Selective glucocorticoid receptor properties of GSK866 analogs with cysteine reactive warheads

Synthetic glucocorticoids (GC) are the mainstay therapy for treatment of acute and chronic inflammatory disorders. Due to the high adverse effects associated with long-term use, GC pharmacology has focused since the nineties on more selective GC ligand-binding strategies, classified as selective glucocorticoid receptor (GR) agonists (SEGRAs) or selective glucocorticoid receptor modulators (SEGRMs). In the current study, GSK866 analogs with electrophilic covalent-binding warheads were developed with potential SEGRA properties to improve their clinical safety profile for long-lasting topical skin disease applications. Since the off-rate of a covalently binding drug is negligible compared to that of a non-covalent drug, its therapeutic effects can be prolonged and typically, smaller doses of the drug are necessary to reach the same level of therapeutic efficacy, thereby potentially reducing systemic side effects. Different analogs of SEGRA GSK866 coupled to cysteine reactive warheads were characterized for GR potency and selectivity in various biochemical and cellular assays. GR-and NF kappa B dependent reporter gene studies show favorable anti-inflammatory properties with reduced GR transactivation of two non-steroidal GSK866 analogs UAMC-1217 and UAMC-1218, whereas UAMC-1158 and UAMC-1159 compounds failed to modulate cellular GR activity. These results were further supported by GR immuno-localization and S211 phospho-GR western analysis, illustrating significant GR phosphoactivation and nuclear translocation upon treatment of GSK866, UAMC-1217, or UAMC-1218, but not in case of UAMC-1158 or UAMC-1159. Furthermore, mass spectrometry analysis of tryptic peptides of recombinant GR ligand-binding domain (LBD) bound to UAMC-1217 or UAMC-1218 confirmed covalent cysteine-dependent GR binding. Finally, molecular dynamics simulations, as well as glucocorticoid receptor ligand-binding domain (GR-LBD) coregulator interaction profiling of the GR-LBD bound to GSK866 or its covalently binding analogs UAMC-1217 or UAMC-1218 revealed subtle conformational differences that might underlie their SEGRA properties. Altogether, GSK866 analogs UAMC-1217 and UAMC-1218 hold promise as a novel class of covalent-binding SEGRA ligands for the treatment of topical inflammatory skin disorders

How does conformational flexibility influence key structural features involved in activation of anaplastic lymphoma kinase?

Anaplastic Lymphoma Kinase (ALK) plays a major role in developing tumor processes and therefore has emerged as a validated therapeutic target. Applying atomistic molecular dynamics simulations on the wild type enzyme and the nine most frequently occurring and clinically important activation mutants we revealed important conformational effects on key interactions responsible for the activation of the enzyme

A Case-By-Case Approach to Pleading Scienter Under the Private Securities Litigation Reform Act of 1995

Securities fraud litigation under Rule lOb-5 threatens all publicly traded companies: according to the Stanford Securities Class Action Clearinghouse, in 1998 a securities fraud lawsuit was filed for nearly every day that the stock markets were open. Some of these lawsuits appear to be frivolous, triggered by inevitable fluctuations in stock prices (so-called fraud by hindsight complaints), while others represent legitimate efforts at private enforcement of the securities laws. Disposition on the pleadings is a critical defense strategy for all securities lawsuits. Securities fraud lawsuits that withstand a 12(b)(6) motion almost always settle, regardless of the actual merits of the case or the probability of success at trial, because of the massive discovery and defense costs associated with such suits. Because Rule lOb-5 requires a showing of scienter, a mental state embracing intent to deceive, manipulate, or defraud, defendants can often successfully dispose of a securities fraud case before being forced to settle by challenging the plaintiff\u27s scienter pleading. For these reasons, the standard for pleading scienter is an appropriate context in which to balance the competing interests of eliminating abusive claims and permitting meritorious ones

Inhibiting ERK Activation with CI-1040 Leads to Compensatory Upregulation of Alternate MAPKs and Plasminogen Activator Inhibitor-1 following Subtotal Nephrectomy with No Impact on Kidney Fibrosis

Extracellular-signal regulated kinase (ERK) activation by MEK plays a key role in many of the cellular processes that underlie progressive kidney fibrosis including cell proliferation, apoptosis and transforming growth factor β1-mediated epithelial to mesenchymal transition. We therefore assessed the therapeutic impact of ERK1/2 inhibition using a MEK inhibitor in the rat 5/6 subtotal nephrectomy (SNx) model of kidney fibrosis. There was a twentyfold upregulation in phospho-ERK1/2 expression in the kidney after SNx in Male Wistar rats. Rats undergoing SNx became hypertensive, proteinuric and developed progressive kidney failure with reduced creatinine clearance. Treatment with the MEK inhibitor, CI-1040 abolished phospho- ERK1/2 expression in kidney tissue and prevented phospho-ERK1/2 expression in peripheral lymphocytes during the entire course of therapy. CI-1040 had no impact on creatinine clearance, proteinuria, glomerular and tubular fibrosis, and α-smooth muscle actin expression. However, inhibition of ERK1/2 activation led to significant compensatory upregulation of the MAP kinases, p38 and JNK in kidney tissue. CI-1040 also increased the expression of plasminogen activator inhibitor-1 (PAI-1), a key inhibitor of plasmin-dependent matrix metalloproteinases. Thus inhibition of ERK1/2 activation has no therapeutic effect on kidney fibrosis in SNx possibly due to increased compensatory activation of the p38 and JNK signalling pathways with subsequent upregulation of PAI-1

The effects of subliminal weapons effect on aggressive behavior in college level students

This study evaluated the difference in displayed aggression in subjects randomly assigned to either a subliminal weapons effect condition or to a controlled condition. 30 students from a small public University in New Jersey signed up for the study to receive credit for their introductory psychology course. The subjects were then randomly assigned to one of the two conditions; the subliminal weapons effect conditioned consisted of a poster of Scarface holding a gun pointed toward the subject on the blackboard (the poster had a sign asking that it not be removed by a professor). The controlled condition had no stimuli on the blackboard. The subjects then completed a survey dealing with their attitude of the dining hall. The results showed that the subjects in the weapons effect condition showed a significantly higher level of aggression than the control group did

Zen and the Art of Jursiprudence

Lawyer bashing is by no means a remarkable phenomenon. It was not remarkable when Shakespeare wrote, [t]he first thing we do, let\u27s kill all the lawyers, and it\u27s not remarkable today. Paul Campos, however, has written a particularly readable example, blending venerable Western lawyer-bashing and pop psychology with unsystematic invocations of Eastern religion. Jurismania is named after Campos\u27s theory that the American legal system has a lot in common with a person suffering from an obsessive-compulsive disorder, an addiction to law that does neither the patient nor those around him much good. In Jurismania, Campos criticizes our insistence on regulating and legalizing every aspect of our lives, and our insistence on exclusive rationality. Campos argues, with regular Taoist allusions, that rationality is not and cannot be the exclusive solution to the questions law raises, and that irrational methods are and should be employed. Campos\u27s intended audience is the general reader whose experience of American law has made him or her wonder if there might not be something wrong with it (pp. vii-viii). Should that audience take Campos\u27s critique seriously, it will strike close to the heart of law and the legal profession. Thus, although they are not the target audience, lawyers ought to think about Jurismania because it reflects and amplifies a perspective that may be common to many nonlawyers who encounter the legal system

Periprostatic Fat Adipokines Expression Correlated with Prostate Cancer Aggressiveness in Men Undergoing Radical Prostatectomy for Clinically Localised Disease

Objectives: To investigate relationship between periprostatic adipose tissue (PPAT) adipokines expression and PCa aggressiveness using both pathological features of radical prostatectomy (RP) and multiparametric MRI parameters.Patients and Methods: Sixty-nine men were recruited to assess immunohistochemical expression of TNFα- and VEGF of periprostatic fat of radical prostate specimens. Percent immunopositivity was quantified on scanned slides using Aperio Positive Pixel Count algorithm for PPAT TNFα, VEGF and androgen receptors. Periprostatic fat volume (PFV) was segmented on contiguous T 1 -weighted axial MRI slices from the level of the prostate base to apex. PFV was normalised to prostate volume (PV) to account for variations in PV (NPFV=PFV/PV). MRI quantitative values (K ep , K trans, and ADC) were measured from PCa primary lesion using OleaSphere software. Patients were stratified into three groups according to RP GS: ≤6, 7(3+4) and 7(4+3) or more. Results: The mean rank of VEGF and TNFα were significantly different between the groups [H(2)= 11.038, p=0.004] and [H(2)=13.086, p=0.001], respectively. Patients with stage pT 3 had higher TNFα (18.2±8.95) positivity than patients with stage pT 2 (13.27±10.66), t (67) =-2.03, p=0.047. TNFα expression significantly correlated with K trans (ρ=0.327, p=0.023). TNFα (p=0.043) and VEGF (p= 0.02) correlates with high-grade PCa (GS≥7) in radical prostatectomy specimens and correlated significantly with upgradation of Gleason score from biopsy to radical prostatectomy histology. Conclusions: Expression level of TNFα and VEGF on immunostaining significantly correlated with aggressivity of PCa. As biomarkers, these suggest the risk of having high-grade PCa in men undergoing RP. This article is protected by copyright. All rights reserved.</p

Impact of the putative cancer stem cell markers and growth factor receptor expression on the sensitivity of ovarian cancer cells to treatment with various forms of the HER inhibitors and cytotoxic drugs

Increased expression and activation of human epidermal growth factor receptor (EGFR) and HER-2 have been reported in numerous cancers. The aim of this study was to determine the sensitivity of a large panel of human ovarian cancer cell lines (OCCLs) to treatment with various forms of small molecule tyrosine kinase inhibitors (TKIs) and cytotoxic drugs. The aim was to see if there was any association between the protein expression of various biomarkers including three putative ovarian cancer stem cell (CSC) markers (CD24, CD44, CD117/c-Kit), P-glycoprotein (P-gp), and HER family members and response to treatment with these agents. The sensitivity of 10 ovarian tumour cell lines to the treatment with various forms of HER TKIs (gefitinib, erlotinib, lapatinib, sapitinib, afatinib, canertinib, neratinib), as well as other TKIs (dasatinib, imatinib, NVP-AEW541, crizotinib) and cytotoxic agents (paclitaxel, cisplatin and doxorubicin), as single agents or in combination, was determined by SRB assay. The effect on these agents on the cell cycle distribution, and downstream signaling molecules and tumour migration were determined using flow cytometry, western blotting, and the IncuCyte Clear View cell migration assay respectively. Of the HER inhibitors, the irreversible pan-TKIs (canertinib, neratinib and afatinib) were the most effective TKIs for inhibiting the growth of all ovarian cancer cells, and for blocking the phosphorylation of EGFR, HER-2, AKT and MAPK in SKOV3 cells. Interestingly, while the majority of cancer cells were highly sensitive to treatment with dasatinib, they were relatively resistant to treatment with imatinib (i.e., IC50 >10 µM). Of the cytotoxic agents, paclitaxel was the most effective for inhibiting the growth of OCCLs, and of various combinations of these drugs, only treatment with a combination of NVP-AEW541 and paclitaxel produced a synergistic or additive anti-proliferative effect in all three cell lines examined (i.e., SKOV3, Caov3, ES2). Finally, of the TKIs, only treatment with afatinib, neratinib and dasatinib were able to reduce the migration of HER-2 overexpressing SKOV3 cells. We did not find any significant association between the expression of putative ovarian CSC marker, HER family members, c-MET, ALK, and IGF-IR and the response to the irreversible HER TKIs. Our results support the need for further investigations of the therapeutic potential of these irreversible HER family blockers in ovarian cancer, and the therapeutic potential of dasatinib when used in combination with the inhibitors of the HER family members in ovarian cancer