US2011244519A1PendingUtilityA1

Use of rna trans-splicing for antibody gene transfer and antibody polypeptide production

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Assignee: VIRXSYS CORPPriority: Oct 8, 2004Filed: Jan 11, 2011Published: Oct 6, 2011
Est. expiryOct 8, 2024(expired)· nominal 20-yr term from priority
Inventors:Gerard J. McgarrityMariano A. Garcia-BlancoMadaiah Puttaraju
A61P 37/06A61P 35/00A61P 31/04A61P 29/00C07K 16/00C12N 15/1027A61K 48/00C07K 16/084C07K 2317/622A61P 19/02A61K 2039/505
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Claims

Abstract

The present invention provides methods and compositions for generating novel nucleic acid molecules through RNA trans-splicing that target a highly expressed pre-mRNA and contain the coding sequence for antibody polypeptide(s). The compositions of the invention include pre-trans-splicing molecules (PTMs) designed to interact with the target precursor messenger RNA molecule (target pre-mRNA) that is abundantly expressed or tumor specific and mediate a trans-splicing reaction resulting in the generation of novel chimeric RNA molecule (chimeric RNA) capable of encoding an antibody polypeptide. The invention provides for the in vivo production of chimeric RNA molecules that encode and result in the production of an antibody polypeptide that is therapeutically effective against, for example, infectious agents, cancer cells, transplantation antigens, rheumatoid arthritis, etc.

Claims

exact text as granted — not AI-modified
1 - 13 . (canceled) 
     
     
         14 . A nucleic acid molecule that encodes an antibody polypeptide wherein said nucleic acid molecule comprises:
 a) one or more target binding domains that target binding of the nucleic acid molecule that encodes the antibody polypeptide to an abundantly expressed target pre-mRNA within the cell;   b) a splice region;   c) a spacer region that separates the splice region from the target binding domain; and   d) a nucleotide sequence encoding the antibody polypeptide to be trans-spliced to the target pre-mRNA;   
       wherein said nucleic acid molecule is recognized by nuclear splicing components within the cell. 
     
     
         15 . The nucleic acid molecule of  claim 14  wherein the abundantly expressed target pre-mRNA is selected from the group consisting of pre-mRNAs encoding albumin, casein, myosin and fibroin. 
     
     
         16 . The nucleic acid molecule of  claim 14  wherein the abundantly expressed target pre-mRNA encodes albumin. 
     
     
         17 . The nucleic acid molecule of  claim 14  wherein the abundantly expressed target pre-mRNA is a tumor-specific or tumor associated transcript. 
     
     
         18 . The nucleic acid molecule of  claim 14  wherein the abundantly expressed target pre-mRNA is a microbial or autoantigen associated transcript. 
     
     
         19 . The nucleic acid molecule of  claim 14  wherein the target pre-mRNA is a viral or yeast associated transcript. 
     
     
         20 . The nucleic acid molecule of  claim 14  wherein the antibody polypeptide is selected from the group consisting of an Ig heavy chain, an Ig light chain, an Ig Fv fragment, an Ig Fab fragment, an Ig Fc fragment, a single chain antibody and combinations thereof. 
     
     
         21 . The nucleic acid molecule of  claim 14  wherein the antibody polypeptide is a single chain antibody. 
     
     
         22 . The nucleic acid molecule of  claim 14  wherein the antibody polypeptide comprises an Ig heavy chain and an Ig light chain. 
     
     
         23 . The nucleic acid molecule of  claim 14  wherein the antibody polypeptide is specific for a tumor-specific or tumor associated antigen. 
     
     
         24 . The nucleic acid molecule of  claim 14  wherein the antibody polypeptide is specific for a microbial or autoantigen associated antigen. 
     
     
         25 . The nucleic acid molecule of  claim 24  wherein the microbial associated antigen is selected from the group consisting of viral and yeast antigens. 
     
     
         26 . The nucleic acid molecule of  claim 14  wherein the nucleic acid molecules further comprises a sequence encoding a cytokine or a growth factor. 
     
     
         27 . A method of producing a chimeric RNA molecule that encodes an antibody molecule in a cell comprising: contacting an abundantly expressed target pre-mRNA within the cell with a nucleic acid molecule encoding the antibody polypeptide that is recognized by nuclear splicing components wherein said nucleic acid molecule comprises:
 a) one or more target binding domains that target binding of the nucleic acid molecule that encodes the antibody polypeptide to the abundantly expressed target pre-mRNA within the cell;   b) a splice region;   c) a spacer region that separates the splice region from the target binding domain; and   d) a nucleotide sequence to be trans-spliced to the target pre-mRNA;   
       wherein the nucleic acid molecule is recognized by nuclear splicing components within the cell. 
     
     
         28 . The method of  claim 27  wherein the abundantly expressed target pre-mRNA is selected from the group consisting of pre-mRNAs encoding albumin, casein, myosin and fibroin. 
     
     
         29 . The method of  claim 27  wherein the abundantly expressed target pre-mRNA encodes albumin. 
     
     
         30 . The method of  claim 27  wherein the abundantly expressed target pre-mRNA is a tumor-specific or tumor associated transcript. 
     
     
         31 . The method of  claim 27  wherein the abundantly expressed target pre-mRNA is a microbial or autoantigen associated transcript. 
     
     
         32 . The method of  claim 27  wherein the abundantly expressed target pre-mRNA is a viral or yeast associated transcript. 
     
     
         33 . The method of  claim 27  wherein the antibody polypeptide is selected from the group consisting of an Ig heavy chain, an Ig light chain, an Ig Fv fragment, an Ig Fab fragment, an Ig Fc fragment, a single chain antibody and combinations thereof. 
     
     
         34 . The method of  claim 27  wherein the antibody polypeptide is a single chain antibody. 
     
     
         35 . The method of  claim 27  wherein the antibody polypeptide comprises an Ig heavy chain and an Ig light chain. 
     
     
         36 . The method of  claim 27  wherein the antibody polypeptide is specific for a tumor-specific or tumor associated antigen. 
     
     
         37 . The method of  claim 27  wherein the antibody polypeptide is specific for a microbial or autoantigen associated antigen. 
     
     
         38 . The method of  claim 37  wherein the microbial associated antigen is selected from the group consisting of viral and yeast antigens. 
     
     
         39 . The method of  claim 27  wherein the nucleic acid molecules further comprises a sequence encoding a cytokine or a growth factor. 
     
     
         40 . A method of producing an antibody polypeptide in a cell comprising: contacting an abundantly expressed target pre-mRNA within the cell with a nucleic acid molecule to produce a chimeric RNA molecule that encodes the antibody polypeptide wherein said nucleic acid molecule comprises:
 a) one or more target binding domains that target binding of the nucleic acid molecule to the abundantly expressed target pre-mRNA within the cell;   b) a splice region;   c) a spacer region that separates the splice region from the target binding domain; and   d) a nucleotide sequence to be trans-spliced to the target pre-mRNA;   
       wherein the nucleic acid molecule is recognized by nuclear splicing components within the cell and wherein the chimeric RNA molecule is translated by the cell to produce the antibody polypeptide. 
     
     
         41 . The method of  claim 40  wherein the abundantly expressed target pre-mRNA is selected from the group consisting of a pre-mRNA encoding albumin, casein, myosin and fibroin. 
     
     
         42 . The method of  claim 40  wherein the abundantly expressed target pre-mRNA encodes albumin. 
     
     
         43 . The method of  claim 40  wherein the abundantly expressed target pre-mRNA is a tumor-specific or tumor associated transcript. 
     
     
         44 . The method of  claim 40  wherein the abundantly expressed target pre-mRNA is a microbial or autoantigen associated transcript. 
     
     
         45 . The method of  claim 40  wherein the abundantly expressed target pre-mRNA is a viral or yeast associated transcript. 
     
     
         46 . The method of  claim 40  wherein the antibody polypeptide is selected from the group consisting of an Ig heavy chain, an Ig light chain, an Ig Fv fragment, an Ig Fab fragment, an Ig Fc fragment, a single chain antibody and combinations thereof. 
     
     
         47 . The method of  claim 40  wherein the antibody polypeptide is a single chain antibody. 
     
     
         48 . The method of  claim 40  wherein the antibody polypeptide comprises an Ig heavy chain and an Ig light chain. 
     
     
         49 . The method of  claim 40  wherein the antibody polypeptide is specific for a tumor-specific or tumor associated antigen. 
     
     
         50 . The method of  claim 40  wherein the antibody polypeptide is specific for a microbial or autoantigen associated antigen. 
     
     
         51 . The method of  claim 50  wherein the microbial associated antigen is selected from the group consisting of viral and yeast antigens. 
     
     
         52 . The method of  claim 40  wherein the nucleic acid molecules further comprises a sequence encoding a cytokine or a growth factor.

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