Human RNase H and compositions and uses thereof

Abstract

The present invention provides polynucleotides and polypeptides encoded thereby of human Type 2 RNase H. Methods of using these polynucleotides and polypeptides in enhancing antisense oligonucleotide therapies are also provided.

What is claimed is:  
     
         1 . An isolated human RNase polypeptide comprising human Type 2 RNase H.  
     
     
         2 . The isolated human RNase polypeptide of  claim 1  wherein the polypeptide comprises SEQ ID NO: 1.  
     
     
         3 . An isolated human RNase polypeptide prepared from a culture of ATCC Deposit No. 98536.  
     
     
         4 . A cloned and expressed human RNase H polypeptide.  
     
     
         5 . The cloned and expressed human RNase H polypeptide of  claim 4  which is a human Type 2 RNase H polypeptide.  
     
     
         6 . The cloned and expressed human RNase H polypeptide of  claim 4  which is a human RNase H1 polypeptide.  
     
     
         7 . The cloned and expressed human RNase H polypeptide of  claim 4  which comprises SEQ ID NO: 1.  
     
     
         8 . The cloned and expressed human RNase H polypeptide of  claim 4  which is prepared from a culture of ATCC Deposit No. 98536.  
     
     
         9 . A composition comprising a cloned and expressed human RNase H polypeptide and a pharmaceutically acceptable carrier.  
     
     
         10 . The composition of  claim 9  wherein the human RNase H polypeptide is a human Type 2 RNase H polypeptide.  
     
     
         11 . The composition of  claim 9  wherein the human RNase H polypeptide is a human RNase H1 polypeptide.  
     
     
         12 . A composition comprising a human RNase H polypeptide and a pharmaceutically acceptable carrier.  
     
     
         13 . The composition of  claim 12  further comprising an antisense oligonucleotide, wherein the human RNase H polypeptide is a human Type 2 polypeptide.  
     
     
         14 . An isolated polynucleotide encoding a human RNase H polypeptide.  
     
     
         15 . The isolated polynucleotide of  claim 14  which is a human Type 2 RNase H.  
     
     
         16 . A vector comprising a nucleic acid encoding a human RNase H polypeptide.  
     
     
         17 . A host cell comprising the vector of  claim 16 .  
     
     
         18 . A composition comprising a vector comprising a nucleic acid encoding a human RNase H polypeptide and a pharmaceutically acceptable carrier.  
     
     
         19 . The composition of  claim 18  further comprising an antisense oligonucleotide, wherein the human RNase H polypeptide is a human Type 2 RNase H polypeptide.  
     
     
         20 . An antibody targeted to a human Type 2 RNase H polypeptide.  
     
     
         21 . A nucleic acid probe capable of hybridizing to a portion of a nucleic acid encoding a human Type 2 RNase H polypeptide.  
     
     
         22 . A human Type 2 RNase H--his-tag fusion polypeptide.  
     
     
         23 . An antisense oligonucleotide capable of eliciting cleavage of its complementary target RNA by a human Type 2 RNase H polypeptide wherein said human Type 2 RNase H polypeptide comprises SEQ ID NO: 1.  
     
     
         24 . A method of enhancing inhibition of expression of a selected protein by an antisense oligonucleotide targeted to an RNA encoding the selected protein comprising: 
 (a) providing an antisense oligonucleotide targeted to an RNA encoding a selected protein whose expression is to be inhibited;    (b) allowing said oligonucleotide and said RNA to hybridize to form an oligonucleotide-RNA duplex;    (c) contacting said oligonucleotide-RNA duplex with a human Type 2 RNase H polypeptide, under conditions in which cleavage of the RNA strand of the oligonucleotide-RNA duplex occurs,    whereby inhibition of expression of the selected protein is enhanced.    
     
     
         25 . The method of  claim 24  wherein the human Type 2 RNase H polypeptide comprises SEQ ID NO: 1.  
     
     
         26 . The method of  claim 25  wherein the antisense oligonucleotide is a chimeric oligonucleotide.  
     
     
         27 . A method of screening oligonucleotides to identify an effective antisense oligonucleotide for inhibition of expression of a selected target protein comprising: 
 (a) contacting a human Type 2 RNase H polypeptide with an RNA encoding the selected target protein and an oligonucleotide complementary to at least a portion of the RNA under conditions in which an oligonucleotide-RNA duplex is formed;    (b) detecting cleavage of the RNA of the oligonucleotide-RNA duplex wherein cleavage is indicative of antisense efficacy.    
     
     
         28 . The method of  claim 27  wherein the human Type 2 RNase H polypeptide is enriched or overexpressed.  
     
     
         29 . The method of  claim 27  wherein the human Type 2 RNase H polypeptide is exogenously added.  
     
     
         30 . The method of  claim 27  wherein the human Type 2 RNase H polypeptide is an isolated, purified human Type 2 RNase H polypeptide.  
     
     
         31 . An effective antisense oligonucleotide identified in accordance with the method of  claim 27 .  
     
     
         32 . The method of  claim 27  further comprising determining the site on the RNA at which cleavage occurs, whereby said site is identified as a Type 2 RNase H-sensitive site.  
     
     
         33 . The method of  claim 32  further comprising identifying an effective antisense oligonucleotide which hybridizes to said Type 2 RNase H-sensitive site.  
     
     
         34 . The method of  claim 27  wherein the oligonucleotide is one of a mixture or library of oligonucleotides.  
     
     
         35 . An effective antisense oligonucleotide identified in accordance with the method of  claim 33 .  
     
     
         36 . A method of making an antisense oligonucleotide which elicits cleavage of its complementary target RNA by a human Type 2 RNase H polypeptide comprising synthesizing an oligonucleotide which is targeted to a selected RNA wherein said oligonucleotide, 
 when hybridized to the selected RNA target to form a duplex, will bind the human Type 2 RNase H polypeptide which thereby cleaves the RNA strand of the duplex.    
     
     
         37 . A method of prognosticating efficacy of antisense therapy of a selected disease comprising measuring the level or activity of a human Type 2 RNase H in a target cell of the antisense therapy.  
     
     
         38 . A method of identifying agents which increase or decrease activity of levels of a human RNase H polypeptide in a host cell comprising: 
 (a) contacting a cell expressing a human RNase H polypeptide with an agent suspected or increasing or decreasing activity or levels of the human RNase H polypeptide; and    (b) measuring the activity or levels of the human RNase H polypeptide in the presence and absence of the agent so that an increase or decrease in the activity or levels of the human RNase H polypeptide can be determined.    
     
     
         39 . A method of identifying agents which increase or decrease activity or levels of an RNase H polypeptide comprising: 
 a) contacting an RNase H polypeptide with an agent suspected of increasing or decreasing activity or levels of said RNase H polypeptide.    b) measuring the activity or levels of the RNase H polypeptide in the presence and absence of the agent so that an increase or decrease in the activity or levels of the human RNAase H polypeptide can be determined.    
     
     
         40 . The method of  claim 39  wherein the RNase H polypeptide is a cloned and expressed RNase H polypeptide.  
     
     
         41 . The method of  claim 39  wherein the RNase H polypeptide is a human RNase H polypeptide.  
     
     
         42 . The method of  claim 39  wherein the RNase H polypeptide is a human RNase H polypeptide having SEQ ID NO: 1.  
     
     
         43 . The method of  claim 39  wherein the RNase H polypeptide is prepared from a culture of ATCC Deposit No. 98536.  
     
     
         44 . A method of making substantially pure human Type 2 RNase H comprising transfecting a host cell with a vector containing a nucleic acid sequence encoding human Type 2 RNase H, wherein said host cells express the human Type 2 RNase H polypeptide, and isolating the human Type 2 RNase H polypeptide.  
     
     
         45 . The method of  claim 44  wherein said human Type 2 RNase polypeptide comprises SEQ ID NO: 1.