Compositions and Methods for Dengue Virus Chimeric Constructs in Vaccines

Abstract

Embodiments herein report compositions, uses and manufacturing of dengue virus constructs and live attenuated dengue viruses. Some embodiments concern a composition that includes, but is not limited to, a tetravalent dengue virus composition. In certain embodiments, compositions can include constructs of one or more serotypes of dengue virus, such as dengue-1 (DEN-1) virus, dengue-2 (DEN-2) virus, dengue-3 (DEN-3) or dengue-4 (DEN-4) virus constructs. In other embodiments, constructs disclosed herein can be combined in a composition to generate a vaccine against more one or more dengue virus constructs that may or may not be subsequently passaged in mammalian cells.

1 - 47 . (canceled) 
     
     
         48 . A polynucleotide molecule encoding a dengue-1/dengue-2 polypeptide chimera, the polynucleotide comprising a first nucleotide sequence encoding nonstructural proteins NS1, NS2A, NS2B, NS3, NS4A, NS4B and NS5 from a modified live, attenuated dengue-2 virus strain PDK-53 comprising at least one mutation to PDK-53, and a second nucleotide sequence encoding at least one structural protein from dengue-1 selected from capsid protein (C), premembrane/membrane protein (prM) and envelope protein (E), wherein the at least one mutation comprises one or more of:
 an adenine to a cytosine mutation at position 3823 encoding a leucine instead of an isoleucine in the chimera at amino acid position 1243 corresponding to NS2A-116;   an adenine to thymine mutation at position 4407 encoding an aspartic acid instead of a glutamic acid in the chimera at amino acid position 1437 corresponding to NS2B-92; and   an adenine to guanine mutation at position 7311.   
     
     
         49 . The polynucleotide molecule of  claim 48 , further comprising at least one additional mutation of:
 a cytosine to thymine mutation at position 7148 encoding an isoleucine instead of a threonine in the chimera at amino acid position 2351 corresponding to NS4B-108; and   a guanine to cytosine mutation at position 2384 encoding an alanine instead of glycine in the chimera at amino acid position 763 corresponding to E-483.   
     
     
         50 . The polynucleotide molecule of  claim 48  comprising the polynucleotide represented by SEQ ID NO: 3. 
     
     
         51 . The polynucleotide molecule of  claim 48  encoding a dengue-1/dengue-2 polypeptide chimera, wherein the polypeptide is represented by SEQ ID NO: 4. 
     
     
         52 . A polypeptide molecule encoded by a polynucleotide molecule according to  claim 48 . 
     
     
         53 . A dengue-1/dengue-2 chimera comprising one or more polypeptide molecules according to  claim 52 . 
     
     
         54 . A pharmaceutical composition comprising a polynucleotide molecule according to  claim 48 , a polypeptide molecule encoded by a polynucleotide molecule according to  claim 48  or a dengue-1/dengue-2 chimera comprising one or more polypeptide molecules encoded by a polynucleotide molecule according to  claim 48 , and a pharmaceutically acceptable excipient. 
     
     
         55 . A method for inducing an immune response in a subject comprising administering a pharmaceutically acceptable amount of the composition of  claim 54 , wherein the composition induces an immune response to dengue virus in the subject. 
     
     
         56 . A vector encoding a polynucleotide molecule of  claim 48 . 
     
     
         57 . A cell comprising the polynucleotide molecule according to  claim 48  or a vector encoding a polynucleotide molecule of  claim 48 . 
     
     
         58 . A polynucleotide molecule encoding a modified live, attenuated dengue-2 virus strain PDK-53 polypeptide molecule, wherein the polynucleotide comprises at least one mutation to PDK-53, wherein the at least one mutation comprises:
 an adenine to guanine mutation at position 592 encoding a glutamic acid instead of a lysine in the polypeptide molecule at amino acid position 166 corresponding to prM-52; and   an adenine to guanine mutation at position 8803 encoding a valine instead of an isoleucine in the polypeptide molecule at amino acid position 2903 corresponding to NS5-412.   
     
     
         59 . The polynucleotide molecule of  claim 58 , further comprising at least one additional mutation of:
 a guanine to cytosine mutation at nucleic acid position 6481 encoding a proline instead of an alanine in the polypeptide molecule at amino acid position 2129 corresponding to NS4A-36; and   a cytosine to thymine mutation at position 7156 encoding a phenylalanine instead of a leucine in the polypeptide molecule at amino acid position 2354 corresponding to NS4B-111.   
     
     
         60 . The polynucleotide molecule according to  claim 58  comprising the polynucleotide represented by SEQ ID NO: 11. 
     
     
         61 . The polynucleotide molecule according to  claim 58  encoding a modified, live attenuated dengue-2 virus strain PDK-53, wherein the polypeptide is represented by SEQ ID NO: 12. 
     
     
         62 . A polypeptide molecule encoded by a polynucleotide molecule according to  claim 58 . 
     
     
         63 . A modified live, attenuated dengue-2 virus strain PDK-53 comprising one or more polypeptide molecules according to  claim 62 . 
     
     
         64 . A pharmaceutical composition comprising a polynucleotide molecule according to  claim 58 , a polypeptide molecule encoded by a polynucleotide molecule according to  claim 58 , or a modified live, attenuated dengue-2 virus strain PDK-53 comprising one or more polypeptide molecules encoded by a polynucleotide molecule according to  claim 58 , and a pharmaceutically acceptable excipient. 
     
     
         65 . A method for inducing an immune response in a subject comprising administering a pharmaceutically acceptable amount of the composition of  claim 64 , wherein the composition induces an immune response to dengue virus in the subject. 
     
     
         66 . A vector encoding a polynucleotide molecule of  claim 58 . 
     
     
         67 . A cell comprising the polynucleotide molecule according to  claim 58  or a vector encoding a polynucleotide molecule of  claim 58 . 
     
     
         68 . A polynucleotide molecule encoding a dengue-3/dengue-2 polypeptide chimera, the polynucleotide comprising a first nucleotide sequence encoding nonstructural proteins NS1, NS2A, NS2B, NS3, NS4A, NS4B and NS5 from a modified live, attenuated dengue-2 virus strain PDK-53 comprising at least one mutation to PDK-53, and a second nucleotide sequence encoding at least one structural protein from dengue-3 selected from capsid protein (C), premembrane/membrane protein (prM) and envelope protein (E), wherein the at least one mutation comprises:
 an adenine to thymine mutation at position 1603 encoding a serine instead of a threonine in the chimera at amino acid position 503 corresponding to E-223; and   an adenine to guanine mutation at position 7620.   
     
     
         69 . The polynucleotide molecule of  claim 68  further comprising an adenine to thymine mutation at position 6436 encoding an asparagine instead of an aspartic acid in the chimera at amino acid position 1243 corresponding to N42A-23. 
     
     
         70 . The polynucleotide molecule according to  claim 68  comprising the polynucleotide represented by SEQ ID NO: 19. 
     
     
         71 . The polynucleotide molecule according to  claim 68  encoding a dengue-3/dengue-2 polypeptide chimera, wherein the polypeptide is represented by SEQ ID NO: 20. 
     
     
         72 . A polypeptide molecule encoded by a polynucleotide according to  claim 68 . 
     
     
         73 . A dengue-3/dengue-2 polypeptide chimera comprising one or more polypeptide molecules according to  claim 72 . 
     
     
         74 . A pharmaceutical composition comprising a polynucleotide molecule according to  claim 68 , a polypeptide molecule encoded by a polynucleotide according to  claim 68 , or a dengue-3/dengue-2 polypeptide chimera comprising one or more polypeptide molecules encoded by a polynucleotide according to  claim 68 , and a pharmaceutically acceptable excipient. 
     
     
         75 . A method for inducing an immune response in a subject comprising administering a pharmaceutically acceptable amount of the composition of  claim 74 , wherein the composition induces an immune response to dengue virus in the subject. 
     
     
         76 . A vector encoding a polynucleotide molecule of  claim 68 . 
     
     
         77 . A cell comprising the polynucleotide molecule according to  claim 68  or a vector encoding a polynucleotide molecule of  claim 68 . 
     
     
         78 . A polynucleotide molecule encoding a dengue-4/dengue-2 polypeptide chimera, the polynucleotide comprising a first nucleotide sequence encoding nonstructural proteins NS1, NS2A, NS2B, NS3, NS4A, NS4B and NS5 from a modified live, attenuated dengue-2 virus strain PDK-53 comprising at least one mutation to PDK-53, and a second nucleotide sequence encoding at least one structural protein from dengue-4 selected from capsid protein (C), premembrane/membrane protein (prM) and envelope protein (E), wherein the at least one mutation comprises:
 an adenine to thymine mutation at position 225;   an adenine to guanine mutation at position 3674 encoding a glycine instead of an aspartic acid in the chimera at amino acid position 1193 corresponding to NS2A-66;   a cytosine to thymine mutation at position 5391;   a cytosine to thymine mutation at position 6437 encoding a valine instead of an alanine in the chimera at amino acid position 2114 corresponding to NS4A-21,   a thymine to cytosine mutation at position 7026, and   an adenine to cytosine mutation at position 9750.   
     
     
         79 . The polynucleotide molecule according to  claim 78 , further comprising an adenine to guanine mutation at position 3773 encoding an arginine instead of a lysine in the chimera at amino acid position 1226 corresponding to NS2A-99. 
     
     
         80 . The polynucleotide molecule according to  claim 78 , further comprising a cytosine to thymine mutation at position 7538 encoding a phenylalanine instead of a serine in the chimera at amino acid position 2481 corresponding to NS4B-238. 
     
     
         81 . The polynucleotide molecule according to  claim 78  comprising the polynucleotide represented by SEQ ID NO: 27. 
     
     
         82 . The polynucleotide molecule according to  claim 78  encoding a dengue-4/dengue-2 polypeptide chimera, wherein the polypeptide is represented by SEQ ID NO: 28. 
     
     
         83 . A polypeptide molecule encoded by a polynucleotide according to  claim 78 . 
     
     
         84 . A dengue-4/dengue-2 polypeptide chimera comprising one or more polypeptide molecules according to  claim 83 . 
     
     
         85 . A pharmaceutical composition comprising a polynucleotide molecule according to  claim 78 , a polypeptide molecule encoded by a polynucleotide molecule according to  claim 78 , or a dengue-4/dengue-2 polypeptide chimera comprising one or more polypeptide molecules encoded by a polynucleotide molecule according to  claim 78 , and a pharmaceutically acceptable excipient. 
     
     
         86 . A method for inducing an immune response in a subject comprising administering a pharmaceutically acceptable amount of the pharmaceutical composition of  claim 85 , wherein the composition induces an immune response to dengue virus in the subject. 
     
     
         87 . A vector encoding a polynucleotide molecule of  claim 78 . 
     
     
         88 . A cell comprising the polynucleotide molecule according to  claim 78  or a vector encoding a polynucleotide molecule of  claim 78 . 
     
     
         89 . An immunogenic composition comprising at least one polynucleotide molecule according to  claim 48  and a pharmaceutically acceptable carrier. 
     
     
         90 . The composition of  claim 89  further comprising:
 a) a polynucleotide molecule encoding a modified live, attenuated dengue-2 virus strain PDK-53 polypeptide molecule, wherein the polynucleotide comprises at least one mutation to PDK-53, wherein the at least one mutation comprises: 
 an adenine to guanine mutation at position 592 encoding a glutamic acid instead of a lysine in the polypeptide molecule at amino acid position 166 corresponding to prM-52; and 
 an adenine to guanine mutation at position 8803 encoding a valine instead of an isoleucine in the polypeptide molecule at amino acid position 2903 corresponding to NS5-412; 
 b) a polynucleotide molecule encoding a dengue-3/dengue-2 polypeptide chimera, the polynucleotide comprising a first nucleotide sequence encoding nonstructural proteins NS1, NS2A, NS2B, NS3, NS4A, NS4B and NS5 from a modified live, attenuated dengue-2 virus strain PDK-53 comprising at least one mutation to PDK-53, and a second nucleotide sequence encoding at least one structural protein from dengue-3 selected from capsid protein (C), premembrane/membrane protein (prM) and envelope protein (E), wherein the at least one mutation comprises: 
 an adenine to thymine mutation at position 1603 encoding a serine instead of a threonine in the chimera at amino acid position 503 corresponding to E-223; and 
 an adenine to guanine mutation at position 7620; and 
 c) a polynucleotide molecule encoding a dengue-4/dengue-2 polypeptide chimera, the polynucleotide comprising a first nucleotide sequence encoding nonstructural proteins NS1, NS2A, NS2B, NS3, NS4A, NS4B and NS5 from a modified live, attenuated dengue-2 virus strain PDK-53 comprising at least one mutation to PDK-53, and a second nucleotide sequence encoding at least one structural protein from dengue-4 selected from capsid protein (C), premembrane/membrane protein (prM) and envelope protein (E), wherein the at least one mutation comprises: 
 an adenine to thymine mutation at position 225; 
 an adenine to guanine mutation at position 3674 encoding a glycine instead of an aspartic acid in the chimera at amino acid position 1193 corresponding to NS2A-66; 
 a cytosine to thymine mutation at position 5391; 
 a cytosine to thymine mutation at position 6437 encoding a valine instead of an alanine in the chimera at amino acid position 2114 corresponding to NS4A-21, 
 a thymine to cytosine mutation at position 7026, and 
 an adenine to cytosine mutation at position 9750. 
 
     
     
         91 . The composition of  claim 90 , further comprising an immunogenic composition for a flavivirus selected from the group consisting of yellow fever virus, tick-borne encephalitis virus, Japanese encephalitis virus, West Nile virus, hepatitis C virus, and a combination of two or more thereof. 
     
     
         92 . A kit comprising at least one polynucleotide molecule according to  claim 48 ; or at least one composition comprising a polynucleotide molecule according to  claim 48 , a polypeptide molecule encoded by a polynucleotide molecule according to  claim 48  or a dengue-1/dengue-2 chimera comprising one or more polypeptide molecules encoded by a polynucleotide molecule according to  claim 48 , and a pharmaceutically acceptable excipient; and a container. 
     
     
         93 . A composition comprising at least one dengue-1/dengue-2 chimera of  claim 53  and a pharmaceutically acceptable carrier. 
     
     
         94 . The composition of  claim 93 , wherein the composition comprises a tetravalent dengue virus composition capable of inducing an immune response in a subject against all four dengue virus serotypes. 
     
     
         95 . The vector according to  claim 56 , wherein the vector is a plasmid vector. 
     
     
         96 . The vector according to  claim 56 , wherein the vector is a cDNA infectious clone.