Methods And Kits For Theranostic Applications

Abstract

The present disclosure is directed to methods and kits for identifying, enriching, and evaluating templated assembly reactants. Some embodiments disclose methods for identifying templated assembly targets by synthesizing templated assembly reactants, hybridizing the templated assembly reactants to target nucleic acids, performing a templated assembly reaction, and identifying the target nucleic acids that hybridized to the templated assembly reactants. Libraries of templated assembly reactants, a kit for identifying templated assembly targets, and a pair of templated assembly targets enriched from a library of chemically-ligated oligonucleotides spatially elicited (CLOSE) products are also disclosed.

1 . A method for identifying templated assembly targets comprising:
 synthesizing a first population of templated assembly reactants and a second population of corresponding templated assembly reactants, wherein the first and second populations of templated assembly reactants comprise oligonucleotide sequences;   hybridizing both populations of templated assembly reactants to target nucleic acids;   performing a templated assembly reaction, wherein the hybridized first population of templated assembly reactants and the hybridized second population of corresponding templated assembly reactants undergo templated assembly; and   identifying the target nucleic acids that hybridized to either the first or second population of templated assembly reactants that underwent templated assembly, wherein the hybridized target nucleic acids are the templated assembly targets.   
     
     
         2 - 14 . (canceled) 
     
     
         15 . A library of templated assembly reactants for identifying templated assembly targets comprising at least a first and second population of templated assembly reactants, wherein the templated assembly reactants comprise an oligonucleotide sequence and a modification for reaction in a templated assembly reaction. 
     
     
         16 . The library of  claim 15 , wherein the templated assembly reactants further comprise either a 5′ or a 3′priming site adjacent to the oligonucleotide sequence. 
     
     
         17 . The library of  claim 15 , wherein the modification in the first population of templated assembly reactants further comprises a 5′-azide and the modification in the second population of templated assembly reactants further comprises a 3′-alkyne group. 
     
     
         18 . The library of  claim 15 , wherein the modification in the first population of templated assembly reactants further comprise a 5′-alkyne and the modification on the second population of templated assembly reactants further comprise a 3′-azide group. 
     
     
         19 . The library of  claim 15 , wherein the templated assembly reaction is selected from the group consisting of a click chemical reaction, a Staudinger reduction, a non-traceless Staudinger ligation, and a native chemical ligation. 
     
     
         20 . The library of  claim 19 , wherein the modification is specific for a traceless Staudinger ligation selected from either a traceless phosphinophenol Staudinger ligation or a traceless phosphinomethanethiol Staudinger ligation. 
     
     
         21 - 22 . (canceled) 
     
     
         23 . The library of  claim 15 , wherein the oligonucleotide sequence further comprises nuclease-resistant phosphodiester backbones or nuclease-resistant sugar moieties. 
     
     
         24 . A kit for identifying templated assembly targets comprising a library of oligonucleotides for identifying templated assembly targets comprising oligonucleotide sequences modified as corresponding templated assembly reactants and reagents. 
     
     
         25 . The kit of  claim 24 , wherein the oligonucleotide sequences further comprise 5′, 3′, or 5′ and 3′ priming site. 
     
     
         26 . The kit of  claim 24 , wherein the oligonucleotide sequences further comprise a 5′ or a 3′ modification for templated assembly. 
     
     
         27 . The kit of  claim 24 , wherein the oligonucleotide sequences comprises random sequences of about 5 to about 100 nucleotides long. 
     
     
         28 . The kit of  claim 24 , wherein the oligonucleotide sequences comprises gene specific sequences of about 5 to about 100 nucleotides long. 
     
     
         29 . A method for enrichment of a pair of templated assembly targets from a library of chemically-ligated oligonucleotides spatially elicited (CLOSE) products comprising:
 obtaining a library of oligonucleotides chemically ligated through templated assembly due to spatial proximity to cellular nucleic acid targets;   amplifying the library of ligated oligonucleotide-cellular nucleic acid targets; and   selectively enriching for ligated oligonucleotide-cellular nucleic acid targets, wherein the ligated targets are selected for relevance to a pathology of an aberrant cell of interest or to a discontinuous hybridization to the cellular nucleic acid targets.   
     
     
         30 . (canceled) 
     
     
         31 . A pair of templated assembly targets enriched from a library of chemically-ligated oligonucleotides spatially elicited (CLOSE) products comprising oligonucleotides chemically ligated due to their spatial proximity through hybridization to cellular nucleic acid templates. 
     
     
         32 . The templated assembly target of  claim 31 , wherein the CLOSE library comprises short PCR product duplexes of the chemically ligated oligonucleotides. 
     
     
         33 . The templated assembly target of  claim 31 , wherein the CLOSE library comprises an amplified library of chemically ligated products. 
     
     
         34 . The templated assembly target of  claim 31 , wherein the CLOSE library comprises an amplified library of rearranged chemically ligated products. 
     
     
         35 . The templated assembly target of  claim 31 , wherein the cellular nucleic acid templates comprise genomic or expressed genes for known oncogenes or tumor suppressors, cell cycle regulators and mediators, transcriptonal regulators and mediators, translational regulators and mediators, telomerases, cytoskeletal components, and kinases. 
     
     
         36 - 37 . (canceled) 
     
     
         38 . A method of evaluating a pair of chemically-ligated oligonucleotides spatially elicited (CLOSE) products for templated assembly comprising:
 modifying the pair of CLOSE products as templated assembly reactants;   transfecting the pair of modified CLOSE products into a target aberrant cell of interest; and   screening for templated assembly of the pair of modified CLOSE products.   
     
     
         39 - 41 . (canceled)