Background-Therapy for chronic systolic heart failure (sHF) has improved over the past 2 decades, but the armamentarium of drugs is limited and consequently sHF remains a leading cause of death and disability. In this investigation, we examined the effects of a novel cardiac myosin activator, omecamtiv mecarbil (formerly CK-1827452) in 2 different models of heart failure. Methods and Results-Two different models of sHF were used: (1) pacing-induced sHF after myocardial infarction (MI-sHF) and (2) pacing-induced sHF after 1 year of chronic pressure overload left ventricular hypertrophy (LVH-sHF).Omecamtiv mecarbil increased systolic function in sHF dogs, chronically instrumented to measure LV pressure, wall thickness, and cardiac output. Omecamtiv mecarbil, infused for 24 hours, induced a sustained increase without desensitization (PϽ0.05) in wall thickening (25Ϯ6.2%), stroke volume (44Ϯ6.5%) and cardiac output (22Ϯ2.8%), and decreased heart rate (15Ϯ3.0%). The major differences between the effect of omecamtiv mecarbil on cardiac function and the effect induced by a catecholamine, for example, dobutamine, is that omecamtiv mecarbil did not increase LV dP/dt but rather increased LV systolic ejection time by 26Ϯ2.9% in sHF. Another key difference is that myocardial O 2 consumption (MVO 2 ), which increases with catecholamines, was not significantly affected by omecamtiv mecarbil. Conclusions-These results demonstrate that chronic infusion of the cardiac myosin activator, omecamtiv mecarbil, improves LV function in sHF without the limitations of progressive desensitization and increased MVO 2. This unique profile may provide a new therapeutic approach for patients with sHF. (Circ Heart Fail. 2010;3:522-527.)

A p53-derived C-terminal peptide induced rapid apoptosis in breast cancer cell lines carrying endogenous p53 mutations or overexpressed wild-type (wt) p53 but was not toxic to nonmalignant human cell lines containing wt p53. Apoptosis occurred through a Fas/APO-1 signaling pathway involving increased extracellular levels of Fas/FasL in the absence of protein synthesis, as well as activation of a Fas/APO-1-specific protease, FLICE. The peptide activity was p53-dependent, and it had no effect in three tumor cell lines with null p53. Furthermore, the C-terminal peptide bound to p53 protein in cell extracts. Thus, p53-dependent, Fas/APO-1 mediated apoptosis can be induced in breast cancer cells with mutant p53 similar to the recently described Fas/APO-1 induced apoptosis by wt p53. However, mutant p53 without p53 peptide does not induce a Fas/ APO-1 activation or apoptosis. Docking of the computed low energy conformations for the C-terminal peptide with those for a recently defined proline-rich regulatory region from the N-terminal domain of p53 suggests a unique low energy complex between the two peptide domains. The selective and rapid induction of apoptosis in cancer cells carrying p53 abnormalities may lead to a novel therapeutic modality.

We investigated the effects of caloric restriction (CR) on growth of tumors and metastases in the 4T1 mammary tumor model and found that CR, compared with normal diet, reduced the growth of mammary tumors and metastases and the total number of metastases that originated both spontaneously from the primary tumor and also experimentally from i.v. injection of the tumor cells. CR also decreased proliferation and angiogenesis and increased apoptosis in tumors. CR reduced levels of insulin, leptin, insulin-like growth factor 1, insulin-like growth factor binding protein 3 and increased adiponectin in tumors. We also demonstrated that tumors from CR mice possessed lower levels of transforming growth factor-β, lower intratumor deposition of collagen IV and reduced invasiveness due to a decrease in tumor secretion of active matrix metalloproteinase 9. Our results suggest that CR-induced metabolic and signaling changes affect the stroma and the tumor cells resulting in a microenvironment that prevents proliferation of breast tumors and their metastases.

Retinoids exert their physiological action by interacting with two families of nuclear receptors, the retinoic acid receptors (RARs) and the retinoid X receptors (RXRs), which regulate gene expression by forming transcriptionally active heterodimeric RAR/RXR or homodimeric RXR/RXR complexes on DNA. Retinoid receptor activity resides in several regions, including the DNA and ligand binding domains, a dimerization interface, and both a ligand-independent (AF-1) and a ligand-dependent (AF-2) transactivation function. While 9-cis retinoic acid (RA) alone is the cognate ligand for the RXRs, both 9-cis RA and all-trans RA (t-RA) compete for binding with high affinity to the RARs. This latter observation suggested to us that the two isomers may interact with a common binding site. Here we report that RARa has two distinct but overlapping binding sites for 9-cis RA and t-RA. Truncation of a human RARa to 419 amino acids yields a receptor that binds both t-RA and 9-cis RA with high affinity, but truncation to amino acid 404 yields a mutant receptor that binds only t-RA with high affinity. Remarkably, this region also defines a C-terminal boundary for AF-2, as addition of amino acids 405 to 419 restores receptor-mediated gene activity to a truncated human RARRa lacking this region. It is interesting to speculate that binding of retinoid stereoisomers to unique sites within an RAR may function with AF-2 to cause differential activation of retinoid-responsive gene pathways.Retinoic acids (RAs) are derivatives of vitamin A (retinol) which affect a wide spectrum of biological activities, including cellular differentiation and vertebrate development (for a review, see reference 48). These compounds exert their biological action by interacting with two families of nuclear receptors, the RA receptors (RARs) (8,18,27,42) (10,34,53), and the peroxisome proliferator-activated receptor (24,26).On the basis of the amino acid homologies, six regions of functional identity have been described for the members of this superfamily of nuclear receptors (16,19). Regions A and B contain a ligand-independent transactivation function, AF-1, that may play a role in specifying activity on particular target genes (38,39 (3,23,32), while the RARs bind both 9-cis RA and all-trans-RA (t-RA) with high affinity (3). We have shown that 9-cis RA and t-RA compete with each other for binding to the RARs (4), indicating that they share a common binding site. Considering the differences in configuration between 9-cis RA and t-RA, we reasoned that there may be subtle differences in the RAR binding pocket to accommodate both of these ligands. Here we report that in fact, 9-cis RA and t-RA interact with distinct binding determinants on RARc. Using C-terminal truncation mutants of human RARo, we defined a 15-amino-acid region that is absolutely required for 9-cis RA binding but not for high-affinity t-RA binding. Furthermore, we show that these same amino acids are required for the ligand-inducible, receptor-mediated transactivation of 3RARE and identify the AF...

Background: Rolling circle amplification of ligated probes is a simple and sensitive means for genotyping directly from genomic DNA. SNPs and mutations are interrogated with open circle probes (OCP) that can be circularized by DNA ligase when the probe matches the genotype. An amplified detection signal is generated by exponential rolling circle amplification (ERCA) of the circularized probe. The low cost and scalability of ligation/ERCA genotyping makes it ideally suited for automated, high throughput methods.