Herpes simplex virus 1 as an oncolytic viral therapy for refractory cancers.

The need for efficacious and non-toxic cancer therapies is paramount. Oncolytic viruses (OVs) are showing great promise and are introducing new possibilities in cancer treatment with their ability to selectively infect tumor cells and trigger antitumor immune responses. Herpes Simplex Virus 1 (HSV-1) is a commonly selected OV candidate due to its large genome, relative safety profile, and ability to infect a variety of cell types. Talimogene laherparevec (T-VEC) is an HSV-1-derived OV variant and the first and only OV therapy currently approved for clinical use by the United States Food and Drug Administration (FDA). This review provides a concise description of HSV-1 as an OV candidate and the genomic organization of T-VEC. Furthermore, this review focuses on the advantages and limitations in the use of T-VEC compared to other HSV-1 OV variants currently in clinical trials. In addition, approaches for future directions of HSV-1 OVs as cancer therapy is discussed.

Utilization of an Abiomed Impella Device as a Rescue Therapy for Acute Ventricular Failure in a Fontan Patient.

Progressive ventricular dysfunction is not uncommon in patients with univentricular hearts as they age. In the acute setting vasoactive support can be employed, but is not always sufficient and patients occasionally require mechanical support. We report the successful implantation and subsequent challenges of a percutaneous Abiomed Impella ventricular assist device as a rescue therapy for a 15-year old-patient with Fontan circulation and severe ventricular dysfunction after cardiac arrest.

Moderate endoplasmic reticulum stress activates a PERK and p38-dependent apoptosis.

The endoplasmic reticulum (ER) has the ability to signal organelle dysfunction via a complex signaling network known as the unfolded protein response (UPR). In this work, hamster fibroblast cells exhibiting moderate levels of ER stress were compared to those exhibiting severe ER stress. Inhibition of N-linked glycosylation was accomplished via a temperature-sensitive mutation in the Dad1 subunit of the oligosaccharyltransferase (OST) complex or by direct inhibition with tunicamycin (Tm). Temperature shift (TS) treatment generated weak activation of ER stress signaling when compared to doses of Tm that are typically used in ER stress studies (500-1000 nM). A dose-response analysis of key ER stress signaling mediators, inositol-requiring enzyme 1 (IRE1) and protein kinase R (PKR)-like endoplasmic reticulum kinase (PERK), revealed 20-40 nM of Tm to generate activation intensity similar to TS treatment. In parental BHK21 cells, moderate (20-40 nM) and high doses (200-1000 nM) of Tm were compared to identify physiological and signaling-based differences in stress response. Inhibition of ER Ca2+ release via ITPR activity with 2-aminoethoxydiphenyl borate (2-APB) or Xestospongin C (XeC) was sufficient to protect against apoptosis induced by moderate but not higher doses of Tm. Analysis of kinase activation over a range of Tm exposures revealed the p38 stress-activated protein kinase (SAPK) to display increasing activation with Tm dosage. Interestingly, Tm induced the extracellular regulated kinases (Erk1/2) only at moderate doses of Tm. Inhibition of ER transmembrane stress sensors (IRE1, PERK) or cytosolic signaling mediators (p38, Jnk1, Erk1/2) was used to evaluate pathways involved in apoptosis activation during ER stress. Inhibition of either PERK or p38 was sufficient to reduce cell death and apoptosis induced by moderate, but not high, doses of Tm. During ER stress, cells exhibited a rapid decline in anti-apoptotic Mcl-1 and survivin proteins. Inhibition of PERK was sufficient to block this affect. This work reveals moderate doses of ER stress to generate patterns of stress signaling that are distinct from higher doses and that apoptosis activation at moderate levels of stress are dependent upon PERK and p38 signaling. Studies exploring ER stress signaling should recognize that this signaling acts as a rheostat rather than a simple switch, behaving distinctively in a dose-dependent manner.

Development of a high-throughput assay for monitoring cAMP levels in cardiac ventricular myocytes.

G-protein-coupled receptors (GPCRs) represent the largest family of transmembrane receptors involved in cell signal transduction. Many of these GPCRs convey their pharmacological actions by regulating intracellular levels of 3',5'-cyclic adenosine monophosphate (cAMP). Although the heart expresses more than 100 GPCRs, drug agonists for approximately one third of these GPCRs have not been identified. The goal of this project was to initiate the development of a high-throughput screening assay for monitoring cAMP in the heart. Neonatal rat cardiac ventricular myocytes were isolated and cultured on coverslips (whole-cell patch clamp recording) or in 96-well plates (fluorescent imaging plate reader measurements). Cells were infected with adenovirus expressing either beta-galactosidase (AdLacZ) or a mutant cyclic nucleotide-gated (CNG) channel containing the double mutation C460W/E583M (AdCNG). Addition of 2 microM forskolin along with 100 microM 3-isobutyl-1-methylxanthine, to increase intracellular cAMP, activated a cation current in myocytes infected with the AdCNG. In myocytes loaded with the fluorescent Ca indicator Fluo-4, stimulation with forskolin, epinephrine, norepinephrine, or the beta-adrenergic receptor agonist isoproterenol increased the fluorescent signal indicative of Ca influx through the CNG channel. In conclusion, CNG channels are readily expressed in cultured cardiac myocytes and may be utilized in high-throughput screening assays of intracellular cAMP.