Characterization of a Recombinant Herpes Simplex Virus 1 Designed To Enter Cells via the IL13Rα2 Receptor of Malignant Glioma Cells

Malignant glioma tumor cells in situ exhibit on their surfaces the interleukin 13 (IL-13) receptor designated IL13R2. To target herpes simplex virus 1 to this receptor, we constructed a recombinant virus (R5111) in which the known heparan sulfate binding sites in glycoproteins B and C were deleted and IL-13 was inserted into both glycoproteins C and D. We also transduced a baby hamster kidney cell line lacking the known viral receptors (J1-1) and Vero cells with a plasmid encoding IL13R2. The J1-1 derivative (J-13R) cell line is susceptible to and replicates the R5111 recombinant virus but not the wild-type parent virus. We report the following. (i) Expression of IL13R2 was rapidly lost from the surface of transduced cells grown in culture. The loss appeared to be related to ligands present in fetal bovine serum in the medium. None of the malignant glioma cell lines cultivated in vitro and tested to date exhibited the IL13R2 receptor. (ii) Soluble IL-13 but not IL-4 or IL-2 blocked the replication of R5111 recombinant virus in J-13R cells. (iii) The endocytosis inhibitor PD98059 blocked the replication in J1-1 cells of a mutant lacking glycoprotein D (gD / ) but not the replication of R5111 in the J-13R cells. We conclude that R5111 enters cells via its interaction with the IL13R2 receptor in a manner that cannot be differentiated from the interaction of wild-type virus with its receptors. The experiments described in this report stemmed from two considerations. In the last decade, several laboratories have attempted to develop herpes simplex virus 1(HSV-1) mutants suitable for cytoreductive therapy of human malignant gliomas. It is estimated that these incurable tumors claim 10,000 lives per year in the United States alone, and current therapeutic modalities seldom produce long-lasting remissions (14). Of the various mutants tested in the mouse model, two such mutants found their way to clinical trials. Both mutants lack the 134.5 genes shown earlier to be essential for viral replication in the central nervous system in experimental animal systems (15, 21). One mutant, G207, lacked in addition the UL39 gene encoding the large subunit of ribonucleotide reductase (15, 16). While both mutants exhibited a satisfactory safety profile within the range of concentrations injected into the tumor mass, neither mutant exhibited unambiguous therapeutic potential as tested. The safety of the 134.5 null mutant hinges on the fact that it is unable to block the shutoff of protein synthesis resulting from the activation of protein kinase R (3, 12). In consequence, viral replication is reduced and the mutant is unable to spread from cell to cell. One potential counterindication for its use is the emergence of second-site mutations that block the activation of protein kinase R and show enhanced capacity to replicate in the central nervous system (2, 11, 18). The introduction of a second deletion reduced the risk of second-site, complementary mutation that partially restored virulence. However, the double mutant is further debilitated and replicates only in dividing cells. In malignant gliomas only a fraction of the tumor cells actively di