Chimeric activators: quantitatively designed protein therapeutics and uses thereof

Abstract

Aspects of the invention provide methods for harnessing the potential of proteins that occur naturally (e.g., in humans) and that have serious but finite toxicity. Aspects of the invention relate to a quantitative systems-biological and structural approach to design a class of chimeric proteins that avoid the toxicity of protein drugs while retaining their desired activities. In particular, chimeric proteins containing a variant form of a natural protein fused to a targeting moiety may be administered to a subject to target a signal (e.g., induction of apoptosis) to particular cells without having a generalized toxic effect in the subject.

What is claimed herein: 
     
         1 . A recombinant protein for selectively targeting a cancer cell, comprising
 a first element a variant of a wild-type tumor necrosis factor alpha (TNFα) protein, the variant comprising at least one amino acid substitution mutation that reduces the binding affinity of the variant for its naturally-occurring cell surface receptors by at least 10-fold relative to the wild-type TNFα protein;   a second element comprising an epidermal growth factor receptor (EGFR) binding protein selected from the group consisting of: a wild-type epidermal growth factor (EGF) protein, MR1-1, and an anti-EGFRde(2-7) antibody or fragment thereof; and   a linker that connects the first and second elements.   
     
     
         2 . The recombinant protein of  claim 1 , wherein when administered to a subject, the second receptor binding protein binds the recombinant protein to the surface of a cancer cell and promotes cell death or a reduction or absence of cell proliferation of the cancer cell, but does not bind the recombinant protein to the surface of a non-cancer cell and does not promote cell death or a reduction or absence of cell proliferation of the non-cancer cell. 
     
     
         3 . The recombinant protein of  claim 1 , wherein the first element is a variant of a wild-type receptor binding protein which comprises at least one amino acid substitution mutation that reduces the binding affinity of the variant for its naturally-occurring cell surface receptors by at least 200-fold relative to the wild-type receptor binding protein. 
     
     
         4 . The recombinant protein of  claim 1 , wherein the variant of a wild-type TNFα is a N39Y, S147Y, or Y87H variant of wild-type TNFα. 
     
     
         5 . The recombinant protein of  claim 1 , wherein the at least one amino acid substitution is N39Y, S147Y, or Y87H relative to wild-type TNFα. 
     
     
         6 . A recombinant protein for selectively targeting a cancer cell, comprising
 a first element a variant of a wild-type tumor interferon alpha (IFNα) protein, the variant comprising at least one amino acid substitution mutation that reduces the binding affinity of the variant for its naturally-occurring cell surface receptors by at least 10-fold relative to the wild-type IFNα protein;   a second element comprising an epidermal growth factor receptor (EGFR) binding protein comprising an anti-EGFRde(2-7) antibody or fragment thereof; and   a linker that connects the first and second elements.   
     
     
         7 . The recombinant protein of  claim 6 , wherein when administered to a subject, the second receptor binding protein binds the recombinant protein to the surface of a cancer cell and promotes cell death or a reduction or absence of cell proliferation of the cancer cell, but does not bind the recombinant protein to the surface of a non-cancer cell and does not promote cell death or a reduction or absence of cell proliferation of the non-cancer cell. 
     
     
         8 . The recombinant protein of  claim 6 , wherein the first element is a variant of a wild-type receptor binding protein which comprises at least one amino acid substitution mutation that reduces the binding affinity of the variant for its naturally-occurring cell surface receptors by at least 200-fold relative to the wild-type receptor binding protein. 
     
     
         9 . The recombinant protein of  claim 6 , wherein the variant of a wild-type IFNα is a K133A, R144A, or R149A variant of wild-type IFNα. 
     
     
         10 . The recombinant protein of  claim 6 , wherein the at least one amino acid substitution is K133A, R144A, or R149A relative to wild-type IFNα. 
     
     
         11 . The recombinant protein of  claim 6 , wherein the anti-EGFRde(2-7) antibody is MR1-1. 
     
     
         12 . A recombinant protein for selectively targeting an adipocyte, comprising
 a first element a variant of a wild-type tumor necrosis factor alpha (TNFα) protein, the variant comprising at least one amino acid substitution mutation that reduces the binding affinity of the variant for its naturally-occurring cell surface receptors by at least 10-fold relative to the wild-type TNFα protein;   a second element comprising a leptin protein; and   a linker that connects the first and second elements.   
     
     
         13 . The recombinant protein of  claim 12 , wherein when administered to a subject, the second receptor binding protein binds the recombinant protein to the surface of an adipocyte and promotes cell death or a reduction or absence of cell proliferation of the adipocyte, but does not bind the recombinant protein to the surface of a non-adipocyte cell and does not promote cell death or a reduction or absence of cell proliferation of the non-adipocyte cell. 
     
     
         14 . The recombinant protein of  claim 12 , wherein the first element is a variant of a wild-type receptor binding protein which comprises at least one amino acid substitution mutation that reduces the binding affinity of the variant for its naturally-occurring cell surface receptors by at least 200-fold relative to the wild-type receptor binding protein. 
     
     
         15 . The recombinant protein of  claim 12 , wherein the variant of a wild-type TNFα is a N39Y, S147Y, or Y87H variant of wild-type TNFα. 
     
     
         16 . The recombinant protein of  claim 12 , wherein the at least one amino acid substitution is N39Y, S147Y, or Y87H relative to wild-type TNFα. 
     
     
         17 . A recombinant protein for selectively targeting a cell infected with HIV, comprising
 a first element a variant of a wild-type tumor interferon alpha (IFNα) protein, the variant comprising at least one amino acid substitution mutation that reduces the binding affinity of the variant for its naturally-occurring cell surface receptors by at least 10-fold relative to the wild-type IFNα protein;   a second element comprising a CD4 protein; and   a linker that connects the first and second elements.   
     
     
         18 . The recombinant protein of  claim 17 , wherein when administered to a subject, the second receptor binding protein binds the recombinant protein to the surface of a cell infected with HIV and promotes cell death or a reduction or absence of cell proliferation of the cell infected with HIV, but does not bind the recombinant protein to the surface of a cell not infected with HIV and does not promote cell death or a reduction or absence of cell proliferation of the cell not infected with HIV. 
     
     
         19 . The recombinant protein of  claim 17 , wherein the first element is a variant of a wild-type receptor binding protein which comprises at least one amino acid substitution mutation that reduces the binding affinity of the variant for its naturally-occurring cell surface receptors by at least 200-fold relative to the wild-type receptor binding protein. 
     
     
         20 . The recombinant protein of  claim 17 , wherein the variant of a wild-type IFNα is a K133A, R144A, or R149A variant of wild-type IFNα. 
     
     
         21 . The recombinant protein of  claim 17 , wherein the at least one amino acid substitution is K133A, R144A, or R149A relative to wild-type IFNα. 
     
     
         22 . A recombinant protein for selectively targeting a cancer cell, comprising
 a first element a variant of a wild-type TRAIL protein, the variant comprising at least one amino acid substitution mutation that reduces the binding affinity of the variant for its naturally-occurring cell surface receptors by at least 10-fold relative to the wild-type TRAIL protein;   a second element comprising an epidermal growth factor receptor (EGFR) binding protein comprising an anti-EGFRde(2-7) antibody or fragment thereof; and   a linker that connects the first and second elements.   
     
     
         23 . The recombinant protein of  claim 22 , wherein when administered to a subject, the second receptor binding protein binds the recombinant protein to the surface of a cancer cell and promotes cell death or a reduction or absence of cell proliferation of the cancer cell, but does not bind the recombinant protein to the surface of a non-cancer cell and does not promote cell death or a reduction or absence of cell proliferation of the non-cancer cell. 
     
     
         24 . The recombinant protein of  claim 22 , wherein the first element is a variant of a wild-type receptor binding protein which comprises at least one amino acid substitution mutation that reduces the binding affinity of the variant for its naturally-occurring cell surface receptors by at least 200-fold relative to the wild-type receptor binding protein. 
     
     
         25 . The recombinant protein of  claim 22 , wherein the variant of a wild-type TRAIL is a Q205A, Y216A, or Y237A variant of wild-type TRAIL. 
     
     
         26 . The recombinant protein of  claim 22 , wherein the at least one amino acid substitution is Q205A, Y216A, or Y237A relative to wild-type TRAIL. 
     
     
         27 . The recombinant protein of  claim 22 , wherein the anti-EGFRde(2-7) antibody is MR1-1.