Developmental expression of GPR3 in rodent cerebellar granule neurons is associated with cell survival and protects neurons from various apoptotic stimuli.

G-protein coupled receptor 3 (GPR3), GPR6, and GPR12 belong to a family of constitutively active Gs-coupled receptors that activate 3'-5'-cyclic adenosine monophosphate (cAMP) and are highly expressed in the brain. Among these receptors, the endogenous expression of GPR3 in cerebellar granule neurons (CGNs) is increased following development. GPR3 is important for neurite outgrowth and neural maturation; however, the physiological functions of GPR3 remain to be fully elucidated.

The Gs-linked receptor GPR3 inhibits the proliferation of cerebellar granule cells during postnatal development.

Background: During postnatal murine and rodent cerebellar development, cerebellar granule precursors (CGP) gradually stop proliferating as they differentiate after migration to the internal granule layer (IGL). Molecular events that govern this program remain to be fully elucidated. GPR3 belongs to a family of Gs-linked receptors that activate cyclic AMP and are abundantly expressed in the adult brain.

Four distinct phases of basket/stellate cell migration after entering their final destination (the molecular layer) in the developing cerebellum.

In the adult cerebellum, basket/stellate cells are scattered throughout the ML, but little is known about the process underlying the cell dispersion. To determine the allocation of stellate/basket cells within the ML, we examined their migration in the early postnatal mouse cerebellum. We found that after entering the ML, basket/stellate cells sequentially exhibit four distinct phases of migration.

ICP0 inhibits the decrease of HSV amplicon-mediated transgene expression.

The herpes simplex virus (HSV) amplicon vector produces an initial host response that limits transgene expression. In this study, we hypothesized that restoration of the HSV gene infected cell protein (ICP0) into the amplicon could circumvent this host response and thus overcome silencing of encoded transgenes. To test this, we constructed an amplicon vector that encodes the ICP0 under control of its native promoter (ICP0+ amplicon).

Neuroblastoma cell lines contain pluripotent tumor initiating cells that are susceptible to a targeted oncolytic virus.

Background: Although disease remission can frequently be achieved for patients with neuroblastoma, relapse is common. The cancer stem cell theory suggests that rare tumorigenic cells, resistant to conventional therapy, are responsible for relapse. If true for neuroblastoma, improved cure rates may only be achieved via identification and therapeutic targeting of the neuroblastoma tumor initiating cell.

Histone Deacetylase Inhibitors Augment Antitumor Efficacy of Herpes-based Oncolytic Viruses.

Replication-conditional (oncolytic) mutants of herpes simplex virus (HSV), are considered promising therapeutic alternatives for human malignancies, and chemotherapeutic adjuvants are increasingly sought to augment their efficacy. Histone deacetylase (HDAC) inhibitors are a new class of antineoplastic agents because of their potent activity in growth arrest, differentiation, and apoptotic death of cancer cells. The ability of the HDAC inhibitors to upregulate exogenous transgene expression and inhibit interferon (IFN) responses prompted our exploration of their use in improving the antitumor efficacy of oncolytic HSV.

Stable transgene expression from HSV amplicon vectors in the brain: potential involvement of immunoregulatory signals.

The herpes simplex virus (HSV) amplicon is a plasmid-based, infectious gene delivery system that carries up to 150 kilobase (kb) of exogenous DNA. We previously characterized early host responses and stability of transgene expression in mice systemically injected with HSV amplicon vectors. Transgene expression was readily detected primarily in the liver but rapidly declined to undetectable levels within 2 weeks.

Lithium inhibits invasion of glioma cells; possible involvement of glycogen synthase kinase-3.

Therapies targeting glioma cells that diffusely infiltrate normal brain are highly sought after. Our aim was to identify novel approaches to this problem using glioma spheroid migration assays. Lithium, a currently approved drug for the treatment of bipolar illnesses, has not been previously examined in the context of glioma migration.

Histone deacetylase inhibitors augment antitumor efficacy of herpes-based oncolytic viruses.

Replication-conditional (oncolytic) mutants of herpes simplex virus (HSV), are considered promising therapeutic alternatives for human malignancies, and chemotherapeutic adjuvants are increasingly sought to augment their efficacy. Histone deacetylase (HDAC) inhibitors are a new class of antineoplastic agents because of their potent activity in growth arrest, differentiation, and apoptotic death of cancer cells. The ability of the HDAC inhibitors to upregulate exogenous transgene expression and inhibit interferon (IFN) responses prompted our exploration of their use in improving the antitumor efficacy of oncolytic HSV.

Tissue inhibitor of metalloproteinase-3 via oncolytic herpesvirus inhibits tumor growth and vascular progenitors.

Malignant solid tumors remain a significant clinical challenge, necessitating innovative therapeutic approaches. Oncolytic viral therapy is a nonmutagenic, biological anticancer therapeutic shown to be effective against human cancer in early studies. Because matrix metalloproteinases (MMP) play important roles in the pathogenesis and progression of cancer, we sought to determine if "arming" an oncolytic herpes simplex virus (oHSV) with an MMP-antagonizing transgene would increase virus-mediated antitumor efficacy.

Early STAT1 activation after systemic delivery of HSV amplicon vectors suppresses transcription of the vector-encoded transgene.

The herpes simplex virus (HSV) amplicon vector is a powerful gene delivery vehicle that can accommodate up to 150 kilobase of exogenous DNA. However, amplicon-mediated transgene expression is often transient outside the nervous system. In order to define the role of host immune responses in silencing amplicon-encoded transgenes, we evaluated the kinetics of cytokine-/chemokine-expression after tail-vein injection of a luciferase-encoding amplicon into mice.

Neural expression of G protein-coupled receptors GPR3, GPR6, and GPR12 up-regulates cyclic AMP levels and promotes neurite outgrowth.

Cyclic AMP regulates multiple neuronal functions, including neurite outgrowth and axonal regeneration. GPR3, GPR6, and GPR12 make up a family of constitutively active G protein-coupled receptors (GPCRs) that share greater than 50% identity and 65% similarity at the amino acid level. They are highly expressed in the central nervous system, and their expression in various cell lines results in constitutive stimulation of cAMP production.

Cyclophosphamide increases transgene expression mediated by an oncolytic adenovirus in glioma-bearing mice monitored by bioluminescence imaging.

Approaches to improve the oncolytic potency of replication-competent adenoviruses include the insertion of therapeutic transgenes into the viral genome. Little is known about the levels and duration of in vivo transgene expression by cells infected with such "armed" viruses. Using a tumor-selective adenovirus encoding firefly luciferase (AdDelta24CMV-Luc) we investigated these questions in an intracranial mouse model for malignant glioma.

DNA-based methods to prepare helper virus-free herpes amplicon vectors and versatile design of amplicon vector plasmids.

The herpes simplex virus (HSV) amplicon vector is a versatile plasmid-based gene delivery vehicle with a large transgene capacity (up to 150 kb) and the ability to infect a broad range of cell types. The vector system was originally developed by Frenkel and her colleagues in 1980. Ever since, a great deal of effort by various investigators has been directed at minimizing the toxicity associated with the inevitable contamination by helper virus.

Plasmid DNA sequences present in conventional herpes simplex virus amplicon vectors cause rapid transgene silencing by forming inactive chromatin.

The herpes simplex virus (HSV)-based amplicon vector, a bacterial-viral-mammalian cell shuttle system, holds promise as a versatile gene delivery vehicle because of its large transgene capacity. However, amplicon-mediated transgene expression is often transient. We hypothesized that the presence of prokaryotic DNA sequences within the packaged vector genome can trigger transcriptional silencing of the entire vector sequence.

Triple combination of oncolytic herpes simplex virus-1 vectors armed with interleukin-12, interleukin-18, or soluble B7-1 results in enhanced antitumor efficacy.

Conditionally replicating herpes simplex virus-1 (HSV-1) vectors are promising therapeutic agents for cancer. Insertion of therapeutic transgenes into the viral genome should confer desired anticancer functions in addition to oncolytic activities. Herein, using bacterial artificial chromosome and two recombinase-mediated recombinations, we simultaneously created four "armed" oncolytic HSV-1, designated vHsv-B7.

Cyclophosphamide allows for in vivo dose reduction of a potent oncolytic virus.

The success of cancer virotherapy depends on its efficacy versus toxicity profile in human clinical trials. Progress towards clinical trials can be hampered by the relatively elevated doses of oncolytic viruses administered in animal models to achieve an anticancer effect and by the even higher doses required in humans to approximate an animal bioequivalent dose. Such elevated doses of injected viral proteins may also lead to undesirable toxicities and are also very difficult to produce in a biotechnological setting.

Volumetric tomography of fluorescent proteins through small animals in vivo.

Volumetric detection and accurate quantification of fluorescent proteins in entire animals would greatly enhance our ability to monitor biological processes in vivo. Here we present a quantitative tomographic technique for visualization of superficial and deep-seated (>2-3 mm) fluorescent protein activity in vivo. We demonstrate noninvasive imaging of lung tumor progression in a murine model, as well as imaging of gene delivery using a herpes virus vector.

Regulation of meiotic prophase arrest in mouse oocytes by GPR3, a constitutive activator of the Gs G protein.

The arrest of meiotic prophase in mouse oocytes within antral follicles requires the G protein G(s) and an orphan member of the G protein-coupled receptor family, GPR3. To determine whether GPR3 activates G(s), the localization of Galpha(s) in follicle-enclosed oocytes from Gpr3(+/+) and Gpr3(-/-) mice was compared by using immunofluorescence and Galpha(s)GFP. GPR3 decreased the ratio of Galpha(s) in the oocyte plasma membrane versus the cytoplasm and also decreased the amount of Galpha(s) in the oocyte.

Brain tumor oncolysis with replication-conditional herpes simplex virus type 1 expressing the prodrug-activating genes, CYP2B1 and secreted human intestinal carboxylesterase, in combination with cyclophosphamide and irinotecan.

The treatment of malignant glioma is currently ineffective. Oncolytic viruses are being explored as a means to selectively lyse tumor cells in the brain. We have engineered a mutant herpes simplex virus type 1 with deletions in the viral UL39 and gamma(1)34.

An oncolytic HSV-1 mutant expressing ICP34.5 under control of a nestin promoter increases survival of animals even when symptomatic from a brain tumor.

Oncolytic herpes simplex virus-1 (HSV-1) mutants possessing mutations in the ICP34.5 and ICP6 genes have proven safe through clinical trials. However, ICP34.

The Gs-linked receptor GPR3 maintains meiotic arrest in mammalian oocytes.

Mammalian oocytes are held in prophase arrest by an unknown signal from the surrounding somatic cells. Here we show that the orphan Gs-linked receptor GPR3, which is localized in the oocyte, maintains this arrest. Oocytes from Gpr3 knockout mice resume meiosis within antral follicles, independently of an increase in luteinizing hormone, and this phenotype can be reversed by injection of Gpr3 RNA into the oocytes.

Gene transfer into hepatocytes mediated by herpes simplex virus-Epstein-Barr virus hybrid amplicons.

Gene transfer into hepatocytes is highly desirable for the long-term goal of replacing deficient proteins and correcting metabolic disorders. Vectors based on herpes simplex virus type-1 (HSV-1) have been demonstrated to mediate efficient gene transfer into hepatocytes both in vitro and in vivo. Large transgene capacity and extrachromosomal persistence make HSV-1/EBV hybrid amplicon vectors an attractive candidate for hepatic gene replacement therapy.

Cadherin activity is required for activity-induced spine remodeling.

Neural activity induces the remodeling of pre- and postsynaptic membranes, which maintain their apposition through cell adhesion molecules. Among them, N-cadherin is redistributed, undergoes activity-dependent conformational changes, and is required for synaptic plasticity. Here, we show that depolarization induces the enlargement of the width of spine head, and that cadherin activity is essential for this synaptic rearrangement.