| Methods and compositions for treating neuropathic pain -> Monitor Keywords |
|
|
  Methods and compositions for treating neuropathic painUSPTO Application #: 20060040880Title: Methods and compositions for treating neuropathic pain Abstract: Compositions and methods for the treatment of neuropathic pain are provided. Compositions of the invention may comprise proteins with a zinc-finger domain fused to a regulatory domain that is capable of either activating or repressing the expression of a target gene involved in neuropathic pain. Alternatively, compositions of the invention may comprise a nucleic acid sequence encoding a protein of the invention, which nucleic acid sequence may optionally be provided as a plasmid or within a virus or other vector for delivery to a target cell or tissue. Methods of treating neuropathic pain involving treatment of subject with the compositions of the invention are also provided. Exemplary target genes for the treatment of neuropathic pain include VR1, NaV1.8, and TrkA. (end of abstract) Agent: Stoel Rives LLP One Utah Center - Salt Lake City, UT, US Inventors: John R.M. Forsayeth, Raymond A. Chavez, Trevor N. Collingwood, Andrew R. McNamara, Yann Jouvenot USPTO Applicaton #: 20060040880 - Class: 514044000 (USPTO) Related Patent Categories: Drug, Bio-affecting And Body Treating Compositions, Designated Organic Active Ingredient Containing (doai), O-glycoside, , Nitrogen Containing Hetero Ring, Polynucleotide (e.g., Rna, Dna, Etc.) The Patent Description & Claims data below is from USPTO Patent Application 20060040880. Brief Patent Description - Full Patent Description - Patent Application Claims
CROSS REFERENCE TO RELATED APPLICATIONS [0001] This application claims priority to U.S. Provisional Patent Application Ser. No. 60/560,535, filed Apr. 8, 2004, and U.S. Provisional Patent Application Ser. No. 60/567,757, filed Jun. 2, 2004, the disclosures of which are herby incorporated by reference in their entireties. BACKGROUND [0002] Neuropathic pain, also referred to as a chronic pain, is a complex disorder resulting from injury to the nerve, spinal cord or brain. There is evidence that nerve fibers in subjects with neuropathic pain develop abnormal excitability, particularly hyper-excitability, Zimmerman (2001) Eur J Pharmacol 429 (1-3):23-37. Although the American Pain Society estimates that nearly 50 million Americans are totally or partially disabled by pain, there are currently very few effective, well-tolerated treatments available. Wetzel et al. (1997) Ann Pharmacother 31 (9):1082-3). Indeed, existing therapeutics cause a range of undesirable side effects primarily due to the difficulty in developing small-molecule drugs capable of specifically targeting the receptor/channel of choice. [0003] Studies have shown the existence of primary sensory neurons that can be excited by noxious heat, mechanical damage, intense pressure or irritant chemicals, but not by innocuous stimuli such as warmth or light touch. These nociceptors selectively detect pain-inducing stimuli and appear to be distinct from other sensory mechanisms. This suggests that by suppressing the molecular mechanism of nociception it might be possible to limit the perception of painful stimuli without compromising general sensory awareness. [0004] Transduction of noxious stimuli in nociception is mediated by cellular receptors that typically include non-selective ion channels (e.g., vanilloid receptor, VR1), sodium ion channels (e.g., PN3/NaV.sub.1.8), tyrosine receptor kinases (e.g., TrkA), and GPCRs (e.g., bradykinin receptors). The majority of these receptors are expressed only in neuronal cells that are involved in both chronic and acute nociception, making them possible targets for therapeutic intervention aimed at limiting the pain response. Conventional therapeutic approaches typically focus on attempting to identify ligands that function as antagonists for these receptors. However, a major barrier to this approach is the cross-reactivity of receptor antagonists with other receptors of similar structure that are distinct from the pain-related targets. [0005] The study of the molecular mechanisms triggering neuropathic pain has identified several genes that are abnormally expressed in sensory neurons of the Dorsal Root Ganglion (DRG) in models of neuropathic pain, including Vanilloid Receptor 1 (VR1), a non-selective cationic channel responding to thermal, pH and capsacin stimulation (Hudson et al. (2001) Eur J Neurosci 13 (11):2105-2114; Walker et al. (2003) J. Pharmacol. Exp Ther 304 (1):56-62; Tyrosine kinase A receptor or high-affinity NGF receptor (TRKA), which has been shown to be upregulated in DRG neurons after chronic spinal cord injury (Qiao et al. (2002) J. Comp Neurol. 449 (3):217-230); (iii) the sodium channel Nav1.8 (also referred to as PN3 or SCN10A) (Coward et al. (2000) Pain 85 (1-2):41-50); and nitric oxide synthase (NOS) (Zimmerman, supra). Lai et al. (2002) Pain 95 (1-2):143-152, showed that reduced levels of Nav1.8 correlate with inhibition of neuropathic pain in the rat spinal nerve injury model of chronic pain. [0006] However, the modulation of genes aberrantly expressed in neuropathic pain has not been previously described. Furthermore, the ability to alter expression of these genes may have utility in treating and/or preventing many forms of pain. SUMMARY [0007] A variety of zinc finger proteins (ZFPs) and methods utilizing such proteins are provided for use in treating neuropathic pain. ZFPs that bind to a target site in genes that are aberrantly expressed in subjects having neuropathic pain are described. In addition, ZFPs that bind to a target site in genes expressed at normal levels in subjects experiencing neuropathic pain, modulation of whose expression results in decreased pain perception, are also provided. For example, using the methods and compositions described herein, genes that are over-expressed in the dorsal root ganglia (DRG) of pain patients (e.g., VR1, TRKA and/or Nav1.8) can be repressed, while genes that are under-expressed in the same populations can be activated. [0008] The ZFPs can be fused to a regulatory domain as part of a fusion protein. By selecting either an activation domain or a repression domain for fusion with the ZFP, one can either activate or repress gene expression. Thus, by appropriate choice of the regulatory domain fused to the ZFP, one can selectively modulate the expression of a target gene and hence various physiological processes correlated with neuropathic pain. [0009] By engineering ZFPs that bind to (and modulate expression of) genes encoding molecular targets involved in neuropathic pain to varying degrees, the extent to which a physiological process (e.g., pain) is modulated can be varied, thereby enabling treatment to be tailored. This can be achieved because multiple target sites (e.g., 9, 12 or 18 base pair target sites) in any given gene can be acted upon by the ZFPs provided herein. Thus, in some methods, a plurality of ZFPs (or fusions comprising these ZFPs) is administered. These ZFPs can then bind to different target sites located within a single target gene (e.g., VR1, TRKA, Nav1.8, etc.). Alternatively, the ZFPs can bind to target sites in different genes (e.g., two or more of VR1, TRKA, NAV1.8, etc.). Such ZFPs can in some instances have a synergistic effect. In certain methods, the plurality of fusion proteins includes different regulatory domains. [0010] Also provided herein are polynucleotides and nucleic acids that encode the ZFPs disclosed herein. Additionally, pharmaceutical compositions containing the nucleic acids and/or ZFPs are also provided. For example, certain compositions include a nucleic acid that encodes one of the ZFPs described herein operably linked to a regulatory sequence, combined with a pharmaceutically acceptable carrier or diluent, wherein the regulatory sequence allows for expression of the nucleic acid in a cell. Protein-based compositions include a ZFP as disclosed herein and a pharmaceutically acceptable carrier or diluent. [0011] These and other embodiments will readily occur to those of ordinary skill in the art in view of the disclosure herein. BRIEF DESCRIPTION OF THE DRAWINGS [0012] FIG. 1 is a graph depicting repression of VR1 gene expression in rat cells transfected with a plasmid encoding a fusion of a KOX repression domain and a VR1-targeted ZFP (designated 6332, 6337, 6338). The fusion proteins are designated 6332-KOX, 6337-KOX, and 6338-KOX. "NTC" refers to a non-transfected control. [0013] FIG. 2 is a graph depicting repression of VR1 gene expression in rat cells transfected with a plasmid encoding a fusion of a KOX repression domain and a VR1-targeted ZFP (designated 6144, 6149, 6150). The fusion proteins are designated 6144-KOX, 6149-KOX, and 6150-KOX. "eGFP" refers to an enhanced Green Fluorescent Protein (GFP) control. [0014] FIG. 3 is a graph depicting results of FACS and shows repression of VR1 protein levels in rat cells transfected with a plasmid encoding 6144-KOX, 6149-KOX, 6150-KOX a fusion of a KOX repression domain and a VR1-targeted ZFP (designated 6144, 6149, 6150). The fusion proteins are designated 6144-KOX, 6149-KOX, 6150-KOX, 6332-KOX, 6337-KOX, and 6338-KOX. "GFP" refers to FACS results obtained with a GFP control. [0015] FIG. 4 is a graph depicting repression of TrkA gene expression by in rat cells transfected with a plasmid encoding a fusion of a KOX repression domain and a TrkA-targeted ZFP (designated 6182, 6297) and a plasmid encoding puromycin resistance. Puromycin selection is used to kill untransfected cells. The fusion proteins are designated 6182-KOX and 6297-KOX. "Puromycin cntrl" refers to controls co-transfected with a control plasmid and the plasmid encoding puromycin resistance. [0016] FIG. 5 is a graph depicting results of FACS and showing repression of TrkA protein levels in rat cells co-transfected with a plasmid encoding 6182-KOX or 6297-KOX and a plasmid encoding puromycin resistance. "Puromycin cntrl" refers to controls co-transfected with a control plasmid and the plasmid encoding puromycin resistance. [0017] FIG. 6 is a graph depicting repression of NAV1.8 in human cells transfected with a plasmid encoding a fusion of a KOX repression domain and a NAV1.8-targeted ZFP (designated 6584, 6585, 6586, 6587, 6590, 6591, 6621, and 6622). The fusion proteins are designated 6584-KOX, 6585-KOX, 6586-KOX, 6587-KOX, 6590-KOX, 6591-KOX, 6621-KOX, and 6622-KOX. "eGFP" refers to an enhanced Green Fluorescent Protein (GFP) control. [0018] FIG. 7 is a graph showing levels of human TrkA mRNA, normalized to human GAPDH mRNA, in K562 cells transfected with plasmids encoding ZFP/KOX fusion proteins. The identity of the encoded protein is shown on the abscissa: EF-1a refers to the promoter controlling expression of the fusion protein; Kox refers to the presence of a KOX repression domain in the encoded protein, and the number refers to the particular TrkA-targeted zinc finger binding domain (see Tables 1 and 5 for DNA target sequences and recognition domain amino acid sequences, respectively, for these zinc finger domains). EF-laGFPKox and pBluescript are control plasmids: EF-laGFPKox lacks an engineered zinc finger binding domain; pBluescript is a vector lacking sequences encoding a fusion protein. Bars show the standard error of the mean for duplicate determinations. [0019] FIG. 8 is an autoradiographic image of a protein blot in which lysates from cells transfected with plasmids encoding TrkA-targeted ZFP/KOX fusion proteins were analyzed. The top panel shows assays for the presence of TrkA and TFIIB. The lower panel shows assays for the presence of the zinc finger/Kox fusion proteins, using a primary mouse anti-FLAG M2 monoclonal antibody and a donkey anti-mouse IgG-horseradish peroxidase secondary antibody. Abbreviations and protein identifications are the same as in FIG. 7. DETAILED DESCRIPTION Continue reading... Full patent description for Methods and compositions for treating neuropathic pain Brief Patent Description - Full Patent Description - Patent Application Claims Click on the above for other options relating to this Methods and compositions for treating neuropathic pain patent application. ###  How KEYWORD MONITOR works... a FREE service from FreshPatents1. Sign up (takes 30 seconds). 2. Fill in the keywords to be monitored. 3. Each week you receive an email with patent applications related to your keywords. Start now! - Receive info on patent apps like Methods and compositions for treating neuropathic pain or other areas of interest. ### Previous Patent Application: Compostions and methods for enhancing delivery of nucleic acids into cells and for modifying expression of target genes in cells Next Patent Application: Methods for treating cancer using anti-wnt2 monoclonal antibodies and sirna Industry Class: Drug, bio-affecting and body treating compositions ### FreshPatents.com Support Thank you for viewing the Methods and compositions for treating neuropathic pain patent info. IP-related news and info Results in 3.89656 seconds Other interesting Feshpatents.com categories: Accenture , Agouron Pharmaceuticals , Amgen , AT&T , Bausch & Lomb , Callaway Golf |
||
