Modular targeted therapeutic agents and methods of making same

Abstract

Provided herein are methods for making targeted therapeutics. In several embodiments, the therapeutics are directed against soluble agents such as toxins, venoms, and/or other factors that alter physiological biopathways as well as methods of using such therapeutics to treat patients or patient populations to reduce, eliminate, or inactivate, detrimental soluble agents that such patients or patient populations have been exposed to. In several embodiments, the therapeutics are directed to patient-specific disease markers. In several embodiments, the methods comprise screening a library comprising proteins linked to their cognate mRNAs to identify mRNA-protein pairs that bind to the diseased cells, isolating one or more proteins from the identified mRNA-protein pairs, and conjugating the isolated protein(s) to a therapeutic agent.

1 . A method for generation of a bifunctional targeted therapeutic that targets an agent of interest, the method comprising:
 identifying a first protein capable of specifically interacting with the agent of interest;   identifying a first mRNA that encodes for said first protein;   identifying non-antibody second portion capable of eliciting an immune response through interaction with one or more components of the immune system;   identifying a second mRNA that encodes for said non-antibody second portion;   generating a first and a second cDNA corresponding to each of said first and said second mRNAs;   fusing said first and said second cDNA to the first and second ends of a bridge cDNA to generate a fused cDNA;   translating said fused cDNA into a corresponding fused protein,   wherein a first portion of said fused protein is capable of interacting with said agent of interest and a second portion of said fused protein is capable of eliciting an immune response,   thereby generating a bifunctional targeted therapeutic that targets said agent of interest.   
     
     
         2 . The method for generation of a bifunctional targeted therapeutic according to  claim 1 , wherein said non-antibody second portion is a second protein. 
     
     
         3 . The method according to  claim 1 , wherein said first protein is identified by screening a library comprising proteins linked to their cognate mRNAs to identify one or more proteins that interact with the agent of interest. 
     
     
         4 . The method according to  claim 1 , further comprising screening the bifunctional targeted therapeutic against said agent of interest and said one or more components of the immune system. 
     
     
         5 . The method according to  claim 1 , wherein said non-antibody second portion has a known mRNA sequence. 
     
     
         6 . The method according to  claim 2 , wherein said second protein is capable of binding to the heavy chain of an antibody. 
     
     
         7 . The method according to  claim 6 , wherein said second protein is capable of binding to the constant region of the heavy chain of said antibody. 
     
     
         8 . The method according to  claim 7 , wherein said second protein is capable of binding to the CH1 region of the heavy chain. 
     
     
         9 . The method according to  claim 1 , wherein said non-antibody second portion is capable of binding to an IgG antibody. 
     
     
         10 . The method according to  claim 1 , wherein agent of interest is a selected from the group consisting of animal toxins, insect toxins, plant toxins, algae-derived toxins, fungi-derived toxins, bacterial-derived toxins, biowarfare agents, and biopathway modulators. 
     
     
         11 . The method according to  claim 1 , wherein said agent of interest targets one or more of the blood, blood vessels, nervous tissue, and muscle tissue,
 wherein said soluble agent targets one or more ion channels,   wherein said soluble agent induces muscle paralysis,   wherein said soluble agent prevents blood clotting, or   wherein said soluble agent induces increased gastrointestinal water secretion.   
     
     
         12 . The method according to  claim 3 , wherein said screening comprises:
 immobilizing said agent of interest on a solid support;   blocking a site on said agent of interest with which said first protein interacts;   exposing a plurality of candidate bifunctional targeted therapeutics to said blocked and immobilized agent of interest;   identifying those candidate bifunctional targeted therapeutics that bind to said blocked and immobilized agent of interest and those that do not bind to said blocked and immobilized agent of interest; and   collecting those candidate bifunctional targeted therapeutic that do not bind to said blocked and immobilized agent of interest.   
     
     
         13 . The method of  claim 12 , further comprising screening said collected candidate bifunctional targeted therapeutics for interaction with one or more components of the immune system. 
     
     
         14 . The method of  claim 13 , wherein screening of said collected bifunctional targeted therapeutics comprises:
 immobilizing a component of the immune system on a solid support;   exposing said collected candidate bifunctional targeted therapeutics to said immobilized component of the immune system;   identifying those candidate bifunctional targeted therapeutics that bind to said immobilized component of the immune system and those that do not bind to said immobilized component of the immune system; and   discarding those candidate bifunctional targeted therapeutics that do not bind to said immobilized component of the immune system and collecting those candidate bifunctional targeted therapeutics that do bind to said component of the immune system.   
     
     
         15 . The method of  claim 13 , wherein said component of the immune system is an antibody. 
     
     
         16 . The method of  claim 15 , wherein said antibody is an IgG antibody. 
     
     
         17 . The method of  claim 16 , wherein said antibody is an IgG isotype 1, 2, 3, or 4 antibody. 
     
     
         18 . A method for treating a subject that has been exposed to a soluble agent, comprising:
 identifying a subject who has been exposed to a soluble agent; and   administering to said subject a bifunctional targeted therapeutic generated by the method of  claim 1 ,   wherein said bifunctional targeted therapeutic is generated to interact with said soluble agent,   wherein said elicited immune response results in clearance of the soluble agent and treatment of said subject.