Calcineurin Aß-Specific Anchoring Confers Isoform-Specific Compartmentation and Function in Pathological Cardiac Myocyte Hypertrophy.

BACKGROUND: The Ca2+/calmodulin-dependent phosphatase calcineurin is a key regulator of cardiac myocyte hypertrophy in disease. An unexplained paradox is how the ß isoform of the calcineurin catalytic A-subunit (CaNAß) is required for induction of pathological myocyte hypertrophy, despite calcineurin Aα expression in the same cells. It is unclear how the pleiotropic second messenger Ca2+ drives excitation-contraction coupling while not stimulating hypertrophy by calcineurin in the normal heart. Elucidation of the mechanisms conferring this selectivity in calcineurin signaling should reveal new strategies for targeting the phosphatase in disease. METHODS: Primary adult rat ventricular myocytes were studied for morphology and intracellular signaling. New Förster resonance energy transfer reporters were used to assay Ca2+ and calcineurin activity in living cells. Conditional gene deletion and adeno-associated virus-mediated gene delivery in the mouse were used to study calcineurin signaling after transverse aortic constriction in vivo. RESULTS: CIP4 (Cdc42-interacting protein 4)/TRIP10 (thyroid hormone receptor interactor 10) was identified as a new polyproline domain-dependent scaffold for CaNAß2 by yeast 2-hybrid screen. Cardiac myocyte-specific CIP4 gene deletion in mice attenuated pressure overload-induced pathological cardiac remodeling and heart failure. Blockade of CaNAß polyproline-dependent anchoring using a competing peptide inhibited concentric hypertrophy in cultured myocytes; disruption of anchoring in vivo using an adeno-associated virus gene therapy vector inhibited cardiac hypertrophy and improved systolic function after pressure overload. Live cell Förster resonance energy transfer biosensor imaging of cultured myocytes revealed that Ca2+ levels and calcineurin activity associated with the CIP4 compartment were increased by neurohormonal stimulation, but minimally by pacing. Conversely, Ca2+ levels and calcineurin activity detected by nonlocalized Förster resonance energy transfer sensors were induced by pacing and minimally by neurohormonal stimulation, providing functional evidence for differential intracellular compartmentation of Ca2+ and calcineurin signal transduction. CONCLUSIONS: These results support a structural model for Ca2+ and CaNAß compartmentation in cells based on an isoform-specific mechanism for calcineurin protein-protein interaction and localization. This mechanism provides an explanation for the specific role of CaNAß in hypertrophy and its selective activation under conditions of pathologic stress. Disruption of CaNAß polyproline-dependent anchoring constitutes a rational strategy for therapeutic targeting of CaNAß-specific signaling responsible for pathological cardiac remodeling in cardiovascular disease deserving of further preclinical investigation.

Masticatory behaviour and chewing difficulties in young adults with temporomandibular disorders.

BACKGROUND: Difficulties and limitations on masticatory function are among the main reasons why patients with temporomandibular disorder (TMD) seek care. OBJECTIVE: To evaluate the masticatory behaviour and perception of chewing difficulties in adults with mild TMD of recent onset, considering the presence of malocclusion. METHODS: Eighty-one young adults were divided into groups according to the presence of TMD and malocclusion: Non-TMD Normal Occlusion (n = 18), Non-TMD Malocclusion (n = 22), TMD Normal Occlusion (n = 18) and TMD Malocclusion (n = 23). TMD was assessed using the TMD Research Diagnostic Criteria and volunteers also answered questionnaires regarding their perception about jaw functional limitation and difficulty to chew foods of different textures. Masticatory and swallowing behaviours were assessed using the Orofacial Myofunctional Evaluation with Scores (OMES) protocol. Chewing time and chewing frequency taken to ingest the test-food were also obtained. Two-way-ANOVA was used to analyse the TMD, occlusion and TMD × occlusion interaction effects. RESULTS: Temporomandibular disorder effect was observed on vertical jaw mobility and jaw function limitation total scores, meaning that groups differed in the perception of opening limitation and mandibular limitation according to TMD status with a medium effect size. Also, more changes in chewing function (OMES-chewing score) and higher chewing frequency was observed in the presence of TMD (P < 0.05). Occlusion effect was only observed on OMES-swallowing score and no TMD × occlusion interaction effect was observed. CONCLUSION: Changes in chewing behaviour, frequency and perception of mandibular limitation was observed in the presence of TMD, pointing out the importance of functional evaluation when planning and establishing a treatment plan.

Histone Methyltransferase G9a Is Required for Cardiomyocyte Homeostasis and Hypertrophy.

BACKGROUND: Correct gene expression programming of the cardiomyocyte underlies the normal functioning of the heart. Alterations to this can lead to the loss of cardiac homeostasis, triggering heart dysfunction. Although the role of some histone methyltransferases in establishing the transcriptional program of postnatal cardiomyocytes during heart development has been shown, the function of this class of epigenetic enzymes is largely unexplored in the adult heart. In this study, we investigated the role of G9a/Ehmt2, a histone methyltransferase that defines a repressive epigenetic signature, in defining the transcriptional program for cardiomyocyte homeostasis and cardiac hypertrophy. METHODS: We investigated the function of G9a in normal and stressed cardiomyocytes with the use of a conditional, cardiac-specific G9a knockout mouse, a specific G9a inhibitor, and high-throughput approaches for the study of the epigenome (chromatin immunoprecipitation sequencing) and transcriptome (RNA sequencing); traditional methods were used to assess cardiac function and cardiovascular disease. RESULTS: We found that G9a is required for cardiomyocyte homeostasis in the adult heart by mediating the repression of key genes regulating cardiomyocyte function via dimethylation of H3 lysine 9 and interaction with enhancer of zeste homolog 2, the catalytic subunit of polycomb repressive complex 2, and MEF2C-dependent gene expression by forming a complex with this transcription factor. The G9a-MEF2C complex was found to be required also for the maintenance of heterochromatin needed for the silencing of developmental genes in the adult heart. Moreover, G9a promoted cardiac hypertrophy by repressing antihypertrophic genes. CONCLUSIONS: Taken together, our findings demonstrate that G9a orchestrates critical epigenetic changes in cardiomyocytes in physiological and pathological conditions, thereby providing novel therapeutic avenues for cardiac pathologies associated with dysregulation of these mechanisms.

Peptidomimetic Targeting of Cavß2 Overcomes Dysregulation of the L-Type Calcium Channel Density and Recovers Cardiac Function.

BACKGROUND: L-type calcium channels (LTCCs) play important roles in regulating cardiomyocyte physiology, which is governed by appropriate LTCC trafficking to and density at the cell surface. Factors influencing the expression, half-life, subcellular trafficking, and gating of LTCCs are therefore critically involved in conditions of cardiac physiology and disease. METHODS: Yeast 2-hybrid screenings, biochemical and molecular evaluations, protein interaction assays, fluorescence microscopy, structural molecular modeling, and functional studies were used to investigate the molecular mechanisms through which the LTCC Cavß2 chaperone regulates channel density at the plasma membrane. RESULTS: On the basis of our previous results, we found a direct linear correlation between the total amount of the LTCC pore-forming Cavα1.2 and the Akt-dependent phosphorylation status of Cavß2 both in a mouse model of diabetic cardiac disease and in 6 diabetic and 7 nondiabetic cardiomyopathy patients with aortic stenosis undergoing aortic valve replacement. Mechanistically, we demonstrate that a conformational change in Cavß2 triggered by Akt phosphorylation increases LTCC density at the cardiac plasma membrane, and thus the inward calcium current, through a complex pathway involving reduction of Cavα1.2 retrograde trafficking and protein degradation through the prevention of dynamin-mediated LTCC endocytosis; promotion of Cavα1.2 anterograde trafficking by blocking Kir/Gem-dependent sequestration of Cavß2, thus facilitating the chaperoning of Cavα1.2; and promotion of Cavα1.2 transcription by the prevention of Kir/Gem-mediated shuttling of Cavß2 to the nucleus, where it limits the transcription of Cavα1.2 through recruitment of the heterochromatin protein 1γ epigenetic repressor to the Cacna1c promoter. On the basis of this mechanism, we developed a novel mimetic peptide that, through targeting of Cavß2, corrects LTCC life-cycle alterations, facilitating the proper function of cardiac cells. Delivery of mimetic peptide into a mouse model of diabetic cardiac disease associated with LTCC abnormalities restored impaired calcium balance and recovered cardiac function. CONCLUSIONS: We have uncovered novel mechanisms modulating LTCC trafficking and life cycle and provide proof of concept for the use of Cavß2 mimetic peptide as a novel therapeutic tool for the improvement of cardiac conditions correlated with alterations in LTCC levels and function.

It's Time for An Epigenomics Roadmap of Heart Failure.

The post-genomic era has completed its first decade. During this time, we have seen an attempt to understand life not just through the study of individual isolated processes, but through the appreciation of the amalgam of complex networks, within which each process can influence others. Greatly benefiting this view has been the study of the epigenome, the set of DNA and histone protein modifications that regulate gene expression and the function of regulatory non-coding RNAs without altering the DNA sequence itself. Indeed, the availability of reference genome assemblies of many species has led to the development of methodologies such as ChIP-Seq and RNA-Seq that have allowed us to define with high resolution the genomic distribution of several epigenetic elements and to better comprehend how they are interconnected for the regulation of gene expression. In the last few years, the use of these methodologies in the cardiovascular field has contributed to our understanding of the importance of epigenetics in heart diseases, giving new input to this area of research. Here, we review recently acquired knowledge on the role of the epigenome in heart failure, and discuss the need of an epigenomics roadmap for cardiovascular disease.

Endometriosis and endometrial cancer: A propensity score-adjusted real-world data study.

Endometriosis is a benign condition characterized by the presence of ectopic endometrial tissue. Our study investigated the effect of endometriosis on the risk of endometrial cancer (EC) and the prognosis of endometriosis-associated EC. In our study, 197,196 patients with endometriosis and without a previous diagnosis of EC were compared with 6,455,556 females encountering health services for examinations, with body mass index (BMI) data, and without endometriosis or EC. A propensity score generated 197,141 matched pairs. In the endometriosis cohort, 875 cases of EC were seen, whereas 558 were in the control group: the hazard ratio (HR) was 1.56 (95% CI 1.40-1.73, p < 0.001). Women with endometriosis were more likely to develop invasive endometrioid (p = 0.005) and clear cell (p < 0.001) EC. There was no difference in overall survival between endometriosis-associated EC and EC without endometriosis. Our epidemiological findings were consistent with the evidence of an association between endometriosis and EC.

p90 ribosomal S6 kinase 3 contributes to cardiac insufficiency in α-tropomyosin Glu180Gly transgenic mice.

Myocardial interstitial fibrosis is an important contributor to the development of heart failure. Type 3 p90 ribosomal S6 kinase (RSK3) was recently shown to be required for concentric myocyte hypertrophy under in vivo pathological conditions. However, the role of RSK family members in myocardial fibrosis remains uninvestigated. Transgenic expression of α-tropomyosin containing a Glu180Gly mutation (TM180) in mice of a mixed C57BL/6:FVB/N background induces a cardiomyopathy characterized by a small left ventricle, interstitial fibrosis, and diminished systolic and diastolic function. Using this mouse model, we now show that RSK3 is required for the induction of interstitial fibrosis in vivo. TM180 transgenic mice were crossed to RSK3 constitutive knockout (RSK3(-/-)) mice. Although RSK3 knockout did not affect myocyte growth, the decreased cardiac function and mild pulmonary edema associated with the TM180 transgene were attenuated by RSK3 knockout. The improved cardiac function was consistent with reduced interstitial fibrosis in the TM180;RSK3(-/-) mice as shown by histology and gene expression analysis, including the decreased expression of collagens. The specific inhibition of RSK3 should be considered as a potential novel therapeutic strategy for improving cardiac function and the prevention of sudden cardiac death in diseases in which interstitial fibrosis contributes to the development of heart failure.

CIP4 is required for the hypertrophic growth of neonatal cardiac myocytes.

BACKGROUND: CIP4 is a scaffold protein that regulates membrane deformation and tubulation, organization of the actin cytoskeleton, endocytosis of growth factor receptors, and vesicle trafficking. Although expressed in the heart, CIP4 has not been studied with regards to its potential function in cardiac myocytes. RESULTS: We now show using RNA interference that CIP4 expression in neonatal rat ventricular myocytes is required for the induction of non-mitotic, hypertrophic growth by the α-adrenergic agonist phenylephrine, the IL-6 cytokine leukemia inhibitor factor, and fetal bovine serum, as assayed using morphometry, immunocytochemistry for the hypertrophic marker atrial natriuretic factor and [3H]leucine incorporation for de novo protein synthesis. This requirement was consistent with the induction of CIP4 expression by hypertrophic stimulation. The inhibition of myocyte hypertrophy by CIP4 small interfering oligonucleotides (siRNA) was rescued by expression of a recombinant CIP4 protein, but not by a mutant lacking the N-terminal FCH domain responsible for CIP4 intracellular localization. CONCLUSIONS: These results imply that CIP4 plays a significant role in the intracellular hypertrophic signal transduction network that controls the growth of cardiac myocytes in heart disease.

Clinical Performance Evaluation of a Hyaluronic Acid Dental Gel for the Treatment of Traumatic Ulcers in Patients with Fixed Orthodontic Appliances: A Randomized Controlled Trial.

BACKGROUND: A newly available gel containing hyaluronic acid (HA) and polyvinylpyrrolidone was tested for efficacy on traumatic oral ulcers (TOU) caused by fixed orthodontic appliances. METHODS: A double-blind RCT was conducted to test the new gel versus a placebo. According to the sample size calculation, a total of 60 patients were considered sufficient and randomly allocated to one of the two groups out of a pool of 100 total patients who initially agreed to participate in the study. A VAS scale test and lesion measurements at T0, T1, and T2 were performed on the patients. RESULTS: A total of 70 patients developed TOU, with 8 drop-outs; the intergroup comparison showed a statistically significant greater dimension of the lesion in the control group at T2 when compared to the test group. The pain experienced by the patients belonging to the test group was significantly lower than the pain in the patients in the control group Conclusions: Under the limitations of the study, the new formula might provide faster healing with less pain experienced by the patient when compared to a placebo.

A comparative MudPIT analysis identifies different expression profiles in heart compartments.

Cardiomyopathies indistinctly affect atrial and ventricular cardiac compartments with alterations of their mechanical and/or electrical activity. To understand the main mechanisms involved in these pathological alterations, a detailed knowledge of the physiology of the healthy heart is critical. In the present work, we utilize multidimensional protein identification technology to characterize the murine left ventricle (LV), right ventricle (RV), and atria (A) proteomes, identifying thousands of distinct proteins. Moreover, using multidimensional algorithm protein map tool, relative abundances of proteins among the heart chambers were investigated. In sum, we found 16 and 55 proteins were more abundant in LV compared to RV and A, respectively; 47 and 60 proteins were more abundant in RV than LV and A, respectively; and, 81 and 74 proteins were more abundant in A than LV and RV, respectively. This detailed characterization of myocardial compartment proteome represents an important advancement in the knowledge of heart physiology, and may contribute to the identification of key features underlying the onset of cardiomyopathy.

Human cord blood CD34+ progenitor cells acquire functional cardiac properties through a cell fusion process.

The efficacy of cardiac repair by stem cell administration relies on a successful functional integration of injected cells into the host myocardium. Safety concerns have been raised about the possibility that stem cells may induce foci of arrhythmia in the ischemic myocardium. In a previous work (36), we showed that human cord blood CD34(+) cells, when cocultured on neonatal mouse cardiomyocytes, exhibit excitation-contraction coupling features similar to those of cardiomyocytes, even though no human genes were upregulated. The aims of the present work are to investigate whether human CD34(+) cells, isolated after 1 wk of coculture with neonatal ventricular myocytes, possess molecular and functional properties of cardiomyocytes and to discriminate, using a reporter gene system, whether cardiac differentiation derives from a (trans)differentiation or a cell fusion process. Umbilical cord blood CD34(+) cells were isolated by a magnetic cell sorting method, transduced with a lentiviral vector carrying the enhanced green fluorescent protein (EGFP) gene, and seeded onto primary cultures of spontaneously beating rat neonatal cardiomyocytes. Cocultured EGFP(+)/CD34(+)-derived cells were analyzed for their electrophysiological features at different time points. After 1 wk in coculture, EGFP(+) cells, in contact with cardiomyocytes, were spontaneously contracting and had a maximum diastolic potential (MDP) of -53.1 mV, while those that remained isolated from the surrounding myocytes did not contract and had a depolarized resting potential of -11.4 mV. Cells were then resuspended and cultured at low density to identify EGFP(+) progenitor cell derivatives. Under these conditions, we observed single EGFP(+) beating cells that had acquired an hyperpolarization-activated current typical of neonatal cardiomyocytes (EGFP(+) cells, -2.24 ± 0.89 pA/pF; myocytes, -1.99 ± 0.63 pA/pF, at -125 mV). To discriminate between cell autonomous differentiation and fusion, EGFP(+)/CD34(+) cells were cocultured with cardiac myocytes infected with a red fluorescence protein-lentiviral vector; under these conditions we found that 100% of EGFP(+) cells were also red fluorescent protein positive, suggesting cell fusion as the mechanism by which cardiac functional features are acquired.

Safety and Effects of the Rapid Maxillary Expander on Temporomandibular Joint in Subjects Affected by Juvenile Idiopathic Arthritis: A Retrospective Study.

BACKGROUND: In Juvenile Idiopathic Arthritis (JIA) temporo-mandibular joints are often affected causing skeletal and dental malocclusions. The most frequent condition is mandibular hypoplasia, that may be associated with maxillary hypoplasia. The aim of this retrospective case control study is to investigate the effects and the safety of rapid maxillary expansion (RME) in growing patients affected by JIA. It was evaluated whether RME could be performed without complications on TMJs of JIA patients using DC/TMD protocol, and naso-maxillary transversal parameters were compared with the ones obtained on healthy patients. METHODS: Twenty-five patients affected by JIA that ceased to manifest TMJ (Temporo-Mandibular Joint) symptoms in the previous year were treated with RME to solve the maxillary transverse hypoplasia. Postero-anterior cephalometric tracings were collected before and after treatment; linear measurements were obtained to study maxillary and nasal cavity modifications. Data were compared to those of a similar group of twenty-five healthy patients. Paired t-test and Independent t-test were used to evaluate changes before and after treatment in each group and to perform a comparison between the groups. RESULTS: All patients demonstrated a statistically significant increase in nasal cavity width, maxillary width and upper and lower intermolar width. No patients presented a worsening of their TMJs condition. Intragroup comparisons revealed significant changes of cephalometric measurements, but no difference was found when comparing JIA and healthy patients. CONCLUSIONS: Growing patients with JIA that ceased to show signs of active TMJ involvement for at least one year could be safely treated with RME, expecting similar benefits to those of healthy patients. Dentists and rheumatologists should be informed of safety and potential benefits of palatal expansion in JIA patients in order to improve the outcome of orthodontic treatment and reduce the indication for more invasive procedures (i.e., Surgical Assisted Rapid Maxillary Expansion).

Akt increases sarcoplasmic reticulum Ca2+ cycling by direct phosphorylation of phospholamban at Thr17.

Cardiomyocytes adapt to physical stress by increasing their size while maintaining cell function. The serine/threonine kinase Akt plays a critical role in this process of adaptation. We previously reported that transgenic overexpression of an active form of Akt (Akt-E40K) in mice results in increased cardiac contractility and cell size, as well as improved sarcoplasmic reticulum (SR) Ca(2+) handling. Because it is not fully elucidated, we decided to study the molecular mechanism by which Akt-E40K overexpression improves SR Ca(2+) handling. To this end, SR Ca(2+) uptake and the phosphorylation status of phospholamban (PLN) were evaluated in heart extracts from wild-type and Akt-E40K mice and mice harboring inducible and cardiac specific knock-out of phosphatidylinositol-dependent kinase-1, the upstream activator of Akt. Moreover, the effect of Akt was assessed in vitro by overexpressing a mutant Akt targeted preferentially to the SR, and by biochemical assays to evaluate potential interaction with PLN. We found that when activated, Akt interacts with and phosphorylates PLN at Thr(17), the Ca(2+)-calmodulin-dependent kinase IIdelta site, whereas silencing Akt signaling, through the knock-out of phosphatidylinositol-dependent kinase-1, resulted in reduced phosphorylation of PLN at Thr(17). Furthermore, overexpression of SR-targeted Akt in cardiomyocytes improved Ca(2+) handling without affecting cell size. Thus, we describe here a new mechanism whereby the preferential translocation of Akt to the SR is responsible for enhancement of contractility without stimulation of hypertrophy.

Assessment of the Stability of the Palatal Rugae in a 3D-3D Superimposition Technique Following Slow Maxillary Expansion (SME).

The Palatal Rugae are considered a useful human identification marker for both orthodontists and forensic personnel. The principal aim of the present study was to evaluate the stability of palatal rugae with a 3D-3D superimposition procedure following Slow Maxillary Expansion (SME), in order to assess whether they kept their uniqueness and validity for human identification, even after a specific dental treatment. For this purpose, a sample of 27 digital dental models - belonging to growing patients (13 males and 14 females), aged between 8.5 and 15 years, who underwent SME therapy - was retrospectively studied and compared with a control group of 27 untreated subjects - (13 males and 14 females). Digital dental models were obtained pre-treatment and at device removal; both were processed by means of an intraoral scanner. A superimposition procedure was thus performed to reach the minimum point-to-point distance between two models of palatal rugae. Intra- and inter-observer differences were statistically analyzed by paired Wilcoxon test and Intra-class Correlation coefficient (ICC), showing values larger than 0.93. There was no difference in Root-Mean-Square (RMS) values between untreated control subjects and subjects treated with Leaf Expander (p = 0.062). A RMS value of 0.43 was the threshold to distinguish the pooled group ("Untreated" and "Leaf") from any mismatch. According to the obtained results, this study failed to reject the null hypothesis and presented no differences between the RMS values of the Test group and the RMS values of the untreated control group. This work highlighted the usefulness of 3D superimposition procedure for purposes of human identification, in subjects undergoing dental treatment. However, keeping in sight the forensic use of this technique as a helpful probation element in court, further studies should be performed to confirm these findings.

Pooled Analysis of Clinical Outcome of Patients with Chemorefractory Metastatic Colorectal Cancer Treated within Phase I/II Clinical Studies Based on Individual Biomarkers of Susceptibility: A Single-Institution Experience.

BACKGROUND: Patients with metastatic colorectal cancer (mCRC) refractory to standard therapies have a poor prognosis. In this setting, recruitment into clinical trials is warranted, and studies driven by selection according to individual tumor molecular characteristics are expected to provide added value. OBJECTIVE: We retrospectively analyzed data from patients with mCRC refractory to or following failure of standard therapies who were enrolled into phase I/II clinical studies at the Niguarda Cancer Center based on the presence of a specific molecular profile expected to represent the target of susceptibility to the experimental drug(s). PATIENTS AND METHODS: From June 2011 to May 2016, 2044 patients with mCRC underwent molecular screening. Eighty patients (3.9%) were enrolled in ad hoc studies; the median age was 60 years (range 36-86) and the median number of previous treatment lines was five (range 2-8). Molecular characteristics exploited within these studies were MGMT promoter hypermethylation (48.7%), HER2 amplification (28.8%), BRAF V600E mutation (20%), and novel gene fusions involving ALK or NTRK (2.5%). RESULTS: One patient (1%) had RECIST (Response Evaluation Criteria In Solid Tumors) complete response (CR), 13 patients (16.5%) experienced a partial response (PR), and 28 (35%) stable disease (SD). Median progression-free survival (PFS) was 2.8 months (range 2.63-3.83), with 24% of patients displaying PFS >5 months. Median growth modulation index (GMI) was 0.85 (range 0-15.61) and 32.5% of patients had GMI >1.33. KRAS exon 2 mutations were found in 38.5% of patients, and among the 78 patients with known KRAS status, those with wild-type tumors had longer PFS than those with mutated tumors (3.80 [95% CI 2.80-5.03] vs. 2.13 months [95% CI 1.77-2.87], respectively, p = 0.001). Median overall survival (OS) was 7.83 months (range 7.17-9.33) for all patients, and patients with KRAS wild-type tumors had longer OS than those with mutated tumors (7.83 [95% CI 7.33-10.80] vs. 7.18 months [95% CI 5.63-9.33], respectively, p = 0.06). CONCLUSIONS: This single-institution retrospective study indicates that in a heavily pretreated population approximately 4% of mCRC tumors display a potential actionable molecular context suitable for therapeutic intervention. Application of molecular selection is challenging but improves clinical outcome even in later lines of treatment.

Bosutinib : a review of preclinical and clinical studies in chronic myelogenous leukemia.

INTRODUCTION: Chronic myeloid leukemia (CML) is a hematopoietic stem cell disease. It is characterized by a Bcr-Abl (breakpoint cluster region-Abelson leukemia virus) tyrosine kinase fusion protein produced from the Philadelphia (Ph) chromosome. The tyrosine kinase inhibitor (TKI) imatinib was the first targeted therapy licensed for patients with chronic-phase CML. In recent years, many other TKIs have been approved for the treatment of patients with CML. For this reason, the choice of the best strategy treatment has become increasingly complex. AREAS COVERED: Bosutinib , a dual Src/Abl kinase inhibitor, has shown potent activity against CML and it has been approved by the US FDA for the treatment of chronic, accelerated or blast-phase Ph+ CML. This review was conducted to describe the preclinical and clinical activity of bosutinib and the safety and tolerability of the drug in the treatment of CML. Included studies were identified through a search of electronic databases in July 2013 and relevant conference proceedings. EXPERT OPINION: Imatinib continues to represent the treatment of choice for CML. However, some patients develop resistance or intolerance to imatinib or to other second-generation TKIs. Bosutinib shows a good therapeutic activity with a benign safety profile, no cardiovascular toxicity, and offers an important therapeutic addition to the armamentarium that physicians can use against resistant CML.

Promoter CpG island hypermethylation of the DNA repair enzyme MGMT predicts clinical response to dacarbazine in a phase II study for metastatic colorectal cancer.

PURPOSE: O(6)-methylguanine-DNA-methyltransferase (MGMT) is a DNA repair protein removing mutagenic and cytotoxic adducts from O(6)-guanine in DNA. Approximately 40% of colorectal cancers (CRC) display MGMT deficiency due to the promoter hypermethylation leading to silencing of the gene. Alkylating agents, such as dacarbazine, exert their antitumor activity by DNA methylation at the O(6)-guanine site, inducing base pair mismatch; therefore, activity of dacarbazine could be enhanced in CRCs lacking MGMT. We conducted a phase II study with dacarbazine in CRCs who had failed standard therapies (oxaliplatin, irinotecan, fluoropyrimidines, and cetuximab or panitumumab if KRAS wild-type). EXPERIMENTAL DESIGN: All patients had tumor tissue assessed for MGMT as promoter hypermethylation in double-blind for treatment outcome. Patients received dacarbazine 250 mg/m(2) intravenously every day for four consecutive days, every 21 days, until progressive disease or intolerable toxicity. We used a Simon two-stage design to determine whether the overall response rate would be 10% or more. Secondary endpoints included association of response, progression-free survival, and disease control rate with MGMT status. RESULTS: Sixty-eight patients were enrolled from May 2011 to March 2012. Patients received a median of three cycles of dacarbazine (range 1-12). Grades 3 and 4 toxicities included: fatigue (41%), nausea/vomiting (29%), constipation (25%), platelet count decrease (19%), and anemia (18%). Overall, two patients (3%) achieved partial response and eight patients (12%) had stable disease. Disease control rate (partial response + stable disease) was significantly associated with MGMT promoter hypermethylation in the corresponding tumors. CONCLUSION: Objective clinical responses to dacarbazine in patients with metastatic CRC are confined to those tumors harboring epigenetic inactivation of the DNA repair enzyme MGMT.

Akt regulates L-type Ca2+ channel activity by modulating Cavalpha1 protein stability.

The insulin IGF-1-PI3K-Akt signaling pathway has been suggested to improve cardiac inotropism and increase Ca(2+) handling through the effects of the protein kinase Akt. However, the underlying molecular mechanisms remain largely unknown. In this study, we provide evidence for an unanticipated regulatory function of Akt controlling L-type Ca(2+) channel (LTCC) protein density. The pore-forming channel subunit Ca(v)alpha1 contains highly conserved PEST sequences (signals for rapid protein degradation), and in-frame deletion of these PEST sequences results in increased Ca(v)alpha1 protein levels. Our findings show that Akt-dependent phosphorylation of Ca(v)beta2, the LTCC chaperone for Ca(v)alpha1, antagonizes Ca(v)alpha1 protein degradation by preventing Ca(v)alpha1 PEST sequence recognition, leading to increased LTCC density and the consequent modulation of Ca(2+) channel function. This novel mechanism by which Akt modulates LTCC stability could profoundly influence cardiac myocyte Ca(2+) entry, Ca(2+) handling, and contractility.