New therapeutic approach to heart failure due to myocardial infarction based on targeting growth hormone-releasing hormone receptor

Background We previously showed that growth hormone-releasing hormone (GHRH) agonists are cardioprotective following myocardial infarction (MI). Here, our aim was to evaluate the in vitro and in vivo activities of highly potent new GHRH agonists, and elucidate their mechanisms of action in promoting cardiac repair. Methods and Results H9c2 cells were cultured in serum-free medium, mimicking nutritional deprivation. GHRH agonists decreased calcium influx and significantly improved cell survival. Rats with cardiac infarction were treated with GHRH agonists or placebo for four weeks. MI size was reduced by selected GHRH agonists (JI-38, MR-356, MR-409); this accompanied an increased number of cardiac c-kit+ cells, cellular mitotic divisions, and vascular density. One week post-MI, MR-409 significantly reduced plasma levels of IL-2, IL-6, IL-10 and TNF-? compared to placebo. Gene expression studies revealed favorable outcomes of MR-409 treatment partially result from inhibitory activity on pro-apoptotic molecules and pro-fibrotic systems, and by elevation of bone morphogenetic proteins. Conclusions Treatment with GHRH agonists appears to reduce the inflammatory responses post-MI and may consequently improve mechanisms of healing and cardiac remod eling by regulating pathways involved in fibrosis, apoptosis and cardiac repair. Patients with cardiac dysfunction could benefit from treatment with novel GHRH agonists

New Clathrin-Based Nanoplatforms for Magnetic Resonance Imaging

Background: Magnetic Resonance Imaging (MRI) has high spatial resolution, but low sensitivity for visualization of molecular targets in the central nervous system (CNS). Our goal was to develop a new MRI method with the potential for non-invasive molecular brain imaging. We herein introduce new bio-nanotechnology approaches for designing CNS contrast media based on the ubiquitous clathrin cell protein. Methodology/Principal Findings: The first approach utilizes three-legged clathrin triskelia modified to carry 81 gadolinium chelates. The second approach uses clathrin cages self-assembled from triskelia and designed to carry 432 gadolinium chelates. Clathrin triskelia and cages were characterized by size, structure, protein concentration, and chelate and gadolinium contents. Relaxivity was evaluated at 0.47 T. A series of studies were conducted to ascertain whether fluorescent-tagged clathrin nanoplatforms could cross the blood brain barriers (BBB) unaided following intranasal, intravenous, and intraperitoneal routes of administration. Clathrin nanoparticles can be constituted as triskelia (18.5 nm in size), and as cages assembled from them (55 nm). The mean chelate: clathrin heavy chain molar ratio was 27.0464.8: 1 fo

Preparation and Reactions of 2-tert-Butyl-1,1,3,3-Tetramethylguanidine: 2,2,6-Trimethylcyclohexen-1-yl Iodide

Derek H. R. Barton, Mi Chen, Joseph Cs. Jászberényi, and Dennis K. Taylo

Enzyme-based Intravascular Defense Against Organophosphorus Neurotoxins: Synergism of Dendritic-enzyme Complexes with 2-PAM and Atropine

Novel, enzyme-complexed, nano-delivery systems have been developed to antagonize the lethal effects of organophosphorus (OP) molecules such as diisopropylfluorophosphate and paraoxon. Polymeric nanocapsules can be used to deliver metabolizing enzymes to the circulation, often increasing the enzyme\u27s efficacy by extending their circulatory life and, in some cases, enhancing their specific activity. The bacterial enzymes organophosphorus hydrolase (OPH) and organophosphorus anhydrolase (OPAA) were encapsulated within a nanocapsule, polyoxazoline-based dendritic polymer carrier and employed in combination with the OP antagonists pralidoxime (2-PAM) and atropine. The effective doses for OPH and OPAA, respectively, were 500–550 and 1500–1650 units/kg mice; the size of the entire complex is approximately 200 nm in diameter. These studies compare the efficacy of the two enzymes as prophylactic systems encapsulated within the dendritic polymer. When used in combination with 2-PAM and atropine, the dendritic encapsuled OPAA provided a 25×LD50protection against DFP intoxication, while the similarly constructed OPH complex showed a more dramatic protection (780×LD50) against paraoxon intoxication in Balb/c mice. The studies demonstrate a synergistic enhancement of the antagonist, since the antidotal protection of 2-PAM+atropine against DFP and paraoxon is approximately 8 and 60×LD50, respectively

Article Navigation Comparing Therapeutic and Prophylactic Protection against the Lethal Effect of Paraoxon

Prophylactic and therapeutic efficacy against organophosphorus (OP) intoxication by pralidoxime (2-PAM) and atropine were studied and compared with sterically stabilized long-circulating liposomes encapsulating recombinant organophosphorus hydrolase (OPH), either alone or in various specific combinations, in paraoxon poisoning. Prophylactic and therapeutic properties of atropine and 2-PAM are diminished when they are used alone. However, their prophylactic effects are enhanced when they are used in combination. Present studies indicate that sterically stabilized liposomes (SL) encapsulating recombinant OPH (SL-OPH) alone can provide much better therapeutic and prophylactic protection than the classic 2-PAM + atropine combination. This protection was even more dramatic when SL-OPH was employed in combination with 2-PAM and/or atropine: the magnitude of prophylactic antidotal protection was an astounding 1022 LD50 [920 mg/kg (LD50 of paraoxon with antagonists)/ 0.95 mg/kg (LD50 of control paraoxon)], and the therapeutic antidotal protection was 156 LD50 [140 mg/kg (LD50 of paraoxon with antagonists)/0.9 mg/kg (LD50 of control paraoxon)]. The current study firmly establishes the value of using liposome encapsulating OPH

Ln[DO3A-N-α-(pyrenebutanamido)propionate] complexes: optimized relaxivity and NIR optical properties†

We have proposed recently that the DO3A-N-α-(amino)propionate chelator and its amide conjugates are leads to targeted, high relaxivity, safe contrast agents for magnetic resonance imaging. In this work we illustrate further the expeditious nature and robustness of the synthetic methodologies developed by preparing the DO3A-N-(α-pyrenebutanamido)propionate chelator. Its Gd3+ chelate retains the optimized water exchange, high stability and inertness of the parent complex. The pyrene moiety imparts concentration- dependent self-assembly properties and aggregation-sensitive fluorescence emission to the Gd3+ complex. The Gd3+ complex displays pyrene-centred fluorescence whilst the Yb3+ and Nd3+ complexes exhibit sensitized lanthanide-centred near-infrared luminescence. The aggregated form of the complex displays high relaxivity (32 mM−1 s−1, 20 MHz, 25 °C) thanks to simultaneous optimization of the rotational correlation time and of the water exchange rate. The relaxivity is however still limited by chelate flexibility. This report demonstrates that the DO3A-N-(α-amino)propionate chelator is a valuable platform for constructing high relaxivity CA using simple design principles and robust chemistries accessible to most chemistry labs.This work was financially supported by Fundação para a Ciência e Tecnologia, Portugal: project PTDC/QUI/70063/2006, including a grant to C.I.O.M., grant SFRH/BD/63994/2009 to M.F.F., grant SFRH/BD/46370/2008 to A.F.M. and sabbatical grant SFRH/BSAB/ 1328/2013 to J.A.M. and Rede Nacional de RMN (REDE/1517/RMN/2005) for the acquisition of the Varian VNMRS 600 NMR spectrometer in Coimbra. The work in France was supported by La Ligue contre le Cancer. This work was carried out in the framework of the COST Actions D38 “Metal Based Systems for Molecular Imaging”, TD1004 “Theragnostic Imaging” and CM1006 “EUFEN: European F-Element Network”. S.P. acknowledges support from the Institut National de la Santé et de la Recherche Médicale (INSERM)

Synthesis of new potent agonistic analogs of growth hormone-releasing hormone (GHRH) and evaluation of their endocrine and cardiac activities

•Analogs of GHRH modified at the N- and/or C-terminus were designed and synthesized.•Endocrine activities and binding affinities were determined.•Two analogs highly activated myocardial repair after induced myocardial infarction.•The relationship between GH-releasing potency and structure is discussed.•Substitutions with N-Me-Tyr1, Arg30-NHCH3, or Apa30-NH2 increased GHRH activities. In view of the recent findings of stimulatory effects of GHRH analogs, JI-34, JI-36 and JI-38, on cardiomyocytes, pancreatic islets and wound healing, three series of new analogs of GHRH(1–29) have been synthesized and evaluated biologically in an endeavor to produce more potent compounds. “Agmatine analogs”, MR-356 (N-Me-Tyr1-JI-38), MR-361(N-Me-Tyr1, D-Ala2-JI-38) and MR-367(N-Me-Tyr1, D-Ala2, Asn8-JI-38), in which Dat in JI-38 is replaced by N-Me-Tyr1, showed improved relative potencies on GH release upon subcutaneous administration in vivo and binding in vitro. Modification with N-Me-Tyr1 and Arg29-NHCH3 as in MR-403 (N-Me-Tyr1, D-Ala2, Arg29-NHCH3-JI-38), MR-406 (N-Me-Tyr1, Arg29-NHCH3-JI-38) and MR-409 (N-Me-Tyr1, D-Ala2, Asn8, Arg29-NHCH3-JI-38), and MR-410 (N-Me-Tyr1, D-Ala2, Thr8, Arg29-NHCH3-JI-38) resulted in dramatically increased endocrine activities. These appear to be the most potent GHRH agonistic analogs so far developed. Analogs with Apa30-NH2 such as MR-326 (N-Me-Tyr1, D-Ala2, Arg29, Apa30-NH2-JI-38), and with Gab30-NH2, as MR-502 (D-Ala2, 5F-Phe6, Ser28, Arg29,Gab30-NH2-JI-38) also exhibited much higher potency than JI-38 upon i.v. administration. The relationship between the GH-releasing potency and the analog structure is discussed. Fourteen GHRH agonists with the highest endocrine potencies were subjected to cardiologic tests. MR-409 and MR-356 exhibited higher potency than JI-38 in activating myocardial repair in rats with induced myocardial infarction. As the previous class of analogs, exemplified by JI-38, had shown promising results in multiple fields including cardiology, diabetes and wound healing, our new, more potent, GHRH agonists should manifest additional efficacy for possible medical applications

Novel GHRH antagonists suppress the growth of human malignant melanoma by restoring nuclear p27 function

Malignant melanoma is the deadliest form of skin cancer; the treatment of advanced and recurrent forms remains a challenge. It has recently been reported that growth hormone-releasing hormone (GHRH) receptor is involved in the pathogenesis of melanoma. Therefore, we investigated the effects of our new GHRH antagonists on a human melanoma cancer cell line. Antiproliferative effects of GHRH antagonists, MIA-602, MIA-606 and MIA-690, on the human melanoma cell line, A-375, were studied in vitro using the MTS assay. The effect of MIA-690 (5 μg/day 28 d) was further evaluated in vivo in nude mice bearing xenografts of A-375. Subcellular localization of p27 was detected with Western blot and immunofluorescent staining. MIA-690 inhibited the proliferation of A-375 cells in a dose-dependent manner (33% at 10 μM, and 19.2% at 5 μM, P < 0 .05 vs. control), and suppressed the growth of xenografted tumors by 70.45% (P < 0.05). Flow cytometric analysis of cell cycle effects following the administration of MIA-690 revealed a decrease in the number of cells in G2/M phase (from 19.7% to 12.9%, P < 0.001). Additionally, Western blot and immunofluorescent studies showed that exposure of A-375 cells to MIA-690 triggered the nuclear accumulation of p27. MIA-690 inhibited tumor growth in vitro and in vivo, and increased the translocation of p27 into the nucleus thus inhibiting progression of the cell cycle. Our findings indicate that patients with malignant melanoma could benefit from treatment regimens, which combine existing chemotherapy agents and novel GHRH-antagonists