Crosstalk between Chemokine Receptor CXCR4 and Cannabinoid Receptor CB2 in Modulating Breast Cancer Growth and Invasion

Cannabinoids bind to cannabinoid receptors CB(1) and CB(2) and have been reported to possess anti-tumorigenic activity in various cancers. However, the mechanisms through which cannabinoids modulate tumor growth are not well known. In this study, we report that a synthetic non-psychoactive cannabinoid that specifically binds to cannabinoid receptor CB(2) may modulate breast tumor growth and metastasis by inhibiting signaling of the chemokine receptor CXCR4 and its ligand CXCL12. This signaling pathway has been shown to play an important role in regulating breast cancer progression and metastasis.We observed high expression of both CB(2) and CXCR4 receptors in breast cancer patient tissues by immunohistochemical analysis. We further found that CB(2)-specific agonist JWH-015 inhibits the CXCL12-induced chemotaxis and wound healing of MCF7 overexpressing CXCR4 (MCF7/CXCR4), highly metastatic clone of MDA-MB-231 (SCP2) and NT 2.5 cells (derived from MMTV-neu) by using chemotactic and wound healing assays. Elucidation of the molecular mechanisms using various biochemical techniques and confocal microscopy revealed that JWH-015 treatment inhibited CXCL12-induced P44/P42 ERK activation, cytoskeletal focal adhesion and stress fiber formation, which play a critical role in breast cancer invasion and metastasis. In addition, we have shown that JWH-015 significantly inhibits orthotopic tumor growth in syngenic mice in vivo using NT 2.5 cells. Furthermore, our studies have revealed that JWH-015 significantly inhibits phosphorylation of CXCR4 and its downstream signaling in vivo in orthotopic and spontaneous breast cancer MMTV-PyMT mouse model systems.This study provides novel insights into the crosstalk between CB(2) and CXCR4/CXCL12-signaling pathways in the modulation of breast tumor growth and metastasis. Furthermore, these studies indicate that CB(2) receptors could be used for developing innovative therapeutic strategies against breast cancer

Standardisation framework for the Maudsley staging method for treatment resistance in depression

Background: Treatment-resistant depression (TRD) is a serious and relatively common clinical condition. Lack of consensus on defining and staging TRD remains one of the main barriers to understanding TRD and approaches to intervention. The Maudsley Staging Method (MSM) is the first multidimensional model developed to define and stage treatment-resistance in “unipolar depression”. The model is being used increasingly in treatment and epidemiological studies of TRD and has the potential to support consensus. Yet, standardised methods for rating the MSM have not been described adequately. The aim of this report is to present standardised approaches for rating or completing the MSM. Method: Based on the initial development of the MSM and a narrative review of the literature, the developers of the MSM provide explicit guidance on how the three dimensions of the MSM–treatment failure, severity of depressive episode and duration of depressive episode– may be rated. Result: The core dimension of the MSM, treatment failure, may be assessed using the Maudsley Treatment Inventory (MTI), a new method developed for the purposes of completing the MSM. The MTI consists of a relatively comprehensive list of medications with options for rating doses and provisions treatment for multiple episodes. The second dimension, severity of symptoms, may be assessed using simple instruments such as the Clinical Global Impression, the Psychiatric Status Rating or checklist from a standard diagnostic checklist. The standardisation also provides a simple rating scale for scoring the third dimension, duration of depressive episode. Conclusion: The approaches provided should have clinical and research utility in staging TRD. However, in proposing this model, we are fully cognisant that until the pathophysiology of depression is better understood, staging methods can only be tentative approximations. Future developments should attempt to incorporate other biological/ pathophysiological dimensions for staging

Targeted Curing of All Lysogenic Bacteriophage from Streptococcus pyogenes Using a Novel Counter-selection Technique

We thank the members of the Laboratory of Microbial Pathogenesis and Immunology, especially Annette Nelkenbaum and Ben Winer for their technical assistance. We also thank Estee Colleen Cervantes and Sutapa Banerjee from Hunter College for their technical contribution to this project. We are grateful to Joseph Ferretti for S. pyogenes strain SF370.Streptococcus pyogenes is a human commensal and a bacterial pathogen responsible for a wide variety of human diseases differing in symptoms, severity, and tissue tropism. The completed genome sequences of >37 strains of S. pyogenes, representing diverse disease-causing serotypes, have been published. The greatest genetic variation among these strains is attributed to numerous integrated prophage and prophage-like elements, encoding several virulence factors. A comparison of isogenic strains, differing in prophage content, would reveal the effects of these elements on streptococcal pathogenesis. However, curing strains of prophage is often difficult and sometimes unattainable. We have applied a novel counter-selection approach to identify rare S. pyogenes mutants spontaneously cured of select prophage. To accomplish this, we first inserted a two-gene cassette containing a gene for kanamycin resistance (KanR) and the rpsL wild-type gene, responsible for dominant streptomycin sensitivity (SmS), into a targeted prophage on the chromosome of a streptomycin resistant (SmR) mutant of S. pyogenes strain SF370. We then applied antibiotic counter-selection for the re-establishment of the KanS/SmR phenotype to select for isolates cured of targeted prophage. This methodology allowed for the precise selection of spontaneous phage loss and restoration of the natural phage attB attachment sites for all four prophage-like elements in this S. pyogenes chromosome. Overall, 15 mutants were constructed that encompassed every permutation of phage knockout as well as a mutant strain, named CEM1ΔΦ, completely cured of all bacteriophage elements (a ~10% loss of the genome); the only reported S. pyogenes strain free of prophage-like elements. We compared CEM1ΔΦ to the WT strain by analyzing differences in secreted DNase activity, as well as lytic and lysogenic potential. These mutant strains should allow for the direct examination of bacteriophage relationships within S. pyogenes and further elucidate how the presence of prophage may affect overall streptococcal survival, pathogenicity, and evolution.Yeshttp://www.plosone.org/static/editorial#pee