Central Nervous System Delivery of Interleukin 4 by a Nonreplicative Herpes Simplex Type 1 Viral Vector Ameliorates Autoimmune Demyelination

Abstract

Multiple sclerosis (MS) is a T cell-mediated organ-specific inflammatory disease leading to central nervous system (CNS) demyelination. On the basis of results obtained in experimental autoimmune encephalomyelitis (EAE) models, MS treatment by administration of antiinflammatory cytokines such as interleukin 4 (IL-4) is promising but is hampered by the limited access of the cytokines to the CNS and by the pleiotropic effects of systemically administered cytokines. We established a cytokine delivery system within the CNS using non-replicative herpes simplex type 1 (HSV-1) viral vectors engineered with cytokine genes. These vectors injected into the cisterna magna (i.c.) of mice diffuse in all ventricular and subarachnoid spaces and infect with high efficiency the ependymal and leptomeningeal cell layers surrounding these areas, without obvious toxic effects. Heterologous genes contained in the vectors are efficiently transcribed in infected ependymal cells, leading to the production of high amounts of the coded proteins. For example, 4.5 ng of interferon gamma (IFN-gamma) per milliliter is secreted into the cerebrospinal fluid (CSF) up to day 28 postinjection (p.i.) and reaches the CNS parenchyma in bioactive form, as demonstrated by upregulation of MHC class I expression on CNS-resident cells. We then exploited the therapeutic potential of the vectors in EAE mice. An HSV-1-derived vector containing the IL-4 gene was injected i.c. in Biozzi AB/H mice at the time of EAE induction. We found the following in treated mice: (1) delayed EAE onset, (2) a significant decrease in clinical score, (3) a significant decrease in perivascular inflammatory infiltrates and in the number of macrophages infiltrating the CNS parenchyma and the submeningeal spaces, and (4) a reduction in demyelinated areas and axonal loss. Peripheral T cells from IL-4-treated mice were not affected either in their antigen-specific proliferative response or in cytokine secretion pattern. Our results indicate that CNS cytokine delivery with HSV-1 vectors is feasible and might represent an approach for the treatment of demyelinating diseases. Advantages of this approach over systemic cytokine administration are the high cytokine level reached in the CNS, the absence of effects on the peripheral immune system, and the long-lasting cytokine production in the CNS after a single vector administration.

Central nervous system gene therapy with interleukin-4 inhibits progression of ongoing relapsing–remitting autoimmune encephalomyelitis in Biozzi AB/H mice

“…delivery of the d120:LacZ:IL-4 vector does not induce blood leak of IL-4 and does not influence peripheral immune system functions such as antigen presentation, T cell activation and cytokine secretion. 11 However, 72 h after CNS delivery of the IL-4 gene we did not observe any significant reduction of the number of CNS-infiltrating mononuclear cells, and in particular, of macrophages that are considered the ultimate effector cells in EAE. 16,17 How can we explain this apparent paradox?…”

“…Within the CNS, vectors redirect the cell machinery of infected CNS-resident cells to produce discrete amounts of the bioactive form of the cytokine encoded by the transgene into the CSF of mice for almost 1 month after injection without causing any CNS and/or peripheral side-effect. 11 We now report that i.c. injection of an HSV-1-derived vector, containing the gene of the prototypic anti-inflammatory cytokine IL-4 (d120:LacZ:IL-4), in Biozzi AB/H mice with established EAE inhibits disease progression and protects mice from EAE-related mortality.…”

“…11,28 Briefly, the IL-4 gene-containing vector accommodates the Escherichia coli Lac-Z gene driven by the human CMV immediate-early promoter and the mouse IL-4 gene driven by the infected cell protein (ICP)4 promoter (d120:IL-4:LacZ vector); the control vector was identical to the d120:IL-4:LacZ but contained only the Lac-Z gene (d120:Lac-Z vector). Expression cassettes containing the heterologous genes (ie LacZ, IL-4) were inserted in both vectors into the thymidine kinase locus on an ICP4-defective HSV-1 virus (d120) that was propagated on a compGene Therapy lementing cell line (E5).…”

“…of mice, as previously described. 11 EAE induction and treatment EAE was induced by subcutaneous immunization into the flanks with 2 mg of spinal cord homogenate (SCH) in IFA (Difco, Detroit, MI, USA) supplemented with 4 mg/ml of Mycobacterium tuberculosis (strain H37Ra; Difco). Immunization was performed in each mouse twice, 7 days apart, as previously described.…”

“…10 We have recently developed a novel system to deliver cytokine genes into the CNS based on the use of HSV type-1-derived non-replicative vectors. 11 We showed that HSV-1 vectors containing cytokine genes, when injected within the CNS via the intracisternal (i.c.) route, diffuse consistently in all cerebrospinal fluid (CSF) spaces and are able to infect efficiently the layer of choroidal, ependymal, and leptomeningeal cells surrounding these spaces.…”