Flowchart: Preparation: Pka
 


                         

                               

                        

Text Box: U(S)3 Text Box: Hdac2
Text Box: Hdac1
 


 

 

 

Text Box: PKA
Text Box: Ras


Text Box: IL13 alpha


Text Box: IIalpha

J Virol. 2006 Apr;80(8):3752-64.

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US3 and US3.5 Protein Kinases of Herpes Simplex Virus 1 Differ with Respect to Their Functions in Blocking Apoptosis and in Virion Maturation and Egress.

Poon AP, Benetti L, Roizman B.

The Marjorie B. Kovler Viral Oncology Laboratories, The University of Chicago, 910 East 58th Street, Chicago, IL 60637. bernard.roizman@bsd.uchicago.edu.

Previously, we reported that the U(S)3 protein kinase blocks apoptosis, that it activates protein kinase A (PKA), that activation of PKA blocks apoptosis in cells infected with a U(S)3 deletion mutant, and that an overlapping transcriptional unit encodes a truncated kinase designated U(S)3.5. Here, we report the properties of the kinases based on comparisons of herpes simplex virus and baculoviruses expressing U(S)3 or U(S)3.5 kinase. Specifically, we report the following. (i) Both kinases mediate the phosphorylation of HDAC1, HDAC2, and the PKA regulatory IIalpha subunit in the absence of other viral proteins. (ii) Both enzymes mediate the phosphorylation of largely identical sets of proteins carrying the phosphorylation consensus site of PKA, but only U(S)3 blocks apoptosis, suggesting that it is U(S)3 and not PKA that is responsible for the phosphorylation of the proteins bearing the shared consensus phosphorylation site and the antiapoptotic activity. (iii) Both kinases cofractionate with mitochondria. Immune depletion of the U(S)3 and U(S)3.5 kinases from the cytoplasm removed the kinases from the supernatant fraction, but not from the mitochondrial fraction, and therefore, if the antiapoptotic activity of the U(S)3 kinase is expressed in mitochondria, the localization signal and the antiapoptotic functions are located on different parts of the protein. (iv) The U(S)3 protein kinase is required for the translocation of virus particles from the nucleus. Although the U(L)31 protein is phosphorylated in cells infected with the mutant expressing U(S)3.5 kinase, the release of virus particles from nuclei was impeded in some cells, suggesting that the U(S)3 kinase affects the modification of the nuclear membrane more efficiently than the U(S)3.5 kinase.

PMID: 16571792 [PubMed - in process]

Proc Natl Acad Sci U S A. 2006 Mar 22; [Epub ahead of print]

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Construction and properties of a herpes simplex virus 1 designed to enter cells solely via the IL-13{alpha}2 receptor.

Zhou G, Roizman B.

The Marjorie B. Kovler Viral Oncology Laboratories, University of Chicago, 910 East 58th Street, Chicago, IL 60637.

Current design of genetically engineered viruses for selective destruction of cancer cells is based on the observation that attenuated viruses replicate better in tumor cells than in normal cells. The ideal virus, however, is one that can infect only cancer cells by virtue of altered host range. Such a virus can be made more robust than the highly attenuated viruses used in clinical trials. Earlier, we reported the construction of a recombinant herpes simplex virus 1 (R5111) in which the capacity to bind heparan sulfate was disabled and which contained a chimeric IL-13-glycoprotein D that enabled the virus to infect cells expressing the IL-13alpha2 receptor (IL-13Ralpha2) commonly found on the surface of malignant glioblastomas or high-grade astrocytomas. In the earlier report, we showed that the recombinant R5111 was able to enter and infect cells via the interaction of the chimeric glycoprotein D with IL-13Ralpha2 but that the virus retained the capacity to bind and replicate in cells expressing the natural viral receptors HveA or nectin-1. Here, we report the construction of a recombinant virus (R5141) that can only enter and replicate in cells that express the IL-13Ralpha2. The recombinant R5141 does not depend on endocytosis to infect cells. It does not infect cells expressing HveA or nectin-1 receptors or cells expressing IL-13Ralpha2 that had been exposed to soluble IL-13 before infection. The studies described here show that the host range of herpes simplex viruses can be altered by genetic manipulation to specifically target cancer cells.

PMID: 16554374 [PubMed - as supplied by publisher]

Mol Ther. 2006 Mar 25; [Epub ahead of print]

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A Mutant Type 2 Herpes Simplex Virus Deleted for the Protein Kinase Domain of the ICP10 Gene Is a Potent Oncolytic Virus.

Fu X, Tao L, Cai R, Prigge J, Zhang X.

Center for Cell and Gene Therapy, Baylor College of Medicine, Houston, TX 77030, USA; Department of Pediatrics, Baylor College of Medicine, Houston, TX 77030, USA.

Replication-selective oncolytic herpes simplex virus (HSV) has shown considerable promise as an antitumor agent. Although the current oncolytic HSVs were exclusively constructed from HSV-1, HSV-2 has several unique features that could be exploited to convert t

he virus to an oncolytic agent. The N-terminus of the HSV-2 ICP10 gene product contains a well-defined serine/threonine protein kinase (PK) domain, which can activate the Ras/MEK/MAPK mitogenic pathway and thus facilitate efficient HSV-2 replication. Because the Ras signaling pathway is a key regulator of normal cell growth and malignant transformation, it is aberrantly activated in many human tumors. Here we report that a mutant HSV-2 (FusOn-H2), constructed by replacing the PK domain of ICP10 with the gene encoding the green fluorescent protein, can selectively replicate in and thus lyse tumor cells. Moreover, infection of FusOn-H2 led to syncytia formation in tumor cells, providing an additional tumor-destroying mechanism. A single moderate-dose injection of FusOn-H2 into established breast cancer xenografts completely eradicated the tumors in more than 80% of the animals, leading to their long-term survival. We conclude that this HSV-2 mutant is a safe and potent oncolytic agent useful for the treatment of malignant solid tumors such as breast cancer.

PMID: 16569513 [PubMed - as supplied by publisher]

Gene Ther. 2006 Mar 23; [Epub ahead of print]

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Oncolysis of pancreatic tumour cells by a gamma34.5-deleted HSV-1 does not rely upon Ras-activation, but on the PI 3-kinase pathway.

Sarinella F, Calistri A, Sette P, Palu G, Parolin C.

1Department of Histology, Microbiology and Medical Biotechnologies, Section of Microbiology and Virology, Medical School, University of Padua, Padua, Italy.

The ability of viruses to selectively target, replicate within, and destroy tumour cells without deleterious effects in normal cells (oncolysis), makes the use of viruses as an attractive tool for cancer treatment. Pancreatic adenocarcinoma, being insensitive to traditional therapy and having a rather poor prognosis, represents a suitable target to evaluate viral oncolysis as a novel therapeutic approach. Herpes simplex virus (HSV) has been reported to produce an oncolytic effect in cells overexpressing Ras. As Ras signalling is frequently aberrant in pancreatic cancer, we compared four pancreatic cell lines (which differ in the presence of mutated or wild-type ras) for their ability to support growth of gamma34.5-replication attenuated HSV-1 (R3616). Our data show that permissiveness to viral replication is neither associated with enhanced Ras signalling nor with defective PKR activity. By contrast, we provide evidence that disregulation of the PI 3-kinase signalling pathway allows conditionally replication-defective R3616 virus to overcome the cellular antiviral activity.Gene Therapy advance online publication, 23 March 2006; doi:10.1038/sj.gt.3302770.