Identification of alternative protein targets of glutamate-ureido-lysine associated with PSMA tracer uptake in prostate cancer cells.

Prostate-specific membrane antigen (PSMA) is highly overexpressed in most prostate cancers and is clinically visualized using PSMA-specific probes incorporating glutamate-ureido-lysine (GUL). PSMA is effectively absent from certain high-mortality, treatment-resistant subsets of prostate cancers, such as neuroendocrine prostate cancer (NEPC); however, GUL-based PSMA tracers are still reported to have the potential to identify NEPC metastatic tumors. These probes may bind unknown proteins associated with PSMA-suppressed cancers. We have identified the up-regulation of PSMA-like aminopeptidase NAALADaseL and the metabotropic glutamate receptors (mGluRs) in PSMA-suppressed prostate cancers and find that their expression levels inversely correlate with PSMA expression and are associated with GUL-based radiotracer uptake. Furthermore, we identify that NAALADaseL and mGluR expression correlates with a unique cell cycle signature. This provides an opportunity for the future study of the biology of NEPC and potential therapeutic directions. Computationally predicting that GUL-based probes bind well to these targets, we designed and synthesized a fluorescent PSMA tracer to investigate these proteins in vitro, where it shows excellent affinity for PSMA, NAALADaseL, and specific mGluRs associated with poor prognosis.

Atypical cell cycle regulation promotes mammary stem cell expansion during mammary development and tumourigenesis.

BACKGROUND: The cell cycle of mammary stem cells must be tightly regulated to ensure normal homeostasis of the mammary gland to prevent abnormal proliferation and susceptibility to tumorigenesis. The atypical cell cycle regulator, Spy1 can override cell cycle checkpoints, including those activated by the tumour suppressor p53 which mediates mammary stem cell homeostasis. Spy1 has also been shown to promote expansion of select stem cell populations in other developmental systems. Spy1 protein is elevated during proliferative stages of mammary gland development, is found at higher levels in human breast cancers, and promotes susceptibility to mammary tumourigenesis when combined with loss of p53. We hypothesized that Spy1 cooperates with loss of p53 to increase susceptibility to tumour initiation due to changes in susceptible mammary stem cell populations during development and drives the formation of more aggressive stem like tumours. METHODS: Using a transgenic mouse model driving expression of Spy1 within the mammary gland, mammary development and stemness were assessed. These mice were intercrossed with p53 null mice to study the tumourigenic properties of Spy1 driven p53 null tumours, as well as global changes in signaling via RNA sequencing analysis. RESULTS: We show that elevated levels of Spy1 leads to expansion of mammary stem cells, even in the presence of p53, and an increase in mammary tumour formation. Spy1-driven tumours have an increased cancer stem cell population, decreased checkpoint signaling, and demonstrate an increase in therapy resistance. Loss of Spy1 decreases tumor onset and reduces the cancer stem cell population. CONCLUSIONS: This data demonstrates the potential of Spy1 to expand mammary stem cell populations and contribute to the initiation and progression of aggressive, breast cancers with increased cancer stem cell populations.

Tuberin levels during cellular differentiation in brain development.

Tuberin is a member of a large protein complex, Tuberous Sclerosis Complex (TSC), and acts as a sensor for nutrient status regulating protein synthesis and cell cycle progression. Mutations in the Tuberin gene, TSC2, permits the formation of tumors that can lead to developmental defects in many organ systems, including the central nervous system. Tuberin is expressed in the brain throughout development and levels of Tuberin have been found to decrease during neuronal differentiation in cell lines in vitro. Our current work investigates the levels of Tuberin at two stages of embryonic development in vivo, and we study the mRNA and protein levels during a time course using immortalized cell lines in vitro. Our results show that total Tuberin levels are tightly regulated through developmental stages in the embryonic brain. At a cell biology level, we show that Tuberin levels are higher when cells are cultured as neurospheres, and knockdown of Tuberin results in a reduction in the number of neurospheres. This functional data supports the hypothesis that Tuberin is an important regulator of stemness and the reduction of Tuberin levels might support functional differentiation in the central nervous system. Understanding how Tuberin expression is regulated throughout neural development is essential to fully comprehend the role of this protein in several developmental and neural pathologies.

Adolescents' and Adults' Perceptions of Sensory-Based Interventions: A Qualitative Analysis.

IMPORTANCE: Adolescents and adults report that their sensory integration and processing differences affect their occupational performance and quality of life, thus requiring effective sensory-focused interventions. Researchers have yet to investigate this population's experience of occupational therapy interventions designed to remediate these challenges. OBJECTIVE: To explore the perceived experience of adolescents and adults with respect to (1) response to intervention, (2) strategies offered to manage sensory differences, and (3) need for services on completion of an intervention. DESIGN: Retrospective, qualitative study. SETTING: Zoom or phone call. PARTICIPANTS: Eleven adolescents and adults with sensory integration and processing differences who had previously completed occupational therapy interventions. INTERVENTION: Sensory-based intervention based on the principles of Ayres Sensory Integration® (ASI) and the Sensory Therapies and Research Frame of Reference. OUTCOMES AND MEASURES: A semistructured interview to obtain data, followed by an in-depth analysis using an inductive coding process to group initial open codes into themes and common subthemes Results: Open codes were grouped into three core themes: (1) therapist-related factors (what the therapist did in treatment); (2) client-related factors (what the client experienced); and (3) follow-up (future needs of the clients). Four main subthemes of the client-therapist relationship emerged: (1) therapeutic alliance; (2) education and knowledge; (3) strategies, tools, and resources; and (4) future needs. CONCLUSIONS AND RELEVANCE: This study provides a perspective on the experience of adolescents and adults specific to the impact of a sensory-focused occupational therapy intervention on their daily lives. This will help occupational therapists when designing interventions for current and future clients. What This Article Adds: This study highlights the need for further research addressing effective sensory-based interventions for adolescents and adults. It also captures which components of intervention clients deemed helpful and identifies potential targets for future intervention.

Understanding Parents' Experiences With Mainstream Schooling for Their Children With Autism Spectrum Disorder: A Meta-Analysis.

IMPORTANCE: Children with autism spectrum disorder (ASD) face school-related occupational challenges. Attending a mainstream school offers benefits for children's learning and their development of social skills; however, parents express frustration with ensuring their child's unique needs and preferences are met. OBJECTIVE: To gain insight into parents' experiences with the mainstream preschool and primary educational system for their children with ASD. DATA SOURCES: Eleven electronic databases were systematically searched, and processes were followed according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses. STUDY SELECTION AND DATA COLLECTION: Study eligibility was determined through the use of selection criteria and paired independent reviewers. Critical appraisal was conducted using a qualitative research hierarchy and a modified version of the Critical Appraisal Skills Programme tool. Two reviewers synthesized the data into themes, following the Joanna Briggs Institute meta-aggregation process. Twenty-six qualitative studies, representing the voices of 397 parents, are presented in this review (spanning 2013-2021). Articles were set in Westernized and non-Westernized settings and represented culturally and linguistically diverse population groups. FINDINGS: Parents of children with ASD reported a lack of awareness and understanding of their children's unique learning needs in the mainstream school system at all levels. CONCLUSIONS AND RELEVANCE: This review has various occupational therapy practice implications for supporting school-age children with ASD, their parents, and school staff. These include adopting family-centered and ecological approaches, raising awareness, influencing policy, and facilitating collaboration. What This Article Adds: This review provides guidance for occupational therapists working in schools about their practice in working at the individual, targeted, and whole-school levels to address occupational barriers faced by children with ASD.

Parents as Adult Learners in Occupational Therapy for Children With Sensory Challenges: A Scoping Review.

IMPORTANCE: Parent training is an essential part of occupational therapy intervention for children with sensory processing and sensory integration (SP-SI) challenges, and parents' learning needs should be considered. OBJECTIVE: To identify the extent to which adult learning needs are considered in occupational therapy literature addressing parent training as a part of intervention for children with SP-SI challenges. DATA SOURCES: Searches were performed of the following databases: MEDLINE, PsycINFO, CINAHL, Web of Science, EMBASE, and ERIC. The date range was limited to 1990 to 2019 to capture literature focused on family-centered care. STUDY SELECTION AND DATA COLLECTION: Using Arksey and O'Malley's framework for scoping reviews and the Preferred Reporting Items for Systematic Reviews and Meta-Analyses Extension for Scoping Reviews Checklist, the following criteria were used to guide the literature searches: population (parents and families of children with SP-SI challenges), intervention (parent training), outcomes (parent learning needs), and concept (parents as adult learners). FINDINGS: Searches produced 133 peer-reviewed articles, and 5 met the criteria for inclusion. Qualitative thematic analysis, including stakeholder interviews, revealed two themes: (1) Parents focus on children's needs, not their own, and (2) parents benefit from connection with peers and learn through shared experiences. CONCLUSIONS AND RELEVANCE: Parent training is an essential component of occupational therapy; however, there is limited occupational therapy evidence examining parents' learning needs, specifically parents of children with SP-SI challenges. Future studies should investigate parents' learning needs in relation to occupational therapy intervention for families of children with SP-SI challenges. What This Article Adds: Parents' learning needs are both rarely considered in the occupational therapy literature and important for best practice in pediatric therapy for children with SP-SI challenges. The results of this scoping review point to the need for further investigation of parent training programs specifically examining parents as adult learners.

Stabilization of c-Myc by the atypical cell cycle regulator, Spy1, decreases efficacy of breast cancer treatments.

PURPOSE: c-Myc is frequently upregulated in breast cancers, however, targeting c-Myc has proven to be a challenge. Targeting of downstream mediators of c-Myc, such as the 'cyclin-like' cell cycle regulator Spy1, may be a viable therapeutic option in a subset of breast cancer subtypes. METHODS: Mouse mammary tumor cells isolated from MMTV-Myc mice and human breast cancer cell lines were used to manipulate Spy1 levels followed by tamoxifen or chemotherapeutic treatment with a variety of endpoints. Patient samples from TNBC patients were obtained and constructed into a TMA and stained for c-Myc and Spy1 protein levels. RESULTS: Over time, MMTV-Myc cells show a decreased response to tamoxifen treatment with increasing levels of Spy1 in the tamoxifen-resistant cells. shRNA against Spy1 re-establishes tamoxifen sensitivity. Spy1 was found to be highly elevated in human TNBC cell and patient samples, correlating to c-Myc protein levels. c-Myc was found to be stabilized by Spy1 and knocking down Spy1 in TNBC cells shows a significant increase in response to chemotherapy treatments. CONCLUSION: Understanding the interplay between protein expression level and response to treatment is a critical factor in developing novel treatment options for breast cancer patients. These data have shown a connection between Spy1 and c-Myc protein levels in more aggressive breast cancer cells and patient samples. Furthermore, targeting c-Myc has proven difficult, these data suggest targeting Spy1 even when c-Myc is elevated can confer an advantage to current chemotherapies.

The cyclin-like protein SPY1 overrides reprogramming induced senescence through EZH2 mediated H3K27me3.

Fully differentiated cells can be reprogrammed through ectopic expression of key transcription factors to create induced pluripotent stem cells. These cells share many characteristics of normal embryonic stem cells and have great promise in disease modeling and regenerative medicine. The process of remodeling has its limitations, including a very low efficiency due to the upregulation of many antiproliferative genes, including cyclin dependent kinase inhibitors CDKN1A and CDKN2A, which serve to protect the cell by inducing apoptotic and senescent programs. Our data reveals a unique cell cycle mechanism enabling mouse fibroblasts to repress cyclin dependent kinase inhibitors through the activation of the epigenetic regulator EZH2 by a cyclin-like protein SPY1. This data reveals that the SPY1 protein is required for reprogramming to a pluripotent state and is capable of increasing reprogramming efficiency.

Structural basis of divergent cyclin-dependent kinase activation by Spy1/RINGO proteins.

Cyclin-dependent kinases (Cdks) are principal drivers of cell division and are an important therapeutic target to inhibit aberrant proliferation. Cdk enzymatic activity is tightly controlled through cyclin interactions, posttranslational modifications, and binding of inhibitors such as the p27 tumor suppressor protein. Spy1/RINGO (Spy1) proteins bind and activate Cdk but are resistant to canonical regulatory mechanisms that establish cell-cycle checkpoints. Cancer cells exploit Spy1 to stimulate proliferation through inappropriate activation of Cdks, yet the mechanism is unknown. We have determined crystal structures of the Cdk2-Spy1 and p27-Cdk2-Spy1 complexes that reveal how Spy1 activates Cdk. We find that Spy1 confers structural changes to Cdk2 that obviate the requirement of Cdk activation loop phosphorylation. Spy1 lacks the cyclin-binding site that mediates p27 and substrate affinity, explaining why Cdk-Spy1 is poorly inhibited by p27 and lacks specificity for substrates with cyclin-docking sites. We identify mutations in Spy1 that ablate its ability to activate Cdk2 and to proliferate cells. Our structural description of Spy1 provides important mechanistic insights that may be utilized for targeting upregulated Spy1 in cancer.

Barriers to translational research in Windsor Ontario: a survey of clinical care providers and health researchers.

BACKGROUND: Translational research is an ideology focussed on streamlining the transition of novel research into clinical practice to ultimately benefit populations. Central to this approach is overcoming barriers to research involvement and interdisciplinary collaboration. Identifying barriers has been the subject of several studies focused on communities with large academic hospitals. The Windsor-Essex region is currently built around community hospitals which have less of an emphasis on research, employ fewer physicians holding academic appointments and generally do not provide incentivised time for research and training. In this study, we surveyed clinicians and researchers working in Windsor-Essex to gain insight into barriers to translational research important to those working in smaller sized, community-based research networks. METHODS: Using an anonymous close-ended Qualtrics survey distributed via email, we surveyed faculty members from The University of Windsor and clinical care providers from Windsor-Essex (n = 68). This included 24 physicians, 14 allied health professionals, and 30 non-clinician researchers. RESULTS: Managing competing interests, lack of time, funding, infrastructure, and networks were identified by greater than 75% of participants as barriers to research involvement. 62% of physicians identified the lack of permanent post-graduate medical trainees as a barrier. Clinicians were consistently less experienced in research skills compared to others; particularly in publishing results and applying for funding (p < 0.001). Schedule incompatibility, funding issues and identifying interested collaborators with overlapping interests were identified as barriers to interdisciplinary collaboration by 80% of participants. Moreover, 46% of those surveyed were unhappy with their research involvement and these individuals were 13% more likely to perceive research as important for their career progression (p = 0.244). CONCLUSIONS: This study identifies several important barriers to translational research in Windsor-Essex and suggests that many motivated researchers are unhappy with their current involvement. These results will inform decision making in the research community of Windsor-Essex and provides insight for communities of similar size and research capacity. Ultimately, enabling the translation of clinical research in all communities is required to ensure equitable access to cutting edge care.

Cyclin-like proteins tip regenerative balance in the liver to favour cancer formation.

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related deaths worldwide. A variety of factors can contribute to the onset of this disease, including viral infection, obesity, alcohol abuse and non-alcoholic fatty liver disease (NAFLD). These stressors predominantly introduce chronic inflammation leading to liver cirrhosis and finally the onset of HCC; however, approximately 20% of HCC cases arise in the absence of cirrhosis via a poorly defined mechanism. The atypical cyclin-like protein Spy1 is capable of overriding cell cycle checkpoints, promoting proliferation and has been implicated in HCC. We hypothesize that Spy1 promotes sustained proliferation making the liver more susceptible to accumulation of deleterious mutations, leading to the development of non-cirrhotic HCC. We report for the first time that elevation of Spy1 within the liver of a transgenic mouse model leads to enhanced spontaneous liver tumourigenesis. We show that the abundance of Spy1 enhanced fat deposition within the liver and decreased the inflammatory response. Interestingly, Spy1 transgenic mice have a significant reduction in fibrosis and sustained rates of hepatocyte proliferation, and endogenous levels of Spy1 are downregulated during the normal fibrotic response. Our results provide support that abnormal regulation of Spy1 protein drives liver tumorigenesis in the absence of elevated fibrosis and, hence, may represent a potential mechanism behind non-cirrhotic HCC. This work may implicate Spy1 as a prognostic indicator and/or potential target in the treatment of diseases of the liver, such as HCC. The cyclin-like protein Spy1 enhances lipid deposition and reduces fibrosis in the liver. Spy1 also promotes increased hepatocyte proliferation and onset of non-cirrhotic hepatocellular carcinoma (HCC). Thus, Spy1 may be used as a potential target in the treatment of HCC.

The atypical cyclin-like protein Spy1 overrides p53-mediated tumour suppression and promotes susceptibility to breast tumourigenesis.

BACKGROUND: Breast cancer is the most common cancer to affect women and one of the leading causes of cancer-related deaths. Proper regulation of cell cycle checkpoints plays a critical role in preventing the accumulation of deleterious mutations. Perturbations in the expression or activity of mediators of cell cycle progression or checkpoint activation represent important events that may increase susceptibility to the onset of carcinogenesis. The atypical cyclin-like protein Spy1 was isolated in a screen for novel genes that could bypass the DNA damage response. Clinical data demonstrates that protein levels of Spy1 are significantly elevated in ductal and lobular carcinoma of the breast. We hypothesized that elevated Spy1 would override protective cell cycle checkpoints and support the onset of mammary tumourigenesis. METHODS: We generated a transgenic mouse model driving expression of Spy1 in the mammary epithelium. Mammary development, growth characteristics and susceptibility to tumourigenesis were studied. In vitro studies were conducted to investigate the relationship between Spy1 and p53. RESULTS: We found that in the presence of wild-type p53, Spy1 protein is held 'in check' via protein degradation, representing a novel endogenous mechanism to ensure protected checkpoint control. Regulation of Spy1 by p53 is at the protein level and is mediated in part by Nedd4. Mutation or abrogation of p53 is sufficient to allow for accumulation of Spy1 levels resulting in mammary hyperplasia. Sustained elevation of Spy1 results in elevated proliferation of the mammary gland and susceptibility to tumourigenesis. CONCLUSIONS: This mouse model demonstrates for the first time that degradation of the cyclin-like protein Spy1 is an essential component of p53-mediated tumour suppression. Targeting cyclin-like protein activity may therefore represent a mechanism of re-sensitizing cells to important cell cycle checkpoints in a therapeutic setting.

Neuro- and hepatic toxicological profile of (S)-2,4-diaminobutanoic acid in embryonic, adolescent and adult zebrafish.

(S)-2,4-Diaminobutanoic acid (DABA) is a noncanonical amino acid often co-produced by cyanobacteria along with ß-N-methylamino-l-alanine (BMAA) in algal blooms. Although BMAA is a well-established neurotoxin, the toxicity of DABA remains unclear. As part of our development of biocompatible materials, we wish to make use of DABA as both a building block and as the end-product of enzymatically induced depolymerization; however, if it is toxic at very low concentrations, this would not be possible. We examined the toxicity of DABA using both in vivo embryonic and adult zebrafish models. At higher sublethal concentrations (700 µm), the fish demonstrated early signs of cardiotoxicity. Adolescent zebrafish were able to tolerate a higher concentration. Post-mortem histological analysis of juvenile zebrafish showed no liver or brain abnormalities associated with hepato- or neurotoxicity. Combined, these results show that DABA exhibits no overt toxicity at concentrations (100-300 µm) within an order of magnitude of those envisioned for its application. This study further highlights the low cost and ease of using zebrafish as an early-stage toxicological screening tool.

Rationale and design of the EXenatide Study of Cardiovascular Event Lowering (EXSCEL) trial.

Exenatide once-weekly is an extended release formulation of exenatide, a glucagon-like peptide-1 receptor agonist, which can improve glycemic control, body weight, blood pressure, and lipid levels in patients with type 2 diabetes mellitus (T2DM). The EXenatide Study of Cardiovascular Event Lowering (EXSCEL) will compare the impact of adding exenatide once-weekly to usual care with usual care alone on major cardiovascular outcomes. EXSCEL is an academically led, phase III/IV, double-blind, pragmatic placebo-controlled, global trial conducted in 35 countries aiming to enrol 14,000 patients with T2DM and a broad range of cardiovascular risk over approximately 5 years. Participants will be randomized (1:1) to receive exenatide once-weekly 2 mg or matching placebo by subcutaneous injections. The trial will continue until 1,360 confirmed primary composite cardiovascular end points, defined as cardiovascular death, nonfatal myocardial infarction, or nonfatal stroke, have occurred. The primary efficacy hypothesis is that exenatide once-weekly is superior to usual care with respect to the primary composite cardiovascular end point. EXSCEL is powered to detect a 15% relative risk reduction in the exenatide once-weekly group, with 85% power and a 2-sided 5% alpha. The primary safety hypothesis is that exenatide once-weekly is noninferior to usual care with respect to the primary cardiovascular composite end point. Noninferiority will be concluded if the upper limit of the CI is <1.30. EXSCEL will assess whether exenatide once-weekly can reduce cardiovascular events in patients with T2DM with a broad range of cardiovascular risk. It will also provide long-term safety information on exenatide once-weekly in people with T2DM. ClinicalTrials.gov Identifier: NCT01144338.

Activated thrombin-activatable fibrinolysis inhibitor (TAFIa) attenuates breast cancer cell metastatic behaviors through inhibition of plasminogen activation and extracellular proteolysis.

BACKGROUND: Thrombin activatable fibrinolysis inhibitor (TAFI) is a plasma zymogen, which can be converted to activated TAFI (TAFIa) through proteolytic cleavage by thrombin, plasmin, and most effectively thrombin in complex with the endothelial cofactor thrombomodulin (TM). TAFIa is a carboxypeptidase that cleaves carboxyl terminal lysine and arginine residues from protein and peptide substrates, including plasminogen-binding sites on cell surface receptors. Carboxyl terminal lysine residues play a pivotal role in enhancing cell surface plasminogen activation to plasmin. Plasmin has many critical functions including cleaving components of the extracellular matrix (ECM), which enhances invasion and migration of cancer cells. We therefore hypothesized that TAFIa could act to attenuate metastasis. METHODS: To assess the role of TAFIa in breast cancer metastasis, in vitro migration and invasion assays, live cell proteolysis and cell proliferation using MDA-MB-231 and SUM149 cells were carried out in the presence of a TAFIa inhibitor, recombinant TAFI variants, or soluble TM. RESULTS: Inhibition of TAFIa with potato tuber carboxypeptidase inhibitor increased cell invasion, migration and proteolysis of both cell lines, whereas addition of TM resulted in a decrease in all these parameters. A stable variant of TAFIa, TAFIa-CIIYQ, showed enhanced inhibitory effects on cell invasion, migration and proteolysis. Furthermore, pericellular plasminogen activation was significantly decreased on the surface of MDA-MB-231 and SUM149 cells following treatment with various concentrations of TAFIa. CONCLUSIONS: Taken together, these results indicate a vital role for TAFIa in regulating pericellular plasminogen activation and ultimately ECM proteolysis in the breast cancer microenvironment. Enhancement of TAFI activation in this microenvironment may be a therapeutic strategy to inhibit invasion and prevent metastasis of breast cancer cells.

A novel model used to detect differential splice junctions as biomarkers in prostate cancer from RNA-Seq data.

BACKGROUND: In cancer alternative RNA splicing represents one mechanism for flexible gene regulation, whereby protein isoforms can be created to promote cell growth, division and survival. Detecting novel splice junctions in the cancer transcriptome may reveal pathways driving tumorigenic events. In this regard, RNA-Seq, a high-throughput sequencing technology, has expanded the study of cancer transcriptomics in the areas of gene expression, chimeric events and alternative splicing in search of novel biomarkers for the disease. RESULTS: In this study, we propose a new two-dimensional peak finding method for detecting differential splice junctions in prostate cancer using RNA-Seq data. We have designed an integrative process that involves a new two-dimensional peak finding algorithm to combine junctions and then remove irrelevant introns across different samples within a population. We have also designed a scoring mechanism to select the most common junctions. CONCLUSIONS: Our computational analysis on three independent datasets collected from patients diagnosed with prostate cancer reveals a small subset of junctions that may potentially serve as biomarkers for prostate cancer. AVAILABILITY: The pipeline, along with their corresponding algorithms, are available upon request.

Paclitaxel-induced transcriptional regulation of Fas signaling pathway is antagonized by dexamethasone.

Chemotherapy agents like paclitaxel are often a first line of therapy for triple-negative breast cancer patients and a last line of therapy for other aggressive breast cancers. While such agents are effective pro-apoptotic treatments for breast cancer cells, they produce many unwanted side effects. Synthetic glucocorticoids like dexamethasone are commonly prescribed during chemotherapy treatment of breast cancer patients for anti-emetic and anti-hypersensitivity purposes. Dexamethasone, however, is known to enhance cell survival in solid tumors. The prevalent use of dexamethasone in conjunction with paclitaxel in the treatment of breast cancer patients merits concern. Previous studies demonstrate dexamethasone-mediated survival in a subset of breast cancer cells in the presence of paclitaxel. This work expanded the types of cell lines studied and explored the molecular mechanism driving this phenotype. Human breast cancer cell lines representative of several subtypes of breast cancer including triple-negative breast cancer, luminal, and HER2 were treated in the presence and absence of paclitaxel, dexamethasone, and under co-treatment conditions. Cell survival was monitored under differing conditions. We then manipulated aspects of the nuclear factor kappa B (NFkappaB) signaling pathway to elucidate the importance of this pathway in overcoming chemotherapy treatment. All cell lines tested were sensitive to paclitaxel and showed dexamethasone-mediated rescue from paclitaxel-induced apoptosis, but the rescue of overall cell numbers was most pronounced in triple-negative breast cancer cell lines. Dexamethasone and paclitaxel inversely regulated the activity of NFkappaB, which is essential to both paclitaxel-mediated apoptosis and dexamethasone-mediated rescue. The transcriptional target of NFkappaB, Fas receptor, is inversely regulated by paclitaxel and dexamethasone and is a downstream target of paclitaxel-activated NFkappaB. Dexamethasone antagonizes paclitaxel-mediated apoptosis through inhibition of NFkappaB transcription of Fas receptor. Pre-treatment of breast cancer patients with dexamethasone may greatly reduce patient response to paclitaxel. Our study elucidates a novel mechanism of paclitaxel-induced apoptosis in breast cancer cell lines and explicates dexamethasone's antagonism of paclitaxel.

Exenatide once weekly versus liraglutide once daily in patients with type 2 diabetes (DURATION-6): a randomised, open-label study.

BACKGROUND: Glucagon-like peptide-1 receptor agonists exenatide and liraglutide have been shown to improve glycaemic control and reduce bodyweight in patients with type 2 diabetes. We compared the efficacy and safety of exenatide once weekly with liraglutide once daily in patients with type 2 diabetes. METHODS: We did a 26 week, open-label, randomised, parallel-group study at 105 sites in 19 countries between Jan 11, 2010, and Jan 17, 2011. Patients aged 18 years or older with type 2 diabetes treated with lifestyle modification and oral antihyperglycaemic drugs were randomly assigned (1:1), via a computer-generated randomisation sequence with a voice response system, to receive injections of once-daily liraglutide (1·8 mg) or once-weekly exenatide (2 mg). Participants and investigators were not masked to treatment assignment. The primary endpoint was change in glycated haemoglobin (HbA(1c)) from baseline to week 26. Analysis was by intention to treat. This trial is registered with ClinicalTrials.gov, number NCT01029886. FINDINGS: Of 912 randomised patients, 911 were included in the intention-to-treat analysis (450 liraglutide, 461 exenatide). The least-squares mean change in HbA(1c) was greater in patients in the liraglutide group (-1·48%, SE 0·05; n=386) than in those in the exenatide group (-1·28%, 0·05; 390) with the treatment difference (0·21%, 95% CI 0·08-0·33) not meeting predefined non-inferiority criteria (upper limit of CI <0·25%). The most common adverse events were nausea (93 [21%] in the liraglutide group vs 43 [9%] in the exenatide group), diarrhoea (59 [13%] vs 28 [6%]), and vomiting 48 [11%] vs 17 [4%]), which occurred less frequently in the exenatide group and with decreasing incidence over time in both groups. 24 (5%) patients allocated to liraglutide and 12 (3%) allocated to exenatide discontinued participation because of adverse events. INTERPRETATION: Both once daily liraglutide and once weekly exenatide led to improvements in glycaemic control, with greater reductions noted with liraglutide. These findings, plus differences in injection frequency and tolerability, could inform therapeutic decisions for treatment of patients with type 2 diabetes. FUNDING: Eli Lilly and Company and Amylin Pharmaceuticals LLC.

Regulation of biological processes by ubiquitin ligases: a focus on the Pagano Lab's contribution.

Protein homeostasis depends on many fundamental processes including mRNA synthesis, translation, post-translational modifications, and proteolysis. In the late 70s and early 80s the discovery that the small 76 amino acid protein ubiquitin could be attached to target proteins via a multi-stage process involving ubiquitin-activating enzymes, ubiquitin conjugating enzymes, and ubiquitin ligases, revealed an exciting new post-translational mechanism to regulate protein degradation. This cellular system was uncovered using biochemical methods by Avram Hershko, who would later won the Nobel prize for this discovery; however, the biological functions of ubiquitin ligases remained unknown for many years. It was initially described that ubiquitin modifies proteins at one or more lysine residues and once a long ubiquitin chain was assembled, proteins were degraded by the proteasome. Now we know that proteins can be mono-, multimono-, homotypic poly-, or heterotypic poly-ubiquitylated, each of which confers a specific signal that goes beyond protein degradation regulating additional key cellular functions such as signal transduction, protein localization, recognition of damaged proteins, etc.