RTP801 regulates motor cortex synaptic transmission and learning.

Background: RTP801/REDD1 is a stress-regulated protein whose upregulation is necessary and sufficient to trigger neuronal death in in vitro and in vivo models of Parkinson's and Huntington's diseases and is up regulated in compromised neurons in human postmortem brains of both neurodegenerative disorders. Indeed, in both Parkinson's and Huntington's disease mouse models, RTP801 knockdown alleviates motor-learning deficits.

Results: We investigated the physiological role of RTP801 in neuronal plasticity and we found RTP801 in rat, mouse and human synapses.

Post-Myocardial Infarction Heart Failure in Closed-chest Coronary Occlusion/Reperfusion Model in Göttingen Minipigs and Landrace Pigs.

The development of heart failure is the most powerful predictor of long-term mortality in patients surviving acute myocardial infarction (MI). There is an unmet clinical need for prevention and therapy of post-myocardial infarction heart failure (post-MI HF). Clinically relevant pig models of post-MI HF are prerequisites for final proof-of-concept studies before entering into clinical trials in drug and medical device development.

siRNA-Mediated Knockdown of the mTOR Inhibitor RTP801 Promotes Retinal Ganglion Cell Survival and Axon Elongation by Direct and Indirect Mechanisms.

Purpose: To investigate, using in vivo and in vitro models, retinal ganglion cell (RGC) neuroprotective and axon regenerative effects and underlying mechanisms of siRTP801, a translatable small-interfering RNA (siRNA) targeting the mTOR negative regulator RTP801.

Methods: Adult rats underwent optic nerve (ON) crush (ONC) followed by intravitreal siRTP801 or control siRNA (siEGFP) every 8 days, with Brn3a+ RGC survival, GFAP+ reactive gliosis, and GAP43+ regenerating axons analyzed immunohistochemically 24 days after injury. Retinal cultures, prepared from uninjured animals or 5 days after ONC to activate retinal glia, were treated with siRTP801/controls in the presence/absence of rapamycin and subsequently assessed for RGC survival and neurite outgrowth, RTP801 expression, glial responses, and mTOR activity.

The Toll-Like Receptor 5 Agonist Entolimod Mitigates Lethal Acute Radiation Syndrome in Non-Human Primates.

There are currently no approved medical radiation countermeasures (MRC) to reduce the lethality of high-dose total body ionizing irradiation expected in nuclear emergencies. An ideal MRC would be effective even when administered well after radiation exposure and would counteract the effects of irradiation on the hematopoietic system and gastrointestinal tract that contribute to its lethality. Entolimod is a Toll-like receptor 5 agonist with demonstrated radioprotective/mitigative activity in rodents and radioprotective activity in non-human primates.

Altered nutrient response of mTORC1 as a result of changes in REDD1 expression: effect of obesity vs. REDD1 deficiency.

Although aberrant mTORC1 signaling has been well established in models of obesity, little is known about its repressor, REDD1. Therefore, the initial goal of this study was to determine the role of REDD1 on mTORC1 in obese skeletal muscle. REDD1 expression (protein and message) and mTORC1 signaling (S6K1, 4E-BP1, raptor-mTOR association, Rheb GTP) were examined in lean vs.

Sestrin2 inhibits uncoupling protein 1 expression through suppressing reactive oxygen species.

Uncoupling protein 1 (Ucp1), which is localized in the mitochondrial inner membrane of mammalian brown adipose tissue (BAT), generates heat by uncoupling oxidative phosphorylation. Upon cold exposure or nutritional abundance, sympathetic neurons stimulate BAT to express Ucp1 to induce energy dissipation and thermogenesis. Accordingly, increased Ucp1 expression reduces obesity in mice and is correlated with leanness in humans.

DDIT4/REDD1/RTP801 is a novel negative regulator of Schwann cell myelination.

Signals that promote myelination must be tightly modulated to adjust myelin thickness to the axonal diameter. In the peripheral nervous system, axonal neuregulin 1 type III promotes myelination by activating erbB2/B3 receptors and the PI3K/AKT/mTOR pathway in Schwann cells. Conversely, PTEN (phosphatase and tensin homolog on chromosome 10) dephosphorylates PtdIns(3,4,5)P3 and negatively regulates the AKT pathway and myelination.

siRNA targeting Hes5 augments hair cell regeneration in aminoglycoside-damaged mouse utricle.

Notch signaling is active during the development of mosaic epithelial sheets and during their turnover and regeneration. After the loss of hair cells in the mosaic sheet of the vestibular sensory epithelium, new hair cells can be spontaneously generated by transdifferentiation of supporting cells. This regenerative process involves downregulation of the Hes5 gene and is known to be limited and incomplete, especially when the lesion is severe.

ASPP1/2 regulate p53-dependent death of retinal ganglion cells through PUMA and Fas/CD95 activation in vivo.

The transcription factor p53 mediates neuronal death in a variety of stress-related and neurodegenerative conditions. The proapoptotic activity of p53 is tightly regulated by the apoptosis-stimulating proteins of p53 (ASPP) family members: ASPP1 and ASPP2. However, whether ASPP1/2 play a role in the regulation of p53-dependent neuronal death in the CNS is currently unknown.

RTP801 is required for ceramide-induced cell-specific death in the murine lung.

Key host responses to the stress induced by environmental exposure to cigarette smoke (CS) are responsible for initiating pathogenic effects that may culminate in emphysema development. CS increases lung ceramides, sphingolipids involved in oxidative stress, structural alveolar cell apoptosis, and inhibition of apoptotic cell clearance by alveolar macrophages, leading to the development of emphysema-like pathology. RTP801, a hypoxia and oxidative stress sensor, is also increased by CS, and has been recently implicated in both apoptosis and inflammation.

Toxicological and pharmacokinetic properties of chemically modified siRNAs targeting p53 RNA following intravenous administration.

We report the toxicological and pharmacokinetic properties of the synthetic, small interfering RNA I5NP following intravenous administration in rodents and nonhuman primates. I5NP is designed to act via the RNA interference (RNAi) pathway to temporarily inhibit expression of the pro-apoptotic protein p53 and is being developed to protect cells from acute ischemia/reperfusion injuries such as acute kidney injury that can occur during major cardiac surgery and delayed graft function that can occur following renal transplantation. Following intravenous administration, I5NP was very rapidly cleared from plasma was distributed predominantly to the kidney, with very low levels in liver and other tissues.

Identification of granulocyte colony-stimulating factor and interleukin-6 as candidate biomarkers of CBLB502 efficacy as a medical radiation countermeasure.

Given an ever-increasing risk of nuclear and radiological emergencies, there is a critical need for development of medical radiation countermeasures (MRCs) that are safe, easily administered, and effective in preventing and/or mitigating the potentially lethal tissue damage caused by acute high-dose radiation exposure. Because the efficacy of MRCs for this indication cannot be ethically tested in humans, development of such drugs is guided by the Food and Drug Administration's Animal Efficacy Rule. According to this rule, human efficacious doses can be projected from experimentally established animal efficacious doses based on the equivalence of the drug's effects on efficacy biomarkers in the respective species.

Prevention and mitigation of acute radiation syndrome in mice by synthetic lipopeptide agonists of Toll-like receptor 2 (TLR2).

Bacterial lipoproteins (BLP) induce innate immune responses in mammals by activating heterodimeric receptor complexes containing Toll-like receptor 2 (TLR2). TLR2 signaling results in nuclear factor-kappaB (NF-κB)-dependent upregulation of anti-apoptotic factors, anti-oxidants and cytokines, all of which have been implicated in radiation protection. Here we demonstrate that synthetic lipopeptides (sLP) that mimic the structure of naturally occurring mycoplasmal BLP significantly increase mouse survival following lethal total body irradiation (TBI) when administered between 48 hours before and 24 hours after irradiation.

CBLB613: a TLR 2/6 agonist, natural lipopeptide of Mycoplasma arginini , as a novel radiation countermeasure.

To date, there are no safe and effective drugs available for protection against ionizing radiation damage. Therefore, a great need exists to identify and develop non-toxic agents that will be useful as radioprotectors or postirradiation therapies under a variety of operational scenarios. We have developed a new pharmacological agent, CBLB613 (a naturally occurring Mycoplasma-derived lipopeptide ligand for Toll-like receptor 2/6), as a novel radiation countermeasure.

Toll-like receptor 5 agonist protects mice from dermatitis and oral mucositis caused by local radiation: implications for head-and-neck cancer radiotherapy.

Purpose: Development of mucositis is a frequent side effect of radiotherapy of patients with head-and-neck cancer. We have recently reported that bacterial flagellin, an agonist of Toll-like receptor 5 (TLR5), can protect rodents and primates from acute radiation syndrome caused by total body irradiation. Here we analyzed the radioprotective efficacy of TLR5 agonist under conditions of local, single dose or fractionated radiation treatment.

Delayed intrathecal delivery of RhoA siRNA to the contused spinal cord inhibits allodynia, preserves white matter, and increases serotonergic fiber growth.

RhoA is a key regulator of the actin cytoskeleton that is upregulated after spinal cord injury (SCI). We analyzed different methods for siRNA delivery and developed siRNAs targeting RhoA (siRhoA) for SCI treatment. Cy 3.

Intravital two-photon microscopy assessment of renal protection efficacy of siRNA for p53 in experimental rat kidney transplantation models.

Renal ischemia-reperfusion (I/R) injury, which is unavoidable in renal transplantation, frequently influences both short- and long-term allograft survival. Despite decades of laboratory and clinical investigations, and the advent of renal replacement therapy, the overall mortality rate due to acute tubular injury has changed little. I/R-induced DNA damage results in p53 activation in proximal tubule cells (PTC), leading to their apoptosis.

Rtp801, a suppressor of mTOR signaling, is an essential mediator of cigarette smoke-induced pulmonary injury and emphysema.

Rtp801 (also known as Redd1, and encoded by Ddit4), a stress-related protein triggered by adverse environmental conditions, inhibits mammalian target of rapamycin (mTOR) by stabilizing the TSC1-TSC2 inhibitory complex and enhances oxidative stress-dependent cell death. We postulated that Rtp801 acts as a potential amplifying switch in the development of cigarette smoke-induced lung injury, leading to emphysema. Rtp801 mRNA and protein were overexpressed in human emphysematous lungs and in lungs of mice exposed to cigarette smoke.

Multiple parameter radiation injury assessment using a nonhuman primate radiation model-biodosimetry applications.

There are urgent needs to establish capability to rapidly assess radiation injury in mass casualty and population monitoring scenarios. This study's objective was to evaluate several currently available biomarkers that can provide early diagnostic triage information after radiation exposure. Hematology and blood chemistry measurements were performed on samples derived from a nonhuman primate (Macaca mulatta; n = 8) total-body irradiation (TBI) model (6.

siRNA targeted to p53 attenuates ischemic and cisplatin-induced acute kidney injury.

Proximal tubule cells (PTCs), which are the primary site of kidney injury associated with ischemia or nephrotoxicity, are the site of oligonucleotide reabsorption within the kidney. We exploited this property to test the efficacy of siRNA targeted to p53, a pivotal protein in the apoptotic pathway, to prevent kidney injury. Naked synthetic siRNA to p53 injected intravenously 4 h after ischemic injury maximally protected both PTCs and kidney function.

RNAi-mediated silencing of nuclear factor erythroid-2-related factor 2 gene expression in non-small cell lung cancer inhibits tumor growth and increases efficacy of chemotherapy.

Nuclear factor erythroid-2-related factor 2 (Nrf2) is a redox-sensitive transcription factor that regulates the expression of electrophile and xenobiotic detoxification enzymes and efflux proteins, which confer cytoprotection against oxidative stress and apoptosis in normal cells. Loss of function mutations in the Nrf2 inhibitor, Kelch-like ECH-associated protein (Keap1), results in constitutive activation of Nrf2 function in non-small cell lung cancer. In this study, we show that constitutive activation of Nrf2 in lung cancer cells promotes tumorigenicity and contributes to chemoresistance by up-regulation of glutathione, thioredoxin, and the drug efflux pathways involved in detoxification of electrophiles and broad spectrum of drugs.

An agonist of toll-like receptor 5 has radioprotective activity in mouse and primate models.

The toxicity of ionizing radiation is associated with massive apoptosis in radiosensitive organs. Here, we investigate whether a drug that activates a signaling mechanism used by tumor cells to suppress apoptosis can protect healthy cells from the harmful effects of radiation. We studied CBLB502, a polypeptide drug derived from Salmonella flagellin that binds to Toll-like receptor 5 (TLR5) and activates nuclear factor-kappaB signaling.

Studies on the cholesterol-free mouse: strong activation of LXR-regulated hepatic genes when replacing cholesterol with desmosterol.

Objective: Characterization of cholesterol homeostasis in male mice with a genetic inactivation of 3beta-hydroxysteroid-delta24-reductase, causing replacement of almost all cholesterol with desmosterol.

Methods And Results: There was an increase in hepatic sterol synthesis and markedly increased fecal loss of neutral sterols. Fecal excretion of bile acids was similar in knockout mice and in controls.

p120 catenin regulates lamellipodial dynamics and cell adhesion in cooperation with cortactin.

The armadillo-family protein, p120 catenin (p120), binds to the juxtamembrane domain of classical cadherins and increases cell-cell junction stability. Overexpression of p120 modulates the activity of Rho family GTPases and augments cell migratory ability. Here we show that down-regulation of p120 in epithelial MCF-7 cells by siRNA leads to a striking decrease in lamellipodial persistence and focal adhesion formation.