Compositions and methods for inducing an enhanced immune response using poxvirus vectors

Abstract

Provided herein are recombinant poxviruses comprising heterologous or native nucleic acids specifying excess double-stranded RNA (dsRNA) early in infection, which may further comprise heterologous nucleic acids encoding one or more costimulatory molecules, and/or heterologous nucleic acids encoding one or more infectious disease-associated antigens or tumor-associated antigens, as well as pharmaceutical compositions comprising such recombinant poxviruses and methods and uses thereof. The recombinant poxviruses provided herein enhance innate and adaptive immune activation in subjects compared to identical recombinant poxviruses lacking heterologous or native transcription units specifying excess early dsRNA.

1 .- 31 . (canceled) 
     
     
         32 . A recombinant poxvirus for inducing an enhanced innate immune response, the recombinant poxvirus comprising heterologous nucleic acids generating excess double stranded RNA (dsRNA) early in infection, wherein the heterologous nucleic acids comprise sequences that encode partially or completely complementary RNA transcripts that anneal after transcription to form dsRNA, wherein the partially or completely complementary RNA transcripts overlap by 100 or more nucleotides, and wherein the dsRNA enhances an innate immune response as compared to a recombinant poxvirus comprising heterologous nucleic acids that do not express excess dsRNA early in infection. 
     
     
         33 . The recombinant poxvirus of  claim 32 , wherein the partially or completely complementary RNA transcripts overlap from 100 nucleotides to 1000 nucleotides. 
     
     
         34 . The recombinant poxvirus of  claim 32 , wherein the partially or completely complementary RNA transcripts overlap from 300 nucleotides to 1000 nucleotides. 
     
     
         35 . The recombinant poxvirus of  claim 32 , wherein the partially or completely complementary RNA transcripts overlap by 700 or more nucleotides. 
     
     
         36 . The recombinant poxvirus of  claim 32 , wherein a sense messenger RNA (mRNA) is transcribed from one of the heterologous nucleic acids and an anti-sense mRNA is transcribed from the other heterologous nucleic acid. 
     
     
         37 . The recombinant poxvirus of  claim 36 , wherein the sense mRNA and the antisense mRNAs are transcribed from both strands of a single heterologous sequence insert. 
     
     
         38 . The recombinant poxvirus of  claim 36 , wherein the sense mRNA and the antisense mRNAs are transcribed from both strands of heterologous nucleic acids sequences inserted on different parts of the poxviral genome. 
     
     
         39 . The recombinant poxvirus of  claim 32 , wherein the heterologous nucleic acids have at least 95% sequence identity within the region that encodes the partially or completely complementary RNA transcripts. 
     
     
         40 . The recombinant poxvirus of  claim 36 , wherein the sense mRNA and the antisense mRNA are transcribed from both strands of a native poxvirus sequence. 
     
     
         41 . The recombinant poxvirus of  claim 40 , wherein the native poxvirus sequence is selected from an early gene and an immediate-early gene. 
     
     
         42 . The recombinant poxvirus of  claim 32 , wherein expression of the partially or completely complementary RNA transcripts is each directed by a poxviral promoter. 
     
     
         43 . The recombinant poxvirus of  claim 42 , wherein the poxviral promoter is selected from an early promoter and an immediate-early promoter. 
     
     
         44 . A method for enhancing an innate immune response, the method comprising administering a recombinant poxvirus to a subject, wherein the recombinant poxvirus comprises heterologous nucleic acids generating excess double stranded RNA (dsRNA) early in infection in the subject, wherein the heterologous nucleic acids comprise sequences that encode partially or completely complementary RNA transcripts that anneal after transcription to form dsRNA, wherein the partially or completely complementary RNA transcripts overlap by 100 or more nucleotides, and wherein the dsRNA enhances the innate immune response in the subject as compared to a recombinant poxvirus comprising heterologous nucleic acids that do not express excess dsRNA early in infection. 
     
     
         45 . The method of  claim 44 , wherein the administration of the recombinant poxvirus enhances production of type 1 interferons (type 1 IFNs) in the subject, as compared to the production of type 1 interferons (type I IFNs) from administration of a recombinant poxvirus that do not express excess dsRNA early in infection. 
     
     
         46 . The method of  claim 44 , wherein the administration of the recombinant poxvirus enhances production of type 1 interferons (type 1 IFNs), cytokines, and chemokines in the subject, as compared to the production of type 1 interferons (type 1 IFNs), cytokines, and chemokines from administration of a recombinant poxvirus comprising heterologous nucleic acids that do not express excess dsRNA early in infection. 
     
     
         47 . The method of  claim 45 , wherein production of type I IFNs comprises transcription of interferon-beta (IFN-β) -encoding messenger RNA (mRNA), and wherein transcription of IFN-β mRNA increases by at least two-fold. 
     
     
         48 . The method of  claim 46 , wherein transcription of IFN-β mRNA increases by at least ten-fold. 
     
     
         49 . The method of  claim 47 , wherein production of type I IFNs comprises secretion of IFN-β protein, and wherein secretion of IFN-β protein increases by at least two-fold. 
     
     
         50 . The method of  claim 47 , wherein secretion of IFN-β protein increases by at least ten-fold. 
     
     
         51 . The method of  claim 46 , wherein administration enhances production of the cytokines IFN-α, IFN-γ, IL-6, and IL-18. 
     
     
         52 . The method of  claim 46 , wherein administration enhances production of the chemokines CXCL1, CCL2 and CCL5. 
     
     
         53 . The method of  claim 44 , wherein a sense messenger RNA (mRNA) is transcribed from one of the heterologous nucleic acids and an anti-sense mRNA is transcribed from the other heterologous nucleic acid. 
     
     
         54 . The method of  claim 53 , wherein the sense mRNA and the antisense mRNAs are transcribed from both strands of a single heterologous sequence insert. 
     
     
         55 . The method of  claim 53 , wherein the sense mRNA and the antisense mRNAs are transcribed from both strands of heterologous nucleic acids sequences inserted on different parts of the poxviral genome. 
     
     
         56 . A recombinant poxvirus for inducing an innate enhanced immune response, the recombinant poxvirus comprising a first heterologous nucleic acid and a second heterologous nucleic acid, wherein the first heterologous nucleic acid comprises sequences that encode a first RNA transcript partially or completely complementary to a second RNA transcript encoded by the second heterologous nucleic acid, wherein the first and second RNA transcripts anneal after transcription to form double stranded RNA (dsRNA) early in infection, wherein the first and second RNA transcripts overlap by 100 or more nucleotides, and wherein the dsRNA enhances an immune response as compared to a recombinant poxvirus comprising heterologous nucleic acids that do not express excess dsRNA early in infection. 
     
     
         57 . The recombinant poxvirus of  claim 56 , wherein a sense messenger RNA (mRNA) is transcribed from first heterologous nucleic acid and an anti-sense mRNA is transcribed from the second heterologous nucleic acid. 
     
     
         58 . The recombinant poxvirus of  claim 57 , wherein the sense mRNA and the antisense mRNAs are transcribed from both strands of a single heterologous sequence insert. 
     
     
         59 . The recombinant poxvirus of  claim 57 , wherein the sense mRNA and the antisense mRNAs are transcribed from both strands of heterologous nucleic acids sequences inserted on different parts of the poxviral genome.