Novel double-stranded ribonucleic acids with rugged physico-chemical structure and highly specific biologic activity

Abstract

A novel form of Rugged dsRNA with a unique composition and physical characteristics was identified with high specificity of binding to TLR3, which conveys an important range of therapeutic opportunities. Unlike the previous known antiviral Ampligen® (poly I, poly C12,U) the new and improved form (poly I, poly C 30 ,U) has a reduced tendency to form branched dsRNA which results in increased bioactivity due to an increased ability to bind TLR3 receptor. Pharmaceutical formulations containing the new nucleic acid as active ingredients and methods of treatment are also provided. The invention also provides a description of the physicochemical properties of this novel form of Rugged dsRNA and a method for its preparation in substantially pure form. DsRNAs acting thru TLR3 receptor activation are potent antiviral compounds as well as anticancer agents; also through secondary immunomodulation they can enhance the bioactivity of vaccines and also treat autoimmune disorders.

What is claimed is: 
     
         1 . An isolated double-stranded ribonucleic acid (dsRNA) which is resistant to denaturation under conditions that are able to separate hybridized poly(riboinosinic acid) and poly(ribocytosinic acid) strands. 
     
     
         2 . The isolated dsRNA of  claim 1 , wherein both strands of said isolated dsRNA comprise one or more uracil or guanine bases that are not based paired to an opposite strand. 
     
     
         3 . An isolated double-stranded ribonucleic acid (dsRNA) which is resistant to denaturation under conditions that are able to separate hybridized poly(riboinosinic acid) and poly(ribocytidylic acid) strands, wherein the isolated dsRNA:
 has an HPLC chromatogram substantially the same as the 5 minute peak of  FIG. 1 ;   is stable to exposure to thermal stress at 40° C.; and   has an increased bioactivity as evidenced by binding to receptor TLR3-ECD as compared to unimproved poly(I):poly(C 12 U).   
     
     
         4 . An isolated double-stranded ribonucleic acid (dsRNA) which is resistant to enzymatic degradation under conditions that are able to degrade poly(riboinosinic acid) and poly(ribocytidylic acid) strands, wherein the isolated dsRNA:
 has an HPLC chromatogram substantially the same as the 5 minute peak of  FIG. 1 ;   has increased stability to exposure to pancreatic ribonuclease A; and   has an increased bioactivity as evidenced by binding to receptor TLR3-ECD as compared to unselected poly(I):poly(C 12 U).   
     
     
         5 . A method of treating a subject with an immunological dysfunction, said method comprising administration to the subject of the isolated dsRNA defined in  claim 1  in a therapeutic amount. 
     
     
         6 . The method according to  claim 5  wherein said immunological dysfunction is an autoimmune disorder. 
     
     
         7 . A method of treating a subject with an incipient or established microbial infection, said method comprising administration to the subject of the isolated dsRNA defined in  claim 1  in a therapeutic amount. 
     
     
         8 . The method according to  claim 7  wherein said infection is a bacterial, protozoan, or viral infection. 
     
     
         9 . A method of treating a subject with chronic fatigue syndrome, said method comprising administration to the subject of the isolated dsRNA defined in  claim 1  in a therapeutic amount. 
     
     
         10 . A method of treating or preventing tumor or neoplasm formation in a subject, said method comprising administration to the subject of the isolated dsRNA defined in  claim 1  in a therapeutic amount. 
     
     
         11 . The method according to  claim 10  wherein said tumor or neoplasm is a carcinoma, sarcoma, leukemia or lymphoma. 
     
     
         12 . The method according to  claim 10  wherein the tumor or neoplasm is a glioma. 
     
     
         13 . A method of inducing an immune enhancing effect in a subject, said method comprising administration to the subject of the isolated dsRNA defined in  claim 1  in a therapeutic amount. 
     
     
         14 . The method according to  claim 13  wherein said isolated dsRNA is administered as a vaccine adjuvant. 
     
     
         15 . The method according to  claim 14  wherein said isolated dsRNA is administered sequentially or concurrently with the vaccine. 
     
     
         16 . The method according to  claim 13  wherein said subject is administered an anti-tumor or anti-microbial vaccine. 
     
     
         17 . The method according to  claim 16  wherein the vaccine is an anti-protozoan, anti-viral or anti-bacterial vaccine. 
     
     
         18 . The method according to  claim 5 , wherein the therapeutic amount of said isolated dsRNA is infused intravenously. 
     
     
         19 . The method according to  claim 5 , wherein the therapeutic amount is injected intradermally, subcutaneously, or intramuscularly; inhaled or delivered intranasally or intratracheally; or applied transdermally, transmucosally, intranasally, intratracheally, oropharyngeally, or sublingually. 
     
     
         20 . The method according to  claim 5 , wherein the therapeutic amount is administered transocularly.