Cyp1b1 nucleic acids and methods of use

Abstract

The present invention provides nucleic acids containing transcriptional units that encode CYP1B1 polypeptides or portions thereof, wherein the transcriptional units lack sequences found in the untranslated region (UTR) of naturally occurring forms of the CYP1B1 transcript. The nucleic acids of the invention lack translational repressor elements and thus provide for a system of enhanced translation of the CYP1B1 polypeptide or portions thereof. Also disclosed are methods of administering nucleic acids to a mammal and use in the treatment of proliferative disorders or cancer.

1 . A nucleic acid comprising a transcriptional unit comprising a coding sequence encoding a polypeptide comprising CYP1B1 or a portion thereof that comprises a peptide that binds to an MHC class I or class II molecule, wherein the transcriptional unit does not contain a translational repressor element operably linked to the coding sequence. 
     
     
         2 . The nucleic acid of  claim 1 , wherein the polypeptide comprises a segment of CYP1B1 that is eight amino acids in length. 
     
     
         3 . The nucleic acid of  claim 2 , wherein the polypeptide comprises SEQ ID NO:22. 
     
     
         4 . The nucleic acid of  claim 3 , wherein the polypeptide is less than 100 amino acids in length. 
     
     
         5 . The nucleic acid of  claim 1 , wherein the transcriptional unit comprises an RNA stabilization sequence. 
     
     
         6 . The nucleic acid of  claim 1 , wherein the nucleic acid comprises an inducible promoter sequence operably linked to the transcriptional unit. 
     
     
         7 . The nucleic acid of  claim 1 , wherein the polypeptide comprises a targeting signal. 
     
     
         8 . A nucleic acid comprising a transcriptional unit comprising a coding sequence encoding a polypeptide comprising CYP1B1 or a portion thereof that comprises a peptide that binds to an MHC class I or class II Molecule, wherein the transcriptional unit does not contain 150 consecutive nucleotides of SEQ ID NO:18 or SEQ ID NO:19. 
     
     
         9 . The nucleic acid of  claim 8 , wherein the transcriptional unit does not contain at least one of SEQ ID NOs:3-9 or 15-17. 
     
     
         10 . The nucleic acid of  claim 9 , wherein the transcriptional unit does not contain any of SEQ ID NOs:3-9 or 15-17. 
     
     
         11 . The nucleic acid of  claim 8 , wherein the transcriptional unit does not contain 50 consecutive nucleotides of SEQ ID NO:18 or SEQ ID NO:19. 
     
     
         12 . The nucleic acid of  claim 8 , wherein the transcriptional unit does not contain 25 consecutive nucleotides of SEQ ID NO:18 or SEQ ID NO:19. 
     
     
         13 . The nucleic acid of  claim 8 , wherein the transcriptional unit does not contain 10 consecutive nucleotides of SEQ ID NO:18 or SEQ ID NO:19. 
     
     
         14 . The nucleic acid of  claim 8 , wherein the transcriptional unit comprises an RNA stabilization sequence. 
     
     
         15 . The nucleic acid of  claim 8 , wherein the nucleic acid comprises an inducible promoter sequence. 
     
     
         16 . The nucleic acid of  claim 8 , wherein the transcriptional unit comprises a translational regulatory sequence operably linked to the coding sequence. 
     
     
         17 . The nucleic acid of  claim 16 , wherein the translational regulatory sequence is an iron responsive sequence. 
     
     
         18 . The nucleic acid of  claim 8 , wherein the polypeptide comprises a segment of CYP1B1 that is eight amino acids in length. 
     
     
         19 . The nucleic acid of  claim 18 , wherein the polypeptide comprises SEQ ID NO:22. 
     
     
         20 . The nucleic acid of  claim 19 , wherein the polypeptide is less than 100 amino acids in length. 
     
     
         21 . The nucleic acid of  claim 8 , wherein the polypeptide comprises a targeting signal. 
     
     
         22 . A nucleic acid comprising a transcriptional unit, wherein the transcriptional unit encodes a hybrid polypeptide comprising a first and a second segment of CYP1B1, wherein the first and second segments are either contiguous or separated by a spacer amino acid or spacer peptide, wherein the first and second segments are each at least eight amino acids in length, and wherein the first and second segments are non-contiguous portions of CYP1B1. 
     
     
         23 . The nucleic acid of  claim 22 , wherein the hybrid polypeptide further comprises a third segment of CYP1B1, wherein the third segment is at least eight amino acids in length, and wherein the first and third and second and third segments are non-contiguous portions of CYP1B1. 
     
     
         24 . The nucleic acid of  claim 22 , wherein the first segment comprises the sequence of SEQ ID NO:22. 
     
     
         25 . The nucleic acid of  claim 22 , wherein the polypeptide comprises a targeting signal. 
     
     
         26 . A composition comprising the nucleic acid of  claim 1  and an immunostimulatory agent. 
     
     
         27 . The composition of  claim 26 , wherein the immunostimulatory agent is a CpG containing oligonucleotide of 18-30 nucleotides in length. 
     
     
         28 . The composition of  claim 26 , wherein the immunostimulatory agent is IL-12 or IFN-gamma. 
     
     
         29 . The composition of  claim 26 , wherein the immunostimulatory agent is a lipid, nucleic acid, carbohydrate, or bacterial polypeptide. 
     
     
         30 . A composition comprising the nucleic acid of  claim 1  and a nucleic acid encoding an immunostimulatory agent. 
     
     
         31 . The composition of  claim 30 , wherein the immunostimulatory agent is IL-12, IFN-gamma, or a bacterial polypeptide. 
     
     
         32 . A therapeutic composition comprising the nucleic acid of  claim 1  and a pharmaceutically acceptable carrier. 
     
     
         33 . A microparticle comprising a polymeric matrix or shell and the nucleic acid of  claim 1 . 
     
     
         34 . A microparticle comprising a polymeric matrix or shell and the nucleic acid of  claim 8 . 
     
     
         35 . A microparticle comprising a polymeric matrix or shell and the nucleic acid of  claim 22 . 
     
     
         36 . A method of inducing an immune response in a mammal, comprising administering the nucleic acid of  claim 1  to the mammal. 
     
     
         37 . The method of  claim 36 , wherein the mammal suffers from or is at risk for cancer. 
     
     
         38 . The method of  claim 36 , wherein the nucleic acid is administered subcutaneously or intramuscularly. 
     
     
         39 . The method of  claim 36 , wherein the immune response is directed to CYP1B1. 
     
     
         40 . The method of  claim 39 , wherein the immune response is a T cell response. 
     
     
         41 . The method of  claim 39 , wherein the immune response is a B cell response. 
     
     
         42 . A method of inducing an immune response in a mammal, comprising administering the microparticle of  claim 33  to the mammal. 
     
     
         43 . A method of generating an immune response, the method comprising:
 detecting expression of CYP1B1 in a tumor of a mammal; and   administering the nucleic acid of  claim 1  to the mammal, wherein the administration results in the generation of an anti-CYP1B1 immune response in the mammal.   
     
     
         44 . A method of reducing tumor growth or tumor activity in a mammal, comprising:
 identifying a mammal having a tumor;   administering the nucleic acid of  claim 1  to the mammal; and   detecting a reduction in the size or activity of the tumor following the administration of the nucleic acid.   
     
     
         45 . The method of  claim 44 , further comprising detecting CYP1B1 expression in the tumor before administering the nucleic acid. 
     
     
         46 . A method of inducing an immune response in a mammal, the method comprising administering to a mammal a nucleic acid encoding a polypeptide comprising CYP1B1 or portion thereof that binds to an MHC class I or class II molecule, wherein the mammal belongs to a first species, and wherein the CYP1B1 or portion thereof is identical to a sequence of a naturally occurring CYP1B1 polypeptide of a second species. 
     
     
         47 . The method of  claim 46 , further comprising identifying the mammal as having a tumor prior to administering the nucleic acid to the mammal. 
     
     
         48 . The method of  claim 46 , wherein the nucleic acid is the nucleic acid of  claim 1 . 
     
     
         49 . The method of  claim 46 , wherein the nucleic acid is the nucleic acid of  claim 22 . 
     
     
         50 . The method of  claim 46 , wherein the first species is a human. 
     
     
         51 . The method of  claim 50 , wherein the second species is a rodent. 
     
     
         52 . The method of  claim 51 , wherein the rodent is a rat or a mouse.