Ultrastructural Analysis of ICP34.5 ⁻ Herpes Simplex Virus 1 Replication in Mouse Brain Cells In Vivo

Abstract

Replication-competent forms of herpes simplex virus 1 (HSV-1) defective in the viral neurovirulence factor infected cell protein 34.5 (ICP34.5) are under investigation for use in the therapeutic treatment of cancer. In mouse models, intratumoral injection of ICP34.5-defective oncolytic HSVs (oHSVs) has resulted in the infection and lysis of tumor cells, an associated decrease in tumor size, and increased survival times. The ability of these oHSVs to infect and lyse cells is frequently characterized as exclusive to or selective for tumor cells. However, the extent to which ICP34.5-deficient HSV-1 replicates in and may be neurotoxic to normal brain cell types in vivo is poorly understood. Here we report that HSV-1 defective in ICP34.5 expression is capable of establishing a productive infection in at least one normal mouse brain cell type. We show that ␥34.5 deletion viruses replicate productively in and induce cellular damage in infected ependymal cells. Further evaluation of the effects of oHSVs on normal brain cells in animal models is needed to enhance our understanding of the risks associated with the use of current and future oHSVs in the brains of clinical trial subjects and to provide information that can be used to create improved oHSVs for future use.Several types of replication-competent neuroattenuated herpes simplex viruses (HSVs) are currently being evaluated in clinical cancer trials for safety and therapeutic activity (32), as well as for vaccine development (20). A critical safety concern associated with the clinical use of these oncolytic HSVs (oHSVs) is their ability to enter, replicate in, and spread to a wide range of cell types in different regions of the nervous system. One potential complication resulting from invasion of the central nervous system by HSV is herpes simplex encephalitis (HSE), an infection that causes lifelong neurological damage or death. A limited number of genes have been demonstrated to contribute to the virus's ability to trigger HSE. The viral gene ␥34.5 encodes the neurovirulence protein infected cell protein 34.5 (ICP34.5) (29). Viruses lacking the ␥34.5 gene (e.g., R3616 and 1716) were found to be 5 logs less neurovirulent than wild-type strains of 19,36), as quantified by the intracranial LD 50 , i.e., the lethal dose in 50% of mice inoculated intracerebroventricularly with the virus. The basis for this neuroattenuation was initially reported to be the inability of the ␥34.5 deletion viruses to infect or replicate in brain cells (4). Subsequent immunohistochemical studies on infected brain tissue of intracerebroventricularly inoculated mice suggested that ␥34.5 deletion viruses retained the ability to infect a wide range of brain cell types and to replicate in and, by day 7, destroy ependymal cells (ECs) (16,21).To create a more neuroattenuated and thus safer virus, the virus G207 was constructed from the ␥34.5 deletion virus R3616 by insertional mutagenesis of the U L 39 gene (25). The U L 39 gene encodes the large subunit of the viral ribonucleotide reductase (vR...

Pre-clinical Assessment of C134, a Chimeric Oncolytic Herpes Simplex Virus, in Mice and Non-human Primates

“…The speed with which the virus undergoes cell-to-cell spread was the focus of studies by Markovitz et al 1, 2. They showed that Δγ 1 34.5 virus is capable of viral replication including synaptic spread but that this is slow and inefficient and that by D6–D8 post-infection, intrinsic and immune-mediated anti-viral responses limit Δγ 1 34.5 replication and clear the infection 1 .…”

“…We previously reported the construction of a chimeric HSV-1, C134, which is a neurovirulent and is deleted for the γ 1 34.5 gene but expresses the human cytomegalovirus (HCMV) IRS1 gene. The IRS1 gene improves Δγ 1 34.5 replication within tumors and improves its anti-tumor activity in murine tumor models, yet C134 remains as safe as its parent Δγ 1 34.5 virus in lethal dose 50% (LD 50 ) and virus recovery studies 1, 2…”

“…By stereotactically injecting virus into the caudate putamen nucleus and recording neurologic dysfunction and mouse deaths over a 28-day period, an objective and quantitative measure of neurovirulence (LD 50 values) can be calculated using the Spearman-Karber calculation 3, 6, 11, 12. Wild-type (WT) HSV replicates more efficiently, generates greater viral progeny, and efficiently undergoes cell-to-cell spread, leading to death of the animal 1, 2, 13, 14, 15, 16…”

Pancreatic Cancer Gene Therapy: From Molecular Targets to Delivery Systems

“…For instance, deletion of the HSV-1 gamma 34.5 gene limited replication in tumor cells, although recent studies have shown some degree of neurotoxicity in normal brain cells [140]. Another oncolytic virus, the FusOn-H2 virus, was generated by introducing mutations in the PK domain of the viral ICP10 gene, in this virus replication was dependent on the activation of the Ras signaling pathway [93].…”

Effect of a caspase inhibitor, zVADfmk, on the inhibition of breast cancer cells by herpes simplex virus type 1

“…[1][2][3] The removal of virulence genes enhances the safety of HSV-1 vectors against serious complications and/or reversion to wild type. [4][5][6] Some of these oncolytic HSV-1 viruses have been approved for clinical trials in breast cancer, 3,7 head and neck carcinoma, 8 glioma, 9 melanoma, 10 non-small-cell lung carcinoma 11 and metastases from colorectal cancer 12 (reviewed in Manservigi et al 13 ). These clinical trials demonstrate that HSV-1 vectors are generally well tolerated at high doses with few side effects.…”