Differential Oligomerization of the Deubiquitinases USP25 and USP28 Regulates Their Activities.

Deubiquitinases have emerged as promising drug targets for cancer therapy. The two DUBs USP25 and USP28 share high similarity but vary in their cellular functions. USP28 is known for its tumor-promoting role, whereas USP25 is a regulator of the innate immune system and, recently, a role in tumorigenesis was proposed. We solved the structures of the catalytic domains of both proteins and established substantial differences in their activities. While USP28 is a constitutively active dimer, USP25 presents an auto-inhibited tetramer. Our data indicate that the activation of USP25 is not achieved through substrate or ubiquitin binding. USP25 cancer-associated mutations lead to activation in vitro and in vivo, thereby providing a functional link between auto-inhibition and the cancer-promoting role of the enzyme. Our work led to the identification of significant differences between USP25 and USP28 and provided the molecular basis for the development of new and highly specific anti-cancer drugs.

Structural basis for the bi-specificity of USP25 and USP28 inhibitors.

The development of cancer therapeutics is often hindered by the fact that specific oncogenes cannot be directly pharmaceutically addressed. Targeting deubiquitylases that stabilize these oncogenes provides a promising alternative. USP28 and USP25 have been identified as such target deubiquitylases, and several small-molecule inhibitors indiscriminately inhibiting both enzymes have been developed. To obtain insights into their mode of inhibition, we structurally and functionally characterized USP28 in the presence of the three different inhibitors AZ1, Vismodegib and FT206. The compounds bind into a common pocket acting as a molecular sink. Our analysis provides an explanation why the two enzymes are inhibited with similar potency while other deubiquitylases are not affected. Furthermore, a key glutamate residue at position 366/373 in USP28/USP25 plays a central structural role for pocket stability and thereby for inhibition and activity. Obstructing the inhibitor-binding pocket by mutation of this glutamate may provide a tool to accelerate future drug development efforts for selective inhibitors of either USP28 or USP25 targeting distinct binding pockets.

Real-world effectiveness of dupilumab in patients with asthma: Findings from the US ADVANTAGE study.

BACKGROUND: Dupilumab is approved as an add-on maintenance therapy for patients (≥6 years) with moderate-to-severe asthma. Better understanding of real-world effectiveness is needed. OBJECTIVE: To characterize the real-world effectiveness of dupilumab in asthma management. METHODS: This retrospective study included patients (≥12 years of age) diagnosed with asthma, initiating dupilumab between November 2018 and September 2020. The study used a US electronic medical record database (TriNetX Dataworks, Cambridge, Massachusetts). Asthma exacerbation rates before and after the initiation of dupilumab were analyzed using generalized estimating equations models with Poisson probabilistic link to estimate incidence rate ratios (IRRs). Sensitivity analyses were conducted based on previous exacerbation data, eosinophil levels, history of atopic dermatitis or chronic rhinosinusitis with nasal polyps, previous use of biologics, and presence of SARS-CoV-2 (COVID-19). RESULTS: A total of 2400 patients initiating dupilumab met all study criteria. After initiation of dupilumab, risk of asthma exacerbation was reduced by 44% (IRR, 0.56; 95% CI, 0.47-0.57; P = <0.0001) and systemic corticosteroid prescriptions by 48% (IRR, 0.52; 95% CI, 0.48, 0.56; P = <0.0001) compared with those before initiation of dupilumab. Adjustment for COVID-19 showed a greater reduction in asthma exacerbations (IRR, 0.50; 95% CI, 0.45-0.55; P = <0.0001). CONCLUSION: Current real-world efficacy evidence indicates that dupilumab reduces asthma exacerbations and total systemic corticosteroid prescriptions in clinical practice. The effectiveness of dupilumab was observed independent of exacerbation history, eosinophil levels, or COVID-19 impact.

Direct to Angiosuite in Acute Stroke with Mobile Stroke Unit.

BACKGROUND: Early reperfusion has the best likelihood for a favorable outcome in acute ischemic stroke (AIS) with large vessel occlusion (LVO). Our experience with mobile stroke unit (MSU) for direct to angiosuite (DTAS) transfer in AIS patients with suspected LVO is presented. METHODS: Retrospective review of prospectively collected data from November 2019 to August 2022, of patients evaluated and transferred by the University of Alberta Hospital MSU and moved to angiosuite for endovascular thrombectomy (EVT). RESULT: A total of 41 cases were included. Nine were chosen for DTAS and 32 were shifted to angiosuite after stopping for computed tomography (CT) angiography of the head and neck (no-DTAS). Stroke severity measured by NIHSS (median with interquartile range (IQR)) was higher in patients of DTAS, 22 (14-24) vs 14.5 (5-25) in no-DTAS (p = 0.001). The non-contrast CT head in MSU showed hyperdense vessels in 8 (88.88%) DTAS vs 11 (34.35%) no-DTAS patients (p = 0.003). The EVT timelines (median with IQR, 90th percentile) including "door to artery puncture time" were 31 (23-50, 49.2) vs 79 (39-264, 112.8) minutes, and "door to recanalization time" was 69 (49-110, 93.2) vs 105.5 (52-178, 159.5) minutes in DTAS vs no-DTAS group, respectively. The workflow times were significantly shorter in the DTAS group (p < 0.001). Eight (88.88%) out of 9 DTAS patients had LVO and underwent thrombectomy. CONCLUSIONS: MSU for DTAS in patients with high NIHSS scores, cortical signs, and CT showing hyperdense vessel is an effective strategy to reduce the EVT workflow time.

Real-world effectiveness of dupilumab versus benralizumab and mepolizumab.

INTRODUCTION: In the United States, this real-world study compared the effectiveness of dupilumab, benralizumab, andmepolizumab in reducing exacerbations and systemic corticosteroid (SCS) prescriptions among patients with asthma. METHODS: Patients (12 years old) who initiated dupilumab, benralizumab, or mepolizumab (index) between November 2018 and September 2020 were identified by using electronic medical record data. Subjects were included if they had greater than or equal to 12 months of data before and after the index date and two or more severe asthma-related exacerbations before the index date. Differences in baseline characteristics were addressed by using inverse probability treatment weighting (IPTW). Pairwisecomparisons between dupilumab and benralizumab, or mepolizumab were conducted by using negative binomial regression, adjusting for baseline rates and unbalance characteristics (greater than or equal to 10% standardized differences) after IPTW. RESULTS: Overall, a total of 1737 subjects met all criteria: 825 dupilumab, 461 benralizumab, and 451 mepolizumab initiators.In the postindex period, dupilumab was associated with a 24% and 28% significant reduction in the risk of severe asthmaexacerbations versus benralizumab (incidence rate ratio [IRR] 0.76 [95% confidence interval {CI}, 0.67-0.86)] and mepolizumab(IRR 0.72 [95% CI, 0.63-0.82]), respectively. In addition, dupilumab treatment significantly reduced SCS prescriptionsby 16% and 25% versus benralizumab and mepolizumab, respectively (p < 0.05). CONCLUSION: This study represents one of the largest real-world comparisons of biologics (dupilumab, benralizumab, and mepolizumab) for asthma in the United States to date. This analysis shows that the use of dupilumab was associated with a significantly greater reduction in both severe asthma exacerbations and SCS prescriptions compared with benralizumab and mepolizumab.

Advances in the management of acute ischemic stroke.

PURPOSE OF REVIEW: This review aims to summarize the therapeutic advances and evidence in the medical management of acute ischemic stroke (AIS). Recent evidence comparing the efficacy and safety of tenecteplase (TNK) with alteplase for intravenous thrombolysis (IVT) in AIS will be highlighted. Recent advances and evidence on improving micro-circulation following endovascular procedures and neuroprotection will be reviewed. RECENT FINDINGS: A significant number of randomized control studies now support the use of tenecteplase for IVT in AIS. TNK 0.25 mg/kg single bolus is as effective and well tolerated as alteplase 0.9 mg/kg infusion for IVT in AIS. Evidence from randomized control trials (RCTs) has shown effective and well tolerated expansion of the therapeutic window of IVT in the wake-up stroke and up to 9 h after last seen well, using advanced neuroimaging with computed tomography perfusion/MRI. Early evidence suggests that intra-arterial alteplase may help improve microcirculation in patients with large vessel occlusion following successful thrombectomy. However, more trials are required to confirm the results. Similarly, early evidence from a recent RCT showed that remote ischemic conditioning confers potential neuroprotection and improves outcomes in AIS. SUMMARY: Converging evidence has demonstrated that for patients with ischemic stroke presenting at under 4.5 h from the onset, TNK is comparable to alteplase. These data along with the practical advantages of TNK have resulted in a shift to replace intravenous TNK as the standard for thrombolysis. Ongoing studies of IVT with TNK are focussed on defining the optimal dose, expanding the time window with multimodal imaging and defining the role of thrombolysis for bridging patients with stroke due to large vessel occlusion.

Thrombolysis in Stroke Mimics: Comprehensive Stroke Centers vs Telestroke Sites.

BACKGROUND: Hyperacute treatment of acute stroke may lead to thrombolysis in stroke mimics (SM). Our aim was to determine the frequency of thrombolysis in SM in primary stroke centers (PSC) dependent on telestroke versus comprehensive stroke centers (CSC). METHOD: Retrospective review of prospectively collected data from the Quality improvement and Clinical Research (QuICR) registry, the Discharge Abstract Database (DAD), and The National Ambulatory Care Reporting System (NACRS) of consecutive patients treated with intravenous thrombolysis for acute ischemic stroke in Alberta (Canada) from April 2016 to March 2021. RESULT: A total of 2471 patients who received thrombolysis were included. Linking the QuICR registry to DAD 169 (6.83%) patients were identified as SM; however, on our review of the records, only 112 (4.53%) were actual SM. SMs were younger with a mean age of 61.66 (±16.15) vs 71.08 (±14.55) in stroke. National Institute of Health Stroke Scale was higher in stroke with a median (IQR) of 10 (5-17) vs 7 (5-10) in SM. Only one patient (0.89 %) in SM groups had a small parenchymal hemorrhage versus 155 (6.57%) stroke patients had a parenchymal hemorrhage. There was no death among patients of thrombolysed SM during hospitalization versus 276 (11.69%) in stroke. There was no significant difference in the rate of SM among thrombolysed patients between PSC 27 (5.36%) versus CSC 85 (4.3%) (P = 0.312). The most responsible diagnosis of SM was migraine/migraine equivalent, functional disorder, seizure, and delirium. CONCLUSION: The diagnosis of SM may not always be correct when the information is extracted from databases. The rate of thrombolysis in SM via telestroke is similar to treatment in person at CSC.

Electro-Ionic Control of Surface Plasmons in Graphene-Layered Heterostructures.

Precise control of light is indispensable to modern optical communication devices especially as the size of such devices approaches the subwavelength scale. Plasmonic devices are suitable for the development of these optical devices due to the extreme field confinement and its ability to be controlled by tuning the carrier density at the metal/dielectric interface. Here, an electro-ionic controlled plasmonic device consisting of Au/graphene/ion-gel is demonstrated as an optical switch, where an external electric field modulates the real part of the electrical conductivity. The graphene layer enhances charge penetration and charge separation at the Au/graphene interface resulting in an increased photoinduced voltage. The ion-gel immobilized on the Au/graphene further enables the electrical tunability of plasmons which modulates the intensity of the reflected laser light. This work paves the way for developing novel plasmonic electro-optic switches for potential applications such as integrated optical devices.

Proteogenomic analysis of Inhibitor of Differentiation 4 (ID4) in basal-like breast cancer.

BACKGROUND: Basal-like breast cancer (BLBC) is a poorly characterised, heterogeneous disease. Patients are diagnosed with aggressive, high-grade tumours and often relapse with chemotherapy resistance. Detailed understanding of the molecular underpinnings of this disease is essential to the development of personalised therapeutic strategies. Inhibitor of differentiation 4 (ID4) is a helix-loop-helix transcriptional regulator required for mammary gland development. ID4 is overexpressed in a subset of BLBC patients, associating with a stem-like poor prognosis phenotype, and is necessary for the growth of cell line models of BLBC through unknown mechanisms. METHODS: Here, we have defined unique molecular insights into the function of ID4 in BLBC and the related disease high-grade serous ovarian cancer (HGSOC), by combining RIME proteomic analysis, ChIP-seq mapping of genomic binding sites and RNA-seq. RESULTS: These studies reveal novel interactions with DNA damage response proteins, in particular, mediator of DNA damage checkpoint protein 1 (MDC1). Through MDC1, ID4 interacts with other DNA repair proteins (γH2AX and BRCA1) at fragile chromatin sites. ID4 does not affect transcription at these sites, instead binding to chromatin following DNA damage. Analysis of clinical samples demonstrates that ID4 is amplified and overexpressed at a higher frequency in BRCA1-mutant BLBC compared with sporadic BLBC, providing genetic evidence for an interaction between ID4 and DNA damage repair deficiency. CONCLUSIONS: These data link the interactions of ID4 with MDC1 to DNA damage repair in the aetiology of BLBC and HGSOC.

Socioeconomic consequences of drug-resistant epilepsy in an adult cohort from southern India.

OBJECTIVE: The objective of this study was to describe the socioeconomic consequences of drug-resistant epilepsy (DRE). METHODS: This study comprised 132 (equal males and females) consecutive patients aged ≥18 years, who fulfilled the International League Against Epilepsy (ILAE) definition for DRE, prospectively seen in a tertiary care center in South India. We used a structured questionnaire to gather relevant information. RESULTS: The mean age was 31 (range 18-70) years. Mean age of onset of epilepsy was 17 years and mean duration of epilepsy 14 years. The most common epilepsy type and etiology were focal epilepsy and gliotic lesions secondary to perinatal insults, respectively. The average out of the pocket expenditure on antiseizure drugs annually amounted to 19% of the gross national product (GNP)/capita, which was borne by family members in more than two-thirds of the subjects. Almost 60% reported epilepsy having affected their education, 40% their employment, and 90% their marital prospects. Female patients were less often employed outside their homes and had more marital problems compared with males. CONCLUSIONS: In addition to high seizure burden, DRE adversely affects the pursuit of higher education, employment, and marriage. Besides the direct cost of epilepsy, these issues augment both the patient and the caregiver's liability. Socioeconomic consequences of DRE are widely prevalent in developing countries, and this study highlights the need to address them.

Selectivity among Anti-σ Factors by Mycobacterium tuberculosis ClpX Influences Intracellular Levels of Extracytoplasmic Function σ Factors.

Extracytoplasmic function σ factors that are stress inducible are often sequestered in an inactive complex with a membrane-associated anti-σ factor. Mycobacterium tuberculosis membrane-associated anti-σ factors have a small, stable RNA gene A (ssrA)-like degron for targeted proteolysis. Interaction between the unfoldase, ClpX, and a substrate with an accessible degron initiates energy-dependent proteolysis. Four anti-σ factors with a mutation in the degron provided a set of natural substrates to evaluate the influence of the degron on degradation strength in ClpX-substrate processivity. We note that a point mutation in the degron (X-Ala-Ala) leads to an order-of-magnitude difference in the dwell time of the substrate on ClpX. Differences in ClpX/anti-σ interactions were correlated with changes in unfoldase activities. Green fluorescent protein (GFP) chimeras or polypeptides with a length identical to that of the anti-σ factor degron also demonstrate degron-dependent variation in ClpX activities. We show that degron-dependent ClpX activity leads to differences in anti-σ degradation, thereby regulating the release of free σ from the σ/anti-σ complex. M. tuberculosis ClpX activity thus influences changes in gene expression by modulating the cellular abundance of ECF σ factors.IMPORTANCE The ability of Mycobacterium tuberculosis to quickly adapt to changing environmental stimuli occurs by maintaining protein homeostasis. Extracytoplasmic function (ECF) σ factors play a significant role in coordinating the transcription profile to changes in environmental conditions. Release of the σ factor from the anti-σ is governed by the ClpXP2P1 assembly. M. tuberculosis ECF anti-σ factors have an ssrA-like degron for targeted degradation. A point mutation in the degron leads to differences in ClpX-mediated proteolysis and affects the cellular abundance of ECF σ factors. ClpX activity thus synchronizes changes in gene expression with environmental stimuli affecting M. tuberculosis physiology.

Sleep Apnea-Predictor of Functional Outcome in Acute Ischemic Stroke.

BACKGROUND: Sleep apnea is increasingly being recognized as 1 of the important, modifiable risk factors of stroke and cardiovascular diseases. Sleep apnea is thought to impair the functional recovery following stroke. Hence, we evaluated the patients with acute ischemic stroke for prevalence of sleep apnea and compared the functional outcomes of patients with and without sleep apnea, at 3rd month of acute ischemic stroke. METHOD: This study was conducted in Kasturba Medical College (KMC) hospital, Manipal, India, between May 2015 and August 2016. We included 102 consecutive patients of acute ischemic stroke with hemiplegic upper limb power of Medical Research Council (MRC) 3 or less. Sleep apnea was diagnosed in these patients using the sleep disordered Questionnaire, Berlin Questionnaire, and Epworth sleepiness scale. Functional outcome was measured using Barthel score on day 7 and at 3rd month following the onset of stroke. RESULT: Out of 102 patients, sleep apnea was present in 31 (30.6%) patients, more in males (67.7%) and elderly. Hypertension was present in 66.6% of patients with sleep apnea. NIHSS score at admission did not differ between the 2 groups. At 3rd month, the Barthel score calculated was better among patient with no apnea, but this was not statistically significant (P = .119). When mean Barthel score at baseline and 3rd month was calculated using repeated measure Analysis of Variance (ANOVA) between the 2 groups, gain in functional independence in no apnea group was statistically significant (P < .001). CONCLUSION: Sleep-disordered breathing is an independent risk factor for stroke, and sleep apnea is also associated with other known stroke risk factors like hypertension. In acute ischemic stroke, sleep apnea has a negative impact on functional recovery. Sleep apnea is amenable to treatment and should be considered in patients with acute ischemic stroke to improve the chance of recovery, and to reduce the risk of recurrence.

Retrospective assessment of patient characteristics and healthcare costs prior to a diagnosis of Alzheimer's disease in an administrative claims database.

BACKGROUND: The objective of this study was to examine patient characteristics and health care resource utilization (HCRU) in the 36 months prior to a confirmatory diagnosis of Alzheimer's disease (AD) compared to a matched cohort without dementia during the same time interval. METHODS: Patients newly diagnosed with AD (with ≥2 claims) were identified between January 1, 2013 to September 31, 2015, and the date of the second claim for AD was defined as the index date. Patients were enrolled for at least 36 months prior to index date. The AD cohort was matched to a cohort with no AD or dementia codes (1:3) on age, gender, race/ethnicity, and enrollment duration prior to the index date. Descriptive analyses were used to summarize patient characteristics, HCRU, and healthcare costs prior to the confirmatory AD diagnosis. The classification and regression tree analysis and logistic regression were used to identify factors associated with the AD diagnosis. RESULTS: The AD cohort (N = 16,494) had significantly higher comorbidity indices and greater odds of comorbid mental and behavioral diagnoses, including mild cognitive impairment, mood and anxiety disorders, behavioral disturbances, and cerebrovascular disease, heart disease, urinary tract infections, and pneumonia than the matched non-AD or dementia cohort (N = 49,482). During the six-month period before the confirmatory AD diagnosis, AD medication use and diagnosis of mild cognitive impairment, Parkinson's disease, or mood disorder were the strongest predictors of a subsequent confirmatory diagnosis of AD. Greater HCRU and healthcare costs were observed for the AD cohort primarily during the six-month period before the confirmatory AD diagnosis. CONCLUSION: The results of this study demonstrated a higher comorbidity burden and higher costs for patients prior to a diagnosis of AD in comparison to the matched cohort. Several comorbidities were associated with a subsequent diagnosis of AD.

Femtosecond laser-pumped plasmonically enhanced near-infrared random laser based on engineered scatterers.

In this Letter, we report on the design, fabrication, and implementation of a novel plasmon-mode-driven low-threshold near-infrared (NIR) random laser (RL) in the 850-900 nm range based on plasmonic ZnS@Au core-shell scatterers. Plasmon modes in the NIR region are used for nanoscale scatterer engineering of ZnS@Au core-shell particles to enhance scattering, as against pristine ZnS. This plasmonic scattering enhancement coupled with femtosecond (fs) laser pumping is shown to cause a three-fold lasing threshold reduction from 325 µJ/cm2 to 100 µJ/cm2 and a mode Q-factor enhancement from 200 to 540 for ZnS@Au-based RL, as compared to pristine ZnS-based RL. Local field enhancement due to plasmonic ZnS@Au scatterers, as evidenced in the finite-difference time-domain simulation, further adds to this enhancement. This work demonstrates a novel scheme of plasmonic mode coupling in the NIR region and fs excitation in a random laser photonic system, overcoming the inherent deficiencies of weak absorption of gain media and poor scattering cross sections of dielectric scatterers for random lasing in the NIR spectrum.

The role of cancer stem cells in tumor heterogeneity and resistance to therapy.

Cancer is a heterogenous disease displaying marked inter- and intra-tumoral diversity. The existence of cancer stem cells (CSCs) has been experimentally demonstrated in a number of cancer types as a subpopulation of tumor cells that drives the tumorigenic and metastatic properties of the entire cancer. Thus, eradication of the CSC population is critical for the complete ablation of a tumor. This is, however, confounded by the inherent resistance of CSCs to standard anticancer therapies, eventually leading to the outgrowth of resistant tumor cells and relapse in patients. The cellular mechanisms of therapy resistance in CSCs are ascribed to several factors including a state of quiescence, an enhanced DNA damage response and active repair mechanisms, up-regulated expression of drug efflux transporters, as well as the activation of pro-survival signaling pathways and inactivation of apoptotic signaling. Understanding the mechanisms underlying the acquisition of resistance to therapy may hold the key to targeting the CSC population.

Toxin Kid uncouples DNA replication and cell division to enforce retention of plasmid R1 in Escherichia coli cells.

Worldwide dissemination of antibiotic resistance in bacteria is facilitated by plasmids that encode postsegregational killing (PSK) systems. These produce a stable toxin (T) and a labile antitoxin (A) conditioning cell survival to plasmid maintenance, because only this ensures neutralization of toxicity. Shortage of antibiotic alternatives and the link of TA pairs to PSK have stimulated the opinion that premature toxin activation could be used to kill these recalcitrant organisms in the clinic. However, validation of TA pairs as therapeutic targets requires unambiguous understanding of their mode of action, consequences for cell viability, and function in plasmids. Conflicting with widespread notions concerning these issues, we had proposed that the TA pair kis-kid (killing suppressor-killing determinant) might function as a plasmid rescue system and not as a PSK system, but this remained to be validated. Here, we aimed to clarify unsettled mechanistic aspects of Kid activation, and of the effects of this for kis-kid-bearing plasmids and their host cells. We confirm that activation of Kid occurs in cells that are about to lose the toxin-encoding plasmid, and we show that this provokes highly selective restriction of protein outputs that inhibits cell division temporarily, avoiding plasmid loss, and stimulates DNA replication, promoting plasmid rescue. Kis and Kid are conserved in plasmids encoding multiple antibiotic resistance genes, including extended spectrum ß-lactamases, for which therapeutic options are scarce, and our findings advise against the activation of this TA pair to fight pathogens carrying these extrachromosomal DNAs.

MicroRNA profiling of the pubertal mouse mammary gland identifies miR-184 as a candidate breast tumour suppressor gene.

INTRODUCTION: The study of mammalian development has offered many insights into the molecular aetiology of cancer. We previously used analysis of mammary morphogenesis to discover a critical role for GATA-3 in mammary developmental and carcinogenesis. In recent years an important role for microRNAs (miRNAs) in a myriad of cellular processes in development and in oncogenesis has emerged. METHODS: microRNA profiling was conducted on stromal and epithelial cellular subsets microdissected from the pubertal mouse mammary gland. miR-184 was reactivated by transient or stable overexpression in breast cancer cell lines and examined using a series of in vitro (proliferation, tumour-sphere and protein synthesis) assays. Orthotopic xenografts of breast cancer cells were used to assess the effect of miR-184 on tumourigenesis as well as distant metastasis. Interactions between miR-184 and its putative targets were assessed by quantitative PCR, microarray, bioinformatics and 3' untranslated region Luciferase reporter assay. The methylation status of primary patient samples was determined by MBD-Cap sequencing. Lastly, the clinical prognostic significance of miR-184 putative targets was assessed using publicly available datasets. RESULTS: A large number of microRNA were restricted in their expression to specific tissue subsets. MicroRNA-184 (miR-184) was exclusively expressed in epithelial cells and markedly upregulated during differentiation of the proliferative, invasive cells of the pubertal terminal end bud (TEB) into ductal epithelial cells in vivo. miR-184 expression was silenced in mouse tumour models compared to non-transformed epithelium and in a majority of breast cancer cell line models. Ectopic reactivation of miR-184 inhibited the proliferation and self-renewal of triple negative breast cancer (TNBC) cell lines in vitro and delayed primary tumour formation and reduced metastatic burden in vivo. Gene expression studies uncovered multi-factorial regulation of genes in the AKT/mTORC1 pathway by miR-184. In clinical breast cancer tissues, expression of miR-184 is lost in primary TNBCs while the miR-184 promoter is methylated in a subset of lymph node metastases from TNBC patients. CONCLUSIONS: These studies elucidate a new layer of regulation in the PI3K/AKT/mTOR pathway with relevance to mammary development and tumour progression and identify miR-184 as a putative breast tumour suppressor.

ID proteins regulate diverse aspects of cancer progression and provide novel therapeutic opportunities.

The inhibitor of differentiation (ID) proteins are helix-loop-helix transcriptional repressors with established roles in stem cell self-renewal, lineage commitment, and niche interactions. While deregulated expression of ID proteins in cancer was identified more than a decade ago, emerging evidence has revealed a central role for ID proteins in neoplastic progression of multiple tumor types that often mirrors their function in physiological stem and progenitor cells. ID proteins are required for the maintenance of cancer stem cells, self-renewal, and proliferation in a range of malignancies. Furthermore, ID proteins promote metastatic dissemination through their role in remodeling the tumor microenvironment and by promoting tumor-associated endothelial progenitor cell proliferation and mobilization. Here, we discuss the latest findings in this area and the clinical opportunities that they provide.

New insights into the role of ID proteins in breast cancer metastasis: a MET affair.

The establishment of lethal metastases depends on the capacity of a small number of cancer cells to regenerate a tumor after entering a target organ. Stankic and colleagues have identified a role for the inhibitor of differentiation protein, ID1, as a critical regulator of breast cancer stem-like properties and metastatic colonization. Under the control of tumor growth factor-beta signaling, ID1 induces mesenchymal-epithelial transition at the metastatic site by antagonizing the activity of the basic helix-loop-helix transcription factor Twist1. This study sheds light on mechanisms that initiate metastatic outgrowth, and strengthens the concept that epithelial-mesenchymal plasticity is crucial at different stages of metastasis.

Profiling the tyrosine phosphoproteome of different mouse mammary tumour models reveals distinct, model-specific signalling networks and conserved oncogenic pathways.

INTRODUCTION: Although aberrant tyrosine kinase signalling characterises particular breast cancer subtypes, a global analysis of tyrosine phosphorylation in mouse models of breast cancer has not been undertaken to date. This may identify conserved oncogenic pathways and potential therapeutic targets. METHODS: We applied an immunoaffinity/mass spectrometry workflow to three mouse models: murine stem cell virus-Neu, expressing truncated Neu, the rat orthologue of human epidermal growth factor receptor 2, Her2 (HER2); mouse mammary tumour virus-polyoma virus middle T antigen (PyMT); and the p53-/- transplant model (p53). Pathways and protein-protein interaction networks were identified by bioinformatics analysis. Molecular mechanisms underpinning differences in tyrosine phosphorylation were characterised by Western blot analysis and array comparative genomic hybridisation. The functional role of mesenchymal-epithelial transition factor (Met) in a subset of p53-null tumours was interrogated using a selective tyrosine kinase inhibitor (TKI), small interfering RNA (siRNA)-mediated knockdown and cell proliferation assays. RESULTS: The three models could be distinguished on the basis of tyrosine phosphorylation signatures and signalling networks. HER2 tumours exhibited a protein-protein interaction network centred on avian erythroblastic leukaemia viral oncogene homologue 2 (Erbb2), epidermal growth factor receptor and platelet-derived growth factor receptor α, and they displayed enhanced tyrosine phosphorylation of ERBB receptor feedback inhibitor 1. In contrast, the PyMT network displayed significant enrichment for components of the phosphatidylinositol 3-kinase signalling pathway, whereas p53 tumours exhibited increased tyrosine phosphorylation of Met and components or regulators of the cytoskeleton and shared signalling network characteristics with basal and claudin-low breast cancer cells. A subset of p53 tumours displayed markedly elevated cellular tyrosine phosphorylation and Met expression, as well as Met gene amplification. Treatment of cultured p53-null cells exhibiting Met amplification with a selective Met TKI abrogated aberrant tyrosine phosphorylation and blocked cell proliferation. The effects on proliferation were recapitulated when Met was knocked down using siRNA. Additional subtypes of p53 tumours exhibited increased tyrosine phosphorylation of other oncogenes, including Peak1/SgK269 and Prex2. CONCLUSION: This study provides network-level insights into signalling in the breast cancer models utilised and demonstrates that comparative phosphoproteomics can identify conserved oncogenic signalling pathways. The Met-amplified, p53-null tumours provide a new preclinical model for a subset of triple-negative breast cancers.