Antibody and antibody mimetic for visualization and ablation of endogenous proteins

Abstract

Provided are compositions and methods for labeling an endogenous protein, in particular, in a live cell, or for ablating an endogenous or target protein. The compositions relate to a fusion protein having a binding moiety such as an antibody, an antigen binding fragment of an antibody or an antibody mimetic that recognizes the endogenous or target protein.

1 - 34 . (canceled) 
     
     
         35 . A method for treating a disease characterized by expression of an endogenous protein in a cell in a subject in need thereof, comprising administering to the subject a polynucleotide comprising:
 (1) a nucleic acid sequence encoding a fusion polypeptide, wherein the fusion polypeptide comprises
 (a) a binding moiety for recognizing an endogenous protein in the cell, and 
 (b) a transcription factor and 
   (2) a transcription regulatory element operatively linked to the nucleic acid sequence,   
       wherein the transcription regulatory element regulates the transcription of the binding moiety and the transcription factor regulates the activity of the transcription regulatory element. 
     
     
         36 . The method of  claim 35 , wherein the endogenous protein is selected from Tau, alpha-Synuclein, prion protein, Huntingtun, Nav1.7, CCRS, HER-2, EGFR, Estrogen receptor, an oncogenes, or a drug resistant gene. 
     
     
         37 - 45 . (canceled) 
     
     
         46 . The method of  claim 35 , wherein the binding moiety is one or more of an antibody mimetic, an intrabody, an antigen binding fragment of an antibody, a natural ligand, a monobody, a linear peptide, a lipocalin scaffold, an affibody scaffold or a DARPin scaffold. 
     
     
         47 . The method of  claim 46 , wherein the intrabody comprises a fibronectin peptide. 
     
     
         48 . The method of  claim 47 , wherein the fibronection peptide comprises a 10 FnIII fragment or a biological equivalent thereof. 
     
     
         49 . The method of  claim 35 , wherein the transcription factor comprises a DNA binding domain. 
     
     
         50 . The method of  claim 35 , wherein the transcription factor is Gal4, LexA or a zinc finger domain. 
     
     
         51 . The method of  claim 35 , wherein the transcription factor further comprises one or more of a regulatory domain, a transcriptional activator or an inducible promoter. 
     
     
         52 . The method of  claim 35 , wherein the transcription factor comprises a DNA binding domain of Gal4 and a regulatory domain of Krab(A). 
     
     
         53 . The method of  claim 35 , wherein the fusion polypeptide further comprises a reporter. 
     
     
         54 . The method of  claim 53 , wherein the reporter is a green fluorescent protein (GFP), a blue fluorescent protein (BFP), a cyan fluorescent protein (CFP), a yellow fluorescent protein (YFP), mCherry, dTomato, mPlum, mOrange, mCitrine, Ypet, Cerulean CFP, luciferase, or β-galactosidase. 
     
     
         55 . The method of  claim 35 , wherein the endogenous protein is a transmembrane protein, a nucleic protein, a cytoplasmic protein, a secreted protein, or an organelle protein. 
     
     
         56 . The method of  claim 35 , wherein the endogenous protein is a transmembrane protein. 
     
     
         57 . The method of  claim 35 , further comprising administering a second nucleic acid sequence encoding a second fusion polypeptide, the second nucleic acid comprising:
 (a) a minimal promoter;   (b) a transcription factor that mediates expression of the binding moiety; and   (c) a nucleic acid encoding an heterologous protein,   wherein the transcription factor regulates the minimal promoter causing expression of the heterologous protein.   
     
     
         58 . The method of  claim 35 , wherein the disease is selected from the group of: cancer, HIV, a neurological disease, or combination thereof. 
     
     
         59 . The method of  claim 35 , further comprising administration of a therapeutic composition. 
     
     
         60 . The method of  claim 59 , wherein the therapeutic composition comprises a polynucleotide, a polypeptide, a DNA construct, a cell, a compound, or combination thereof. 
     
     
         61 . The method of  claim 35 , wherein the fusion protein comprises a protein degradation signal. 
     
     
         62 . The method of  claim 61 , wherein the protein degradation signal induces protein degradation of a polypeptide encoded by the polynucleotide. 
     
     
         63 . The method of  claim 62 , wherein the protein degradation is through ubiquitination, lysosomal degradation, or autophagy. 
     
     
         64 . The method of  claim 62 , wherein the protein degradation is through ubiquitination. 
     
     
         65 . The method of  claim 61 , wherein the protein degradation signal comprises a ubiquitin ligase, a HECT domain of an E6 protein, C20-WW-HECT, a Ring domain of Der3/Hrd1, a B-box domain of a TRIM protein, a U-box domain, KFERQ (SEQ ID NO: 1), Arg12 or Atg8/LC3. 
     
     
         66 . The method of  claim 65 , wherein the ubiquitin ligase comprises one or more of the group: a Ring domain of a protein X-linked mammalian inhibitor of apoptosis (XIAP), MDM2 or HDM2.