for TBXA2R expression. N = the number of tumors with correlating clinical patient data. HR = hazard ratio. P value calculated using Log-Rank test.

Efferocytosis is the process by which apoptotic cells are cleared from tissue by phagocytic cells. The removal of apoptotic cells prevents them from undergoing secondary necrosis and releasing their inflammation-inducing intracellular contents. Efferocytosis also limits tissue damage by increasing immunosuppressive cytokines and leukocytes and maintains tissue homeostasis by promoting tolerance to antigens derived from apoptotic cells. Thus, tumor cell efferocytosis following cytotoxic cancer treatment could impart tolerance to tumor cells evading treatment-induced apoptosis with deleterious consequences in tumor residual disease. We report here that efferocytosis cleared apoptotic tumor cells in residual disease of lapatinib-treated HER2+ mammary tumors in MMTV-Neu mice, increased immunosuppressive cytokines, myeloid-derived suppressor cells (MDSC), and regulatory T cells (Treg). Blockade of efferocytosis induced secondary necrosis of apoptotic cells, but failed to prevent increased tumor MDSCs, Treg, and immunosuppressive cytokines. We found that efferocytosis stimulated expression of IFN-γ, which stimulated the expression of indoleamine-2,3-dioxegenase (IDO) 1, an immune regulator known for driving maternal-fetal antigen tolerance. Combined inhibition of efferocytosis and IDO1 in tumor residual disease decreased apoptotic cell- and necrotic cell-induced immunosuppressive phenotypes, blocked tumor metastasis, and caused tumor regression in 60% of MMTV-Neu mice. This suggests that apoptotic and necrotic tumor cells, via efferocytosis and IDO1, respectively, promote tumor ‘homeostasis’ and progression.Significance:These findings show in a model of HER2+ breast cancer that necrosis secondary to impaired efferocytosis activates IDO1 to drive immunosuppression and tumor progression.

Supplemental Figure S6. Tumors harvested on treatment d 7 were assessed by IHC for FoxP3 and Arg-1. Representative images are shown. Original magnification = 400X. Asterisks represent areas of acellular debris. Black arrows indicate tumor infiltrating lymphocytes (TILs). Yellow arrows indicate hyper-condensed nuclei characteristic of apoptotic bodies / apoptotic debris.

Immunohistochemistry was used to measure Ki67 and TUNEL+ cells in tumors harvested at day 5. Representative Images are shown. N = 5. Immunohistochemistry was used to measure Ki67 and TUNEL+ cells in tumors harvested at day 5. Representative Images are shown. N = 5. Western analysis of whole cell lysates collected 16 hours after transfection using antibodies indicated at the left of each panel. MCF7 Cells were transfected, and after 16 h, total RNA was assessed by RT-qPCR to measure expression of the indicated genes involved in pyroptosis. Each point represents the average of three experimental replicates, N = 3. Midlines are average {plus minus} S.D. Student's T-test.

Cancer immunotherapies that remove checkpoint restraints on adaptive immunity are gaining clinical momentum but have not achieved widespread success in breast cancers, a tumor type considered poorly immunogenic and which harbors a decreased presence of tumor-infiltrating lymphocytes. Approaches that activate innate immunity in breast cancer cells and the tumor microenvironment are of increasing interest, based on their ability to induce immunogenic tumor cell death, type I IFNs, and lymphocyte-recruiting chemokines. In agreement with reports in other cancers, we observe loss, downregulation, or mutation of the innate viral nucleotide sensor retinoic acid-inducible gene I (RIG-I/DDX58) in only 1% of clinical breast cancers, suggesting potentially widespread applicability for therapeutic RIG-I agonists that activate innate immunity. This was tested using an engineered RIG-I agonist in a breast cancer cell panel representing each of three major clinical breast cancer subtypes. Treatment with RIG-I agonist resulted in upregulation and mitochondrial localization of RIG-I and activation of proinflammatory transcription factors STAT1 and NF-κB. RIG-I agonist triggered the extrinsic apoptosis pathway and pyroptosis, a highly immunogenic form of cell death in breast cancer cells. RIG-I agonist also induced expression of lymphocyte-recruiting chemokines and type I IFN, confirming that cell death and cytokine modulation occur in a tumor cell–intrinsic manner. Importantly, RIG-I activation in breast tumors increased tumor lymphocytes and decreased tumor growth and metastasis. Overall, these findings demonstrate successful therapeutic delivery of a synthetic RIG-I agonist to induce tumor cell killing and to modulate the tumor microenvironment in vivo.Significance: These findings describe the first in vivo delivery of RIG-I mimetics to tumors, demonstrating a potent immunogenic and therapeutic effect in the context of otherwise poorly immunogenic breast cancers. Cancer Res; 78(21); 6183–95. ©2018 AACR.

Background: Dysregulated inflammation is associated with poor outcomes in COVID-19. We aimed to assess the efficacy of namilumab (a granulocyte-macrophage colony stimulating factor inhibitor) and infliximab (a tumour necrosis factor inhibitor) in hospitalised patients with COVID-19, to prioritise agents for phase 3 trials. Methods: In this randomised, multicentre, multi-arm, multistage, parallel-group, open-label, adaptive, phase 2, proof-of-concept trial (CATALYST), we recruited patients (aged ≥16 years) admitted to hospital with COVID-19 pneumonia and C-reactive protein (CRP) concentrations of 40 mg/L or greater, at nine hospitals in the UK. Participants were randomly assigned with equal probability to usual care or usual care plus a single intravenous dose of namilumab (150 mg) or infliximab (5 mg/kg). Randomisation was stratified by care location within the hospital (ward vs intensive care unit [ICU]). Patients and investigators were not masked to treatment allocation. The primary endpoint was improvement in inflammation, measured by CRP concentration over time, analysed using Bayesian multilevel models. This trial is now complete and is registered with ISRCTN, 40580903. Findings: Between June 15, 2020, and Feb 18, 2021, we screened 299 patients and 146 were enrolled and randomly assigned to usual care (n=54), namilumab (n=57), or infliximab (n=35). For the primary outcome, 45 patients in the usual care group were compared with 52 in the namilumab group, and 29 in the usual care group were compared with 28 in the infliximab group. The probabilities that the interventions were superior to usual care alone in reducing CRP concentration over time were 97% for namilumab and 15% for infliximab; the point estimates for treatment–time interactions were –0·09 (95% CI –0·19 to 0·00) for namilumab and 0·06 (–0·05 to 0·17) for infliximab. 134 adverse events occurred in 30 (55%) of 55 patients in the namilumab group compared with 145 in 29 (54%) of 54 in the usual care group. 102 adverse events occurred in 20 (69%) of 29 patients in the infliximab group compared with 112 in 17 (50%) of 34 in the usual care group. Death occurred in six (11%) patients in the namilumab group compared with ten (19%) in the usual care group, and in four (14%) in the infliximab group compared with five (15%) in the usual care group. Interpretation: Namilumab, but not infliximab, showed proof-of-concept evidence for reduction in inflammation—as measured by CRP concentration—in hospitalised patients with COVID-19 pneumonia. Namilumab should be prioritised for further investigation in COVID-19.

The objective of the study was to find out the prevalence of oral squamous cell carcinoma (OSCC) with oral submucous fibrosis (OSF) in patients with OSF.Of 48,757 patients, we found 300 OSF subjects. Three hundred patients of OSF were checked for OSCC. Both OSF and OSCC with OSF (OSCCwOSF) were diagnosed histopathologically. The prevalence of OSCCwOSF was calculated. Descriptive analysis was done. Chi-square test and t-test were calculated for proportions and mean, respectively, to check any difference among OSF and OSCCwOSF groups. Age-specific relative risk was calculated in OSF and OSCCwOSF groups. Multiple logistic regression analysis was done among odd ratios of the different variable between OSF and OSCCwOSF groups.The prevalence of OSCCwOSF among OSF was 13.7% over a period of 1 year. The mean age of OSCCwOSF group was 43.95 ± 10.22 years in comparison to the OSF group that was 35.51 ± 11.26 years (P0.00). The mean habit duration was significantly less in the OSF group when compared to OSCCwOSFgroup for mishri (P = 0.002). Age-specific adjusted relative risk of OSCC in OSF patient increases from 0.33 (18-34 years) to 3.86 (≥65 years).It could be concluded that a 13.7% prevalence rate of OSCCwOSF in OSF patients should alert the clinician. Clinicians should, therefore, anticipate OSSC in OSF patients. This awareness could lead to the early diagnosis and management of such OSCC.

// Michelle M. Williams 1 , David L. Elion 1 , Bushra Rahman 2 , Donna J. Hicks 2 , Violeta Sanchez 3 and Rebecca S. Cook 1 , 2 , 4 , 5 1 Program in Cancer Biology, Vanderbilt University, Nashville, TN 37232, USA 2 Department of Cell and Developmental Biology, Vanderbilt University, Nashville, TN 37232, USA 3 Department of Medicine, Vanderbilt University Medical Center, Nashville, TN 37232, USA 4 Department of Biomedical Engineering, Vanderbilt University, Nashville TN 37232, USA 5 The Vanderbilt-Ingram Cancer Center, Vanderbilt University Medical Center, Nashville, TN 37232, USA Correspondence to: Rebecca S. Cook, email: Rebecca.cook@vanderbilt.edu Keywords: Mcl-1; ABT-263 resistance; mTORC1 signaling; luminal breast cancers Received: April 23, 2019 Accepted: June 14, 2019 Published: September 10, 2019 ABSTRACT Cancers often overexpress anti-apoptotic Bcl-2 proteins for cell death evasion, a recognized hallmark of cancer progression. While estrogen receptor (ER)-α+ breast cancers express high levels of three anti-apoptotic Bcl-2 family members (Bcl-2, Bcl-xL, and Mcl-1), pharmacological inhibition of Bcl-2 and/or Bcl-xL fails to induce cell death in ERα+ breast cancer cell lines, due to rapid and robust Mcl-1 upregulation. The mechanisms of acute Mcl-1 upregulation in response to Bcl-2/Bcl-xL inhibition remain undefined in in ERα+ breast cancers. We report here that blockade of Bcl-2 or Bcl-xL, alone or together, rapidly induced mTOR signaling in ERα+ breast cancer cells, rapidly increasing cap-dependent Mcl-1 translation. Cells treated with a pharmacological inhibitor of cap-dependent translation, or with the mTORC1 inhibitor RAD001/everolimus, displayed reduced protein levels of Mcl-1 under basal conditions, and failed to upregulate Mcl-1 protein expression following treatment with ABT-263, a pharmacological inhibitor of Bcl-2 and Bcl-xL. Although treatment with ABT-263 alone did not sustain apoptosis in tumor cells in culture or in vivo , ABT-263 plus RAD001 increased apoptosis to a greater extent than either agent used alone. Similarly, combined use of the selective Mcl-1 inhibitor VU661013 with ABT-263 resulted in tumor cell apoptosis and diminished tumor growth in vivo . These findings suggest that rapid Mcl-1 translation drives ABT-263 resistance, but can be combated directly using emerging Mcl-1 inhibitors, or indirectly through existing and approved mTOR inhibitors.

Although the slit diaphragm proteins in podocytes are uniquely organized to maintain glomerular filtration assembly and function, little is known about the underlying mechanisms that participate in trafficking these proteins to the correct location for development and homeostasis. Identifying these mechanisms will likely provide novel targets for therapeutic intervention to preserve podocyte function following glomerular injury. Analysis of structural variation in cases of human nephrotic syndrome identified rare heterozygous deletions of EXOC4 in two patients. This suggested that disruption of the highly-conserved eight-protein exocyst trafficking complex could have a role in podocyte dysfunction. Indeed, mRNA profiling of injured podocytes identified significant exocyst down-regulation. To test the hypothesis that the exocyst is centrally involved in podocyte development/function, we generated homozygous podocyte-specific Exoc5 (a central exocyst component that interacts with Exoc4) knockout mice that showed massive proteinuria and died within 4 weeks of birth. Histological and ultrastructural analysis of these mice showed severe glomerular defects with increased fibrosis, proteinaceous casts, effaced podocytes, and loss of the slit diaphragm. Immunofluorescence analysis revealed that Neph1 and Nephrin, major slit diaphragm constituents, were mislocalized and/or lost. mRNA profiling of Exoc5 knockdown podocytes showed that vesicular trafficking was the most affected cellular event. Mapping of signaling pathways and Western blot analysis revealed significant up-regulation of the mitogen-activated protein kinase and transforming growth factor-β pathways in Exoc5 knockdown podocytes and in the glomeruli of podocyte-specific Exoc5 KO mice. Based on these data, we propose that exocyst-based mechanisms regulate Neph1 and Nephrin signaling and trafficking, and thus podocyte development and function.

Unconventional myosins, linked to deafness, are also proposed to play a role in retinal cell physiology. However, their direct role in photoreceptor function remains unclear. We demonstrate that systemic loss of the unconventional myosin MYO1C in mice, specifically causes rhodopsin mislocalization, leading to impaired visual function. Electroretinogram analysis of Myo1c knockout (Myo1c-KO) mice showed a progressive loss of photoreceptor function. Immunohistochemistry and binding assays demonstrated MYO1C localization to photoreceptor inner and outer segments (OS) and identified a direct interaction of rhodopsin with MYO1C. In Myo1c-KO retinas, rhodopsin mislocalized to rod inner segments (IS) and cell bodies, while cone opsins in OS showed punctate staining. In aged mice, the histological and ultrastructural examination of the phenotype of Myo1c-KO retinas showed progressively shorter photoreceptor OS. These results demonstrate that MYO1C is important for rhodopsin localization to the photoreceptor OS, and for normal visual function.

Our aim was to evaluate the impact of OSF on psychological stress. Ninety OSF cases and age and sex-matched controls, enrolled from relatives or accompanying person were included in the study. Psychological stress was evaluated by the Psychological General Well Being Index short version (PGWBI-S). Sets of the psychological component were generated by principal component analysis (PCA). Association between components was accommodated for confounder and interaction was evaluated by conditional stepwise logistic regression analysis. Psychological component generated was component 1 (depressed mood, lack of positive well being, low vitality, anxiety, low vitality, and low self-control). The odds ratio (OR) of low score of component 1 for OSF was 3.66. Depressed mood, lack of positive well being, low vitality, anxiety, low vitality, and low self-control were associated with OSF. Psychological intervention should, therefore, be included in the management of OSF.

Unconventional myosins linked to deafness are also proposed to play a role in retinal cell physiology. However, their direct role in photoreceptor function remains unclear. We demonstrate that systemic loss of the unconventional myosin MYO1C in mice specifically affected opsin trafficking, leading to loss of visual function. Electroretinogram analysis of Myo1c knockout (Myo1c-KO) mice showed a progressive loss of photoreceptor function. Immunohistochemistry and binding assays demonstrated MYO1C localization to photoreceptor inner and outer segments (OS) and identified a direct interaction of rhodopsin with the MYO1C cargo domain. In Myo1c-KO retinas, rhodopsin mislocalized to rod inner segments (IS) and cell bodies, while cone opsins in OS showed punctate staining. In aged mice, the histological and ultrastructural examination of the phenotype of Myo1c-KO retinas showed progressively shorter photoreceptor OS. These results demonstrate that MYO1C is critical for opsin trafficking to the photoreceptor OS and for normal visual function.

When activated, slit diaphragm proteins NEPHRIN and NEPH1 enable signaling pathways leading to podocyte actin cytoskeleton reorganization, which is critical for podocyte recovery from injury. However, the mechanisms through which these proteins are activated remain unknown. This study presents a novel concept showing ligand-induced activation of NEPHRIN and NEPH1. We first identified phosphatase SHP-2, which directly dephosphorylated these proteins. We next identified HGF, a known SHP-2 modulator, as a rapid inducer of NEPHRIN and NEPH1 phosphorylation. Using baculovirus expressed recombinant purified proteins, SPR (surface plasma resonance), molecular modeling and peptide binding approaches, we show that HGF directly binds NEPHRIN and NEPH1 extracellular domains. Further, using cultured podocytes and Drosophila nephrocytes, we demonstrate that while HGF treatment repaired injured podocytes, the addition of inhibitory NEPH1 or NEPHRIN peptides blocked HGF-induced recovery. Overall, this study shows novel activation and deactivation mechanisms for NEPHRIN and NEPH1 that are required for their function.

Hematological malignancies express high levels of CD47 as a mechanism of immune evasion. CD47-SIRPα triggers a cascade of events that inhibit phagocytosis. Preclinical research supports several models of antibody-mediated blockade of CD47-SIRPα resulting in cell death signaling, phagocytosis of cells bearing stress signals, and priming of tumor-specific T cell responses. Four different antibody molecules designed to target the CD47-SIRPα interaction in malignancy are currently being studied in clinical trials: Hu5F9-G4, CC-90002, TTI-621, and ALX-148. Hu5F9-G4, a humanized anti-CD47 blocking antibody is currently being studied in four different Phase I trials. These studies may lay the groundwork for therapeutic bispecific antibodies. Bispecific antibody (CD20-CD47SL) fusion of anti-CD20 (Rituximab) and anti-CD47 also demonstrated a synergistic effect against lymphoma in preclinical models. This review summarizes the large body of preclinical evidence and emerging clinical data supporting the use of antibodies designed to target the CD47-SIRPα interaction in leukemia, lymphoma and multiple myeloma.

Phosphorylation (activation) and dephosphorylation (deactivation) of the slit diaphragm proteins NEPHRIN and NEPH1 are critical for maintaining the kidney epithelial podocyte actin cytoskeleton and, therefore, proper glomerular filtration. However, the mechanisms underlying these events remain largely unknown. Here we show that NEPHRIN and NEPH1 are novel receptor proteins for hepatocyte growth factor (HGF) and can be phosphorylated independently of the mesenchymal epithelial transition receptor in a ligand-dependent fashion through engagement of their extracellular domains by HGF. Furthermore, we demonstrate SH2 domain-containing protein tyrosine phosphatase-2-dependent dephosphorylation of these proteins. To establish HGF as a ligand, purified baculovirus-expressed NEPHRIN and NEPH1 recombinant proteins were used in surface plasma resonance binding experiments. We report high-affinity interactions of NEPHRIN and NEPH1 with HGF, although NEPHRIN binding was 20-fold higher than that of NEPH1. In addition, using molecular modeling we constructed peptides that were used to map specific HGF-binding regions in the extracellular domains of NEPHRIN and NEPH1. Finally, using an in vitro model of cultured podocytes and an ex vivo model of Drosophila nephrocytes, as well as chemically induced injury models, we demonstrated that HGF-induced phosphorylation of NEPHRIN and NEPH1 is centrally involved in podocyte repair. Taken together, this is the first study demonstrating a receptor-based function for NEPHRIN and NEPH1. This has important biological and clinical implications for the repair of injured podocytes and the maintenance of podocyte integrity.

Chimeric antigen receptor modified T cells targeting CD19 and CD20 have shown activity in Phase I, II trials of patients with hematological malignancies. We conducted a systematic review and meta-analysis of all published clinical trials studying the role of efficacy as well as safety of CD-19 and CD-20 chimeric antigen receptor-T therapy for B-cell hematologic malignancies. A total of 16 studies with 195 patients were identified. The pooled analysis showed an overall response rate of 61% (118/195) with complete response of 42% (81/195) and partial response of 19% (37/195). Major adverse events were cytokine release syndrome 33%, neurotoxicity 33% and B-cell aplasia 54%. Collectively, the results indicate encouraging response in relapsed/refractory B lymphoma and leukemia, especially in acute lymphoblastic leukemia (ALL) patients.

An estimated 40,000 deaths will be attributed to breast cancer in 2016, underscoring the need for improved therapies. Evading cell death is a major hallmark of cancer, driving tumor progression and therapeutic resistance. To evade apoptosis, cancers use antiapoptotic Bcl-2 proteins to bind to and neutralize apoptotic activators, such as Bim. Investigation of antiapoptotic Bcl-2 family members in clinical breast cancer datasets revealed greater expression and more frequent gene amplification of MCL1 as compared with BCL2 or BCL2L1 (Bcl-xL) across three major molecular breast cancer subtypes, Luminal (A and B), HER2-enriched, and Basal-like. While Mcl-1 protein expression was elevated in estrogen receptor α (ERα)-positive and ERα-negative tumors as compared with normal breast, Mcl-1 staining was higher in ERα+ tumors. Targeted Mcl-1 blockade using RNAi increased caspase-mediated cell death in ERα+ breast cancer cells, resulting in sustained growth inhibition. In contrast, combined blockade of Bcl-2 and Bcl-xL only transiently induced apoptosis, as cells rapidly acclimated through Mcl-1 upregulation and enhanced Mcl-1 activity, as measured in situ using Mcl-1/Bim proximity ligation assays. Importantly, MCL1 gene expression levels correlated inversely with sensitivity to pharmacologic Bcl-2/Bcl-xL inhibition in luminal breast cancer cells, whereas no relationship was seen between the gene expression of BCL2 or BCL2L1 and sensitivity to Bcl-2/Bcl-xL inhibition. These results demonstrate that breast cancers rapidly deploy Mcl-1 to promote cell survival, particularly when challenged with blockade of other Bcl-2 family members, warranting the continued development of Mcl-1–selective inhibitors for targeted tumor cell killing. Implications: Mcl-1 levels predict breast cancer response to inhibitors targeting other Bcl-2 family members, and demonstrate the key role played by Mcl-1 in resistance to this drug class. Mol Cancer Res; 15(3); 259–68. ©2016 AACR.

Podocytes have a unique structure that supports glomerular filtration function, and many glomerular diseases result in loss of this structure, leading to podocyte dysfunction and ESRD (end stage renal disease). These structural and functional changes involve a complex set of molecular and cellular mechanisms that remain poorly understood. To understand the molecular signature of podocyte injury, we performed transcriptome analysis of cultured human podocytes injured either with PAN (puromycin aminonucleoside) or doxorubicin/adriamycin (ADR). The pathway analysis through DE (differential expression) and gene-enrichment analysis of the injured podocytes showed Tumor protein p53 (P53) as one of the major signaling pathways that was significantly upregulated upon podocyte injury. Accordingly, P53 expression was also up-regulated in the glomeruli of nephrotoxic serum (NTS) and ADR-injured mice. To further confirm these observations, cultured podocytes were treated with the P53 inhibitor pifithrin-&alpha, which showed significant protection from ADR-induced actin cytoskeleton damage. In conclusion, signaling pathways that are involved in podocyte pathogenesis and can be therapeutically targeted were identified by high-throughput transcriptomic analysis of injured podocytes.

Although new drug discoveries are revolutionizing cancer treatments, repurposing existing drugs would accelerate the timeline and lower the cost for bringing treatments to cancer patients. Our goal was to repurpose CPI211, a potent and selective antagonist of the thromboxane A2-prostanoid receptor (TPr), a G-protein–coupled receptor that regulates coagulation, blood pressure, and cardiovascular homeostasis. To identify potential new clinical indications for CPI211, we performed a phenome-wide association study (PheWAS) of the gene encoding TPr, TBXA2R, using robust deidentified health records and matched genomic data from more than 29,000 patients. Specifically, PheWAS was used to identify clinical manifestations correlating with a TBXA2R single-nucleotide polymorphism (rs200445019), which generates a T399A substitution within TPr that enhances TPr signaling. Previous studies have correlated 200445019 with chronic venous hypertension, which was recapitulated by this PheWAS analysis. Unexpectedly, PheWAS uncovered an rs200445019 correlation with cancer metastasis across several cancer types. When tested in several mouse models of metastasis, TPr inhibition using CPI211 potently blocked spontaneous metastasis from primary tumors, without affecting tumor cell proliferation, motility, or tumor growth. Further, metastasis following intravenous tumor cell delivery was blocked in mice treated with CPI211. Interestingly, TPr signaling in vascular endothelial cells induced VE-cadherin internalization, diminished endothelial barrier function, and enhanced transendothelial migration by tumor cells, phenotypes that were decreased by CPI211. These studies provide evidence that TPr signaling promotes cancer metastasis, supporting the study of TPr inhibitors as antimetastatic agents and highlighting the use of PheWAS as an approach to accelerate drug repurposing.

Podocytes have limited ability to recover from injury. Here, we demonstrate that increased mitochondrial biogenesis, to meet the metabolic and energy demand of a cell, accelerates podocyte recovery from injury. Analysis of events induced during podocyte injury and recovery showed marked upregulation of peroxisome proliferator-activated receptor-γ coactivator–1α (PGC-1α), a transcriptional co-activator of mitochondrial biogenesis, and key components of the mitochondrial electron transport chain. To evaluate our hypothesis that increasing mitochondrial biogenesis enhanced podocyte recovery from injury, we treated injured podocytes with formoterol, a potent, specific, and long-acting β2-adrenergic receptor agonist that induces mitochondrial biogenesis in vitro and in vivo. Formoterol increased mitochondrial biogenesis and restored mitochondrial morphology and the injury-induced changes to the organization of the actin cytoskeleton in podocytes. Importantly, β2-adrenergic receptors were found to be present on podocyte membranes. Their knockdown attenuated formoterol-induced mitochondrial biogenesis. To determine the potential clinical relevance of these findings, mouse models of acute nephrotoxic serum nephritis and chronic (Adriamycin [doxorubicin]) glomerulopathy were used. Mice were treated with formoterol post-injury when glomerular dysfunction was established. Strikingly, formoterol accelerated the recovery of glomerular function by reducing proteinuria and ameliorating kidney pathology. Furthermore, formoterol treatment reduced cellular apoptosis and increased the expression of the mitochondrial biogenesis marker PGC-1α and multiple electron transport chain proteins. Thus, our results support β2-adrenergic receptors as novel therapeutic targets and formoterol as a therapeutic compound for treating podocytopathies.

Efferocytosis is the process by which apoptotic cells are cleared from tissue by phagocytic cells. The removal of apoptotic cells prevents them from undergoing secondary necrosis and releasing their inflammation-inducing intracellular contents. Efferocytosis also limits tissue damage by increasing immunosuppressive cytokines and leukocytes and maintains tissue homeostasis by promoting tolerance to antigens derived from apoptotic cells. Thus, tumor cell efferocytosis following cytotoxic cancer treatment could impart tolerance to tumor cells evading treatment-induced apoptosis with deleterious consequences in tumor residual disease. We report here that efferocytosis cleared apoptotic tumor cells in residual disease of lapatinib-treated HER2+ mammary tumors in MMTV-Neu mice, increased immunosuppressive cytokines, myeloid-derived suppressor cells (MDSC), and regulatory T cells (Treg). Blockade of efferocytosis induced secondary necrosis of apoptotic cells, but failed to prevent increased tumor MDSCs, Treg, and immunosuppressive cytokines. We found that efferocytosis stimulated expression of IFN-γ, which stimulated the expression of indoleamine-2,3-dioxegenase (IDO) 1, an immune regulator known for driving maternal-fetal antigen tolerance. Combined inhibition of efferocytosis and IDO1 in tumor residual disease decreased apoptotic cell- and necrotic cell-induced immunosuppressive phenotypes, blocked tumor metastasis, and caused tumor regression in 60% of MMTV-Neu mice. This suggests that apoptotic and necrotic tumor cells, via efferocytosis and IDO1, respectively, promote tumor ‘homeostasis’ and progression. Significance: These findings show in a model of HER2+ breast cancer that necrosis secondary to impaired efferocytosis activates IDO1 to drive immunosuppression and tumor progression.

Introduction: This study is designed to determine whether a full randomised controlled trial (RCT) examining the clinical effectiveness and safety of total knee replacement surgery with or without a tourniquet is warranted and feasible.\ud \ud Method and analysis: Single centre, patient-blinded and assessor-blinded RCT. A computer-generated randomisation service will allocate 50 participants into one of two trial treatments, surgery with or without a tourniquet. The primary objective is to estimate recruitment, crossovers and follow-up of patients. All patients will have an MRI scan of their brain preoperatively and day 1 or 2 postoperatively to identify ischaemic cerebral emboli (primary clinical outcome). Oxford Cognitive Screen, Montreal Cognitive Assessment and Mini-Mental State Examination will be evaluated as outcome tools for measuring cognitive impairment at days 1, 2 and 7 postoperatively. Thigh pain, blood transfusion requirements, venous thromboembolism, revision surgery, surgical complications, mortality and Oxford knee and five-level EuroQol-5D scores will be collected over 12 months. Integrated qualitative research study: 30 trial patients and 20 knee surgeons will take part in semistructured interviews. Interviews will capture views regarding the pilot trial and explore barriers and potential solutions to a full trial. Multicentre cohort study: UK National Joint Registry data will be linked to Hospital Episode Statistics to estimate the relationship between tourniquet use and venous thromboembolic event, length of hospital stay, risk of revision surgery and death. The study will conclude with a multidisciplinary workshop to reach a consensus on whether a full trial is warranted and feasible.\ud \ud Ethics and dissemination: National Research Ethics Committee (West Midlands-Edgbaston) approved this study on 27 January 2016 (15/WM/0455). The study is sponsored by University of Warwick and University Hospitals Coventry and Warwickshire. The results will be disseminated via high-impact peer-reviewed publication.\ud \ud Trial registration number ISRCTN20873088; Pre-results.

Cancer immunotherapies that remove checkpoint restraints on adaptive immunity are gaining clinical momentum but have not achieved widespread success in breast cancers, a tumor type considered poorly immunogenic and which harbors a decreased presence of tumor-infiltrating lymphocytes. Approaches that activate innate immunity in breast cancer cells and the tumor microenvironment are of increasing interest, based on their ability to induce immunogenic tumor cell death, type I IFNs, and lymphocyte-recruiting chemokines. In agreement with reports in other cancers, we observe loss, downregulation, or mutation of the innate viral nucleotide sensor retinoic acid-inducible gene I (RIG-I/DDX58) in only 1% of clinical breast cancers, suggesting potentially widespread applicability for therapeutic RIG-I agonists that activate innate immunity. This was tested using an engineered RIG-I agonist in a breast cancer cell panel representing each of three major clinical breast cancer subtypes. Treatment with RIG-I agonist resulted in upregulation and mitochondrial localization of RIG-I and activation of proinflammatory transcription factors STAT1 and NF-κB. RIG-I agonist triggered the extrinsic apoptosis pathway and pyroptosis, a highly immunogenic form of cell death in breast cancer cells. RIG-I agonist also induced expression of lymphocyte-recruiting chemokines and type I IFN, confirming that cell death and cytokine modulation occur in a tumor cell–intrinsic manner. Importantly, RIG-I activation in breast tumors increased tumor lymphocytes and decreased tumor growth and metastasis. Overall, these findings demonstrate successful therapeutic delivery of a synthetic RIG-I agonist to induce tumor cell killing and to modulate the tumor microenvironment in vivo. Significance: These findings describe the first in vivo delivery of RIG-I mimetics to tumors, demonstrating a potent immunogenic and therapeutic effect in the context of otherwise poorly immunogenic breast cancers. Cancer Res; 78(21); 6183–95. ©2018 AACR.

Estrogen receptor-α positive (ERα+) breast cancer accounts for approximately 70–80% of the nearly 25,0000 new cases of breast cancer diagnosed in the US each year. Endocrine-targeted therapies (those that block ERα activity) serve as the first line of treatment in most cases. Despite the proven benefit of endocrine therapies, however, ERα+ breast tumors can develop resistance to endocrine therapy, causing disease progression or relapse, particularly in the metastatic setting. Anti-apoptotic Bcl-2 family proteins enhance breast tumor cell survival, often promoting resistance to targeted therapies, including endocrine therapies. Herein, we investigated whether blockade of anti-apoptotic Bcl-2 family proteins could sensitize luminal breast cancers to anti-estrogen treatment. We used long-term estrogen deprivation (LTED) of human ERα+ breast cancer cell lines, an established model of sustained treatment with and acquired resistance to aromatase inhibitors (AIs), in combination with Bcl-2/Bcl-xL inhibition (ABT-263), finding that ABT-263 induced only limited tumor cell killing in LTED-selected cells in culture and in vivo. Interestingly, expression and activity of the Bcl-2-related factor Mcl-1 was increased in LTED cells. Genetic Mcl-1 ablation induced apoptosis in LTED-selected cells, and potently increased their sensitivity to ABT-263. Increased expression and activity of Mcl-1 was similarly seen in clinical breast tumor specimens treated with AI + the selective estrogen receptor downregulator fulvestrant. Delivery of Mcl-1 siRNA loaded into polymeric nanoparticles (MCL1 si-NPs) decreased Mcl-1 expression in LTED-selected and fulvestrant-treated cells, increasing tumor cell death and blocking tumor cell growth. These findings suggest that Mcl-1 upregulation in response to anti-estrogen treatment enhances tumor cell survival, decreasing response to therapeutic treatments. Therefore, strategies blocking Mcl-1 expression or activity used in combination with endocrine therapies would enhance tumor cell death.

Nowadays, embedded linux is widely used operating system due to its compatibility with variety of systems like super computer, work station, personal computer, system on chip (SoC), server and custom computer. The major reason of using linux is high portability and open source code. Also, it supports different type of architectures like- x86, PowerPC, MIPS, H8, SPARC, or ARM and many more. This paper deals with design, implementation and testing of character device driver for the GPIO pins of ARM Cortex based platform. Device driver is a piece of code written in C language which is responsible for controlling the hardware and part of the kernel. ARM based platform Raspberry Pi is used in this work, supports the Linux OS. Due to missing of dependencies and patches, the option of cross compilation is available such as tool chain which contains GCC compiler, assembler, linker and debugger etc.

During pregnancy, as the mammary gland prepares for synthesis and delivery of milk to newborns, a luminal mammary epithelial cell (MEC) subpopulation proliferates rapidly in response to systemic hormonal cues that activate STAT5A. While the receptor tyrosine kinase ErbB4 is required for STAT5A activation in MECs during pregnancy, it is unclear how ErbB3, a heterodimeric partner of ErbB4 and activator of phosphatidyl inositol-3 kinase (PI3K) signaling, contributes to lactogenic expansion of the mammary gland. We assessed mRNA expression levels by expression microarray of mouse mammary glands harvested throughout pregnancy and lactation. To study the role of ErbB3 in mammary gland lactogenesis, we used transgenic mice expressing WAP-driven Cre recombinase to generate a mouse model in which conditional ErbB3 ablation occurred specifically in alveolar mammary epithelial cells (aMECs). Profiling of RNA from mouse MECs isolated throughout pregnancy revealed robust Erbb3 induction during mid-to-late pregnancy, a time point when aMECs proliferate rapidly and undergo differentiation to support milk production. Litters nursed by ErbB3 KO dams weighed significantly less when compared to litters nursed by ErbB3 WT dams. Further analysis revealed substantially reduced epithelial content, decreased aMEC proliferation, and increased aMEC cell death during late pregnancy. Consistent with the potent ability of ErbB3 to activate cell survival through the PI3K/Akt pathway, we found impaired Akt phosphorylation in ErbB3 KO samples, as well as impaired expression of STAT5A, a master regulator of lactogenesis. Constitutively active Akt rescued cell survival in ErbB3-depleted aMECs, but failed to restore STAT5A expression or activity. Interestingly, defects in growth and survival of ErbB3 KO aMECs as well as Akt phosphorylation, STAT5A activity, and expression of milk-encoding genes observed in ErbB3 KO MECs progressively improved between late pregnancy and lactation day 5. We found a compensatory upregulation of ErbB4 activity in ErbB3 KO mammary glands. Enforced ErbB4 expression alleviated the consequences of ErbB3 ablation in aMECs, while combined ablation of both ErbB3 and ErbB4 exaggerated the phenotype. These studies demonstrate that ErbB3, like ErbB4, enhances lactogenic expansion and differentiation of the mammary gland during pregnancy, through activation of Akt and STAT5A, two targets crucial for lactation.

The Higher Secondary Certificate (HSC) Examination in Bangladesh was very crucial for students' future, as well as job lives. As the performance of this examination was influenced by some factors, the main scope of this research was to identify these influencing factors with highest possible precision. In this research, an experimental methodology was adopted to generate a dataset which was constructed from students who passed HSC. Several Data mining methods were used to analyze data, extracting models, and develop knowledge to make decision and also used some feature selection methods to remove unnecessary and redundant features which were helped to improve the performance of the classifiers. Here applied several existent feature selection algorithms and a proposed approach to find the relevant features. The results of the empirical analysis were obtained by removing these irrelevant features with some bayes based, function based, lazy based, rule base and tree base classifiers. And finally the results showed, feature reduction had some effect on performance and for overall LMT classifier with our selected features performed better than others.

Vitamin A (all-trans retinol) plays critical roles in mammalian development and vision. Since vitamin A is food-derived, tissue-specific uptake and storage mechanism are needed. In the eye, uptake of RBP4-retinol is mediated by the receptor Stra6, whereas the receptor mediating RBP4 binding and retinol transport into the liver has just recently been discovered. Here we examined the role of zebrafish retinol binding protein receptor 2 (Rbpr2) for RBP4-retinol uptake in developing embryos, using eye development and vision as sensitive readouts. In cultured cells, Rbpr2 localized to membranes and promoted RBP4-retinol uptake. In larvae, Rbpr2 expression was detected in developing intestinal enterocytes and liver hepatocytes. Two rbpr2 mutant zebrafish lines, each resulting in Rbpr2 deficiency, exhibit a small eye defect, and systemic malformations including hydrocephaly and cardiac edema, phenotypes associated with vitamin A deficiency. In the retina, Rbpr2 loss resulted in shorter photoreceptor outer segments, mislocalization and decrease in visual pigments, decreased expression of retinoic acid-responsive genes and photoreceptor cell loss, overall leading to a reduction of visual function. Together, these results demonstrate that Rbpr2-mediated RBP4-retinol uptake in developing liver and intestine is necessary to provide sufficient substrate for ocular retinoid production required for photoreceptor cell maintenance and visual function.

Gastric cancer (GC) is a leading cause of cancer-related deaths worldwide. The Tff1 knockout (KO) mouse model develops gastric lesions that include low-grade dysplasia (LGD), high-grade dysplasia (HGD), and adenocarcinomas. In this study, we used Affymetrix microarrays gene expression platforms for analysis of molecular signatures in the mouse stomach [Tff1-KO (LGD) and Tff1 wild-type (normal)] and human gastric cancer tissues and their adjacent normal tissue samples. Combined integrated bioinformatics analysis of mouse and human datasets indicated that 172 genes were consistently deregulated in both human gastric cancer samples and Tff1-KO LGD lesions (P

To ignore noisy, skewed, correlated, imbalanced and unnecessary features from real life problems, the feature subset selection with learning algorithm was faced some problems of selecting these relevant features. Several factors like-skewed, high kurtosis valued, dependence or correlation influenced features as well as the classifiers performance. A Dropout technique (feature subset selection method) was used to select relevant and important features and mixed with a classifier named as Probabilistic Neural Network (PNN) to find the performance. This study conducted Dropout technique with PNN to prevent above irrelevant features to get better performance. The main task of this method was to apply several techniques (by omitting some fixed features like-skewed, high kurtosis valued, dependent or correlated and Dropout) with PNN in several datasets and noticed that the overall performance for all datasets was significantly improved or unchanged for Dropout with PNN(DPNN) and DPNN always performed better than others.

RIG-I like receptors, RNA helicases that sense viral oligonucleotide motifs and activate innate immunity, are gaining interest in cancer therapy, given their ability to redirect immune responses within the tumor microenvironment (TME), and increase efficacy of experimental cancer vaccines. RIG-I agonists are not well studied in breast cancers, a type of cancer that is often considered immunologically “silent.” We recently reported that therapeutic delivery of RIG-I agonists increase tumor-infiltrating leukocytes (TILs) and expression of proinflammatory Th1 cytokines in the 4T1 mouse model of aggressive, metastatic breast cancer through tumor cell-intrinsic mechanisms. However, these studies do not rule out the importance of myeloid immune responders (e.g., macrophages and dendritic cells) in propagating the effects of RIG-I agonists against tumor cells in vivo, nor do they rule out the impact of RIG-I agonists on adaptive antitumor immunity, a subject that is relatively understudied. We assessed the effects of the RIG-I agonist SLR20 on the the activity of effector T-lymphocytes (TEff) and regulatory T-lymphocytes (TReg) in the TME. Interestingly, SLR20 treatment of mouse and human breast tumor cells increased expression of FAS and MHC-I on tumor cells, and caused tumor cells to express T-cell chemoattractants (e.g., CXCL10, RANTES), potentially increasing T-cells recruitment to tumors, and increasing tumor cell susceptibility to TEff recognition and killing. Using an ex vivo co-culture assay in which 4T1 mouse mammary tumor cells were co-cultured with CD8+ T-cells harvested from mice pre-inoculated with SLR20-treated 4T1 tumor cells, we measured the rate of CD8+-mediated tumor cell killing. This approach revealed that T-cells harvested from mice inoculated with SLR20-treated cells caused greater tumor cell killing than what was seen by CD8+ T-cells harvested from untreated mice. We also found that conditioned media harvested from 4T1 cells treated with SLR20 increased clonal expansion of CD3/CD28-activated T-cells above what was seen with conditioned media harvested from 4T1 cells treated with a control oligonucleotide or from untreated 4T1 cells. TGFβ-mediated differentiation of CD4+ T-cells into tolerogenic and immunosuppressive TRegs was measured in cultures of CD4+ T-cells treated with cultured media derived from SLR20-treated 4T1 cells. These studies showed that cultured media harvested from 4T1 cells treated with SLR20, but not from untreated 4T1 cells or 4T1 cells treated with a control ligand, diminished TReg differentiation, and decreased CD4+ T-cells surface expression of PD-1, CTLA4, and CCR8. Importantly, in vivo experiments assessing therapeutic treatment of 4T1 tumors with SLR20 revealed greater tumor growth inhibition when SLR20 was combined with PD-L1 targeting antibodies. Taken together, these findings indicate that therapeutic activation of RIG-I signaling operates at the interface of innate and adaptive immunity within breast tumors to redirect the TME from an immunosuppressed state to one that is immunogenic and receptive to clinically relevant checkpoint inhibitors. Citation Format: David L. Elion, Max E. Jacobson, Donna J. Hicks, Bushra Rahman, Violeta Sanchez, Paula I Gonzales-Ericsson, Olga Fedorova, Anna M. Pyle, John T. Wilson, Rebecca S. Cook. RIG-I agonists reinforce antitumor adaptive immunity and decrease Treg activity in breast cancer [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr A187.