¹Molecular Neuro-Oncology Laboratories, Neurosurgical Service, Massachusetts General Hospital, Charlestown, MA, USA

Correspondence: Dr Y Saeki, Dardinger Laboratory for Neuro-oncology and Neurosciences, Department of Neurological Surgery, The Ohio State University Medical Center, 385B Wiseman Hall, 400 West 12th Avenue, Columbus, OH 43210, USA. E-mail: saeki.6@osu.edu

²Current address: Department of Neurological Surgery, Okayama University Graduate School of Medicine and Dentistry, Okayama, Japan.

³Current address: Department of Neurosurgery, School of Medicine, Tokyo Medical and Dental University, Tokyo, Japan.

⁴Current address: Dardinger Laboratory for Neuro-oncology and Neurosciences, Department of Neurological Surgery, The Ohio State University Medical Center, Columbus, OH 43210, USA.

Received 2 October 2005; Accepted 30 November 2005; Published online 10 January 2006.

Abstract

Replication-conditional herpes simplex virus (HSV)-based vectors have great potential in the treatment of various types of cancers including brain tumors. HSV mutants lacking the U_L39 gene and both copies of the ₁34.5 gene (e.g. MGH1, G207) have been demonstrated to possess oncolytic effects as well as potent anticancer vaccination effects without compromising safety. Such mutants thus provide optimal templates to produce novel oncolytic HSV vectors for cancer gene therapy applications. In order to accomplish quick and efficient construction of oncolytic HSV vectors, a novel BAC-based method designated as 'HSVQuik system' was developed. This system sequentially utilizes two different site-specific recombination systems to introduce virtually any transgene cassettes of interest into the deleted U_L39 locus (Flp-FRT in Escherichia coli) and to release the vector genome sequence from the procaryotic plasmid backbone (Cre-loxP in Vero cells). Taking advantage of the HSVQuik system, we constructed three oncolytic HSV vectors that express mouse IL4, CD40 ligand and 6CK, respectively. In vivo therapeutic experiments using two luciferase-labeled syngeneic mouse brain tumor models revealed that expression of these immunomodulators significantly enhanced antitumor efficacy of oncolytic HSV. The HSVQuik system, together with luciferase-labeled tumor models, should expedite the process of generating and evaluating oncolytic HSV vectors for cancer gene therapy applications.