Method for delivering genes
Methods of delivering transgenes to target cells using plasmids comprising viral inverted terminal repeat (ITR) sequences are described. Such plasmids are capable of directing sustained transgene expression in target cells in rats provided that at least one adeno-associated virus (AAV) ITR sequence is present in the plasmid, regardless of whether that ITR is located upstream or downstream of the transgene. In a particular embodiment, plasmids comprising one or more AAV ITR sequence and an IL-10 transgene are shown to be effective in sustained reversal of pain in an animal model of neuropathic pain.
1. A method of treating neuropathic pain in a vertebrate subject comprising intrathecally administering to the nervous system of said subject a plasmid DNA encoding at least one adeno-associated virus (AAV) inverted terminal repeat (ITR) and further wherein the DNA encodes IL-10, operably linked to expression control elements, under conditions that result in expression of said IL-10 in vivo to reduce pain, wherein the plasmid DNA is encapsulated by a non-liposomal particulate carrier, and further wherein a single intrathecal administration of said encapsulated plasmid DNA to the vertebrate subject is capable of providing expression of IL-10 in vivo for at least 10 days.
2. The method of claim 1 wherein the plasmid DNA molecule comprises a single AAV ITR.
3. The method of claim 1 , wherein the particulate carrier is a poly(lactide-co-glycolide) carrier.
4. The method of claim 1 , wherein the particulate carrier is a polymethyl methacrylate carrier, or a poly(lactide) carrier.
5. The method of claim 1 , wherein a single intrathecal administration of said encapsulated plasmid DNA to the vertebrate subject is capable of providing expression of IL-10 in vivo for at least 30 days.
6. A method of treating neuropathic pain in a vertebrate subject comprising intrathecally administering to the nervous system of said subject plasmid DNA encoding (1) two adeno-associated virus (AAV) inverted terminal repeats (ITRs) and (2) IL-10, operably linked to expression control elements, under conditions that result in expression of said IL-10 in vivo to reduce pain, wherein the plasmid DNA is encapsulated by a non-liposomal particulate carrier, and further wherein a single intrathecal administration of said encapsulated plasmid DNA to the vertebrate subject is capable of providing expression of IL-10 in vivo for at least 10 days.
7. The method of claim 6 , wherein the particulate carrier is a poly(lactide-co-glycolide) carrier.
8. The method of claim 6 , wherein the particulate carrier is a polymethyl methacrylate polymer, or a poly(lactide) carrier.
9. The method of claim 6 , wherein a single intrathecal administration of said encapsulated plasmid DNA to the vertebrate subject is capable of providing expression of IL-10 in vivo for at least 30 days.