Enhancing nucleic acid polymerization by aromatic compounds

Abstract

The present disclosure relates to compounds, methods, and compositions for improving nucleic acid polymerization, including DNA replication by in vitro primer extension to produce polymers for, e.g., nanopore-based single molecule sequencing of DNA templates. Provided are nucleic acid polymerase reaction compositions with polymerization-enhancing moieties that allow for enhanced DNA polymerase activity on nucleotide analogs, thereby improving the length of primer extension products for sequencing applications.

Description

[0001] Cross-references to related applications

[0002] This disclosure claims the filing date interests of U.S. Provisional Application No. 63 / 614,144, filed December 22, 2023; U.S. Provisional Application No. 63 / 614,218, filed December 22, 2023; U.S. Provisional Application No. 63 / 680,104, filed August 7, 2024; U.S. Provisional Application No. 63 / 687,453, filed August 27, 2024; and U.S. Provisional Application No. 63 / 688,911, filed August 30, 2024, the disclosure of which is hereby incorporated herein by reference in its entirety. Technical Field

[0003] This disclosure relates to new chemical entities, and more specifically to new organic molecules (including compositions thereof) optionally having inorganic components, and methods for their manufacture and utilization (particularly in relation to their effects on enzyme performance).

[0004] sequence list

[0005] The contents of the electronic sequence listing (P39808-WO sequence listing.xml; size: 15,170 bytes; and creation date: December 18, 2024) are incorporated herein by reference in their entirety. Background Technology

[0006] The measurement of biomolecules is fundamental to modern medicine, widely used in medical research, more specifically in diagnosis and treatment, and drug development. Nucleic acids encode the information necessary for the activities and reproduction of living organisms, essentially serving as a blueprint for life. Determining such a blueprint is useful in both pure research and applied sciences. In medicine, sequencing can be used to diagnose and develop treatments for a wide range of conditions, including cancer, heart disease, autoimmune diseases, multiple sclerosis, and obesity. In industry, sequencing can be used to design improved enzymatic processes or synthetic biology. For example, in biology, this tool can be used to study the health of ecosystems, thus having a wide range of applications. Similarly, the measurement of proteins and other biomolecules provides biomarkers and insights into the spread of diseases and pathogens.

[0007] An individual's unique DNA sequence provides valuable information about their susceptibility to certain diseases. It also offers patients the opportunity to screen for early detection and / or receive preventative treatment. Furthermore, based on a patient's individual profile, clinicians will be able to administer personalized therapies to maximize drug efficacy and / or minimize the risk of adverse drug reactions. Similarly, identifying pathogen profiles can provide new treatments for infectious diseases and more robust pathogen surveillance. Low-cost whole-genome DNA sequencing will lay the foundation for modern medicine. To achieve this, sequencing technologies must continuously advance in throughput, accuracy, and read length.

[0008] Over the past decade, a plethora of next-generation DNA sequencing technologies have become commercially available, significantly reducing the cost of whole-genome sequencing. These include sequencing-by-synthesis (“SBS”) platforms (Illumina, Inc., 454 LifeSciences, Ion Torrent, Pacific Biosciences) and similar connection-based platforms (CompleteGenomics, Life Technologies Corporation). Many other technologies are under development, utilizing a wide variety of sample handling and detection methods. For example, GnuBio (Cambridge, Mass.) uses picoliter reaction vessels to control millions of carefully selected probes in sequencing reactions, while Halcyon Molecular (Redwood City, Calif.) is exploring techniques for direct DNA measurement using transmission electron microscopy.

[0009] Nanopore-based nucleic acid sequencing is a compelling method that has been extensively studied. Kasianowicz et al. (Proc. Natl. Acad. Sci. USA 93: 13770-13773, 1996) characterized single-stranded polynucleotides (SNPs) during electrical translocation through α-hemolysin nanopores embedded in a lipid bilayer. It has been demonstrated that partial blockage of the nanopore pore opening can be measured as a measure of the reduction in ionic current during SNP translocation. However, SNP sequencing in nanopores suffers from small signal differences immersed in significant background noise due to the need to resolve closely spaced bases (0.34 nm). The challenge of achieving single-base resolution in nanopores becomes even more demanding due to the observed rapid translocation rate of SNPs, typically around one base per microsecond. The translocation rate can be reduced by adjusting operating parameters such as voltage, salt composition, pH, temperature, and viscosity. However, such adjustments cannot reduce the translocation rate to a level that allows for single-base resolution.

[0010] Stratos Genomics has developed a method called extended sequencing (“SBX”), which uses a biochemical process to transcribe DNA sequences onto a measurable polymer called “Xpandomer” (Kokoris et al., U.S. Patent No. 7,939,259, “High Throughput Nucleic Acid Sequencing by Expansion”). The transcribed sequence is encoded along the Xpandomer backbone in high signal-to-noise ratio reporters spaced approximately 10 nm apart and designed for high signal-to-noise ratio, well-differentiated responses. These differences provide significant enhancements in read efficiency and accuracy of Xpandomer relative to native DNA. Xpandomer enables a variety of next-generation DNA sequencing detection technologies and is well-suited for nanopore sequencing.

[0011] Xpandomer is produced from a non-natural nucleotide analogue called XNTP, characterized by long substituents that allow the Xpandomer backbone to extend post-synthesis (see published PCT application WO2016 / 081871 by Kokoris et al., incorporated herein by reference in its entirety). Due to their atypical structure, XNTPs and other nucleotide analogues (e.g., nucleotide analogues partially modified with detectable markers) present new challenges as substrates for currently available DNA polymerases. Published PCT applications WO2017 / 087281 and WO2018 / 204717 by Kokoris et al. (incorporated herein by reference in its entirety) describe engineered DP04 polymerase variants with enhanced primer extension activity using non-natural, large nucleotide analogues as substrates.

[0012] Within the DNA template itself, certain nucleotide sequence motifs are known to pose additional replication challenges to DNA polymerases. Particularly important is the operation of homopolymers or short repetitive DNA sequences, which can trigger slippage, or “replication slip.” Replication slip is thought to encompass the following steps: (i) copying of the first repetitive sequence via replication mechanisms, (ii) replication pause and dissociation of the polymerase from the newly synthesized end, (iii) unpairing of the newly synthesized strand and its pairing with the second repetitive sequence, and (iv) restart of DNA synthesis. Therefore, stagnation of replication mechanisms within repetitive regions can lead to primer and template misalignment. In vivo, misalignment of the two DNA strands during replication can result in DNA rearrangements, such as deletions or duplications of different lengths. In vitro, replication slip can lead to replication errors at the site of the slippage event. This reduction in the polymerase’s sustained synthetic capacity or accuracy significantly impairs specific applications or desired genetic manipulation.

[0013] Therefore, novel methods and compositions for enhancing polymerase reactions under conditions including one or more reagents with atypical structures are necessary (e.g., in extended sequencing (SBX) and other applications in biotechnology and biomedicine (e.g., DNA amplification, conventional sequencing, labeling, detection, cloning, etc.)) and will be valuable in the art. This disclosure addresses these needs and provides further related advantages.

[0014] All topics discussed in the Background section are not necessarily prior art and should not be assumed to be prior art simply because they are discussed in the Background section. Following these lines of thought, unless explicitly stated otherwise, any awareness of problems in prior art discussed in the Background section or related to such topics should not be considered prior art. Instead, the discussion of any topic in the Background section should be considered as part of the inventor's method for solving a particular problem, and may itself be inventive. Summary of the Invention

[0015] This disclosure provides compounds, compositions, and uses thereof for enhancing nucleic acid polymerase activity. In some embodiments, polymerase activity is enhanced during polymerization reactions under conditions that introduce one or more challenges to the polymerase, such as conditions including non-natural nucleotide analog substrates or template motifs that impair the polymerase's ability to synthesize continuously. This enhancement is achieved by supplementing the polymerization reaction with one or more compounds of this disclosure, which may be referred to herein as "polymerase-enhancing molecules" ("PEMs").

[0016] One aspect of this disclosure is a compound having formula (I), such as PEM:

[0017] (I),

[0018] Or its solvates, hydrates, tautomers, chelates or salts,

[0019] in

[0020] a is 0 or an integer in the range of 1 to 4;

[0021] a' is 0 or an integer in the range of 1 to 4;

[0022] m is 1, 2, or 3;

[0023] m' is 1, 2, or 3;

[0024] n is 0, 1, or 2;

[0025] p is 0, 1, or 2;

[0026] Z is a 5- or 6-membered heteroaromatic ring containing one, two, three or four heteroatoms selected from O, N or S;

[0027] L is a linking group;

[0028] M is independently selected from hydrogen, halogens, and C1–C4 alkyl groups each time it appears;

[0029] Ar1, each time it appears, is independently selected from optionally substituted phenyl, pyridine, bipyridine, tripyridine, pyrazine, pyridazine, furan, dibenzofuran, thiophene, pyrrole, selenophene, naphthalene, fluorene, phenanthrene, cyclophosphine, phthalazine, quinazoline, quinoxaline, naphthidine, phenanthrene-rholine, purine, and carbazole.

[0030] The substituents of Ar1 are independently selected each time they appear from halogens, –OH, –CN, –NO2, C1–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, C1–C6 cycloalkyl, –OR 0 –CONH2, –C(O)NR 1 R 1' –C(O)(CH2) a NR 1 R 1' –NR 1 R 1' –NR 1 C(O)R 3 –C(O)SR 3 –COR 3 –CO(CH2) a OC(O)R 3 –OC(O)R 3 –C(O)OR 3 –C–O–R3 , thiols, -R 4 –H、–SOR 1 –S(O)2R 1 –S(O)2NR 1 R 1' –CH2–NR 1 S(O)2R 3 –NR 1 S(O)2R 3 and –C(CH3)=N–(phenyl)–O–CH2–C–CH;

[0031] R 0 Each time it appears, it is independently selected from C1–C6 alkyl, C1–C6 haloalkyl, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl.

[0032] R 1 and R 1' Each time it appears, it is independently selected from H, hydroxyl, C1–C6 alkyl, C1–C6 haloalkyl, C1–C 20 Heteroalkyl, C1–C 10 Heteroalkyl –NH2, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic (e.g., via one or more R... 3 Substituent groups), substituted or unsubstituted aryl groups, substituted or unsubstituted arylalkyl groups, and substituted or unsubstituted heteroaryl groups, benzyl groups, –C(=NH)NH2, –CH2CO2R 0 –C(H)(CH) a (COOH))2, –CH2C(O)NHCH2CO2H, –CH2CH2OH, –CH2CH2NHC(O)R 3 –(CH2) a C(O)NH(CH2) a C(O)OR 3 –(CH2) a –CN, spermine, –(CH2) a –NH2、–C(O)R 3 –(CH2) a C(O)OR 3 –(CH2) a OR 3 –C(H)((CH2) a OH) a' –CHCH(OH)(CH2)a OH, –(CH2) a OH, C1–C6–C(O)OH, can be substituted (e.g., via R...) 3 (Substituted group) or unsubstituted –(CH2) a - Heterocyclic rings

[0033] , ;and

[0034] Where R 1 and R 1' Together they form substituted or unsubstituted heterocycles, including but not limited to aza-butanes, pyrrolidines, piperidines, piperazines, morpholines, etc.

[0035] or ;

[0036] R 2 Each time it appears, it is independently selected from C2–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, and substituted or unsubstituted haloalkoxy.

[0037] R 3 Each time it appears, it is independently selected from H, C1–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, –C1–C6–OH, –C1–C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine;

[0038] R 4 Each time it appears, it is independently selected from alkylene groups interrupted by one or more heteroatoms, wherein the heteroatoms are O, S, NH or a combination thereof;

[0039] Y is independently selected from Ar2, –C(O)–Ar2, and –(CH2) each time it appears. a Ar2, –(CH2)3PO(OEt)2 or –CH2CO2Me;

[0040] Ar2 is independently selected each time it appears from: (i) substituted or unsubstituted 5- and 6-membered monocyclic aromatic or heterocyclic rings; (ii) substituted or unsubstituted 9- and 10-membered fused bicyclic rings comprising two monocyclic rings, wherein at least one of the two monocyclic rings is an aromatic or heterocyclic ring; and (iii) substituted or unsubstituted 13- and 14-membered fused tricyclic rings comprising three monocyclic rings together, wherein at least one of the three monocyclic rings is an aromatic or heterocyclic ring.

[0041] Each Ar2 is independently transmitted via G 1 G 2 G 3 G 4 and G 5 Replace, of which:

[0042] When Ar2 is monosubstituted, G 1 Each time it appears, it is independently selected from oxo, C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–C(R) 1 (R) 1' (R) 3 –NH2, –NO2, –SO3R 3 SO3O – 、–SO3N(H)(R 1 ), –E–C(O)R 3 , –E–CO2H, –B(OH)2, –C(O)NR 1 R 1' –E–PO(OR) 1 )2, and through G 2 G 3 G 4 and G 5 Substituted aryl groups;

[0043] G 2 G 3 G 4 and G 5 Each time it appears, it is independently selected from groups that are absent or selected from groups containing the following: halogen, –CN, –NO2, –SO3R. 3 C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–(CH2) a C(O)R 3 , –E–CO2H, –E–CHO, –E–C(O)R 3 , –E–C(O)NH(OH), –E–C(O)NHR1 –E–C(O)N(H)C(H)(R 1 (R) 1' ), –E–C(O)N(R 1 (CH2) a P(O)(O)OH)2、–E–C(O)N(R 1 (CH2) a SO3H), –E–C(O)NR 1 R 1' –E–NR 1 R 1' –E–OR 2 –C(O)–heterocyclic alkyl (where the heterocyclic alkyl is substituted or unsubstituted), –C(O)–N(H)–CH(COOH)((CH2) a – heteroaryl) (wherein the heteroaryl group is substituted or unsubstituted), wherein E is independently selected from direct bond and C1–C6 alkylene group each time it appears.

[0044] In some implementations, the compound of formula (I) is in the form of a salt.

[0045] In some embodiments, Z is not a triazole. For example, in some embodiments, Z is not a 1,2,3-triazole or a 1,2,4-triazole.

[0046] In some embodiments, Z is a diazole (e.g., imidazole, pyrazole, pyrimidine, pyridazine, pyrazine). In other embodiments, Z is a pyrrole, thiophene, or triazole (e.g., 1,2,3-triazole; 1,2,4-triazole). In still other embodiments, Z is a thiazole. In yet still other embodiments, Z is an isoxazole. In still further embodiments, Z is a tetraazole.

[0047] In some implementations, each Z is independently replaced by one "M" portion. In other implementations, each Z is independently replaced by two "M" portions. In still other implementations, each Z is independently replaced by three "M" portions.

[0048] In some embodiments, L is a heteroalkylene group with a length of 2 to 10 carbon atoms. In other embodiments, L is a heteroalkylene group with a length of 2 to 10 carbon atoms, wherein one or more carbon atoms are replaced by at least one heteroatom selected from oxygen, nitrogen, and sulfur.

[0049] Non-limiting examples of suitable L groups bridging two Ar1 groups are shown below:

[0050]

[0051] Non-limiting examples of compounds having formula (I) are shown in Table 1 of this document.

[0052] Another aspect of this disclosure is a polymer, copolymer, or metal gel having any of the compounds of formula (I).

[0053] Another aspect of this disclosure is a composition comprising molecules of any one of two or more compounds having formula (I).

[0054] Another aspect of this disclosure is a compound having the formula (IC):

[0055] (IC),

[0056] Or its solvates, hydrates, tautomers, chelates or salts, wherein:

[0057] a is 0 or an integer in the range of 1 to 4;

[0058] a' is 0 or an integer in the range of 1 to 4;

[0059] m is 1, 2, or 3;

[0060] m' is 1, 2, or 3;

[0061] n is 0, 1, or 2;

[0062] p is 0, 1, or 2;

[0063] When X is C, W is N, or when X is N, W is C;

[0064] It is a single bond or a double bond, wherein the double bond begins at either W or X where the carbon is present;

[0065] L is a linking group;

[0066] M is independently selected from hydrogen, halogens, and C1–C4 alkyl groups each time it appears;

[0067] Ar1, each time it appears, is independently selected from optionally substituted phenyl, pyridine, bipyridine, tripyridine, pyrazine, pyridazine, furan, dibenzofuran, thiophene, pyrrole, selenophene, naphthalene, fluorene, phenanthrene, cyclophosphine, phthalazine, quinazoline, quinoxaline, naphthidine, phenanthrene-rholine, purine, and carbazole.

[0068] The substituents of Ar1 are independently selected each time they appear from halogens, –OH, –CN, –NO2, C1–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, C1–C6 cycloalkyl, –OR 0 –CONH2, –C(O)NR 1 R 1' –C(O)(CH2) a NR 1 R 1' –NR 1 R 1' –NR 1 C(O)R 3 –C(O)SR 3 –COR 3 –CO(CH2) a OC(O)R 3 –OC(O)R 3 –C(O)OR 3 –C–O–R 3 , thiols, -R 4 -H、–SOR 1 –S(O)2R 1 –S(O)2NR 1 R 1' –CH2–NR 1 S(O)2R 3 –NR 1 S(O)2R 3 and –C(CH3)=N–(phenyl)–O–CH2–C–CH;

[0069] R 0 Each time it appears, it is independently selected from C1–C6 alkyl, C1–C6 haloalkyl, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl.

[0070] R 1 and R 1' Each time it appears, it is independently selected from H, hydroxyl, C1–C6 alkyl, C1–C6 haloalkyl, C1–C 20 Heteroalkyl, C1–C 10 Heteroalkyl –NH2, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic (e.g., via one or more R... 3Substituent groups), substituted or unsubstituted aryl groups, substituted or unsubstituted arylalkyl groups, and substituted or unsubstituted heteroaryl groups, benzyl groups, –C(=NH)NH2, –CH2CO2R 0 –C(H)(CH) a (COOH))2, –CH2C(O)NHCH2CO2H, –CH2CH2OH, –CH2CH2NHC(O)R 3 –(CH2) a C(O)NH(CH2) a C(O)OR 3 –(CH2) a –CN, spermine, –(CH2) a –NH2、–C(O)R 3 –(CH2) a C(O)OR 3 –(CH2) a OR 3 –C(H)((CH2) a OH) a' –CHCH(OH)(CH2) a OH, –(CH2) a OH, C1–C6–C(O)OH, can be substituted (e.g., via R...) 3 (Substituted group) or unsubstituted –(CH2) a - Heterocyclic rings

[0071]

[0072] Where R 1 and R 1' Together they form substituted or unsubstituted heterocycles, including but not limited to aza-butanes, pyrrolidines, piperidines, piperazines, morpholines, etc.

[0073]

[0074] R 2 Each time it appears, it is independently selected from C2–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, and substituted or unsubstituted haloalkoxy.

[0075] R 3Each time it appears, it is independently selected from H, C1–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, –C1–C6–OH, –C1–C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine;

[0076] R 4 Each time it appears, it is independently selected from alkylene groups interrupted by one or more heteroatoms, wherein the heteroatoms are O, S, NH or a combination thereof;

[0077] Y is independently selected from Ar2, –C(O)–Ar2, and –(CH2) each time it appears. a Ar2, –(CH2)3PO(OEt)2 or –CH2CO2Me;

[0078] Ar2 is independently selected each time it appears from: (i) substituted or unsubstituted 5- and 6-membered monocyclic aromatic or heterocyclic rings; (ii) substituted or unsubstituted 9- and 10-membered fused bicyclic rings comprising two monocyclic rings, wherein at least one of the two monocyclic rings is an aromatic or heterocyclic ring; and (iii) substituted or unsubstituted 13- and 14-membered fused tricyclic rings comprising three monocyclic rings together, wherein at least one of the three monocyclic rings is an aromatic or heterocyclic ring.

[0079] Each Ar2 is independently transmitted via G 1 G 2 G 3 G 4 and G 5 Replace, of which:

[0080] When Ar2 is monosubstituted, G 1 Each time it appears, it is independently selected from oxo, C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–C(R) 1 (R) 1' (R) 3 –NH2, –NO2, –SO3R 3 SO3O – 、–SO3N(H)(R 1 ), –E–C(O)R 3 , –E–CO2H, –B(OH)2, –C(O)NR 1 R 1' –E–PO(OR) 1 )2, and through G 2 G 3 G 4 and G 5 Substituted aryl groups;

[0081] G 2 G 3 G 4 and G 5 Each time it appears, it is independently selected from groups that are absent or selected from groups containing the following: halogen, –CN, –NO2, –SO3R. 3 C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–(CH2) a C(O)R 3 , –E–CO2H, –E–CHO, –E–C(O)R 3 , –E–C(O)NH(OH), –E–C(O)NHR 1 –E–C(O)N(H)C(H)(R 1 (R) 1' ), –E–C(O)N(R 1 (CH2) a P(O)(O)OH)2、–E–C(O)N(R 1 (CH2) a SO3H), –E–C(O)NR 1 R 1' –E–NR 1 R 1' –E–OR 2 –C(O)–heterocyclic alkyl (where the heterocyclic alkyl is substituted or unsubstituted), –C(O)–N(H)–CH(COOH)((CH2) a – heteroaryl) (wherein the heteroaryl group is substituted or unsubstituted), wherein E is independently selected from direct bond and C1–C6 alkylene group each time it appears.

[0082] Another aspect of this disclosure is a polymer, copolymer, or metal gel of any of the compounds having formula (IC).

[0083] Another aspect of this disclosure is a composition comprising molecules of any one of two or more compounds having formula (IC).

[0084] Another aspect of this disclosure is a method for enhancing a nucleic acid polymerase reaction, the method comprising the steps of: forming a nucleic acid polymerase reaction composition comprising a template nucleic acid, a nucleic acid polymerase, a mixture of nucleotides and / or nucleotide analogs, and at least one PEM (such as those compounds having any of the formulas (I), (IA), (IB), (IC), and / or (ID) as shown herein); and incubating the nucleic acid polymerase reaction composition under conditions that allow for nucleic acid polymerization. It is believed that the at least one PEM increases the sustained synthetic capacity, rate, and / or fidelity of the nucleic acid polymerase reaction. In some embodiments, the at least one PEM increases the length of the resulting nucleic acid product compared to a nucleic acid polymerase reaction lacking the at least one PEM.

[0085] In some embodiments, the nucleic acid polymerase is a DNA polymerase. In some embodiments, the DNA polymerase is DPO4 or a variant thereof. In other embodiments, the mixture of nucleotides or nucleotide analogs is a mixture of nucleotide analogs comprising nucleoside triaminophosphate, wherein each nucleoside triaminophosphate comprises a nucleobase selected from the group consisting of adenine, guanine, thymine, and cytosine, and a polymer chain portion, wherein a first end of the polymer chain portion is attached to the nucleobase, and a second end of the polymer chain portion is attached to the α-phosphate of the nucleoside triaminophosphate to provide nucleotide analog expansion by cleavage of the aminophosphate bond.

[0086] In some embodiments, the nucleic acid polymerization reaction produces a scalable polymer of nucleotide analogs, wherein the scalable polymer encodes the nucleobase sequence information of the template nucleic acid. In other embodiments, the conditions allowing the nucleic acid polymerization reaction include a suitable polymerization buffer and oligonucleotide primers. In a further embodiment, the suitable buffer comprises one or more components selected from MnCl2, buffer solutions, salts, sugars, single-stranded binding proteins (SSBs), imidazoles, pyrazoles, triazoles, betaine, molecular crowding agents, dimethyl sulfoxide (DMSO), alkyl glycols, glycerols, N-methyl-2-pyrrolidone (NMP), acetamide, butylated hydroxyanisole (BHA), polyphosphates, and / or ureas. In other embodiments, the reaction mixture further comprises a nucleic acid inserter. In other embodiments, the reaction mixture further comprises a polyanion recognition moiety. In a further embodiment, the mixture of nucleotides or nucleotide analogs comprises a nucleotide analog containing a detectable marker. In yet another embodiment, the detectable marker is an optically detectable marker selected from the group consisting of luminescent, chemiluminescent, fluorescent, fluorescent, chromogenic, or chromogenic markers.

[0087] Another aspect of this disclosure is the use of PEMs (including any of the compounds disclosed herein having any of the formulas (I), (IA), (IB), (IC), and / or (ID) shown herein) in enhancing nucleic acid polymerase reactions.

[0088] Another aspect of this disclosure is a composition comprising at least one PEM (including any of the compounds disclosed herein having any of the formulas (I), (IA), (IB), (IC), and / or (ID) shown herein) and a mixture of nucleotide analogs. This composition is believed to be useful, for example, when combined with a polymerase, wherein the at least one PEM increases the quantity and accuracy of nucleotide analogs incorporated into daughter chains during template-dependent polymerization reactions relative to the presence of the at least one PEM. In other embodiments, the at least one PEM comprises multiple PEMs.

[0089] Optionally, the mixture of nucleotide analogs comprises nucleoside triaminophosphates, wherein each nucleoside triaminophosphate comprises a nucleobase selected from the group consisting of adenine, guanine, thymine, and cytosine, and a polymer chain moiety, wherein a first end of the polymer chain moiety is attached to a nucleobase, and a second end of the polymer chain moiety is attached to an α-phosphate of the nucleoside triaminophosphate to provide nucleotide analog expansion by cleavage of the aminophosphate bond. In other embodiments, the composition further comprises a buffer containing one or more components selected from MnCl2, buffer solutions, salts, sugars, single-chain binding proteins (SSBs), imidazoles, pyrazoles, triazoles, betaine, molecular congestion agents, dimethyl sulfoxide (DMSO), alkyl glycols, glycerols, N-methyl-2-pyrrolidone (NMP), acetamide, butylated hydroxyanisole (BHA), polyphosphates, and / or ureas. In other embodiments, the composition further comprises single-chain binding proteins (SSBs). In other embodiments, the composition further comprises urea. In some embodiments, the mixture of nucleotide analogs includes nucleotide analogs containing a detectable marker. In some embodiments, the detectable marker is an optically detectable marker selected from the group consisting of luminescent, chemiluminescent, fluorescent, fluorescent, chromogenic, or chromogenic markers.

[0090] Another aspect of this disclosure is a method for sequencing a DNA or RNA template, the method comprising the steps of: forming a DNA polymerase reaction composition comprising a DNA or RNA template, a replication primer complexed with the template, a DNA polymerase, a mixture of nucleotides or nucleotide analogs, and at least one PEM (including any of the compounds disclosed herein having any of the formulas (I), (IA), (IB), (IC), and / or (ID) shown herein); and incubating the DNA polymerase reaction composition under conditions allowing DNA polymerization, wherein the at least one PEM enhances the rate, fidelity, or sustained synthesis capability of the DNA polymerase reaction. In some embodiments, the method may further comprise determining the sequence of the nucleotide or nucleotide analog in the resulting nucleotide or nucleotide analog polymer. In some embodiments, the at least one PEM is a compound having any of the formulas (I), (IA), (IB), (IC), and (ID). In other embodiments, the at least one PEM is any of the compounds shown herein in Table 1. In other embodiments, the mixture of nucleotide analogs comprises nucleoside triaminophosphate, wherein each nucleoside triaminophosphate comprises a nucleobase selected from the group consisting of adenine, guanine, thymine, and cytosine, and a polymer chain portion, wherein a first end of the polymer chain portion is attached to a nucleobase, and a second end of the polymer chain portion is attached to an α-phosphate of the nucleoside triaminophosphate to provide nucleotide analog expansion by cleavage of the aminophosphate bond. In other embodiments, the DNA polymerase is DPO4 or a variant thereof. In other embodiments, the resulting polymer of nucleotide analog is a scalable polymer. In other embodiments, the method further comprises the step of contacting the scalable polymer with an aminophosphate cleaving agent to produce an expanded polymer of the nucleotide analog. In some embodiments, the polymer chain portion of each nucleotide analog comprises a reporter portion specific to the nucleobase of the analog. In other embodiments, the reporter portion generates a characteristic electronic signal. In yet another embodiment, the step of determining the nucleotide analog sequence comprises the step of translocating the expanded polymer of the nucleotide analog through a nanopore.

[0091] Another aspect of this disclosure is the use of PEMs (such as those compounds having any of the formulas (I), (IA), (IB), (IC), and / or (ID) shown herein) in sequencing DNA or RNA templates.

[0092] Another aspect of this disclosure is a composition comprising PEM (such as those compounds having any of the formulas (I), (IA), (IB), (IC) and / or (ID) shown herein) and a polynucleotide.

[0093] Another aspect of this disclosure is a composition comprising PEM (such as those compounds having any of the formulas (I), (IA), (IB), (IC) and / or (ID) shown herein) and polypeptides (e.g., polypeptides such as enzymes, wherein the enzyme may be a nucleic acid polymerase).

[0094] The following are exemplary, non-limiting numbering embodiments of this disclosure. Furthermore, unless otherwise specifically stated, each atom identified in a chemical formula can be any isotope of that atom. For example, the name C (carbon) includes... 12 C 13 C or 14 C and its mixtures, especially mixtures of naturally occurring abundance isotopes, while H (hydrogen) includes 1 H, 2 H and 3 H and its mixtures, and O (oxygen) includes 16 O and 18 O and its mixtures, and N (nitrogen) includes 14 N and 15 N and its mixtures, etc. Other atoms are similar.

[0095] Implementation Scheme 1) A compound having formula (I):

[0096] (I),

[0097] Or its solvates, hydrates, tautomers, chelates or salts,

[0098] in

[0099] a is 0 or an integer in the range of 1 to 4;

[0100] a' is 0 or an integer in the range of 1 to 4;

[0101] m is 1, 2, or 3;

[0102] m' is 1, 2, or 3;

[0103] n is 0, 1, or 2;

[0104] p is 0, 1, or 2;

[0105] Z is a 5- or 6-membered heteroaromatic ring containing one, two, three or four heteroatoms selected from O, N or S, wherein each Z may be independently substituted with one or more halogens and / or one or more C1-C4 alkyl groups;

[0106] L is a linking group;

[0107] M is independently selected from hydrogen, halogens, and C1-C4 alkyl groups each time it appears;

[0108] Ar1, each time it appears, is independently selected from optionally substituted phenyl, pyridine, bipyridine, tripyridine, pyrazine, pyridazine, furan, dibenzofuran, thiophene, pyrrole, selenophene, naphthalene, fluorene, phenanthrene, cyclophosphine, phthalazine, quinazoline, quinoxaline, naphthidine, phenanthrene-rholine, purine, and carbazole.

[0109] The substituents of Ar1 are independently selected each time they appear from halogens, –OH, –CN, –NO2, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 heteroalkyl, C1-C6 cycloalkyl, –OR 0 –CONH2, –C(O)NR 1 R 1' –C(O)(CH2) a NR 1 R 1' –NR 1 R 1' –NR 1 C(O)R 3 –C(O)SR 3 –COR 3 –CO(CH2) a OC(O)R 3 –OC(O)R 3 –C(O)OR 3 –C–O–R 3 , thiols, -R 4 -H、–SOR 1 –S(O)2R 1 –S(O)2NR 1 R 1' –CH2–NR 1 S(O)2R 3 –NR 1 S(O)2R 3 and –C(CH3)=N–(phenyl)-O-CH2-C-CH;

[0110] R 0Each time it appears, it is independently selected from C1-C6 alkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl.

[0111] R 1 and R 1' Each time it appears, it is independently selected from H, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C 20 Heteroalkyl, C1-C 10 Heteroalkyl –NH2, C2-C6 alkenyl, C2-C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic (e.g., via one or more R... 3 Substituent groups), substituted or unsubstituted aryl groups, substituted or unsubstituted arylalkyl groups, and substituted or unsubstituted heteroaryl groups, benzyl groups, –C(=NH)NH2, –CH2CO2R 0 –C(H)(CH) a (COOH))2, -CH2C(O)NHCH2CO2H, -CH2CH2OH, -CH2CH2NHC(O)R 3 –(CH2) a C(O)NH(CH2) a C(O)OR 3 –(CH2) a –CN, spermine, –(CH2) a –NH2、–C(O)R 3 –(CH2) a C(O)OR 3 –(CH2) a OR 3 –C(H)((CH2) a OH) a' –CHCH(OH)(CH2) a OH, –(CH2) a OH, C1-C6–C(O)OH, can be substituted (e.g., via R) 3 (Substituted group) or unsubstituted –(CH2) a - Heterocyclic rings

[0112] , ;and

[0113] Where R 1 and R 1'Together they form substituted or unsubstituted heterocycles, including but not limited to aza-butanes, pyrrolidines, piperidines, piperazines, morpholines, etc.

[0114] or ;

[0115] R 2 Each time it appears, it is independently selected from C2-C6 alkyl, C1-C6 haloalkyl, C1-C6 heteroalkyl, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, and substituted or unsubstituted haloalkoxy.

[0116] R 3 Each time it appears, it is independently selected from H, C1-C6 alkyl, C1-C6 haloalkyl, C1–C6 heteroalkyl, –C1–C6–OH, –C1–C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine;

[0117] R 4 Each time it appears, it is independently selected from alkylene groups interrupted by one or more heteroatoms, wherein the heteroatoms are O, S, NH or a combination thereof;

[0118] Y is independently selected from Ar2, –C(O)–Ar2, and –(CH2) each time it appears. a Ar2, –(CH2)3PO(OEt)2 or –CH2CO2Me;

[0119] Ar2 is independently selected each time it appears from: (i) substituted or unsubstituted 5- and 6-membered monocyclic aromatic or heterocyclic rings; (ii) substituted or unsubstituted 9- and 10-membered fused bicyclic rings comprising two monocyclic rings, wherein at least one of the two monocyclic rings is an aromatic or heterocyclic ring; and (iii) substituted or unsubstituted 13- and 14-membered fused tricyclic rings comprising three monocyclic rings together, wherein at least one of the three monocyclic rings is an aromatic or heterocyclic ring.

[0120] Each Ar2 is independently transmitted via G 1 G 2 G 3 G 4 and G 5 Replace, of which:

[0121] When Ar2 is monosubstituted, G 1Each time it appears, it is independently selected from oxo, C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–C(R) 1 (R) 1' (R) 3 –NH2, –NO2, –SO3R 3 SO3O – 、–SO3N(H)(R 1 ), –E–C(O)R 3 , –E–CO2H, –B(OH)2, –C(O)NR 1 R 1' –E–PO(OR) 1 )2, and through G 2 G 3 G 4 and G 5 Substituted aryl groups;

[0122] G 2 G 3 G 4 and G 5 Each time it appears, it is independently selected from groups that are absent or selected from groups containing the following: halogen, –CN, –NO2, –SO3R. 3 C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–(CH2) a C(O)R 3 , –E–CO2H, –E–CHO, –E–C(O)R 3 , –E–C(O)NH(OH), –E–C(O)NHR 1 –E–C(O)N(H)C(H)(R 1 (R) 1' ), –E–C(O)N(R 1 (CH2) a P(O)(O)OH)2、–E–C(O)N(R 1 (CH2) a SO3H), –E–C(O)NR 1 R 1' –E–NR 1 R 1' –E–OR 2 –C(O)–heterocyclic alkyl (where the heterocyclic alkyl is substituted or unsubstituted), –C(O)–N(H)–CH(COOH)((CH2) a– heteroaryl) (wherein the heteroaryl group is substituted or unsubstituted), wherein E is independently selected from direct bond and C1–C6 alkylene group each time it appears.

[0123] Implementation Scheme 2) The compound according to Implementation Scheme 1, wherein the compound of formula (I) has either formula (IA) or (IB):

[0124] (IA) or

[0125] (IB),

[0126] Or its salts, solvates, hydrates, tautomers, or chelates,

[0127] Ar1, Ar2, Z, and Y are as defined above.

[0128] Implementation Scheme 3: The compound according to Implementation Scheme 1, wherein the compound of formula (I) has formula (IC):

[0129] (IC),

[0130] Or its solvates, hydrates, tautomers, chelates or salts,

[0131] in

[0132] When X is C, W is N, or when X is N, W is C;

[0133] It is a single bond or a double bond, wherein the double bond begins at either W or X where the carbon atom is present; and

[0134] m, m', n, p, L, and Y are as defined above.

[0135] Implementation Scheme 4) The compound according to any one of Implementation Schemes 1 to 3, wherein Ar1 is a monocyclic heterocyclic aryl group.

[0136] Implementation Scheme 5) The compound according to any one of Implementation Schemes 1 to 3, wherein Ar1 is selected from:

[0137]

[0138] In formula (I), (IA) or (IB), each group “Z” is located at position “k” on Ar1; or in formula (IC), each triazole is located at position “k” on Ar1.

[0139] Implementation Scheme 6) The compound according to any one of Implementation Schemes 1 to 3, wherein Ar1 is a bicyclic aryl group.

[0140] Implementation Scheme 7) The compound according to any one of Implementation Schemes 1 to 3, wherein Ar1 is a bicyclic carbocyclic aryl group selected from:

[0141] , , , , , and

[0142] In formula (I), (IA) or (IB), each group “Z” is located at position “k” on Ar1; or in formula (IC), each triazole is located at position “k” on Ar1.

[0143] Implementation Scheme 8) The compound according to any one of Implementation Schemes 1 to 3, wherein Ar1 is a bicyclic heterocyclic aryl group selected from:

[0144] , , , , , and

[0145] In formula (I), (IA) or (IB), each group “Z” is located at position “k” on Ar1; or in formula (IC), each triazole is located at position “k” on Ar1.

[0146] Implementation Scheme 9) The compound according to any one of Implementation Schemes 1 to 3, wherein Ar1 is a tricyclic aryl group.

[0147] Implementation Scheme 10) The compound according to Implementation Scheme 9, wherein the tricyclic aryl group is selected from:

[0148] and .

[0149] Implementation Scheme 11) The compound according to Implementation Scheme 9, wherein the tricyclic aryl group is selected from:

[0150] , , , , , and ,

[0151] In formula (I), (IA) or (IB), each group “Z” is located at position “k” on Ar1; or in formula (IC), each triazole is located at position “k” on Ar1.

[0152] Implementation Scheme 12) The compound according to Implementation Scheme 1, wherein Ar1 is a tricyclic heteroaryl group selected from...

[0153] , , and

[0154] In formula (I), (IA) or (IB), each group “Z” is located at position “k” on Ar1; or in formula (IC), each triazole is located at position “k” on Ar1.

[0155] Implementation Scheme 13) The compound according to any one of the preceding embodiments, wherein Ar2 is a substituted or unsubstituted 5-membered monocyclic aromatic ring selected from the group consisting of: thiophene, 1,2-thiazole, 1,3-thiazole, furan, 1,2-oxazole, 1,3-oxazole, 1H-pyrrole, 1H-pyrazole, oxadiazole, thiadiazole, 1,2,4-triazole, 1,2,3-triazole and 1H-imidazole.

[0156] Implementation Scheme 14) The compound according to any one of Implementation Schemes 1 to 12, wherein Ar2 is a substituted or unsubstituted 6-membered monocyclic aromatic ring selected from the group consisting of benzene, pyridine, pyridazine, pyrimidine and pyrazine.

[0157] Implementation Scheme 15) The compound according to any one of Implementation Schemes 1 to 12, wherein Ar2 is a substituted or unsubstituted 9-membered fused bicyclic aromatic ring system selected from the group consisting of: benzofuran, 1,3-benzoxazole, furano[3,2-b]pyridine, furano[3,2-c]pyridine, furano[2,3-c]pyridine, furano[2,3-b]pyridine, indole, 1H-benzimidazole, 1H-pyrrolo[3,2-b]pyridine ... [c]pyridine, 1H-pyrrolo[2,3-c]pyridine, 1H-pyrrolo[2,3-b]pyridine, benzothiophene, 1,3-benzothiazole, thieno[3,2-b]pyridine, thieno[3,2-c]pyridine, thieno[2,3-c]pyridine, benzoxadiazole, benzothiadiazole, benzoisoxazole, benzotriazole and thieno[2,3-b]pyridine.

[0158] Implementation Scheme 16) The compound according to any one of Implementation Schemes 1 to 12, wherein Ar2 is a substituted or unsubstituted 10-membered fused bicyclic aromatic ring system selected from the group consisting of: naphthylene, quinoline, quinazoline, quinoxaline, 1,5-naphthidine, 1,6-naphthidine, 1,7-naphthidine, 1,8-naphthidine, isoquinoline, phthalazine, 2,6-naphthidine and 2,7-naphthidine.

[0159] Implementation Scheme 17) The compound according to any one of Implementation Schemes 1 to 12, wherein Ar2 is a substituted or unsubstituted pyridinyl ring selected from...

[0160] , and ;

[0161] The substituent G (where G can be G as defined above) 1 G 2 G 3 G 4 and G 5 (Any of the following) appears 0, 1 or 2 times on the pyridyl ring.

[0162] Implementation Scheme 18) The compound according to any one of Implementation Schemes 1 to 12, wherein Ar2 is a phenyl ring having the following formula,

[0163]

[0164] The substituent G (or G as defined herein) 1 G 2 G 3 G 4 and G 5 The phenyl group appears 0, 1, or 2 times on the phenyl ring. In one embodiment, G is a derivative of G. 2 G 3 G 4 and G 5 Substituted aryl groups (e.g., phenyl).

[0165] Implementation Scheme 19) The compound according to any one of Implementation Schemes 1 to 12, wherein Ar2 is a phenyl ring selected from...

[0166]

[0167] Implementation Scheme 20) The compound according to Implementation Schemes 1 to 18, wherein Ar2 contains one or more amino substituents.

[0168] Implementation Scheme 21) The compound according to Implementation Schemes 1 to 18, wherein Ar2 contains one or more methoxy substituents.

[0169] Implementation Scheme 22) The compound according to Implementation Schemes 1 to 18, wherein Ar2 contains one or more carboxylic acid substituents.

[0170] Implementation Scheme 23) The compound according to Implementation Schemes 1 to 18, wherein Ar2 contains one or more –CH2–CO2–CH3 substituents.

[0171] Implementation Scheme 24) The compound according to Implementation Schemes 1 to 18, wherein Ar2 contains one or more trifluoromethyl substituents.

[0172] Implementation Scheme 25) The compound according to Implementation Schemes 1 to 18, wherein Ar2 contains one or more hydroxyl substituents.

[0173] Implementation Scheme 26) The compound according to Implementation Schemes 1 to 18, wherein Ar2 contains at least two substituents, wherein the first substituent is a carboxylic acid and the second substituent is a hydroxyl group.

[0174] Implementation Scheme 27) The compound according to Implementation Schemes 1 to 18, wherein Ar2 contains at least two substituents, wherein the first substituent is a carboxylic acid and the second substituent is a trifluoromethyl group.

[0175] Implementation Scheme 28) The compound according to Implementation Schemes 1 to 18, wherein Ar2 contains at least one sulfonic acid group (–SO3H) or a salt thereof (–SO2O) - ).

[0176] Implementation Scheme 29) The compound according to Implementation Schemes 1 to 19, wherein Ar2 comprises two substituents, wherein one of the two substituents is a sulfonic acid group (–SO3H) or a salt thereof (–SO2O). - ).

[0177] Implementation Scheme 30) The compound according to any one of the preceding implementation schemes, wherein the compound is in the form of a chelate.

[0178] Implementation Scheme 31) The compound according to Implementation Scheme 30, wherein the chelate is a copper chelate.

[0179] Implementation Scheme 32) The compound according to any one of the foregoing implementation schemes, wherein the compound is in the form of a salt.

[0180] Implementation Scheme 33) The compound according to any one of the preceding implementation schemes, wherein the compound has a logP of at least 4.9.

[0181] Implementation Scheme 34) The compound according to Implementation Scheme 1, wherein the compound has one of the following formulas:

[0182]

[0183] Or its salts or hydrates.

[0184] Implementation Scheme 35) The compound according to Implementation Scheme 1, wherein the compound has one of the following formulas:

[0185]

[0186] Or its salts or hydrates.

[0187] Implementation Scheme 36) The compound according to Implementation Scheme 1, wherein Ar2 contains at least two substituents selected from hydroxyl, carboxylic acid formamide and trifluoromethyl.

[0188] Implementation Scheme 37) The compound according to Implementation Scheme 1, wherein the compound has one of the following formulas:

[0189]

[0190] Or its salts or hydrates.

[0191] Implementation Scheme 38) The compound according to Implementation Scheme 1, wherein the compound has one of the following formulas:

[0192] , , , or .

[0193] Implementation Scheme 39) The compound according to Implementation Scheme 1, wherein the compound has one of the following formulas:

[0194]

[0195] Or its salts or hydrates.

[0196] Implementation Scheme 40) The compound according to Implementation Scheme 1, wherein the compound has one of the following formulas:

[0197] , or .

[0198] Implementation Scheme 41) The compound according to Implementation Scheme 1, wherein the compound has one of the following formulas:

[0199] , or .

[0200] Implementation Scheme 42) The compound according to Implementation Scheme 1, wherein the compound has one of the following formulas:

[0201] , , , , , or .

[0202] Implementation Scheme 43) The compound according to Implementation Scheme 1, wherein the compound has one of the following formulas:

[0203] , or .

[0204] Implementation Scheme 44) The compound according to Implementation Scheme 1, wherein the compound has one of the following formulas:

[0205] , or .

[0206] Implementation Scheme 45) The compound according to Implementation Scheme 1, wherein the compound has one of the following formulas:

[0207] , , or .

[0208] Implementation Scheme 46) The compound according to Implementation Scheme 1, wherein the compound has one of the following formulas:

[0209]

[0210] Or its salts or hydrates.

[0211] Implementation Scheme 47) The compound according to Implementation Scheme 1, wherein the compound has one of the following formulas:

[0212]

[0213] (Embodiment 48) A composition comprising: a compound according to any one of embodiments 1 to 47; and a molecular crowding agent.

[0214] Implementation Scheme 49) The composition according to Implementation Scheme 48, wherein the molecular crowding agent is a polyalkylene glycol.

[0215] (Embodiment 50) A composition comprising: a compound according to any one of embodiments 1 to 47; and an aqueous buffer solution.

[0216] Implementation Scheme 51) The composition according to Implementation Scheme 50, wherein the aqueous buffer is TrisHCl.

[0217] (Embodiment 52) ​​A composition comprising: a compound according to any one of embodiments 1 to 47; and a polynucleotide.

[0218] Implementation Scheme 53) The composition according to Implementation Scheme 52, wherein the polynucleotide is a 20-60mer oligonucleotide.

[0219] Implementation Scheme 54) The composition according to Implementation Scheme 52, wherein the polynucleotide is a 25-50mer oligonucleotide.

[0220] (Embodiment 55) A composition comprising: a compound according to any one of embodiments 1 to 47; and a protein.

[0221] Implementation Scheme 56) The composition according to Implementation Scheme 55, wherein the protein is a DNA polymerase.

[0222] (Embodiment 57) A composition comprising: a compound according to any one of embodiments 1 to 47; and a mixture of nucleotides or nucleotide analogs.

[0223] (Embodiment 58) A composition for enhancing the sustained synthesis capacity, fidelity, or rate of a DNA polymerase reaction, comprising: at least one compound according to any one of embodiments 1 to 47; and a mixture of nucleotide analogs.

[0224] (Embodiment 59) A composition comprising a mixture of at least one compound according to any one of embodiments 1 to 47 and a nucleotide analogue, wherein, relative to the same polymerization reaction in which at least one compound according to any one of embodiments 1 to 47 is absent, the at least one compound according to any one of embodiments 1 to 47 increases the quantity and accuracy of nucleotide analogues incorporated into daughter chains during template-dependent polymerization.

[0225] (Effective 60) The composition according to Effect 59, wherein the mixture of nucleotide analogs comprises nucleoside triaminophosphate, wherein each nucleoside triaminophosphate comprises a nucleobase selected from the group consisting of adenine, guanine, thymine, and cytosine, and a polymer chain portion, wherein a first end of the polymer chain portion is attached to the nucleobase, and a second end of the polymer chain portion is attached to the α-phosphate of the nucleoside triaminophosphate to provide nucleotide analog extension by cleavage of the aminophosphate bond.

[0226] Implementation Scheme 61) The composition according to Implementation Scheme 59 further comprises a buffer solution containing one or more components selected from MnCl2, buffer solution, salt, sugar, single-chain binding protein (SSB), imidazole, pyrazole, triazole, betaine, molecular crowding agent, dimethyl sulfoxide (DMSO), alkyl glycol, glycerol, N-methyl-2-pyrrolidone (NMP), acetamide, butylated hydroxyanisole (BHA), polyphosphate and / or urea.

[0227] Implementation Scheme 62) The composition according to Implementation Scheme 59 further comprises a single-chain binding protein.

[0228] (Scheme 63) The composition according to Scheme 59 further comprises urea.

[0229] Implementation Scheme 64) The composition according to Implementation Scheme 59, wherein the mixture of nucleotide analogs comprises a nucleotide analog containing a detectable marker.

[0230] (Scheme 65) The composition according to Scheme 64, wherein the detectable marker is an optically detectable marker selected from the group consisting of luminescent, chemiluminescent, fluorescent, fluorescent, chromogenic, or chromogenic markers.

[0231] Implementation Scheme 66) A kit for sequencing a nucleic acid template, the kit comprising at least one composition according to any one of Implementation Schemes 48 to 65.

[0232] Implementation Scheme 67) A method for enhancing nucleic acid polymerase reactions, the method comprising:

[0233] a. Forming a nucleic acid polymerase reaction composition comprising:

[0234] i. Template nucleic acid,

[0235] ii. Nucleic acid polymerase,

[0236] iii. A mixture of nucleotides or nucleotide analogs, and

[0237] iv. At least one compound according to any one of embodiments 1-47; and

[0238] b. Incubating a nucleic acid polymerase reaction composition under conditions that allow for nucleic acid polymerization, wherein the at least one compound according to any one of embodiments 1 to 47 increases the sustained synthesis capacity, rate, or fidelity of the nucleic acid polymerase reaction.

[0239] Implementation Scheme 68) The method according to Implementation Scheme 67, wherein the compound according to any one of Implementation Schemes 1 to 47 increases the length of the resulting nucleic acid product compared to a nucleic acid polymerase reaction lacking the compound according to any one of Implementation Schemes 1 to 47.

[0240] (Scheme 69) The method according to Scheme 67, wherein the at least one compound according to any one of Schemes 1 to 47 comprises a plurality of compounds according to any one of Schemes 1 to 47.

[0241] Implementation Scheme 70) The method according to Implementation Scheme 67, wherein the nucleic acid polymerase is a polymerase.

[0242] Implementation Scheme 71) The method according to Implementation Scheme 67, wherein the nucleic acid polymerase is a DNA polymerase.

[0243] Implementation Scheme 72) The method according to Implementation Scheme 71, wherein the DNA polymerase is DPO4 or a variant thereof.

[0244] Implementation Scheme 73) The method according to Implementation Scheme 67, wherein the mixture of nucleotides or nucleotide analogs is a mixture of nucleotide analogs comprising nucleoside triaminophosphate, wherein each nucleoside triaminophosphate comprises a nucleobase selected from the group consisting of adenine, guanine, thymine, and cytosine and a polymer chain portion, wherein a first end of the polymer chain portion is attached to the nucleobase and a second end of the polymer chain portion is attached to the α-phosphate of the nucleoside triaminophosphate to provide nucleotide analog extension by cleavage of the aminophosphate bond.

[0245] Implementation Scheme 74) The method according to Implementation Scheme 67, wherein the nucleic acid polymerization reaction produces an expandable polymer of nucleotide analogs, wherein the expandable polymer encodes the nucleobase sequence information of the template nucleic acid.

[0246] Implementation Scheme 75) The method according to Implementation Scheme 67, wherein the conditions for promoting the nucleic acid polymerization reaction include a suitable polymerization buffer and oligonucleotide primers.

[0247] Implementation Scheme 76) The method according to Implementation Scheme 67, wherein the suitable buffer comprises one or more components selected from MnCl2, buffer, salt, sugar, single-stranded binding protein (SSB), imidazole, pyrazole, triazole, betaine, molecular crowding agent, dimethyl sulfoxide (DMSO), alkyl diol, glycerol, N-methyl-2-pyrrolidone (NMP), acetamide, butylated hydroxyanisole (BHA), polyphosphate and / or urea.

[0248] Implementation Scheme 77) The method according to Implementation Scheme 67, wherein the reaction mixture further comprises a single-chain binding protein.

[0249] Implementation Scheme 78) The method according to Implementation Scheme 67, wherein the reaction mixture further comprises urea.

[0250] Implementation Scheme 79) The method according to Implementation Scheme 67, wherein the mixture of nucleotides or nucleotide analogs comprises a nucleotide analog containing a detectable marker.

[0251] Implementation Scheme 80) The method according to Implementation Scheme 73, wherein the detectable marker is an optically detectable marker selected from the group consisting of luminescent, chemiluminescent, fluorescent, fluorescent, chromogenic, or chromogenic markers.

[0252] Implementation Scheme 81) A method for sequencing a DNA or RNA template, the method comprising the following steps:

[0253] a. Forming a DNA polymerase reaction composition comprising:

[0254] i. DNA or RNA template,

[0255] ii. Replication primers compounded with the template,

[0256] iii. DNA polymerase,

[0257] iv. A mixture of nucleotides or nucleotide analogs,

[0258] v. At least one compound according to any one of embodiments 1-43,

[0259] b. Incubating the DNA polymerase reaction composition under conditions that allow DNA polymerization, wherein at least one compound according to any one of embodiments 1 to 47 increases the rate, fidelity, or sustained synthesis capability of the DNA polymerase reaction; and

[0260] c. Determine the sequence of the nucleotide or nucleotide analog in the resulting nucleotide or nucleotide analog polymer.

[0261] Implementation Scheme 82) The method according to Implementation Scheme 81, wherein the mixture of nucleotide analogs comprises nucleoside triaminophosphate, wherein each nucleoside triaminophosphate comprises a nucleobase selected from the group consisting of adenine, guanine, thymine, and cytosine, and a polymer chain portion, wherein a first end of the polymer chain portion is connected to the nucleobase, and a second end of the polymer chain portion is connected to the α-phosphate of the nucleoside triaminophosphate to provide nucleotide analog extension by cleavage of the aminophosphate bond.

[0262] Implementation Scheme 83) The method according to any one of Implementation Schemes 81 or 82, wherein the DNA polymerase is DPO4 or a variant thereof.

[0263] Implementation Scheme 84) The method according to any one of Implementation Schemes 81 or 82, wherein the polymer of the resulting nucleotide analog is a scalable polymer.

[0264] (Embodiment 85) The method according to any one of embodiments 81 further includes the step of contacting the expandable polymer with an aminophosphate cleaving agent to produce an expanded polymer of nucleotide analogs.

[0265] (Effective Implementation 86) The method according to any one of Effects 81 or 82, wherein the polymer chain portion of each of the nucleotide analogs comprises a reporter portion characteristic of the nucleobases of the analog.

[0266] Implementation Scheme 87) The method according to Implementation Scheme 86, wherein the reporting portion generates a characteristic electronic signal.

[0267] Implementation Scheme 88) The method according to Implementation Scheme 81, wherein the step of determining the nucleotide analog sequence includes the step of shifting the extended polymer of the nucleotide analog through a nanopore.

[0268] (Embodiment 89) A polymer, copolymer or oligomer derived from one or more of the compounds according to any one of embodiments 1 to 47.

[0269] (Embodiment 90) A composition comprising one or more of the compounds according to any one of embodiments 1 to 47.

[0270] (Embodiment 91) The composition according to embodiment 90, wherein the composition comprises at least two molecules of a compound having any one of embodiments 1 to 47.

[0271] Implementation Scheme 92) A compound having the following structure:

[0272] ,

[0273] in

[0274] a is 0 or an integer in the range of 1 to 4;

[0275] a' is 0 or an integer in the range of 1 to 4;

[0276] Z is a 5- or 6-membered heteroaromatic ring containing one, two, three or four heteroatoms selected from O, N or S;

[0277] Ar1, each time it appears, is independently selected from optionally substituted phenyl, pyridine, bipyridine, tripyridine, pyrazine, pyridazine, furan, dibenzofuran, thiophene, pyrrole, selenophene, naphthalene, fluorene, phenanthrene, cyclophosphine, phthalazine, quinazoline, quinoxaline, naphthidine, phenanthrene-rholine, purine, and carbazole.

[0278] The substituents of Ar1 are independently selected each time they appear from halogens, –OH, –CN, –NO2, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 heteroalkyl, C1-C6 cycloalkyl, –OR0 –CONH2, –C(O)NR 1 R 1' –C(O)(CH2) a NR 1 R 1' –NR 1 R 1' –NR 1 C(O)R 3 –C(O)SR 3 –COR 3 –CO(CH2) a OC(O)R 3 –OC(O)R 3 –C(O)OR 3 –C–O–R 3 , thiols, -R 4 -H、–SOR 1 –S(O)2R 1 –S(O)2NR 1 R 1' –CH2–NR 1 S(O)2R 3 –NR 1 S(O)2R 3 and –C(CH3)=N–(phenyl)-O-CH2-C-CH;

[0279] R 0 Each time it appears, it is independently selected from C1-C6 alkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl.

[0280] R 1 and R 1' Each time it appears, it is independently selected from H, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C 20 Heteroalkyl, C1-C 10 Heteroalkyl –NH2, C2-C6 alkenyl, C2-C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic (e.g., via one or more R... 3 Substituent groups), substituted or unsubstituted aryl groups, substituted or unsubstituted arylalkyl groups, and substituted or unsubstituted heteroaryl groups, benzyl groups, –C(=NH)NH2, –CH2CO2R 0 –C(H)(CH) a(COOH))2, -CH2C(O)NHCH2CO2H, -CH2CH2OH, -CH2CH2NHC(O)R 3 –(CH2) a C(O)NH(CH2) a C(O)OR 3 –(CH2) a –CN, spermine, –(CH2) a –NH2、–C(O)R 3 –(CH2) a C(O)OR 3 –(CH2) a OR 3 –C(H)((CH2) a OH) a' –CHCH(OH)(CH2) a OH, –(CH2) a OH, C1-C6–C(O)OH, can be substituted (e.g., via R) 3 (Substituted group) or unsubstituted –(CH2) a - Heterocyclic rings

[0281] , ;and

[0282] Where R 1 and R 1' Together they form substituted or unsubstituted heterocycles, including but not limited to aza-butanes, pyrrolidines, piperidines, piperazines, morpholines, etc.

[0283] or ;

[0284] R 3 Each time it appears, it is independently selected from H, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 heteroalkyl, –C1-C6–OH, –C1-C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine;

[0285] R 4 Each time it appears, it is independently selected from alkylene groups interrupted by one or more heteroatoms, wherein the heteroatoms are O, S, NH or a combination thereof;

[0286] G 1Each time it appears, it is independently selected from oxo, C1-C6 alkyl, C1-C6 haloalkyl, –E–O–R 3 –E–C(R) 1 (R) 1' (R) 3 –NH2, –NO2, –SO3R 3 SO3O - 、–SO3N(H)(R 1 ), –E–C(O)R 3 , -E-CO2H, –B(OH)2, –C(O)NR 1 R 1' -E-PO(OR) 1 )2, and through G 2 G 3 G 4 and G 5 Substituted aryl; and

[0287] G 2 G 3 G 4 and G 5 Each time it appears, it is independently selected from groups that are absent or selected from groups containing the following: halogen, –CN, –NO2, –SO3R. 3 C1-C6 alkyl, C1-C6 haloalkyl, –E–O–R 3 –E–(CH2) a C(O)R 3 , –E–CO2H, –E–CHO, –E–C(O)R 3 , –E–C(O)NH(OH), –E–C(O)NHR 1 –E–C(O)N(H)C(H)(R 1 (R) 1' ), –E–C(O)N(R 1 (CH2) a P(O)(O)OH)2、–E–C(O)N(R 1 (CH2) a SO3H), –E–C(O)NR 1 R 1' –E–NR 1 R 1' –E–OR 2 –C(O)–heterocyclic alkyl (where the heterocyclic alkyl is substituted or unsubstituted), –C(O)–N(H)–CH(COOH)((CH2) a– heteroaryl) (wherein the heteroaryl group is substituted or unsubstituted), wherein E is independently selected from direct bond and C1-C6 alkylene group each time it appears.

[0288] Implementation Scheme 93) The compound according to Implementation Scheme 92, wherein each substituent of Ar1 is independently selected from –C(O)-CH3, –C(O)-NH2, –C(O)O-CH2CH3, -CF3, –C(O)O-tert-butyl, –C(O)-N(CH3)2, –C(O)-N(H)CH3, –C(O)-N(H)CH2CH3, –C(O)-N(H) (cycloalkyl), –C(O)–N(H) (CH2)4, –C(O)–N(CH2CH3)2, –C(O)–N(H) (tert-butyl), –C(O)–N(H) (C6-cycloalkyl), –C(O)–N(H) (CH2)3, –C(O)–N(H) (C5-cycloalkyl), –C(O –N(H)(C4–cycloalkyl), –C(O)–N(H)(phenyl), –C(O)–N(H)(CH2CH2)N(H)C(O)–CH3, –C(O)–N(H)C(NH)(NH2), –C(O)–N(CH2CH2CH3)2, –C(O)–N–(C(H)(CH3)(CH2CH3))2, –C(O)–N(CH2CH2CH2CH3)2, –C(O)–N(H)(benzyl), –C(O)–N(H)–CH2CH2OH, –C(O)–S–CH2CH3, –C(O)N(CH2)2, –N(H)S(O)(O)–phenyl , –C(O)N(H)CH2CH2CF3, –C(O)N(H)CH3, –C(O)OCH2CH3, –C(O)–N(H)–CH2CH2O–CH2CH2O–CH3, –C(O)–N(H)–(CH2CH2O)8–CH3, –C(O)N(H)–CH2 –CH2–CH2–N(H)–CH2–CH2–CH2–N(H)–CH2–CH2–CH2–NH2, –C(O)N(H)–CH2–CH2–NH2, –C(O)N(H)C(O)CH3, –C(O)N(H)–CH2–CH3, –S(O)(O)N(H) –CH2–CH3, –S(O)(O)N–(CH2–CH3)2, –S(O)(O)N(H)–(C3–cycloalkyl), –S(O)(O)N(H)–(CH2CH2)OH, –S(O)(O)OH, –C(O)N(H)C(H)(CH2OH)2, –C(O)N( CH2CH2OH)2, –C(O)N(H)CH2CH2=CH2, –C(O)CH2N(CH3)(CH2)(COOH), –S(O)(O)(CH2CH3), –S(O)(O)N(H)CH2–CH(OH)(CH2OH), –S(O)(O)–NH2 and -S(O)(O)N(H)CH2–C(O)(O–tert-butyl).

[0289] Implementation Scheme 94) A compound having the following structure:

[0290] ,

[0291] a is 0 or an integer in the range of 1 to 4;

[0292] a' is 0 or an integer in the range of 1 to 4;

[0293] Ar1, each time it appears, is independently selected from optionally substituted phenyl, pyridine, bipyridine, tripyridine, pyrazine, pyridazine, furan, dibenzofuran, thiophene, pyrrole, selenophene, naphthalene, fluorene, phenanthrene, cyclophosphine, phthalazine, quinazoline, quinoxaline, naphthidine, phenanthrene-rholine, purine, and carbazole.

[0294] The substituents of Ar1 are independently selected each time they appear from –C(O)-CH3, –C(O)-NH2, –C(O)O-CH2CH3, -CF3, –C(O)O-tert-butyl, –C(O)-N(CH3)2, –C(O)-N(H)CH3, –C(O)-N(H)CH2CH3, –C(O)-N(H) (cycloalkyl), –C(O)–N(H) (CH2)4, –C(O)–N(CH2CH3)2, –C(O)–N(H) (tert-butyl), –C(O)–N(H) (C6-cycloalkyl), –C(O)–N(H) (CH2)3, –C(O)–N(H) (C5-cycloalkyl), –C(O)–N(H) (C4–cycloalkyl), –C(O)–N(H)(phenyl), –C(O)–N(H)(CH2CH2)N(H)C(O)–CH3, –C(O)–N(H)C(NH)(NH2), –C(O)–N(CH2CH2CH3)2, –C(O)–N–(C(H)(CH3)(CH2CH3))2, –C(O)–N(CH2CH2CH2CH3)2, –C(O)–N(benzyl), –C(O)–N(H)–CH2CH2OH, –C(O)–S–CH2CH3, –C(O)N(CH2)2, –N(H)S(O)(O)–phenyl, –C(O)N(H)CH2 CH2CF3, –C(O)N(H)CH3, –C(O)OCH2CH3, –C(O)–N(H)–CH2CH2O–CH2CH2O–CH3, –C(O)–N(H)–(CH2CH2O)8–CH3, –C(O)N(H)–CH2–CH2–CH2–N(H)–CH2 –CH2–CH2–N(H)–CH2–CH2–CH2–NH2, –C(O)N(H)–CH2–CH2–NH2, –C(O)N(H)C(O)CH3, –C(O)N(H)–CH2–CH3, –S(O)(O)N(H)–CH2–CH3, –S(O)(O)N–(C H2–CH3)2, –S(O)(O)N(H)–(C3–cycloalkyl), –S(O)(O)N(H)–(CH2CH2)OH, –S(O)(O)OH, –C(O)N(H)C(H)(CH2OH)2, –C(O)N(CH2CH2OH)2, –C(O)N(H)CH2CH 2=CH2, –C(O)CH2N(CH3)(CH2)(COOH), –S(O)(O)(CH2CH3), –S(O)(O)N(H)CH2–CH(OH)(CH2OH), –S(O)(O)–NH2, –S(O)(O)N(H)CH2–C(O)(O–tert-butyl);

[0295] And among them G1 G 2 G 3 G 4 and G 5 As defined in this article.

[0296] R 0 Each time it appears, it is independently selected from C1-C6 alkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl.

[0297] R 1 and R 1' Each time it appears, it is independently selected from H, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C 20 Heteroalkyl, C1-C 10 Heteroalkyl –NH2, C2-C6 alkenyl, C2-C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic (e.g., via one or more R... 3 Substituent groups), substituted or unsubstituted aryl groups, substituted or unsubstituted arylalkyl groups, and substituted or unsubstituted heteroaryl groups, benzyl groups, –C(=NH)NH2, –CH2CO2R 0 –C(H)(CH) a (COOH))2, -CH2C(O)NHCH2CO2H, -CH2CH2OH, -CH2CH2NHC(O)R 3 –(CH2) a C(O)NH(CH2) a C(O)OR 3 –(CH2) a –CN, spermine, –(CH2) a –NH2、–C(O)R 3 –(CH2) a C(O)OR 3 –(CH2) a OR 3 –C(H)((CH2) a OH) a' –CHCH(OH)(CH2) a OH, –(CH2) a OH, C1-C6–C(O)OH, can be substituted (e.g., via R) 3 (Substituted group) or unsubstituted –(CH2) a - Heterocyclic rings

[0298] , ;and

[0299] Where R 1 and R 1' Together they form substituted or unsubstituted heterocycles, including but not limited to aza-butanes, pyrrolidines, piperidines, piperazines, morpholines, etc.

[0300] or ;

[0301] R 3 Each time it appears, it is independently selected from H, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 heteroalkyl, –C1-C6–OH, –C1-C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine;

[0302] R 4 Each time it appears, it is independently selected from alkylene groups interrupted by one or more heteroatoms, wherein the heteroatoms are O, S, NH or a combination thereof;

[0303] G 1 Each time it appears, it is independently selected from oxo, C1-C6 alkyl, C1-C6 haloalkyl, –E–O–R 3 –E–C(R) 1 (R) 1' (R) 3 –NH2, –NO2, –SO3R 3 SO3O - 、–SO3N(H)(R 1 ), –E–C(O)R 3 , -E-CO2H, –B(OH)2, –C(O)NR 1 R 1' -E-PO(OR) 1 )2, and through G 2 G 3 G 4 and G 5 Substituted aryl; and

[0304] G 2 G 3 G 4 and G 5Each time it appears, it is independently selected from groups that are absent or selected from groups containing the following: halogen, –CN, –NO2, –SO3R. 3 C1-C6 alkyl, C1-C6 haloalkyl, –E–O–R 3 –E–(CH2) a C(O)R 3 , –E–CO2H, –E–CHO, –E–C(O)R 3 , –E–C(O)NH(OH), –E–C(O)NHR 1 –E–C(O)N(H)C(H)(R 1 (R) 1' ), –E–C(O)N(R 1 (CH2) a P(O)(O)OH)2、–E–C(O)N(R 1 (CH2) a SO3H), –E–C(O)NR 1 R 1' –E–NR 1 R 1' –E–OR 2 –C(O)–heterocyclic alkyl (where the heterocyclic alkyl is substituted or unsubstituted), –C(O)–N(H)–CH(COOH)((CH2) a – heteroaryl) (wherein the heteroaryl group is substituted or unsubstituted), wherein E is independently selected from direct bond and C1-C6 alkylene group each time it appears.

[0305] Implementation Scheme 95) A compound having either formula (IB) or (ID):

[0306] (IB),

[0307] (ID),

[0308] in

[0309] in

[0310] a is 0 or an integer in the range of 1 to 4;

[0311] a' is 0 or an integer in the range of 1 to 4;

[0312] Z is a 5- or 6-membered heteroaromatic ring containing one, two, three or four heteroatoms selected from O, N or S;

[0313] M is independently selected from hydrogen, halogens, and C1-C4 alkyl groups each time it appears;

[0314] Ar1, each time it appears, is independently selected from optionally substituted phenyl, pyridine, bipyridine, tripyridine, pyrazine, pyridazine, furan, dibenzofuran, thiophene, pyrrole, selenophene, naphthalene, fluorene, phenanthrene, cyclophosphine, phthalazine, quinazoline, quinoxaline, naphthidine, phenanthrene-rholine, purine, and carbazole.

[0315] The substituents of Ar1 are independently selected each time they appear from halogens, –OH, –CN, –NO2, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 heteroalkyl, C1-C6 cycloalkyl, –OR 0 –CONH2, –C(O)NR 1 R 1' –C(O)(CH2) a NR 1 R 1' –NR 1 R 1' –NR 1 C(O)R 3 –C(O)SR 3 –COR 3 –CO(CH2) a OC(O)R 3 –OC(O)R 3 –C(O)OR 3 –C–O–R 3 , thiols, -R 4 -H、–SOR 1 –S(O)2R 1 –S(O)2NR 1 R 1' –CH2–NR 1 S(O)2R 3 –NR 1 S(O)2R 3 and –C(CH3)=N–(phenyl)-O-CH2-C-CH;

[0316] R 0 Each time it appears, it is independently selected from C1-C6 alkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl.

[0317] R 1 and R 1'Each time it appears, it is independently selected from H, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C 20 Heteroalkyl, C1-C 10 Heteroalkyl –NH2, C2-C6 alkenyl, C2-C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic (e.g., via one or more R... 3 Substituent groups), substituted or unsubstituted aryl groups, substituted or unsubstituted arylalkyl groups, and substituted or unsubstituted heteroaryl groups, benzyl groups, –C(=NH)NH2, –CH2CO2R 0 –C(H)(CH) a (COOH))2, -CH2C(O)NHCH2CO2H, -CH2CH2OH, -CH2CH2NHC(O)R 3 –(CH2) a C(O)NH(CH2) a C(O)OR 3 –(CH2) a –CN, spermine, –(CH2) a –NH2、–C(O)R 3 –(CH2) a C(O)OR 3 –(CH2) a OR 3 –C(H)((CH2) a OH) a' –CHCH(OH)(CH2) a OH, –(CH2) a OH, C1-C6–C(O)OH, can be substituted (e.g., via R) 3 (Substituted group) or unsubstituted –(CH2) a - Heterocyclic rings

[0318] , ;and

[0319] Where R 1 and R 1' Together they form substituted or unsubstituted heterocycles, including but not limited to aza-butanes, pyrrolidines, piperidines, piperazines, morpholines, etc.

[0320] or ;

[0321] R 3Each time it appears, it is independently selected from H, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 heteroalkyl, –C1-C6–OH, –C1-C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine;

[0322] R 4 Each time it appears, it is independently selected from alkylene groups interrupted by one or more heteroatoms, wherein the heteroatoms are O, S, NH or a combination thereof;

[0323] Ar2 is independently selected each time it appears from: (i) substituted or unsubstituted 5- and 6-membered monocyclic aromatic or heterocyclic rings; (ii) substituted 9- and 10-membered fused bicyclic rings comprising two substituted or unsubstituted monocyclic rings, wherein at least one of the two monocyclic rings is an aromatic or heterocyclic ring; and (iii) substituted or unsubstituted 13- and 14-membered fused tricyclic rings comprising three monocyclic rings together, wherein at least one of the three monocyclic rings is an aromatic or heterocyclic ring.

[0324] Each Ar2 is independently transmitted via G 1 G 2 G 3 G 4 and G 5 Replace, of which:

[0325] When Ar2 is monosubstituted, G 1 Each time it appears, it is independently selected from oxo, C1-C6 alkyl, C1-C6 haloalkyl, –E–O–R 3 –E–C(R) 1 (R) 1' (R) 3 –NH2, –NO2, –SO3R 3 SO3O – 、–SO3N(H)(R 1 ), –E–C(O)R 3 , -E-CO2H, –B(OH)2, –C(O)NR 1 R 1' -E-PO(OR) 1 )2, and through G 2 G 3 G 4 and G 5 Substituted aryl groups;

[0326] G2 G 3 G 4 and G 5 Each time it appears, it is independently selected from groups that are absent or selected from groups containing the following: halogen, –CN, –NO2, –SO3R. 3 C1-C6 alkyl, C1-C6 haloalkyl, –E–O–R 3 –E–(CH2) a C(O)R 3 -E-CO2H, -E-CHO, -EC(O)R 3 -EC(O)NH(OH), -EC(O)NHR 1 –E–C(O)N(H)C(H)(R 1 (R) 1' ), –E–C(O)N(R 1 (CH2) a P(O)(O)OH)2、–E–C(O)N(R 1 (CH2) a SO3H), -EC(O)NR 1 R 1' -E-NR 1 R 1' -E-OR 2 –C(O)-heterocyclic alkyl (where the heterocyclic alkyl is substituted or unsubstituted), –C(O)–N(H)–CH(COOH)((CH2) a – heteroaryl) (wherein the heteroaryl group is substituted or unsubstituted), wherein E is independently selected from direct bond and C1-C6 alkylene group each time it appears.

[0327] (Implementation Scheme 96) A composition comprising molecules of two or more compounds having any of the following formulas:

[0328]

[0329] in

[0330] a is 0 or an integer in the range of 1 to 4;

[0331] a' is 0 or an integer in the range of 1 to 4;

[0332] Z is a 5- or 6-membered heteroaromatic ring containing one, two, three or four heteroatoms selected from O, N or S;

[0333] Ar1, each time it appears, is independently selected from optionally substituted phenyl, pyridine, bipyridine, tripyridine, pyrazine, pyridazine, furan, dibenzofuran, thiophene, pyrrole, selenophene, naphthalene, fluorene, phenanthrene, cyclophosphine, phthalazine, quinazoline, quinoxaline, naphthidine, phenanthrene-rholine, purine, and carbazole.

[0334] The substituents of Ar1 are independently selected each time they appear from halogens, –OH, –CN, –NO2, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 heteroalkyl, C1-C6 cycloalkyl, –OR 0 –CONH2, –C(O)NR 1 R 1' –C(O)(CH2) a NR 1 R 1' –NR 1 R 1' –NR 1 C(O)R 3 –C(O)SR 3 –COR 3 –CO(CH2) a OC(O)R 3 –OC(O)R 3 –C(O)OR 3 –C–O–R 3 , thiols, -R 4 -H、–SOR 1 –S(O)2R 1 –S(O)2NR 1 R 1' –CH2–NR 1 S(O)2R 3 –NR 1 S(O)2R 3 and –C(CH3)=N–(phenyl)-O-CH2-C-CH;

[0335] R 0 Each time it appears, it is independently selected from C1-C6 alkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl.

[0336] R 1 and R 1' Each time it appears, it is independently selected from H, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C 20 Heteroalkyl, C1-C 10Heteroalkyl –NH2, C2-C6 alkenyl, C2-C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic (e.g., via one or more R... 3 Substituent groups), substituted or unsubstituted aryl groups, substituted or unsubstituted arylalkyl groups, and substituted or unsubstituted heteroaryl groups, benzyl groups, –C(=NH)NH2, –CH2CO2R 0 –C(H)(CH) a (COOH))2, -CH2C(O)NHCH2CO2H, -CH2CH2OH, -CH2CH2NHC(O)R 3 –(CH2) a C(O)NH(CH2) a C(O)OR 3 –(CH2) a –CN, spermine, –(CH2) a –NH2、–C(O)R 3 –(CH2) a C(O)OR 3 –(CH2) a OR 3 –C(H)((CH2) a OH) a' –CHCH(OH)(CH2) a OH, –(CH2) a OH, C1-C6–C(O)OH, can be substituted (e.g., via R) 3 (Substituted group) or unsubstituted –(CH2) a - Heterocyclic rings

[0337] , ;and

[0338] Where R 1 and R 1' Together they form substituted or unsubstituted heterocycles, including but not limited to aza-butanes, pyrrolidines, piperidines, piperazines, morpholines, etc.

[0339] or ;

[0340] R 3Each time it appears, it is independently selected from H, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 heteroalkyl, –C1-C6–OH, –C1-C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine; and

[0341] R 4 Each occurrence is independently selected from alkylene groups interrupted by one or more heteroatoms, wherein the heteroatoms are O, S, NH, or combinations thereof.

[0342] Implementation Scheme 97) The compound according to Implementation Scheme 96, wherein Z is selected from imidazole, pyrazole, pyrimidine, pyridazine, pyrazine, pyrrole, thiophene, triazole (e.g., 1,2,3-triazole; 1,2,4-triazole, thiazole and).

[0343] Implementation Scheme 98) The compound according to Implementation Scheme 96, wherein each Z comprises one, two or three “M” parts.

[0344] Implementation Scheme 99) The compound according to Implementation Scheme 96, wherein Ar1 is substituted by the following: –C(O)–CH3, –C(O)–NH2, –C(O)O–CH2CH3, –CF3, –C(O)O–tert-butyl, –C(O)–N(CH3)2, –C(O)–N(H)CH3, –C(O)–N(H)CH2CH3, –C(O)–N(H) (cycloalkyl), –C(O)–N(H) (CH2)4, –C(O)–N(CH2CH3)2, –C(O)–N(H) (tert-butyl), –C(O)–N(H) (C6–cycloalkyl), –C(O)–N(H) (CH2)3, –C(O)–N(H) (C5–cycloalkyl), –C(O)–N( H)(C4–cycloalkyl), –C(O)–N(H)(phenyl), –C(O)–N(H)(CH2CH2)N(H)C(O)–CH3, –C(O)–N(H)C(NH)(NH2), –C(O)–N(CH2CH2CH3)2, –C(O)–N–(C(H)(CH3)(CH2CH3))2, –C(O)–N(CH2CH2CH2CH3)2, –C(O)–N(H)(benzyl), –C(O)–N(H)–CH2CH2OH, –C(O)–S–CH2CH3, –C(O)N(CH2)2, –N(H)S(O)(O)–phenyl, –C (O)N(H)CH2CH2CF3, –C(O)N(H)CH3, –C(O)OCH2CH3, –C(O)–N(H)–CH2CH2O–CH2CH2O–CH3, –C(O)–N(H)–(CH2CH2O)8–CH3, –C(O)N(H)–CH2–C H2–CH2–N(H)–CH2–CH2–CH2–N(H)–CH2–CH2–CH2–NH2, –C(O)N(H)–CH2–CH2–NH2, –C(O)N(H)C(O)CH3, –C(O)N(H)–CH2–CH3, –S(O)(O)N(H)– CH2–CH3, –S(O)(O)N–(CH2–CH3)2, –S(O)(O)N(H)–(C3–cycloalkyl), –S(O)(O)N(H)–(CH2CH2)OH, –S(O)(O)OH, –C(O)N(H)C(H)(CH2OH)2, –C(O)N( CH2CH2OH)2, –C(O)N(H)CH2CH2=CH2, –C(O)CH2N(CH3)(CH2)(COOH), –S(O)(O)(CH2CH3), –S(O)(O)N(H)CH2–CH(OH)(CH2OH), –S(O)(O)–NH2 Or S(O)(O)N(H)CH2–C(O)(O–tert-butyl).

[0345] Implementation Scheme 100) A compound having formula (I):

[0346] (I),

[0347] Or its solvates, hydrates, tautomers, chelates or salts,

[0348] in

[0349] a is 0 or an integer in the range of 1 to 4;

[0350] a' is 0 or an integer in the range of 1 to 4;

[0351] m is 1, 2, or 3;

[0352] m' is 1, 2, or 3;

[0353] n is 0, 1, or 2;

[0354] p is 0, 1, or 2;

[0355] Z is a 5- or 6-membered heteroaromatic ring containing one, two, three or four heteroatoms selected from O, N or S;

[0356] Y is independently selected from Ar2, –C(O)–Ar2, and –(CH2) each time it appears. a Ar2, –(CH2)3PO(OEt)2 or –CH2CO2Me;

[0357] L is a linking group;

[0358] M is independently selected from hydrogen, halogens, and C1–C4 alkyl groups each time it appears;

[0359] Ar1, each time it appears, is independently selected from optionally substituted phenyl, pyridine, bipyridine, tripyridine, pyrazine, pyridazine, furan, dibenzofuran, thiophene, pyrrole, selenophene, naphthalene, fluorene, phenanthrene, cyclophosphine, phthalazine, quinazoline, quinoxaline, naphthidine, phenanthrene-rholine, purine, and carbazole.

[0360] The substituents of Ar1 are independently selected each time they appear from halogens, –OH, –CN, –NO2, C1–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, C1–C6 cycloalkyl, –OR 0 –CONH2, –C(O)NR 1 R 1' –C(O)(CH2) a NR 1 R 1'–NR 1 R 1' –NR 1 C(O)R 3 –C(O)SR 3 –COR 3 –CO(CH2) a OC(O)R 3 –OC(O)R 3 –C(O)OR 3 –C–O–R 3 , thiols, -R 4 -H、–SOR 1 –S(O)2R 1 –S(O)2NR 1 R 1' –CH2–NR 1 S(O)2R 3 –NR 1 S(O)2R 3 and –C(CH3)=N–(phenyl)–O–CH2–C–CH;

[0361] R 0 Each time it appears, it is independently selected from C1–C6 alkyl, C1–C6 haloalkyl, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl.

[0362] R 1 and R 1' Each time it appears, it is independently selected from H, hydroxyl, C1–C6 alkyl, C1–C6 haloalkyl, C1–C 20 Heteroalkyl, C1–C 10 Heteroalkyl –NH2, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted arylalkyl, and substituted or unsubstituted heteroaryl, benzyl, –C(=NH)NH2, –CH2CO2R 0 –C(H)(CH) a (COOH))2, –CH2C(O)NHCH2CO2H, –CH2(CH2) a OH、–CH2CH2NHC(O)R 3 –(CH2) a C(O)NH(CH2) a C(O)OR 3 –(CH2) a–CN, spermine, –(CH2) a –NH2、–C(O)R 3 –(CH2) a C(O)OR 3 –(CH2) a OR 3 –C(H)((CH2) a OH) a' –CHCH(OH)(CH2) a OH, –(CH2) a OH, C1–C6–C(O)OH, and –(CH2) can be substituted or unsubstituted. a - Heterocyclic rings

[0363] , ;and

[0364] Where R 1 and R 1' Together they form substituted or unsubstituted heterocycles, including but not limited to aza-butanes, pyrrolidines, piperidines, piperazines, morpholines, etc.

[0365] or ;

[0366] R 2 Each time it appears, it is independently selected from C2–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, and substituted or unsubstituted haloalkoxy.

[0367] R 3 Each time it appears, it is independently selected from H, C1–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, –C1–C6–OH, –C1–C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine;

[0368] R 4 Each time it appears, it is independently selected from alkylene groups interrupted by one or more heteroatoms, wherein the heteroatoms are O, S, NH or a combination thereof;

[0369] Ar2 is independently selected each time it appears from: (i) substituted or unsubstituted 5- and 6-membered monocyclic aromatic or heterocyclic rings; (ii) substituted or unsubstituted 9- and 10-membered fused bicyclic rings comprising two monocyclic rings, wherein at least one of the two monocyclic rings is an aromatic or heterocyclic ring; and (iii) substituted or unsubstituted 13- and 14-membered fused tricyclic rings comprising three monocyclic rings together, wherein at least one of the three monocyclic rings is an aromatic or heterocyclic ring.

[0370] Each Ar2 is independently transmitted via G 1 G 2 G 3 G 4 and G 5 Replace, of which:

[0371] When Ar2 is monosubstituted, G 1 Each occurrence is independently selected from oxo, C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–C(R) 1 (R) 1' (R) 3 –NH2, –NO2, –SO3R 3 SO3O - 、–SO3N(H)(R 1 ), –E–C(O)R 3 , –E–CO2H, –B(OH)2, –C(O)NR 1 R 1' –E–PO(OR) 1 )2, and through G 2 G 3 G 4 and G 5 Substituted aryl; and

[0372] G 2 G 3 G 4 and G 5 Each time it appears, it is independently selected from groups that are absent or selected from groups containing the following: halogen, –CN, –NO2, –SO3R. 3 C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–(CH2) a C(O)R 3 , –E–CO2H, –E–CHO, –E–C(O)R 3 , –E–C(O)NH(OH), –E–C(O)NHR1 –E–C(O)N(H)C(H)(R 1 (R) 1' ), –E–C(O)N(R 1 (CH2) a P(O)(O)OH)2、–E–C(O)N(R 1 (CH2) a SO3H), –E–C(O)NR 1 R 1' –E–NR 1 R 1' –E–OR 2 –C(O)–heterocyclic alkyl (where the heterocyclic alkyl is substituted or unsubstituted), –C(O)–N(H)–CH(COOH)((CH2) a – heteroaryl) (wherein the heteroaryl group is substituted or unsubstituted), wherein E is independently selected from direct bond and C1–C6 alkylene group each time it appears;

[0373] Z is not a triazole.

[0374] Implementation Scheme 101) The compound according to Implementation Scheme 100, wherein Z contains two heteroatoms.

[0375] Implementation Scheme 102) The compound according to Implementation Scheme 100, wherein Z is a diazole.

[0376] (Scheme 103) The compound according to Scheme 102, wherein the diazole is selected from the group consisting of imidazole, pyrazole, pyrimidine, pyridazine and pyrazine.

[0377] Implementation Scheme 104) The compound according to Implementation Scheme 100, wherein Z contains three heteroatoms.

[0378] Implementation Scheme 105) The compound according to Implementation Scheme 100, wherein Z contains four heteroatoms.

[0379] Implementation Scheme 106) The compound according to Implementation Scheme 100, wherein Z is selected from the group consisting of: pyrrole, thiophene, thiazole, isoxazole and tetrazolium.

[0380] Implementation Scheme 107) The compound according to any one of Implementation Schemes 100 to 106, wherein Ar1 is phenyl.

[0381] Implementation Scheme 108) The compound according to any one of Implementation Schemes 100 to 106, wherein Ar1 is pyridine.

[0382] Implementation Scheme 109) The compound according to any one of Implementation Schemes 100 to 106, wherein Ar1 is furan.

[0383] Implementation Scheme 110) The compound according to any one of Implementation Schemes 100 to 106, wherein Ar1 is carbazole.

[0384] Implementation Scheme 111) The compound according to any one of Implementation Schemes 100 to 106, wherein Ar1 is naphthyl.

[0385] Implementation Scheme 112) The compound according to any one of Implementation Schemes 100 to 106, wherein Ar1 is a pyridazine.

[0386] Implementation Scheme 113) The compound according to any one of Implementation Schemes 100 to 106, wherein Ar1 is thiophene.

[0387] Implementation Scheme 114) The compound according to any one of Implementation Schemes 100 to 106, wherein Ar1 is pyrrole.

[0388] Implementation Scheme 115) The compound according to any one of Implementation Schemes 100 to 106, wherein Ar1 is a dibenzofuran.

[0389] Implementation Scheme 116) The compound according to any one of Implementation Schemes 100 to 106, wherein Ar1 is naphthidine.

[0390] Implementation Scheme 117) The compound according to any one of Implementation Schemes 100 to 116,Each substituent of Ar1 is independently selected from –C(O)–CH3, –C(O)–NH2, –C(O)O–CH2CH3, –CF3, –C(O)O–tert-butyl, –C(O)–N(CH3)2, –C(O)–N(H)CH3, –C(O)–N(H)CH2CH3, –C(O)–N(H) (cycloalkyl), –C(O)–N(H) (CH2)4, –C(O)–N(CH2CH3)2, –C(O)–N(H) (tert-butyl), –C(O)–N(H) (C6–cycloalkyl), –C(O)–N(H) (CH2)3, –C(O)–N(H) (C5–cycloalkyl), – C(O)–N(H)(C4–cycloalkyl), –C(O)–N(H)(phenyl), –C(O)–N(H)(CH2CH2)N(H)C(O)–CH3, –C(O)–N(H)C(NH)(NH2), –C(O)–N(CH2CH2CH3)2, –C(O)–N–(C(H)(CH3)(CH2CH3))2, –C(O)–N(CH2CH2CH2CH3)2, –C(O)–N(H)(benzyl), –C(O)–N(H)–CH2CH2OH, –C(O)–S–CH2CH3, –C(O)N(CH2)2, –N(H)S(O)(O)– Phenyl, –C(O)N(H)CH2CH2CF3, –C(O)N(H)CH3, –C(O)OCH2CH3, –C(O)–N(H)–CH2CH2O–CH2CH2O–CH3, –C(O)–N(H)–(CH2CH2O)8–CH3, –C(O)N(H)–C H2–CH2–CH2–N(H)–CH2–CH2–CH2–N(H)–CH2–CH2–CH2–NH2, –C(O)N(H)–CH2–CH2–NH2, –C(O)N(H)C(O)CH3, –C(O)N(H)–CH2–CH3, –S(O)(O)N(H )–CH2–CH3, –S(O)(O)N–(CH2–CH3)2, –S(O)(O)N(H)–(C3–cycloalkyl), –S(O)(O)N(H)–(CH2CH2)OH, –S(O)(O)OH, –C(O)N(H)C(H)(CH2OH)2, –C(O)N (CH2CH2OH)2, –C(O)N(H)CH2CH2=CH2, –C(O)CH2N(CH3)(CH2)(COOH), –S(O)(O)(CH2CH3), –S(O)(O)N(H)CH2–CH(OH)(CH2OH), –S(O)(O)–NH2 and –S(O)(O)N(H)CH2–C(O)(O–tert-butyl). ,

[0391] Implementation Scheme 118) The compound according to any one of Implementation Schemes 100 to 116, wherein Ar1 is via at least one –SO2–N(H)(R 1 ) group substitution.

[0392] Implementation Scheme 119) The compound according to Implementation Scheme 118, wherein R 1 It is a C1 to C6 alkyl group.

[0393] Implementation Scheme 120) The compound according to Implementation Scheme 119, wherein the C1 to C6 alkyl group is methyl or ethyl.

[0394] Implementation Scheme 121) The compound according to any one of Implementation Schemes 100 to 120, wherein Ar2 is selected from the group consisting of benzene, pyridine, pyridazine, pyrimidine and pyrazine.

[0395] (Scheme 122) The compound according to any one of Schemes 100 to 121, wherein each Ar2 is partially substituted by one or more of the following groups: –NH2, –C(O)OH, –OH, –OCH3, –C(O)NHOH, –C(O)NH2, –(CH2)3C(O)OH, –CF3, –Cl, –P(O)(OH)2, –F, –C(O)N(H)CH2CH3, –C(O)N(H)CH2C(O)OCH3, –C(O)N(H)CH2CF2CF2CF3 -C(O)N(H)CH2C(O)OH, -C(O)N(H)C(H)(C(O)OOH)CH2C(O)OH, -C(O)N(H)CH2C(O)N(H)CH2C(O)OH, -SO3H, -NO2, -C(O)N(H)CH2PO3H2, -C(O)N(H)CH23O3H, -B(OH)2, -C(OH)(CF3)(CF3), -C(O)-5-membered heterocyclic alkyl-C(O)OH, -C(O)N(H)C(H)(C(O)OH)CH2-imidazole, -S(O)(O)N(H)-CH2COOH, -C(O)N(H)CH(C(O)OH)(CH2CH2C(O)OH), -C(O)N(H)CH2CH2SO3H, -SO2O – –C(O)N(H)–imidazole–C(O)OH and –C(O)N(CH3)2.

[0396] (Scheme 123) The compound according to any one of Schemes 100 to 121, wherein each Ar2 is derived from at least one –SO3H moiety or at least one –SO3- Partial replacement.

[0397] (Scheme 124) The compound according to any one of Schemes 100 to 121, wherein each Ar2 is derived from at least two –SO3H moieties or at least two –SO3 - Partial replacement.

[0398] (Embodiment 125) The compound according to any one of embodiments 100 to 121, wherein each Ar2 is substituted with at least two moieties, wherein the first moieties of said at least two moieties comprise a –SO3H moiety or a –SO3 moiety. - One of the parts.

[0399] (Scheme 126) The compound according to Scheme 125, wherein the second part of the at least two parts is selected from the group consisting of: –NO2, –CF3, –F, –Cl, –I, methyl, ethyl, propyl and butyl.

[0400] (Scheme 127) The compound according to any one of Schemes 100 to 121, wherein the compound has either formula (IA) or (IB):

[0401] (IA) or

[0402] (IB).

[0403] (Scheme 128) The compound according to any one of Schemes 100 to 121, wherein the compound has the following formula:

[0404] .

[0405] (Scheme 129) The compound according to any one of Schemes 100 to 121, wherein the compound has any one of the following formulas:

[0406]

[0407] (Scheme 130) The compound according to any one of embodiments 100 to 121, wherein the compound has any one of the following formulas:

[0408]

[0409] Implementation Scheme 131) A compound having either formula (IA) or (IB):

[0410] (IA) or

[0411] (IB),

[0412] Or its solvates, hydrates, tautomers, chelates or salts,

[0413] in

[0414] a is 0 or an integer in the range of 1 to 4;

[0415] a' is 0 or an integer in the range of 1 to 4;

[0416] m is 1, 2, or 3;

[0417] m' is 1, 2, or 3;

[0418] n is 0, 1, or 2;

[0419] p is 0, 1, or 2;

[0420] Z is a 5- or 6-membered heteroaromatic ring containing one, two, three or four heteroatoms selected from O, N or S;

[0421] Y is independently selected from Ar2, –C(O)–Ar2, and –(CH2) each time it appears. a Ar2, –(CH2)3PO(OEt)2 or –CH2CO2Me;

[0422] M is independently selected from hydrogen, halogens, and C1-C4 alkyl groups each time it appears;

[0423] Ar1, each time it appears, is independently selected from optionally substituted phenyl, pyridine, bipyridine, tripyridine, pyrazine, pyridazine, furan, dibenzofuran, thiophene, pyrrole, selenophene, naphthalene, fluorene, phenanthrene, cyclophosphine, phthalazine, quinazoline, quinoxaline, naphthidine, phenanthrene-rholine, purine, and carbazole.

[0424] Ar1 is converted to –OH and –C(O)NR 1 R 1' –C(O)(CH2) a NR 1 R 1' –CO(CH2) a OC(O)R 3 –OC(O)R 3 –S(O)2R 1 –S(O)2NR 1 R1' –CH2–NR 1 S(O)2R 3 Or –NR 1 S(O)2R 3 Replace at least one of them;

[0425] R 0 Each time it appears, it is independently selected from C1–C6 alkyl, C1–C6 haloalkyl, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl.

[0426] R 1 and R 1' Each time it appears, it is independently selected from H, hydroxyl, C1–C6 alkyl, C1–C6 haloalkyl, C1–C 20 Heteroalkyl, C1–C 10 Heteroalkyl –NH2, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted arylalkyl, and substituted or unsubstituted heteroaryl, benzyl, –C(=NH)NH2, –CH2CO2R 0 –C(H)(CH) a (COOH))2, –CH2C(O)NHCH2CO2H, –CH2(CH2) a OH、–CH2CH2NHC(O)R 3 –(CH2) a C(O)NH(CH2) a C(O)OR 3 –(CH2) a –CN, spermine, –(CH2) a –NH2、–C(O)R 3 –(CH2) a C(O)OR 3 –(CH2) a OR 3 –C(H)((CH2) a OH) a' –CHCH(OH)(CH2) a OH, –(CH2) a OH, C1–C6–C(O)OH, and –(CH2) can be substituted or unsubstituted. a - Heterocyclic rings

[0427] , ;and

[0428] Where R 1 and R 1' Together they form substituted or unsubstituted heterocycles, including but not limited to aza-butanes, pyrrolidines, piperidines, piperazines, morpholines, etc.

[0429] or ;

[0430] R 2 Each time it appears, it is independently selected from C2–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, and substituted or unsubstituted haloalkoxy.

[0431] R 3 Each time it appears, it is independently selected from H, C1–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, –C1–C6–OH, –C1–C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine;

[0432] Ar2 is independently selected each time it appears from: (i) substituted or unsubstituted 5- and 6-membered monocyclic aromatic or heterocyclic rings; (ii) substituted or unsubstituted 9- and 10-membered fused bicyclic rings comprising two monocyclic rings, wherein at least one of the two monocyclic rings is an aromatic or heterocyclic ring; and (iii) substituted or unsubstituted 13- and 14-membered fused tricyclic rings comprising three monocyclic rings together, wherein at least one of the three monocyclic rings is an aromatic or heterocyclic ring.

[0433] Each Ar2 is independently transmitted via G 1 G 2 G 3 G 4 and G 5 Replace, of which:

[0434] When Ar2 is monosubstituted, G 1 Each time it appears, it is independently selected from oxo, C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–C(R) 1 (R) 1' (R) 3 –NH2, –NO2, –SO3R3 SO3O - 、–SO3N(H)(R 1 ), –E–C(O)R 3 , –E–CO2H, –B(OH)2, –C(O)NR 1 R 1' –E–PO(OR) 1 )2, and through G 2 G 3 G 4 and G 5 Substituted aryl groups;

[0435] G 2 G 3 G 4 and G 5 Each time it appears, it is independently selected from groups that are absent or selected from groups containing the following: halogen, –CN, –NO2, –SO3R. 3 C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–(CH2) a C(O)R 3 , –E–CO2H, –E–CHO, –E–C(O)R 3 , –E–C(O)NH(OH), –E–C(O)NHR 1 –E–C(O)N(H)C(H)(R 1 (R) 1' ), –E–C(O)N(R 1 (CH2) a P(O)(O)OH)2、–E–C(O)N(R 1 (CH2) a SO3H), –E–C(O)NR 1 R 1' –E–NR 1 R 1' –E–OR 2 –C(O)–heterocyclic alkyl (where the heterocyclic alkyl is substituted or unsubstituted), –C(O)–N(H)–CH(COOH)((CH2) a – heteroaryl) (wherein the heteroaryl group is substituted or unsubstituted), wherein E is independently selected from direct bond and C1–C6 alkylene group each time it appears.

[0436] Implementation Scheme 132) The compound according to Implementation Scheme 131, wherein at least one Ar1 substituent is selected from the group consisting of: –OH, –C(O)NR 1R 1' –C(O)(CH2) a NR 1 R 1' –CO(CH2) a OC(O)R 3 –OC(O)R 3 –S(O)2R 1 and –S(O)2NR 1 R 1' .

[0437] Implementation scheme 133). The compound according to claim 131, wherein Ar1 is via at least one –C(O)(CH2). a NR 1 R 1' replace.

[0438] Implementation Scheme 134) The compound according to Implementation Scheme 133, wherein R 1 For H.

[0439] Implementation Scheme 135) The compound according to Implementation Scheme 134, wherein R 1' It is a C1–C6 alkyl group.

[0440] (Scheme 136) The compound according to Scheme 135, wherein the C1–C6 alkyl group is branched.

[0441] (Scheme 137) The compound according to Scheme 135, wherein the C1–C6 alkyl group comprises at least one substituent.

[0442] Implementation Scheme 138) The compound according to Implementation Scheme 136, wherein the at least one substituent is a hydroxyl group.

[0443] Implementation Scheme 139) The compound according to Implementation Scheme 133, wherein R 1 and R 1' Together they form cycloalkyl groups or heterocycloalkyl groups.

[0444] Implementation Scheme 140) The compound according to Implementation Scheme 131, wherein Ar1 is via at least one –S(O)2NR 1 R 1' replace.

[0445] Implementation Scheme 141) The compound according to Implementation Scheme 140, wherein R 1 For H.

[0446] Implementation Scheme 142) The compound according to Implementation Scheme 141, wherein R 1' It is a C1–C6 alkyl group.

[0447] (Scheme 143) The compound according to Scheme 142, wherein the C1–C6 alkyl group is branched.

[0448] (Scheme 144) The compound according to Scheme 142, wherein the C1–C6 alkyl group comprises at least one substituent.

[0449] (Scheme 145) The compound according to Scheme 144, wherein the at least one substituent is a hydroxyl group.

[0450] Implementation Scheme 146) The compound according to Implementation Scheme 140, wherein R 1 and R 1' Together they form cycloalkyl groups or heterocycloalkyl groups.

[0451] (Scheme 147) The compound according to any one of Schemes 131 to 146, wherein each Ar2 is derived from at least one –SO3H moiety or at least one –SO3 - Partial replacement.

[0452] Implementation Scheme 148) The compound according to any one of Implementation Schemes 131 to 146, wherein each Ar2 is derived from at least two –SO3H moieties or at least two –SO3 - Partial replacement.

[0453] (Scheme 149) The compound according to any one of Schemes 131 to 146, wherein each Ar2 is substituted with at least two moieties, wherein the first moieties of the at least two moieties comprise a –SO3H moiety or a –SO3 moiety. - One of the parts.

[0454] (Scheme 150) The compound according to Scheme 149, wherein the second part of the at least two parts is selected from the group consisting of: –NO2, –CF3, –F, –Cl, –I, methyl, ethyl, propyl and butyl.

[0455] Implementation Scheme 151) The compound according to any one of Implementation Schemes 131 to 145, wherein the compound has any one of the following formulas:

[0456]

[0457] (Scheme 152) The compound according to any one of Schemes 131 to 145, wherein the compound has any one of the following formulas:

[0458]

[0459] Implementation Scheme 153) The compound according to Implementation Scheme 131, wherein the compound has the following formula:

[0460] .

[0461] Implementation Scheme 154) The compound according to Implementation Scheme 131, wherein the compound has the following formula:

[0462] .

[0463] (Implementation Scheme 155) The compound according to any one of Implementation Schemes 131 to 154, wherein Z is a diazole.

[0464] (Scheme 156) The compound according to any one of Schemes 131 to 154, wherein Z is selected from the group consisting of imidazole, pyrazole, pyrimidine, pyridazine and pyrazine.

[0465] Implementation Scheme 157) The compound according to any one of Implementation Schemes 131 to 154, wherein Z is not a triazole.

[0466] (Scheme 158) The compound according to any one of Schemes 131 to 146, wherein the compound has any one of the following formulas:

[0467]

[0468] Implementation Scheme 159) A compound having any of the following formulas:

[0469]

[0470] Or its solvates, hydrates, tautomers, chelates or salts,

[0471] in

[0472] Z is a 5- or 6-membered heteroaromatic ring containing one, two, three or four heteroatoms selected from O, N or S;

[0473] M is independently selected from hydrogen, halogens, and C1–C4 alkyl groups each time it appears;

[0474] Y is independently selected from Ar2, –C(O)–Ar2, and –(CH2) each time it appears. a Ar2, –(CH2)3PO(OEt)2 or –CH2CO2Me;

[0475] Ar1 is independently selected each time it appears from optionally substituted phenyl, pyridine, bipyridine, tripyridine, pyrazine, pyridazine, furan, dibenzofuran, thiophene, pyrrole, selenophene, naphthalene, fluorene, phenanthrene, cyclophosphine, phthalazine, quinazoline, quinoxaline, naphthidine, phenanthrene-rholine, purine, and carbazole;

[0476] Ar2 is independently selected each time it appears from: (i) substituted or unsubstituted 5- and 6-membered monocyclic aromatic or heterocyclic rings; (ii) substituted or unsubstituted 9- and 10-membered fused bicyclic rings comprising two monocyclic rings, wherein at least one of the two monocyclic rings is an aromatic or heterocyclic ring; and (iii) substituted or unsubstituted 13- and 14-membered fused tricyclic rings comprising three monocyclic rings together, wherein at least one of the three monocyclic rings is an aromatic or heterocyclic ring.

[0477] Each Ar2 is independently transmitted via G 1 G 2 G 3 G 4 and G 5 Replace, of which:

[0478] When Ar2 is monosubstituted, G 1 Each time it appears, it is independently selected from oxo, C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–C(R) 1 (R) 1' (R) 3 –NH2, –NO2, –SO3R 3 SO3O – 、–SO3N(H)(R 1 ), –E–C(O)R 3 , –E–CO2H, –B(OH)2, –C(O)NR 1 R 1' –E–PO(OR) 1 )2, and through G 2 G 3 G 4 and G 5 Substituted aryl groups;

[0479] G 2 G 3 G 4 and G 5 Each time it appears, it is independently selected from groups that are absent or selected from groups containing the following: halogen, –CN, –NO2, –SO3R. 3 C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–(CH2) a C(O)R 3 , –E–CO2H, –E–CHO, –E–C(O)R 3 , –E–C(O)NH(OH), –E–C(O)NHR 1 –E–C(O)N(H)C(H)(R 1 (R) 1' ), –E–C(O)N(R 1 (CH2) a P(O)(O)OH)2、–E–C(O)N(R 1 (CH2) a SO3H), –E–C(O)NR 1 R 1' –E–NR 1 R 1' –E–OR 2 –C(O)–heterocyclic alkyl (where the heterocyclic alkyl is substituted or unsubstituted), –C(O)–N(H)–CH(COOH)((CH2) a – heteroaryl) (wherein the heteroaryl group is substituted or unsubstituted), wherein E is independently selected from direct bond and C1–C6 alkylene group each time it appears;

[0480] R 0 Each time it appears, it is independently selected from C1–C6 alkyl, C1–C6 haloalkyl, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl.

[0481] R 1 and R 1' Each time it appears, it is independently selected from H, hydroxyl, C1–C6 alkyl, C1–C6 haloalkyl, C1–C 20 Heteroalkyl, C1–C 10Heteroalkyl –NH2, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted arylalkyl, and substituted or unsubstituted heteroaryl, benzyl, –C(=NH)NH2, –CH2CO2R 0 –C(H)(CH) a (COOH))2, –CH2C(O)NHCH2CO2H, –CH2(CH2) a OH、–CH2CH2NHC(O)R 3 –(CH2) a C(O)NH(CH2) a C(O)OR 3 –(CH2) a –CN, spermine, –(CH2) a –NH2、–C(O)R 3 –(CH2) a C(O)OR 3 –(CH2) a OR 3 –C(H)((CH2) a OH) a' –CHCH(OH)(CH2) a OH, –(CH2) a OH, C1–C6–C(O)OH, and –(CH2) can be substituted or unsubstituted. a - Heterocyclic rings

[0482] , ;and

[0483] Where R 1 and R 1' Together they form substituted or unsubstituted heterocycles, including but not limited to aza-butanes, pyrrolidines, piperidines, piperazines, morpholines, etc.

[0484] or ;

[0485] R 2 Each time it appears, it is independently selected from C2–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, and substituted or unsubstituted haloalkoxy; and

[0486] R 3Each time it appears, it is independently selected from H, C1–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, –C1–C6–OH, –C1–C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine.

[0487] Implementation Scheme 160) The compound according to Implementation Scheme 159, wherein Z contains two heteroatoms.

[0488] Implementation Scheme 161) The compound according to Implementation Scheme 160, wherein Z is a diazole.

[0489] Implementation Scheme 162) The compound according to Implementation Scheme 161, wherein the diazole is selected from the group consisting of imidazole, pyrazole, pyrimidine, pyridazine and pyrazine.

[0490] Implementation Scheme 163) The compound according to Implementation Scheme 159, wherein Z contains three heteroatoms.

[0491] Implementation Scheme 164) The compound according to Implementation Scheme 159, wherein Z contains four heteroatoms.

[0492] Implementation Scheme 165) The compound according to Implementation Scheme 159, wherein Z is selected from the group consisting of: pyrrole, thiophene, thiazole, isoxazole and tetrazolium.

[0493] Implementation Scheme 166) The compound according to Implementation Scheme 159, wherein Z is a triazole.

[0494] Implementation Scheme 167) The compound according to Implementation Scheme 159, wherein Z is not a triazole.

[0495] Implementation Scheme 168) The compound according to any one of Implementation Schemes 159 to 167, wherein Ar1 is phenyl.

[0496] Implementation Scheme 169) The compound according to any one of Implementation Schemes 159 to 167, wherein Ar1 is pyridine.

[0497] Implementation Scheme 170) The compound according to any one of Implementation Schemes 159 to 167, wherein Ar1 is furan.

[0498] Implementation Scheme 171) The compound according to any one of Implementation Schemes 159 to 167, wherein Ar1 is carbazole.

[0499] Implementation Scheme 172) The compound according to any one of Implementation Schemes 159 to 167, wherein Ar1 is naphthyl.

[0500] Implementation Scheme 173) The compound according to any one of Implementation Schemes 159 to 167, wherein Ar1 is a pyridazine.

[0501] Implementation Scheme 174) The compound according to any one of Implementation Schemes 159 to 167, wherein Ar1 is thiophene.

[0502] (Scheme 175) The compound according to any one of Schemes 159 to 167, wherein Ar1 is pyrrole.

[0503] Implementation Scheme 176) The compound according to any one of Implementation Schemes 159 to 167, wherein Ar1 is a dibenzofuran.

[0504] Implementation Scheme 177) The compound according to any one of Implementation Schemes 159 to 167, wherein Ar1 is naphthidine.

[0505] (Scheme 178) The compound according to any one of Schemes 159 to 167, wherein each Ar2 is partially substituted by one or more of the following groups: –NH2, –C(O)OH, –OH, –OCH3, –C(O)NHOH, –C(O)NH2, –(CH2)3C(O)OH, –CF3, –Cl, –P(O)(OH)2, –F, –C(O)N(H)CH2CH3, –C(O)N(H)CH2C(O)OCH3, –C(O)N(H)CH2CF2CF2CF3 -C(O)N(H)CH2C(O)OH, -C(O)N(H)C(H)(C(O)OOH)CH2C(O)OH, -C(O)N(H)CH2C(O)N(H)CH2C(O)OH, -SO3H, -NO2, -C(O)N(H)CH2PO3H2, -C(O)N(H)CH23O3H, -B(OH)2, -C(OH)(CF3)(CF3), -C(O)-5-membered heterocyclic alkyl-C(O)OH, -C(O)N(H)C(H)(C(O)OH)CH2-imidazole, -S(O)(O)N(H)-CH2COOH, -C(O)N(H)CH(C(O)OH)(CH2CH2C(O)OH), -C(O)N(H)CH2CH2SO3H, -SO2O -–C(O)N(H)–imidazole–C(O)OH and –C(O)N(CH3)2.

[0506] (Scheme 179) The compound according to any one of Schemes 159 to 177, wherein each Ar2 is derived from at least one –SO3H moiety or at least one –SO3 - Partial replacement.

[0507] (Scheme 180) The compound according to any one of Schemes 159 to 177, wherein each Ar2 is derived from at least two –SO3H moieties or at least two –SO3 - Partial replacement.

[0508] (Embodiment 181) The compound according to any one of embodiments 159 to 177, wherein each Ar2 is substituted with at least two moieties, wherein the first moieties of said at least two moieties comprise a –SO3H moiety or a –SO3 moiety. - One of the parts.

[0509] (Scheme 182) The compound according to Scheme 181, wherein the second part of the at least two parts is selected from the group consisting of: –NO2, –CF3, –F, –Cl, –I, methyl, ethyl, propyl and butyl.

[0510] Implementation Scheme 183) A compound having any of the following formulas:

[0511]

[0512] Or its solvates, hydrates, tautomers, chelates or salts,

[0513] in

[0514] Ar1 is independently selected each time it appears from optionally substituted phenyl, pyridine, bipyridine, tripyridine, pyrazine, pyridazine, furan, dibenzofuran, thiophene, pyrrole, selenophene, naphthalene, fluorene, phenanthrene, cyclophosphine, phthalazine, quinazoline, quinoxaline, naphthidine, phenanthrene-rholine, purine, and carbazole;

[0515] R 1 and R 1' Each time it appears, it is independently selected from H, hydroxyl, C1–C6 alkyl, C1–C6 haloalkyl, C1–C 20 Heteroalkyl, C1–C 10Heteroalkyl –NH2, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted arylalkyl, and substituted or unsubstituted heteroaryl, benzyl, –C(=NH)NH2, –CH2CO2R 0 –C(H)(CH) a (COOH))2, –CH2C(O)NHCH2CO2H, –CH2(CH2) a OH、–CH2CH2NHC(O)R 3 –(CH2) a C(O)NH(CH2) a C(O)OR 3 –(CH2) a –CN, spermine, –(CH2) a –NH2、–C(O)R 3 –(CH2) a C(O)OR 3 –(CH2) a OR 3 –C(H)((CH2) a OH) a' –CHCH(OH)(CH2) a OH, –(CH2) a OH, C1–C6–C(O)OH, and –(CH2) can be substituted or unsubstituted. a - Heterocyclic rings

[0516] , ;and

[0517] Where R 1 and R 1' Together they form substituted or unsubstituted heterocycles, including but not limited to aza-butanes, pyrrolidines, piperidines, piperazines, morpholines, etc.

[0518] or ;

[0519] R 2 Each time it appears, it is independently selected from C2–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, and substituted or unsubstituted haloalkoxy.

[0520] R 3Each time it appears, it is independently selected from H, C1–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, –C1–C6–OH, –C1–C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine; and

[0521] G 2 and G 3 Each time it appears, it is independently selected from groups that are absent or selected from groups containing the following: halogen, –CN, –NO2, –SO3R. 3 C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–(CH2) a C(O)R 3 , –E–CO2H, –E–CHO, –E–C(O)R 3 , –E–C(O)NH(OH), –E–C(O)NHR 1 –E–C(O)N(H)C(H)(R 1 (R) 1' ), –E–C(O)N(R 1 (CH2) a P(O)(O)OH)2、–E–C(O)N(R 1 (CH2) a SO3H), –E–C(O)NR 1 R 1' –E–NR 1 R 1' –E–OR 2 –C(O)–heterocyclic alkyl (where the heterocyclic alkyl is substituted or unsubstituted), –C(O)–N(H)–CH(COOH)((CH2) a – heteroaryl) (wherein the heteroaryl group is substituted or unsubstituted), wherein E is independently selected from direct bond and C1–C6 alkylene group each time it appears.

[0522] Implementation Scheme 184) The compound according to Implementation Scheme 183, wherein the compound has any of the following formulas:

[0523]

[0524] (Scheme 185) The compound according to Scheme 183, wherein the compound has any of the following formulas:

[0525]

[0526] Implementation Scheme 186) The compound according to any one of Implementation Schemes 183 to 184, wherein R 1 For H.

[0527] Implementation Scheme 187) The compound according to Implementation Scheme 186, wherein R 1' It is a C1–C6 alkyl group.

[0528] (Scheme 188) The compound according to Scheme 187, wherein the C1–C6 alkyl group is branched.

[0529] (Scheme 189) The compound according to Scheme 187, wherein the C1–C6 alkyl group comprises at least one substituent.

[0530] (Scheme 190) The compound according to Scheme 189, wherein the at least one substituent is a hydroxyl group.

[0531] Implementation Scheme 191) The compound according to any one of Implementation Schemes 183 to 184, wherein R 1 and R 1' Together they form cycloalkyl groups or heterocycloalkyl groups.

[0532] Implementation Scheme 192) The compound according to any one of Implementation Schemes 185 to 191, wherein G 2 and G 3 Each is represented by H.

[0533] Implementation Scheme 193) The compound according to any one of Implementation Schemes 183 to 191, wherein G 2 or G 3 One of them is –SO3H or –SO3 - .

[0534] Implementation Scheme 194) A compound having the following formula:

[0535] ,

[0536] Or its solvates, hydrates, tautomers, chelates or salts,

[0537] in

[0538] Z is a 5- or 6-membered heteroaromatic ring containing one, two, three or four heteroatoms selected from O, N or S;

[0539] M is independently selected from hydrogen, halogens, and C1–C4 alkyl groups each time it appears;

[0540] Y is independently selected from Ar2, –C(O)–Ar2, and –(CH2) each time it appears. a Ar2, –(CH2)3PO(OEt)2 or –CH2CO2Me;

[0541] Ar1 is independently selected each time it appears from optionally substituted phenyl, pyridine, bipyridine, tripyridine, pyrazine, pyridazine, furan, dibenzofuran, thiophene, pyrrole, selenophene, naphthalene, fluorene, phenanthrene, cyclophosphine, phthalazine, quinazoline, quinoxaline, naphthidine, phenanthrene-rholine, purine, and carbazole;

[0542] Ar2 is independently selected each time it appears from: (i) substituted or unsubstituted 5- and 6-membered monocyclic aromatic or heterocyclic rings; (ii) substituted or unsubstituted 9- and 10-membered fused bicyclic rings comprising two monocyclic rings, wherein at least one of the two monocyclic rings is an aromatic or heterocyclic ring; and (iii) substituted or unsubstituted 13- and 14-membered fused tricyclic rings comprising three monocyclic rings together, wherein at least one of the three monocyclic rings is an aromatic or heterocyclic ring.

[0543] Each Ar2 is independently denoted by –C(O)OR 3 –C1–C6 haloalkyl, –NO2 and –SO3H substitution;

[0544] R 3 Each time it appears, it is independently selected from H, C1–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, –C1–C6–OH, –C1–C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine.

[0545] (Scheme 195) The compound according to Scheme 194, wherein the –C1-C6 haloalkyl group is –CF3.

[0546] Implementation Scheme 196) The compound according to Implementation Scheme 194, wherein the –C(O)OR 3 It is –C(O)OH.

[0547] Implementation Scheme 197) The compound according to Implementation Scheme 194, wherein Ar2 contains two substituents.

[0548] Implementation Scheme 198) The compound according to Implementation Scheme 194, wherein Ar2 contains three substituents.

[0549] Implementation Scheme 199) The compound according to Implementation Scheme 194, wherein Ar2 contains four substituents.

[0550] Implementation Scheme 200) The compound according to Implementation Scheme 194, wherein the compound comprises a –C(O)OR 3 A portion and a –C1–C6 haloalkyl moiety.

[0551] Implementation Scheme 201) The compound according to Implementation Scheme 194, wherein the compound comprises a –NO2 moiety and a –SO3H moiety.

[0552] (Scheme 202) The compound according to Scheme 194, wherein the compound comprises a –SO3H moiety and a –C1–C6 haloalkyl moiety.

[0553] Implementation Scheme 203) The compound according to Implementation Scheme 194, wherein the compound comprises a –C(O)OR 3 One part and one –NO2 part.

[0554] (Scheme 204) The compound according to Scheme 194, wherein the compound comprises at least one –SO3H moiety.

[0555] (Embodiment 205) A polymer, copolymer or metal gel comprising any of the compounds described in embodiments 100 to 204.

[0556] (Embodiment 206) A composition comprising at least two molecules of any one of the compounds described in embodiments 100 to 204.

[0557] Implementation Scheme 207) A method for enhancing nucleic acid polymerase reactions, the method comprising:

[0558] a. Forming a nucleic acid polymerase reaction composition comprising:

[0559] i. Template nucleic acid,

[0560] ii. Nucleic acid polymerase,

[0561] iii. A mixture of nucleotides or nucleotide analogs, and

[0562] iv. at least one of the compounds according to any one of embodiments 1 to 105; and

[0563] b. Incubating a nucleic acid polymerase reaction composition under conditions that allow for nucleic acid polymerization, wherein the at least one compound according to any one of embodiments 100 to 204 increases the sustained synthesis capacity, rate, or fidelity of the nucleic acid polymerase reaction.

[0564] Implementation Scheme 208) The method according to Implementation Scheme 207, wherein the nucleic acid polymerase is a DNA polymerase.

[0565] (Implementation Scheme 209) The method according to Implementation Scheme 208, wherein the DNA polymerase is DPO4 or a variant thereof.

[0566] (Implementation Scheme 210) The method according to Implementation Scheme 207, wherein the mixture of nucleotides or nucleotide analogs is a mixture of nucleotide analogs comprising nucleoside triaminophosphate, wherein each nucleoside triaminophosphate comprises a nucleobase selected from the group consisting of adenine, guanine, thymine, and cytosine and a polymer chain portion, wherein a first end of the polymer chain portion is connected to the nucleobase and a second end of the polymer chain portion is connected to the α-phosphate of the nucleoside triaminophosphate to provide extension of the nucleotide analog by cleavage of the aminophosphate bond.

[0567] Implementation Scheme 211) The method according to Implementation Scheme 207, wherein the nucleic acid polymerization reaction produces an expandable polymer of nucleotide analogs, wherein the expandable polymer encodes the nucleobase sequence information of the template nucleic acid.

[0568] Implementation Scheme 212) The method according to Implementation Scheme 207, wherein the conditions allowing the nucleic acid polymerization reaction include a suitable polymerization buffer and oligonucleotide primers.

[0569] (Implementation Scheme 213) The method according to Implementation Scheme 212, wherein the suitable polymerization buffer comprises one or more components selected from MnCl2, buffer, salt, sugar, single-chain binding protein (SSB), imidazole, pyrazole, triazole, betaine, molecular crowding agent, dimethyl sulfoxide (DMSO), alkyl glycol, glycerol, N-methyl-2-pyrrolidone (NMP), acetamide, butylated hydroxyanisole (BHA), polyphosphate and / or urea.

[0570] (Scheme 214) The method according to Scheme 207, wherein the nucleic acid polymerase reaction composition further comprises a single-stranded binding protein.

[0571] (Scheme 215) The method according to Scheme 207, wherein the nucleic acid polymerase reaction composition further comprises urea.

[0572] Implementation Scheme 216) The method according to Implementation Scheme 207, wherein the mixture of nucleotides or nucleotide analogs comprises a nucleotide analog containing a detectable marker.

[0573] Implementation Scheme 217) The method according to Implementation Scheme 216, wherein the detectable marker is an optically detectable marker selected from the group consisting of luminescent, chemiluminescent, fluorescent, fluorescent, chromogenic, or chromogenic markers.

[0574] (Implementation Scheme 218) Use of the compound according to any one of Implementation Schemes 100 to 204 in a nucleic acid polymerase reaction.

[0575] Implementation Scheme 219) A method for sequencing a DNA or RNA template, the method comprising the following steps:

[0576] a. Forming a DNA polymerase reaction composition comprising:

[0577] i. DNA or RNA template,

[0578] ii. Replication primers compounded with the template,

[0579] iii. DNA polymerase,

[0580] iv. A mixture of nucleotides or nucleotide analogs,

[0581] v. At least one of the compounds according to any one of embodiments 100 to 204;

[0582] b. Incubating the DNA polymerase reaction composition under conditions that allow DNA polymerization, wherein at least one compound according to any one of embodiments 100 to 204 increases the rate, fidelity, or sustained synthesis capability of the DNA polymerase reaction; and

[0583] c. Determine the sequence of the nucleotide or nucleotide analog in the resulting nucleotide or nucleotide analog polymer.

[0584] (Implementation Scheme 220) The method according to Implementation Scheme 219, wherein the mixture of nucleotide analogs comprises nucleoside triaminophosphate, wherein each nucleoside triaminophosphate comprises a nucleobase selected from the group consisting of adenine, guanine, thymine, and cytosine, and a polymer chain portion, wherein a first end of the polymer chain portion is connected to the nucleobase, and a second end of the polymer chain portion is connected to the α-phosphate of the nucleoside triaminophosphate to provide nucleotide analog extension by cleavage of the aminophosphate bond.

[0585] Implementation Scheme 221) The method according to Implementation Scheme 220, wherein the DNA polymerase is DPO4 or a variant thereof.

[0586] Implementation Scheme 222) The method according to Implementation Scheme 219, wherein the polymer of the resulting nucleotide analog is a scalable polymer.

[0587] (Implementation Scheme 223) The method according to any one of Implementation Schemes 219 to 222 further includes the step of contacting the expandable polymer with an aminophosphate cleaving agent to produce an expanded polymer of a nucleotide analog.

[0588] (Implementation Scheme 224) The method according to Implementation Scheme 220, wherein the polymer chain portion of each of the nucleotide analogs comprises a reporter portion specific to the nucleobases of the analog.

[0589] (Implementation Scheme 225) According to the method of implementation scheme 224, each of the reporting portions generates a characteristic electronic signal.

[0590] Implementation Scheme 226) The method according to Implementation Scheme 219, wherein the step of determining the nucleotide analog sequence includes the step of shifting the extended polymer of the nucleotide analog through a nanopore.

[0591] (Implementation Scheme 227) Use of the compound according to any one of Implementation Schemes 100 to 204 in sequencing DNA or RNA templates.

[0592] (Embodiment 228) A composition comprising: any one of the compounds described in embodiments 1 to 105; and a molecular crowding agent.

[0593] (Scheme 229) The composition according to Scheme 228, wherein the molecular crowding agent is a polyalkylene glycol.

[0594] (Embodiment 230) A composition comprising: any one of the compounds described in embodiments 100 to 204; and a buffer solution.

[0595] Implementation Scheme 231) The composition according to Implementation Scheme 230, wherein the buffer comprises one or more components selected from MnCl2, buffer, salt, sugar, single-chain binding protein (SSB), imidazole, pyrazole, triazole, betaine, molecular crowding agent, dimethyl sulfoxide (DMSO), alkyl diol, glycerol, N-methyl-2-pyrrolidone (NMP), acetamide, butylated hydroxyanisole (BHA), polyphosphate and / or urea.

[0596] (Scheme 232) The composition according to Scheme 231, wherein the salt is selected from the group consisting of: NaCl, NaBr, NaOAc, NaF, sodium formate, sodium dihydrogen phosphate, disodium hydrogen phosphate, NaSO4, sodium carbonate, sodium bicarbonate, sodium hexanoate, monosodium glutamate, sodium perchlorate, CsCl, LiCl, LiOAc, LiF, lithium carbonate, LiPO4, KCl, KOAc, KF, KSO4, potassium dihydrogen phosphate, dipotassium hydrogen phosphate, potassium carbonate, potassium bicarbonate, potassium glutamate, NH4Cl, NH4F, NH4OAc, NH4SO4, NH4Br, ammonium citrate, ammonium carbonate, ammonium bicarbonate, ammonium sulfite, ammonium glutamate, ammonium dihydrogen phosphate, tetramethylammonium chloride (TMACl), trimethylamine N-oxide (TMAO), tetraethylammonium chloride (TEACl), guanidine chloride, guanidine thiocyanate, and guanidine carbonate.

[0597] (Scheme 233) The composition according to Scheme 231, wherein the salt is an inorganic salt.

[0598] (Scheme 234) The composition according to Scheme 231, wherein the salt is NaCl or KCl.

[0599] The composition according to embodiment 132, wherein the sugar is maltose, trehalose, cellobiose, or sucrose.

[0600] (Scheme 235) The composition according to Scheme 231, wherein the imidazole is a derivative or analog of an imidazole selected from the following: imidazole chloride, imidazole acetate, 1-methylimidazolium, 2-methylimidazolium, 1-ethylimidazolium, 1-ethyl-3-methylimidazolium chloride, 2-methyl-2-imidazoline, 1-butyl-3-methylimidazolium chloride, 1-methylimidazolium chloride, 1-hexyl-3-methylimidazolium salt, 3-octyl-1-methylimidazolium salt, and 1-decyl-3-methylimidazolium salt.

[0601] (Scheme 236) The composition according to Scheme 231, wherein the alkanediol is ethylene glycol, propylene glycol or butanediol, such as propylene glycol.

[0602] (Embodiment 237) The composition according to embodiment 231, wherein the propylene glycol is 1,2-propanediol or 1,3-propanediol, such as 1,2-propanediol. In some embodiments, the butanediol is 1,2-butanediol, 1,3-butanediol, 1,4-butanediol, 2,3-butanediol, 2,4-butanediol or 3,4-butanediol, such as 1,2-butanediol.

[0603] (Embodiment 238) The composition according to embodiment 231, wherein the molecular crowding agent is PEG, such as PEG4k to PEG25k or PEG4k to PEG10k, such as PEG5k, PEG8k or PEG10k.

[0604] (Scheme 239) The composition according to Scheme 231, wherein the polyphosphate is a tripolyphosphate, tetrapolyphosphate, pentapolyphosphate, hexapolyphosphate, trimetaphosphate (TMP), hexametaphosphate (HMP), or polyphosphate 60.

[0605] (Embodiment 240) A composition comprising: any one of the compounds described in embodiments 100 to 204; and a polynucleotide.

[0606] Implementation Scheme 241) The composition according to Implementation Scheme 240, wherein the polynucleotide is a 20 to 60 mer oligonucleotide.

[0607] (Embodiment 242) A composition comprising: any one of the compounds according to embodiments 100 to 204; and a protein.

[0608] (Scheme 243) The composition according to Scheme 242, wherein the protein is a DNA polymerase.

[0609] (Embodiment 244) A composition comprising: any one of the compounds described in embodiments 100 to 204; and a mixture of nucleotides or nucleotide analogs.

[0610] (Embodiment 245) A composition for enhancing the sustained synthesis capacity, fidelity, or rate of a DNA polymerase reaction, comprising: at least one compound according to any one of embodiments 100 to 204; and a mixture of nucleotide analogs.

[0611] (Embodiment 246) A composition comprising a mixture of at least one compound according to any one of embodiments 100 to 204 and a nucleotide analogue, wherein, relative to the same polymerization reaction in which at least one compound according to any one of embodiments 100 to 204 is absent, the at least one compound according to any one of embodiments 100 to 204 increases the quantity and accuracy of nucleotide analogues incorporated into daughter chains during template-dependent polymerization.

[0612] (Effective Implementation 247) The composition according to Effective Implementation 246, wherein the mixture of nucleotide analogs comprises nucleoside triaminophosphate, wherein each nucleoside triaminophosphate comprises a nucleobase selected from the group consisting of adenine, guanine, thymine, and cytosine, and a polymer chain portion, wherein a first end of the polymer chain portion is attached to the nucleobase, and a second end of the polymer chain portion is attached to the α-phosphate of the nucleoside triaminophosphate to provide nucleotide analog extension by cleavage of the aminophosphate bond.

[0613] (Embodiment 248) The composition according to embodiment 247 further comprises at least one buffer component selected from the following: Tris OAc, NH4OAc, PEG, water-miscible organic solvent, polyphosphate 60, NMS and MnCl2.

[0614] (Scheme 249) The composition according to Scheme 247 further comprises a single-chain binding protein.

[0615] (Embodiment 250) The composition according to embodiment 247 further comprises urea.

[0616] Implementation Scheme 251) The composition according to Implementation Scheme 247, wherein the mixture of nucleotide analogs comprises a nucleotide analog containing a detectable marker.

[0617] (Scheme 252) The composition according to Scheme 251, wherein the detectable marker is an optically detectable marker selected from the group consisting of luminescent, chemiluminescent, fluorescent, fluorescent, chromogenic, or chromogenic markers.

[0618] The foregoing and additional features of this disclosure, and how they are obtained, will become apparent, and this disclosure will be best understood by referring to the following more detailed description. All references disclosed herein are incorporated herein by reference in their entirety as if each reference were incorporated separately.

[0619] This brief overview is provided to introduce certain concepts in a simplified form, which will be described in further detail below. Unless otherwise expressly stated, this brief overview is not intended to identify key or essential features of the claimed subject matter, nor is it intended to limit the scope of the claimed subject matter.

[0620] Details of one or more embodiments are set forth in the following description. Features shown or described in conjunction with an exemplary embodiment may be combined with features of other embodiments. Thus, any of the various embodiments described herein may be combined to provide further embodiments. If necessary, aspects of the embodiments may be modified to incorporate concepts from various patents, applications, and disclosures as identified herein to provide other further embodiments. Other features, objects, and advantages will be apparent from the description, drawings, and claims. Attached Figure Description

[0621] Exemplary features, their nature, and various advantages of this disclosure will become apparent from the following detailed description of the accompanying drawings and various embodiments. Non-limiting and non-exhaustive embodiments are described with reference to the accompanying drawings, wherein, unless otherwise stated, the same labels or reference numerals refer to the same parts in the various views. The dimensions and relative positions of the elements in the drawings are not necessarily drawn to scale. For example, the shapes of various elements are selected, enlarged, and positioned to improve the readability of the drawings. Specific shapes of the drawn elements have been selected for ease of identification in the drawings.

[0622] Figures 1A, 1B, 1C, and 1D are simplified diagrams illustrating the key features of generalized XNTPs and their use in extended sequencing (SBX).

[0623] Figure 2 is a schematic diagram illustrating more details of one implementation of XNTP.

[0624] Figure 3 is a schematic diagram illustrating one embodiment of Xpandomer passing through biological nanopores.

[0625] Figure 4 shows the gel containing the primer extension product.

[0626] Figure 5 shows the gel containing the primer extension product.

[0627] Figures 6A and 6B are histograms showing the alignment read groups of nanopore-derived sequences.

[0628] Figures 7A and 7B are histograms showing the alignment read groups of nanopore-derived sequences.

[0629] Figure 8 shows the gel containing the primer extension product.

[0630] Figure 9 shows the gel containing the primer extension product.

[0631] Figure 10 shows the gel containing the primer extension product.

[0632] Figure 11 shows the gel containing the primer extension product.

[0633] Figure 12 shows the gel containing the primer extension product.

[0634] Figure 13 shows the gel containing the primer extension product.

[0635] Figure 14 shows the gel containing the primer extension product.

[0636] Figure 15 shows a series of gels displaying primer extension products from a variety of exemplary PEM compounds, and in particular PEM compounds 277 (lane 2), 51 (lane 3), 73 (lane 4), 114 (lane 5), 311 (lane 6), 301 (lane 7), 331 (lane 8), 273 (lane 9), and 367 (lane 10) (according to the compound numbers listed in Table 1 herein). Detailed Implementation

[0637] It should also be understood that, unless expressly indicated otherwise, in any method claimed herein that includes more than one step or action, the order of the steps or actions of the method is not necessarily limited to the order in which the steps or actions of the method are described.

[0638] As used herein, unless the context clearly indicates otherwise, the singular forms “a / an” and “the” include multiple referents. Similarly, unless the context clearly indicates otherwise, the word “or” is intended to include “and”. The term “include” is defined as inclusive, such as “include A or B” meaning to include A, B, or A and B.

[0639] As used herein in the specification and claims, “or” should be understood to have the same meaning as “and / or” as defined above. For example, when separating items in a list, “or” or “and / or” should be interpreted as inclusive, meaning it includes several elements or at least one element in the list, but also includes more than one element, and optionally includes additional unlisted items. Only terms indicating the opposite, such as “only one of” or “exactly one of”, or “consisting of” as used in the claims, will refer to the inclusion of several elements or exactly one element in the list. In general, the term “or” as used herein should only be interpreted as indicating an exclusive alternative (i.e., “one or the other, but not both”) when preceded by exclusive terms such as “or,” “one of,” “only one,” or “exactly one.” “Constitutes substantially of” as used in the claims should have the ordinary meaning used in the field of patent law.

[0640] As used herein, the terms “comprising,” “including,” and “having” are used interchangeably and have the same meaning. Similarly, “comprising,” “including,” and “having” are used interchangeably and have the same meaning. Specifically, the definition of each term is consistent with the usual definition of “comprising” under U.S. patent law, and therefore each term can be understood as an open-ended term meaning “at least the following” and can also be interpreted as not excluding additional features, limitations, aspects, etc. Thus, for example, “an apparatus having components a, b, and c” means that the apparatus includes at least components a, b, and c. Similarly, the phrase “a method relating to steps a, b, and c” means that the method includes at least steps a, b, and c. Furthermore, although the steps and processes may be outlined in a specific order herein, those skilled in the art will recognize that this order of steps and processes may vary.

[0641] As used herein in the specification and claims, with respect to a list of one or more elements, the phrase “at least one” should be understood to mean at least one element selected from any one or more elements in the list, but does not necessarily include at least one of each element specifically listed in the list, nor exclude any combination of elements in the list. In addition to the elements specifically identified in the list of elements referred to by the phrase “at least one,” this definition also allows for the optional presence of other elements, whether or not they are related to the specifically identified elements. Thus, as a non-limiting example, “at least one of A and B” (or equivalently, “at least one of A or B,” or equivalently, “at least one of A and / or B”) in one embodiment may refer to at least one, optionally including more than one A and the absence of B (and optionally including elements other than B); in another embodiment, it refers to at least one, optionally including more than one B and the absence of A (and optionally including elements other than A); in yet another embodiment, it refers to at least one, optionally including more than one A, and at least one, optionally including more than one B (and optionally including other elements), etc.

[0642] Throughout this specification, references to "an embodiment," "a particular embodiment," etc., indicate that a specific feature, structure, or characteristic described in connection with that embodiment is included in at least one embodiment. Therefore, the phrases "in one embodiment" or "in a particular embodiment" appearing in different places throughout this specification do not necessarily refer to the same embodiment. Furthermore, specific features, structures, or characteristics may be combined in any suitable manner in one or more embodiments.

[0643] This disclosure can be more readily understood by referring to the following detailed description of embodiments of this disclosure and examples included herein. Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains.

[0644] In one aspect of this disclosure, compounds having formula (I), such as PEM compounds, are provided:

[0645] (I),

[0646] Or its solvates, hydrates, tautomers, chelates or salts,

[0647] in

[0648] a is 0 or an integer in the range of 1 to 4;

[0649] a' is 0 or an integer in the range of 1 to 4;

[0650] m is 1, 2, or 3;

[0651] m' is 1, 2, or 3;

[0652] n is 0, 1, or 2;

[0653] p is 0, 1, or 2;

[0654] Z is a 5- or 6-membered heteroaromatic ring containing one, two, three or four heteroatoms selected from O, N or S, wherein each Z may be independently substituted with one or more halogens and / or one or more C1-C4 alkyl groups;

[0655] L is a linking group;

[0656] M is independently selected from hydrogen, halogens, and C1-C4 alkyl groups each time it appears;

[0657] Ar1, each time it appears, is independently selected from optionally substituted phenyl, pyridine, bipyridine, tripyridine, pyrazine, pyridazine, furan, dibenzofuran, thiophene, pyrrole, selenophene, naphthalene, fluorene, phenanthrene, cyclophosphine, phthalazine, quinazoline, quinoxaline, naphthidine, phenanthrene-rholine, purine, and carbazole.

[0658] The substituents of Ar1 are independently selected each time they appear from halogens, –OH, –CN, –NO2, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 heteroalkyl, C1-C6 cycloalkyl, –OR 0 –CONH2, –C(O)NR 1 R 1' –C(O)(CH2) a NR 1 R 1' –NR 1 R 1' –NR 1 C(O)R 3 –C(O)SR 3 –COR 3 –CO(CH2) a OC(O)R 3 –OC(O)R 3 –C(O)OR 3 –C–O–R 3 , thiols, -R 4 -H、–SOR 1 –S(O)2R 1 –S(O)2NR 1 R 1' –CH2–NR1 S(O)2R 3 –NR 1 S(O)2R 3 and –C(CH3)=N–(phenyl)-O-CH2-C-CH;

[0659] R 0 Each time it appears, it is independently selected from C1-C6 alkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl.

[0660] R 1 and R 1' Each time it appears, it is independently selected from H, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C 20 Heteroalkyl, C1-C 10 Heteroalkyl –NH2, C2-C6 alkenyl, C2-C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic (e.g., via one or more R... 3 Substituent groups), substituted or unsubstituted aryl groups, substituted or unsubstituted arylalkyl groups, and substituted or unsubstituted heteroaryl groups, benzyl groups, –C(=NH)NH2, –CH2CO2R 0 –C(H)(CH) a (COOH))2, -CH2C(O)NHCH2CO2H, -CH2CH2OH, -CH2CH2NHC(O)R 3 –(CH2) a C(O)NH(CH2) a C(O)OR 3 –(CH2) a –CN, spermine, –(CH2) a –NH2、–C(O)R 3 –(CH2) a C(O)OR 3 –(CH2) a OR 3 –C(H)((CH2) a OH) a' –CHCH(OH)(CH2) a OH, –(CH2) a OH, C1-C6–C(O)OH, can be substituted (e.g., via R) 3 (Substituted group) or unsubstituted –(CH2) a - Heterocyclic rings

[0661] , ;and

[0662] Where R 1 and R 1' Together they form substituted or unsubstituted heterocycles, including but not limited to aza-butanes, pyrrolidines, piperidines, piperazines, morpholines, etc.

[0663] or ;

[0664] R 2 Each time it appears, it is independently selected from C2-C6 alkyl, C1-C6 haloalkyl, C1-C6 heteroalkyl, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, and substituted or unsubstituted haloalkoxy.

[0665] R 3 Each time it appears, it is independently selected from H, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 heteroalkyl, –C1-C6–OH, –C1-C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine;

[0666] R 4 Each time it appears, it is independently selected from alkylene groups interrupted by one or more heteroatoms, wherein the heteroatoms are O, S, NH or a combination thereof;

[0667] Y is independently selected from Ar2, –C(O)–Ar2, and –(CH2) each time it appears. a Ar2, –(CH2)3PO(OEt)2 or –CH2CO2Me;

[0668] Ar2 is independently selected each time it appears from: (i) substituted or unsubstituted 5- and 6-membered monocyclic aromatic or heterocyclic rings; (ii) substituted or unsubstituted 9- and 10-membered fused bicyclic rings comprising two monocyclic rings, wherein at least one of the two monocyclic rings is an aromatic or heterocyclic ring; and (iii) substituted or unsubstituted 13- and 14-membered fused tricyclic rings comprising three monocyclic rings together, wherein at least one of the three monocyclic rings is an aromatic or heterocyclic ring.

[0669] Each Ar2 is independently transmitted via G 1 G2 G 3 G 4 and G 5 Replace, of which:

[0670] When Ar2 is monosubstituted, G 1 Each time it appears, it is independently selected from oxo, C1-C6 alkyl, C1-C6 haloalkyl, –E–O–R 3 –E–C(R) 1 (R) 1' (R) 3 –NH2, –NO2, –SO3R 3 SO3O – 、–SO3N(H)(R 1 ), –E–C(O)R 3 , -E-CO2H, –B(OH)2, –C(O)NR 1 R 1' -E-PO(OR) 1 )2, and through G 2 G 3 G 4 and G 5 Substituted aryl groups;

[0671] G 2 G 3 G 4 and G 5 Each time it appears, it is independently selected from groups that are absent or selected from groups containing the following: halogen, –CN, –NO2, –SO3R. 3 C1-C6 alkyl, C1-C6 haloalkyl, –E–O–R 3 –E–(CH2) a C(O)R 3 , –E–CO2H, –E–CHO, –E–C(O)R 3 , –E–C(O)NH(OH), –E–C(O)NHR 1 –E–C(O)N(H)C(H)(R 1 (R) 1' ), –E–C(O)N(R 1 (CH2) a P(O)(O)OH)2、–E–C(O)N(R 1 (CH2) a SO3H), –E–C(O)NR 1 R 1' –E–NR 1 R 1'–E–OR 2 –C(O)–heterocyclic alkyl (where the heterocyclic alkyl is substituted or unsubstituted), –C(O)–N(H)–CH(COOH)((CH2) a – heteroaryl) (wherein the heteroaryl group is substituted or unsubstituted), wherein E is independently selected from direct bond and C1-C6 alkylene group each time it appears.

[0672] In some embodiments, Z is not a triazole. In some embodiments, Z is not a 1,2,3-triazole or a 1,2,4-triazole.

[0673] In some embodiments, Z is a diazole (e.g., imidazole, pyrazole, pyrimidine, pyridazine, pyrazine). In other embodiments, Z is a pyrrole, thiophene, or triazole (e.g., 1,2,3-triazole; 1,2,4-triazole). In still other embodiments, Z is a thiazole. In still other embodiments, Z is an isoxazole. In yet still further embodiments, Z is a tetraazole.

[0674] In a further implementation scheme, Z is selected from imidazole, thiazole, and isoxazole.

[0675] In some embodiments, each Z is independently substituted with one “M” moiety. In other embodiments, each Z is independently substituted with two “M” moiety. In still other embodiments, each Z is independently substituted with three “M” moiety. In some embodiments, one or more of the compounds having formula (I) can be polymerized (e.g., via cationic polymerization, such as by incubating one or more PEMs with KCl). In some embodiments, this disclosure provides a composition comprising molecules of two or more compounds having any of formulas (I), (IA), (IB), (IC), and / or (ID).

[0676] In some embodiments, the compound of formula (I) has either formula (IA) or (IB):

[0677] (IA) or

[0678] (IB),

[0679] Or its solvates, hydrates, tautomers, chelates or salts,

[0680] Ar1, Ar2, M, Z, and Y are as defined above. In some embodiments, Z is not a triazole. In some embodiments, Z is not a 1,2,3-triazole or a 1,2,4-triazole. In other embodiments, Z is a triazole. In some embodiments, one or more compounds having any of formula (IA) or (IB) can be polymerized (e.g., via cationic polymerization, such as by incubating one or more PEMs with KCl). In some embodiments, this disclosure provides a composition comprising molecules of two or more compounds having any of formula (IA) or (IB).

[0681] In some embodiments, the compound of formula (I) has formula (IC):

[0682] (IC)

[0683] Or its solvates, hydrates, tautomers, chelates or salts,

[0684] in:

[0685] When X is C, W is N, or when X is N, W is C;

[0686] It is a single bond or a double bond, wherein the double bond begins at either W or X where the carbon atom is present; and

[0687] m, m', n, p, Ar1, L, and Y are as defined above.

[0688] In some embodiments, one or more of the compounds having the formula (IC) can be polymerized (e.g., via cationic polymerization, such as by incubating one or more PEMs with KCl). In some embodiments, this disclosure provides a composition comprising molecules of two or more compounds having the formula (IC).

[0689] In some embodiments, compounds having formula (IC) have the structure of formula (ID):

[0690] (ID),

[0691] Ar1, Ar2, and M are as defined above.

[0692] In some embodiments, one or more of the compounds having formula (ID) can be polymerized (e.g., via cationic polymerization, such as by incubating one or more PEMs with KCl). In some embodiments, this disclosure provides a composition comprising molecules of two or more compounds having formula (ID).

[0693] As used in the specification and appended claims, unless otherwise stated, the following terms have the meanings indicated.

[0694] Some of the chemical groups named in this article are preceded by an abbreviation indicating the total number of carbon atoms found in the specified chemical group. For example, C1-C4 alkyl can be alternatively written as C 1-4 Alkyl groups describe alkyl groups having at least one to up to four carbon atoms, while C4-C 12 Cycloalkylalkyl (also denoted as C) 4-12 Cycloalkyl (C1-C6) describes a cycloalkyl group having a total of 4 to 12 carbon atoms. The total number of carbons in the abbreviation does not include carbons that may be present in substituents of the group. For example, C1-C6 alkyl refers to an alkyl group containing one to six carbon atoms; C1-C6 haloalkyl refers to a haloalkyl group containing one to six carbon atoms; and C1-C6 alkylene refers to an alkylene binary radical containing one to six carbon atoms.

[0695] In addition to the foregoing, as used in the specification and appended claims, unless otherwise stated, the following terms shall have the meanings indicated:

[0696] As used herein, unless otherwise stated, the term "alkyl" itself, or as part of another substituent, refers to a straight (i.e., unbranched) or branched chain or a combination thereof, which may be a fully saturated, monounsaturated, or polyunsaturated group, and may include a group having a specified number of carbon atoms (i.e., C1-C1). 10 Alkyl groups refer to divalent and polyvalent groups (1 to 10 carbons). "Alkyl" is uncyclized. Examples of saturated hydrocarbon groups include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl, isobutyl, sec-butyl, (cyclohexyl)methyl, and homologues and isomers of n-pentyl, n-hexyl, n-heptyl, and n-octyl. Unsaturated alkyl groups are alkyl groups having one or more double or triple bonds. Examples of unsaturated alkyl groups include, but are not limited to, vinyl, 2-propenyl, crotonyl, 2-isopentenyl, 2-butadienyl, 2,4-pentadienyl, 3-(1,4-pentadienyl), ethynyl, 1-propynyl, 3-propynyl, 3-butynyl, and higher homologues and isomers. An alkoxy group is an alkyl group attached to the rest of the molecule via an oxygen atom (–O–).

[0697] As used herein, unless otherwise stated, the term "heteroalkyl" on its own or in combination with another term refers to a stable straight or branched chain consisting of at least one carbon atom and at least one heteroatom selected from the group consisting of O, N, P, Si, and S, wherein nitrogen, phosphorus, and sulfur atoms may optionally be oxidized, and the nitrogen heteroatom may optionally be quaternized. The heteroatoms O, N, P, S, and Si may be located at any internal position within the heteroalkyl group or at a position where the alkyl group is attached to the remainder of the molecule. The heteroalkyl group is uncyclized. Examples include, but are not limited to: —CH2—CH2—O—CH3, —CH2—CH2—NH—CH3, —CH2—CH2—N(CH3)—CH3, —CH2—S—CH2—CH3, —CH2—O—CH3, —S(O)—CH3, —CH2—CH2—S(O)2—CH3, —CH═CH—O—CH3, —Si(CH3)3, —CH2—CH═N—OCH3, —CH═CH—N(CH3)—CH3, —O—CH2—CH3, and —CN. At most two heteroatoms can be consecutive, for example, —CH2—NH—OCH3.

[0698] "Alkenyl" refers to a straight-chain or branched hydrocarbon chain group consisting only of carbon and hydrogen atoms, containing at least one double bond, optionally having a specified number of carbon atoms, such as two to twelve carbon atoms, two to eight carbon atoms, two to six carbon atoms, or two to four carbon atoms, and connected to the rest of the molecule by single bonds, such as vinyl, propenyl, butenyl, pentenyl, penta-1,4-dienyl, etc.

[0699] "Alynyl" refers to a straight-chain or branched hydrocarbon chain group consisting only of carbon and hydrogen atoms, containing at least one triple bond, optionally having a specified number of carbon atoms, such as two to twelve, two to eight, two to six, or two to four carbon atoms, and connected to the rest of the molecule by single bonds, such as vinyl, propenyl, butenyl, pentenyl, penta-1,4-dienyl, etc. "Halogenated" refers to bromine, chlorine, fluorine, or iodine.

[0700] "Haloalkyl" refers to an alkyl group as defined above, substituted with one or more halogenated groups as defined above, such as trifluoromethyl, difluoromethyl, trichloromethyl, 2,2,2-trifluoroethyl, 1-fluoromethyl-2-fluoroethyl, 3-bromo-2-fluoropropyl, 1-bromomethyl-2-bromoethyl, etc. Similarly, "haloalkenyl" refers to an alkenyl group as defined herein, substituted with one or more halogenated groups as defined herein, and "haloalkynyl" refers to an alkynyl group as defined herein, substituted with one or more halogenated groups as defined herein.

[0701] "alkylene" or "alkylene chain" refers to a straight or branched divalent hydrocarbon chain in which the remainder of a molecule is connected to a group consisting only of carbon and hydrogen, is unsaturated, and optionally has a specified number of carbon atoms. Examples include methylene, ethylene, propylene, n-butene, etc. The alkylene chain is connected to the remainder of the molecule by a single bond and to the group by a single bond. The connection point between the alkylene chain and the remainder of the molecule and to the group can be one carbon or any two carbons within the chain. Similar to alkyl groups, unsaturation can be introduced into the alkylene chain to provide an unsaturated alkylene chain. When unsaturation is introduced into the alkylene chain, the resulting group may be referred to as an unsaturated alkylene group or chain, wherein the unsaturated alkylene chain is typically referred to as an alkenyl group (having at least one carbon-carbon double bond) and an alkyne group (having at least one carbon-carbon triple bond). In one embodiment, and when specified, the alkylene chain in the compounds of this disclosure may be or include an unsaturated alkylene chain.

[0702] A "denylene" or "denylene chain" refers to a straight-chain or branched divalent hydrocarbon chain that connects the rest of a molecule to a group consisting only of carbon and hydrogen, containing at least one double bond and optionally having a specified number of carbon atoms (e.g., two to twelve carbon atoms). Examples of denylene groups include vinylene, propenylene, n-butenylene, etc. The denylene chain is connected to the rest of the molecule by a single bond and to the group by a double or single bond. The connection point between the denylene chain and the rest of the molecule, and to the group, can be one carbon or any two carbons within the chain.

[0703] "Aryl" refers to a cyclic group comprising at least 5 ring atoms, optionally 1-6 heterocyclic atoms selected from O, S, and N, and at least one aromatic ring. A 5-membered monocyclic aromatic ring contains 5 ring atoms selected from carbon and heteroatoms, while a 6-membered monocyclic aromatic ring contains 6 ring atoms selected from carbon and heteroatoms. An exemplary monocyclic aromatic ring with 5 members is pyrrole, and an exemplary monocyclic aromatic ring with 6 members is pyridine. Aryl groups can be, for example, monocyclic, bicyclic, tricyclic, or tetracyclic ring systems, which may include fused or bridged ring systems. Carbocyclic aryl groups contain only carbon atoms on the ring atoms, examples of which include, but are not limited to, aryl groups derived from anthracene, acenaphthene, phenanthrene, anthracene, azulene, benzene, benzoxene, fluoranthene, fluorene, asymmetric indole, symmetric indole, indene, naphthalene, phenanthracene, heptamethrin, pyrene, and triphenylene. In one embodiment, the aryl group is phenyl or naphthyl, and in another embodiment, it is phenyl. When the aryl group includes a non-carbon ring atom, such as oxygen, sulfur, or nitrogen, the aryl group may be referred to as a heteroaryl group. A heteroaryl group can be, for example, a monocyclic, bicyclic, tricyclic, or tetracyclic ring system, which may include fused or bridged ring systems. The nitrogen, carbon, or sulfur atom in the heteroaryl group may optionally be oxidized; the nitrogen atom may optionally be quaternized.

[0704] An "arylalkyl" group is an alkyl, alkenyl, or alkynyl group as defined above, wherein its hydrogen atoms are replaced by an aryl group as defined above. Exemplary arylalkyl groups include benzyl (−CH2phenyl), phenylethyl (−CH2CH2phenyl), and phenylethylene (−CH=CH–phenyl) groups and fused (cycloalkylaryl) alkyl groups such as 4-ethyl-indenyl. The arylalkyl group may be substituted on the aryl moiety, the alkyl, alkenyl, or alkynyl moiety, or both.

[0705] "Fused rings" refers to ring systems where rings are fused together, meaning they share two adjacent ring atoms. A fused ring containing two fused 5- or / or 6-membered monocyclic rings is a bicyclic ring system, where each ring is a monocyclic ring and independently has 5 or 6 ring atoms, and the two rings are fused because they share two ring atoms. For example, naphthalene is a 10-membered fused ring system formed by the fusion of two 6-membered monocyclic rings (benzene). Naphthalene is bicyclic because it contains two (bi = 2) rings. As another example, 1,3-benzothiazole is a 9-membered fused ring system formed by the fusion of a 6-membered ring (benzene) and a 5-membered ring (1,3-thiazole). 1,3-benzothiazole is bicyclic because it contains two rings.

[0706] "Carbocyclic group" refers to a stable 3- to 18-membered aromatic or non-aromatic cyclic group consisting of 3 to 18 carbon atoms. Unless otherwise specified in the specification, the carbocyclic group can be a monocyclic, bicyclic, tricyclic, or tetracyclic system, which may include fused or bridged ring systems, and may be partially or fully saturated. Non-aromatic carbocyclic groups include cycloalkyl groups, while aromatic carbocyclic groups include aryl groups.

[0707] "Cycloalkyl" refers to a stable non-aromatic monocyclic or polycyclic hydrocarbon group consisting only of carbon and hydrogen atoms. It may include fused or bridged ring systems, have three to fifteen carbon atoms, preferably three to ten, and be saturated or unsaturated, connected to the rest of the molecule by a single bond. Monocyclic groups include, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl. Polycyclic groups include, for example, adamantyl, norbornyl, decahydronaphthyl, 7,7-dimethyl-bicyclo-[2.2.1]heptyl, etc.

[0708] "Heterocyclic group" refers to a stable 3- to 18-membered aromatic or non-aromatic ring group, consisting of two to twelve carbon atoms and one to six heteroatoms selected from the group consisting of nitrogen, oxygen, and sulfur. Unless otherwise specified in the specification, the heterocyclic group can be a monocyclic, bicyclic, tricyclic, or tetracyclic ring system, which may include fused or bridged ring systems; the nitrogen, carbon, or sulfur atoms in the heterocyclic group may optionally be oxidized; the nitrogen atom may optionally be quaternized; and the heterocyclic group may be partially or completely saturated. Examples of non-aromatic heterocyclic groups include, but are not limited to, dioxolane, thiophene[1,3]dithiaalkyl, decahydroisoquinolinyl, imidazolinyl, imidazoalkyl, isothiazolyl, isoxazolyl, morpholinyl, octahydroindolyl, octahydroisoindolyl, 2-oxopiperazinyl, 2-oxopiperidinyl, 2-oxopiperidinyl, oxazolyl, piperidinyl, piperazinyl, 4-piperidinyl, pyrrolyl, pyrazolyl, pyrazolpyrimidinyl, quininecycloyl, thiazoalkyl, tetrahydrofuranyl, trioxoxalyl, trithiaalkyl, triazinanyl, tetrahydropyranyl, thiomorpholinyl, thiomorpholinyl, 1-oxo-thiomorpholinyl, and 1,1-dioxo-thiomorpholinyl.

[0709] Optionally, but only in the context of detailed description, each of the alkyl, alkenyl, alkylene, alkenylene, carbocyclic, cycloalkyl, aryl, heterocyclic, and heteroaryl groups in the PEM compounds of this disclosure may be substituted by one or more unsubstituted (e.g., the alkyl substituent on the alkyl group is not further substituted, i.e., the alkyl substituent is an unsubstituted alkyl) substituents selected from the group consisting of: alkyl, alkenyl, halogenated, haloalkyl, haloalkenyl, cyano, oxo, thio, nitro, aryl, aralkyl, cycloalkyl, cycloalkylalkyl, heterocyclic, heterocyclic alkyl, heteroaryl, heteroaryl, heteroarylalkyl, –R b —OR a –R b –OC(O)—R a –R b –N(R a )2、–R b –C(O)R a –R b –C(O)OR a –R b –C(O)N(R a )2、–R b –N(R a )C(O)OR c –R b –N(R a )C(O)R c –R b –N(R a S(O) t Rc (where t is 1 to 2), –R b —N=C(OR a )R a –R b –S(O) t OR c (where t is 1 to 2), –R b –S(O) s R c (where s is 0 to 2) and –R b –S(O) t N(R a )2 (where t is 1 to 2), where each R a Independently, it is hydrogen, alkyl, alkenyl, haloalkyl, cycloalkyl, cycloalkylalkyl, aryl, aralkyl, heterocyclic, heterocyclic alkyl, heteroaryl, or heteroarylalkyl; R b Each is independently a direct bond or a straight-chain or branched alkylene or alkenylene chain; and each R c It can be alkyl, alkenyl, haloalkyl, cycloalkyl, cycloalkylalkyl, aryl, aralkyl, heterocyclic, heterocyclic alkyl, heteroaryl or heteroarylalkyl. In some embodiments, the substituents may be independently selected from alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, cycloalkynyl, aryl, heteroaryl, heterocycloalkyl, aralkyl, heteroarylalkyl, (heterocycloalkyl)alkyl, hydroxyl, protected hydroxyl, alkoxy, aryloxy, acyl, mercapto, alkylthio, arylthio, cyano, cyanate, halogen, thiocarbonyl, O-carbamoyl, N-carbamoyl, O-thiocarbamoyl, N-thiocarbamoyl, C-amide, N-amide, S-sulfonamide, N-sulfonamide, C-carboxyl, protected C-carboxyl, O-carboxyl, isocyanothio, cyanothio, isothiocyanothio, nitro, silyl, thio, sulfinyl, sulfonyl, haloalkyl, haloalkoxy, trihalomethanesulfonyl, trihalomethanesulfonamide, ether, amino (e.g., monosubstituted or disubstituted amino) and their protected derivatives. Any of the above groups may include one or more heteroatoms, including O, N, or S. For example, when partially substituted with an alkyl group, the alkyl group may contain a heteroatom selected from O, N, or S (e.g., –(CH2–CH2–O–CH2–CH3)).

[0710] "Amino" refers to the -NH2 group. "Cyano" refers to the -CN group. "Hydroxy" refers to the -OH group. "Nitro" refers to the -NO2 group. "Oxo" refers to the =O substituent. "Thio" refers to the =S substituent. "Trifluoromethyl" refers to the -CF3 group. "Trifluoromethoxy" refers to the -OCF3 group. Thiols, also known as mercapto groups, refer to the -SH group.

[0711] "Acyl" refers to the group –C(O)R, which can also be written as –C(=O)R, where R is an alkyl, aralkyl, carbocyclic, aryl, heteroaryl, or heterocyclic group. For example, when R is methyl, the acyl group can be called an acetyl group.

[0712] "Alkoxy" refers to a group of the formula –OR, where R is an alkyl or haloalkyl group. In one embodiment, the alkoxy group contains up to six carbon atoms. Exemplary alkoxy groups include methoxy and ethoxy groups. Halogen-substituted alkoxy groups may be referred to herein as haloalkoxy groups, which include, for example, trifluoromethoxy, trichloromethoxy, etc.

[0713] "Heteroeneyl" or "heteroeneyl chain" refers to a straight or branched divalent hydrocarbon chain in which the rest of the molecule is attached to a group consisting of carbon and hydrogen and at least one heteroatom selected from N, O and S.

[0714] "Haloalkoxy" refers to an alkoxy group that is substituted by one or more halo groups as defined above, such as trifluoromethoxy, difluoromethoxy, trichloromethoxy, 2,2,2-trifluoroethoxy, 3-bromo-2-fluoropropoxy, etc. The alkoxy moiety of the haloalkoxy group may be optionally substituted, as defined above.

[0715] "N-Heterocyclic group" refers to a heterocyclic group containing at least one nitrogen atom. The N-heterocyclic group may optionally be substituted, as described above for heterocyclic groups.

[0716] "Heterocyclic alkyl" refers to the formula –R b R h The group, wherein R b It is an alkylene chain as defined above and R h It is a heterocyclic group as defined above, and if the heterocyclic group is a nitrogen-containing heterocyclic group, the heterocyclic group may be attached to an alkyl group on the nitrogen atom. The alkylene chain of the heterocyclic alkyl group may optionally be substituted, as defined above for the alkylene chain. The heterocyclic moiety of the heterocyclic alkyl group may optionally be substituted, as defined above for the heterocyclic group.

[0717] "N-Heteroaryl" refers to a heteroaryl group as defined above containing at least one nitrogen atom, wherein the connection point between the heteroaryl group and the rest of the molecule is through a nitrogen atom in the heteroaryl group. The N-heteroaryl group may optionally be substituted, as described above for heteroaryl groups.

[0718] "Heteroarylalkyl" refers to the formula –R b R i The group, wherein R bIt is an alkylene chain as defined above and R i A heteroaryl group is defined herein as such. The heteroaryl moiety of a heteroarylalkyl group may optionally be substituted, as defined herein as a heteroaryl group. The alkylene chain moiety of a heteroarylalkyl group may optionally be substituted, as defined herein as an alkylene chain. Similarly, an arylalkyl group refers to a heteroarylalkyl group in which the heteroaryl moiety is replaced by the corresponding carbocyclic aryl group, i.e., the heteroatom is replaced by a carbon atom, and adjustments are made as needed for hydrogen substitution.

[0719] "Hydroxyalkyl" refers to the formula –R b OH groups, where R b It is an alkylene chain as defined herein. The –OH (hydroxyl, also known as hydroxyl) group may be attached to any carbon atom in the alkylene chain. The alkylene chain portion of the heteroarylalkyl group may optionally be substituted, as defined above for the alkylene chain.

[0720] The PEM compounds with acidic or basic groups described herein are generally used as free acids or free bases. Alternatively, PEM compounds with acidic or basic groups can be used in the form of salts, such as acid or base addition salts. Acid addition salts of free amino compounds can be prepared by methods known in the art and can be formed from organic and inorganic acids. Suitable organic acids include maleic acid, fumaric acid, benzoic acid, ascorbic acid, succinic acid, methanesulfonic acid, acetic acid, trifluoroacetic acid, oxalic acid, propionic acid, tartaric acid, salicylic acid, citric acid, gluconic acid, lactic acid, mandelic acid, cinnamic acid, aspartic acid, stearic acid, palmitic acid, glycolic acid, glutamic acid, and benzenesulfonic acid. Suitable inorganic acids include hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, and nitric acid. Base addition salts include those that form with formic acid anions and those that form with organic and inorganic cations, such as those selected from alkali metals and alkaline earth metals (e.g., lithium, sodium, potassium, magnesium, barium, and calcium), as well as ammonium ions and their substituted derivatives (e.g., dibenzylammonium, benzylammonium, 2-hydroxyethylammonium, etc.). Therefore, the term "salt" in the description of PEM compounds herein is intended to encompass any and all salt forms.

[0721] The PEM compounds disclosed herein may be in the form of chelates. A chelate is a compound containing an organic ligand (e.g., a triazole-Ar group) bonded to a central metal atom at two or more sites.

[0722] Regarding stereoisomers, the PEM compounds described herein may have one or more chiral (or asymmetric) centers, thus producing enantiomers, diastereomers, and other stereoisomers that can be defined according to absolute stereochemistry as (R)- or (S)-. When the compounds described herein contain an alkene double bond or other geometrically asymmetric centers, unless otherwise stated, it is intended to indicate that these compounds simultaneously contain both E and Z geometric isomers (e.g., cis or trans). Similarly, unless otherwise stated, all possible isomers, as well as their racemic and optically pure forms, and also all tautomers, are included. Thus, various stereoisomers and mixtures thereof are contemplated, including “enantiomers,” which refer to two stereoisomers whose molecules are non-overlapping mirror images of each other. Therefore, compounds may exist in any isomeric form, including racemates, racemic mixtures, and as individual enantiomers or diastereomers.

[0723] Furthermore, some crystalline forms of PEM compounds can exist as polymorphs, which are also considered herein. Additionally, some PEM compounds may also form solvates with water or other organic solvents. Such solvates are similarly included within the scope of the compounds described herein.

[0724] As those skilled in the art will understand, any of the above-described compounds can be doped with radioactive isotopes. Therefore, compounds using the same isotopic labels as those described herein, wherein one or more atoms are replaced by atoms with atomic masses or mass numbers different from those commonly found in nature, are also contemplated. Examples of isotopes that can be doped into these compounds include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, fluorine, and chlorine. Thus, reference to an element, such as hydrogen (H) or carbon (C), is intended to encompass all its isotopes. For example, the name C (carbon) includes... 12 C 13 C or 14 C and its mixtures, while H (hydrogen) includes 1 H, 2 H and 3 H and its mixtures, as well as O (oxygen) including 16 O and 18 O and its mixtures, as well as N (nitrogen) including 14 N and 15 N and its mixtures, and so on, as well as other atoms. Isotopically labeled PEM compounds can be used to trace PEM compounds or portions thereof in processes such as assays.

[0725] In some embodiments, the PEM compound of any of formulas (I), (IA), (IB), (IC), and (ID) comprises an Ar1 moiety, wherein, in some embodiments, Ar1 is an aryl group, also referred to as an aromatic moiety. The aromatic moiety may be a carbocyclic or heterocyclic aromatic moiety, wherein each aromatic ring atom is a carbon in the carbocyclic aromatic moiety, and at least one of the aromatic ring atoms is nitrogen, oxygen, or sulfur in the heterocyclic aromatic moiety.

[0726] In some embodiments, Ar1 comprises 1 to 6 rings, wherein up to six ring atoms are selected from oxygen, sulfur, and nitrogen, and the remainder are carbon atoms. Optionally, the Ar1 moiety may comprise 1 to 5 rings, wherein up to five ring atoms are selected from oxygen, sulfur, and nitrogen. In other embodiments, the Ar1 group may comprise 1 to 4 rings, wherein up to four ring atoms are selected from oxygen, sulfur, and nitrogen. As yet another embodiment, the Ar1 moiety may comprise 1 to 3 rings, wherein up to three ring atoms are selected from oxygen, sulfur, and nitrogen. As another example, Ar1 may comprise 1 to 2 rings, wherein up to three ring atoms are selected from oxygen, sulfur, and nitrogen. As described herein, each ring may be independently a five-membered ring (i.e., five ring atoms forming a ring), a six-membered ring, or a seven-membered ring, and in some embodiments, each ring is a five-membered ring or a six-membered ring.

[0727] In some embodiments, A1 is a carbocyclic aromatic moiety. In some embodiments, the carbocyclic moiety may contain one (e.g., benzene), two (e.g., naphthalene, azulene), three (e.g., acenaphthene, fluorene), four (e.g., fluoranthene, acethracene), five (e.g., pentanebenzene, styrene), or six (e.g., hexabenzene) aromatic rings. For convenience, the Ar1 group is exemplified herein by naming it in its unsubstituted form (e.g., benzene), although in the compounds disclosed herein, the Ar1 group is the corresponding group, for example, when m is 2 and Ar1 is additionally unsubstituted, with two ring hydrogens substituted by a triazole group. For example, Ar1 can be a monocyclic carbocyclic moiety, i.e., phenyl, also known as a C6 aromatic moiety. As another example, Ar1 can be a bicyclic carbocyclic moiety, such as naphthyl, which is C6. 10 Fang ethnic group section.

[0728] Non-limiting examples of the aromatic Ar1 portion are heterocyclic aromatic portions, which may also be referred to as heteroaryl groups. In some embodiments, any heterocyclic portion may contain one, two, three, four, five, or six aromatic rings in addition to containing one, two, three, four, five, or six heteroatoms (i.e., atoms selected from nitrogen, sulfur, and oxygen other than carbon). Optionally, the heteroatoms (if present) are nitrogen, oxygen, sulfur, or selenium. For example, the aromatic portion may be a monocyclic heterocyclic portion, such as pyridyl (which is a six-membered C5 aromatic portion) or pyrazinyl (which is a six-membered C4 aromatic portion). As another example, the aromatic portion may be a bicyclic heterocyclic portion, such as quinolinyl or isoquinolinyl (which are ten-membered C9 aromatic portions) or 1,5-naphthodiyl, 2,6-naphthodiyl, or 2,7-naphthodiyl (which are exemplary ten-membered C8 aromatic portions).

[0729] In some embodiments, the heteroaryl group is an aromatic ring compound containing five or more ring members, one or more of which are heteroatoms, such as, but not limited to, N, O, S, and Se. In some embodiments, a heteroaryl group designated as C2-heteroaryl can be a 5-membered ring having two carbon atoms and three heteroatoms, a 6-membered ring having two carbon atoms and four heteroatoms, etc. Similarly, a C4-heteroaryl can be a 5-membered ring having one heteroatom, a 6-membered ring having two heteroatoms, etc. The sum of the number of carbon atoms and the number of heteroatoms equals the total number of ring atoms. Heteroaryl groups include, but are not limited to, groups such as pyrrole, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, isoxazolyl, thiazolyl, pyridinyl, phenylthio, benzobenzylthio, benzofuranyl, indole, azaindole, indole, benzimidazolyl, azabenzimidazolyl, benzoxazolyl, benzothiazolyl, benzothiadiazolyl, imidazopyridyl, isoxazolylpyridyl, thianaphthalenyl, purine, xanthocyanidinyl, adenine, guanine, quinolinyl, isoquinolinyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl, quinoxalinyl, and quinazolinyl groups. Therefore, the terms "heteroaryl" and "heteroaryl group" include fused-ring compounds, such as those in which at least one ring, but not necessarily all rings, are aromatic, including tetrahydroquinolinyl, tetrahydroisoquinolinyl, indole, and 2,3-dihydroindoleyl.

[0730] In some embodiments, when m is 2, Ar1 must be substituted with two Z-Ar2 moieties (as in formulas (I), (IA), and (IB)) or two triazole-Ar2 moieties (as in formulas (IC) and (ID)), and any two carbons of the Ar1 moieties can be substituted with one of these two Z-Ar2 moieties or the triazole-Ar2 moieties. For example, when Ar1 is a substituted benzene, Ar1 can be substituted at the ortho, meta, or para positions, as shown below, where k represents the position where substitution may occur in the aromatic moieties:

[0731]

[0732] In other embodiments, when Ar1 is a substituted naphthalene and m is 2, Ar1 can be substituted on any two naphthyl carbon atoms, where the following structures show substitution options, where k indicates "Z" group substitution (e.g., Z-Ar2). Or triazole-Ar2) may occur at the following positions on the aromatic moieties:

[0733]

[0734] The aforementioned embodiments use the carbocyclic Ar1 moiety as an illustrative Ar1 moiety to illustrate the "Z" group (e.g., triazole) on Ar1. However, the same principle applies to the substitution of the "Z" group (e.g., triazole substitution) on the Ar1 group of heterocyclic aromatic compounds. For example, when Ar1 is a substituted pyridine and m is 2, the two "Z" groups (e.g., triazole groups) of Z-Ar2 (e.g., triazole-Ar2) can be located at either of the following positions on the pyridine ring, where k is used to specify the position where the "Z" group (e.g., triazole group) can be located:

[0735]

[0736] In some implementations, Ar1 has a monocyclic heteroaromatic structure, selected from...

[0737]

[0738] The ring of the "Z" group (e.g., the triazole ring) is substituted at position k on Ar1.

[0739] In other embodiments, Ar1 is a monocyclic carbon ring structure selected from the following:

[0740]

[0741] The ring of the "Z" group (e.g., the triazole ring) is substituted at position k on Ar1.

[0742] In some other embodiments, Ar1 is selected from the following bicyclic carbon ring structures:

[0743]

[0744] The ring of the "Z" group (e.g., the triazole ring) is substituted at position k on Ar1.

[0745] In a further embodiment, Ar1 is a polycyclic heterocyclic structure having two six-membered rings, one five-membered ring, and one nitrogen ring atom selected from:

[0746]

[0747] The ring of the "Z" group (e.g., the triazole ring) is substituted at position k on Ar1.

[0748] In a further embodiment, Ar1 is a polycyclic heterocyclic structure having three six-membered rings and two nitrogen ring atoms selected from:

[0749]

[0750] The ring of the "Z" group (e.g., the triazole ring) is substituted at position k on Ar1.

[0751] In some embodiments, Ar1 comprises substituted and unsubstituted portions as described herein. In other embodiments, Ar1 is a substituted aromatic portion. In still other embodiments, Ar1 is an unsubstituted aromatic portion. In a substituted Ar1 portion, one or more hydrogen atoms that would otherwise be bonded to a ring atom have been substituted by a substituent; for example, optionally 1, 2, 3, 4, 5, or 6 hydrogen atoms may be substituted by a substituent (or partially). Substituents on Ar1 do not refer to Z–Ar2 (e.g., triazole–Ar2) portions that are necessarily present when m equals 1, or two Z–Ar2 (e.g., triazole–Ar2) portions that are necessarily present when m equals 2, or three Z–Ar2 (e.g., triazole–Ar2) portions that are necessarily present when m equals 3.

[0752] In some embodiments, the substituents on Ar1 include atoms selected from deuterium, halogens (F, Cl, Br, I), carbon, nitrogen, oxygen, and sulfur, and optionally also contain hydrogen, and further atoms forming counterions (if present). Deuterium and halides are considered monovalent atoms, while carbon, nitrogen, oxygen, and sulfur are considered polyvalent atoms because they are capable of forming more than one covalent bond simultaneously. In addition to monovalent atoms, the substituents on Ar1 may have multiple polyvalent atoms, such as 1 to 25, 1 to 22, 1 to 15, 1 to 10, or 1 to 5 polyvalent atoms, optionally selected from carbon, nitrogen, oxygen, and sulfur. Descriptions of substituents having up to 10 polyvalent atoms are provided below. Other substituents, including those having up to 25 polyvalent atoms, are known to those skilled in the art.

[0753] In some embodiments, the substituents on Ar1 contain zero polyvalent atoms. In this embodiment, the hydrogen atoms bonded to the ring atoms are replaced by another monovalent atom (such as deuterium, fluorine, chlorine, bromine, or iodine).

[0754] In other embodiments, the substituents on Ar1 contain one polyvalent atom. In this embodiment, one or more hydrogen atoms bonded to the ring atoms of Ar1 are replaced by a single polyvalent atom, wherein the open valence of the polyvalent atom is filled by one or more monovalent atoms, such as hydroxyl (OH), mercapto (SH), amino (NH2), methyl (CH3), and methylene (=CH2), including their fully or partially halogenated and deuterated forms, such as CF3.

[0755] In yet other embodiments, the substituent on Ar1 contains two polyvalent atoms. In these embodiments, one or more hydrogen atoms bonded to the ring atom of Ar1 are replaced by a first polyvalent atom, which in turn bonds to a second polyvalent atom, thereby providing a substituent formed of two polyvalent atoms, wherein the open valences of the polyvalent atoms are filled by one or more monovalent atoms. Examples of such substituents are well known to those skilled in the art. Specific examples include ethyl (CH2CH3), ethylene (CH=CH2), ethynyl (C≡CH), ethylene (=CHCH3), aminomethyl (CH2NH2), aminomethylene (=CHNH2), thiomethylene (=CHSH), hydroxymethylene (=CHOH), hydroxymethyl (CH2OH), mercaptomethyl (CH2SH), N-methylamine (NHCH3), methyl sulfide (SCH3), methoxy (OCH3), nitriles (CN), formyl (C(O)H), thioaldehyde (C(S)H), N-hydroxy (N-OH), hydroxylamine (ONH2), hydrazine (NH2NH2), diazine (N=NH), and diazo compounds (N≡N), including their fully or partially halogenated and deuterated versions, such as OCF3 and CH2CD3.

[0756] In some other embodiments, the substituent on Ar1 contains three polyvalent atoms. In this embodiment, one or more hydrogen atoms bonded to the ring atoms of Ar1 are replaced by a first polyvalent atom, which in turn is bonded directly or indirectly to each of the second and third polyvalent atoms; thus, the first polyvalent atom is bonded to the second polyvalent atom, and the third polyvalent atom is bonded to one or both of the first and second polyvalent atoms, thereby providing a substituent formed of three polyvalent atoms, wherein the open valences on the polyvalent atoms are filled by one or more monovalent atoms. Examples of such substituents are well known to those skilled in the art and are provided herein, such as nitro, methyl ketone, and carboxyl groups.

[0757] In yet another embodiment, the substituent on Ar1 contains four polyvalent atoms. In this embodiment, one or more hydrogen atoms bonded to the ring atom of Ar1 are replaced by a first polyvalent atom, which in turn is directly or indirectly bonded to each of the second, third, and fourth polyvalent atoms, thus providing a substituent formed of four polyvalent atoms, wherein the open valences on the polyvalent atoms are filled by one or more monovalent atoms. Examples of such substituents are well known to those skilled in the art and are provided herein, such as methyl ester (CO2CH3), N-methylformamide (C(O)NHCH3), and acetamide (NHC(O)CH3).

[0758] In some other embodiments, the substituents on Ar1 contain five polyvalent atoms. In this embodiment, one or more hydrogen atoms bonded to the ring atoms of Ar1 are replaced by a first polyvalent atom, which in turn is directly or indirectly bonded to each of the second, third, fourth, and fifth polyvalent atoms, thus providing a substituent formed of five polyvalent atoms, wherein the open valences on the polyvalent atoms are filled by one or more monovalent atoms. Examples of such substituents are well known to those skilled in the art and are provided herein, such as ethyl esters (CO2CH2CH3), S-ethyl thiocarbonate (C(O)SCH2CH3), N-ethylformamide (C(O)NHCH2CH3), and N,N-dimethylformamide (C(O)N(CH3)2).

[0759] In some other embodiments, the substituents on Ar1 contain six polyvalent atoms. In this embodiment, one or more hydrogen atoms bonded to the ring atoms of Ar1 are replaced by a first polyvalent atom, which in turn is directly or indirectly bonded to each of the second, third, fourth, fifth, and sixth polyvalent atoms, thus providing a substituent formed of six polyvalent atoms, wherein the open valences on the polyvalent atoms are filled by one or more monovalent atoms. Examples of such substituents are well known to those skilled in the art and are provided herein, such as N-cyclopropylformamide (C(O)NH-cyclopropyl), N-propylformamide (C(O)NHCH2CH2CH3), N-(2-hydroxyethyl)formamide (C(O)NHCH2CH2OH), and N-formamidinamide (C(O)NHC(=NH)NH2).

[0760] In yet another embodiment, the substituent on Ar1 contains seven polyvalent atoms. In this embodiment, one or more hydrogen atoms bonded to the ring atom of Ar1 are replaced by a first polyvalent atom, which in turn is directly or indirectly bonded to each of the second, third, fourth, fifth, sixth, and seventh polyvalent atoms, thus providing a substituent formed of seven polyvalent atoms, wherein the open valences on the polyvalent atoms are filled by one or more monovalent atoms. Examples of such substituents are well known to those skilled in the art and are provided herein, such as N-(n-butyl)formamide (C(O)NHCH2CH2CH2CH3), N-(tert-butyl)formamide (C(O)NHC(CH3)3), N,N-diethylformamide (C(O)N(CH2CH3)2), and N-cyclobutylformamide (C(O)NH(cyclobutyl)).

[0761] In yet another embodiment, the substituent on Ar1 contains eight polyvalent atoms. In this embodiment, one or more hydrogen atoms bonded to the ring atom of Ar1 are replaced by a first polyvalent atom, which in turn is directly or indirectly bonded to each of the second, third, fourth, fifth, sixth, seventh, and eighth polyvalent atoms, thus providing a substituent formed of eight polyvalent atoms, wherein the open valences on the polyvalent atoms are filled by one or more monovalent atoms. Examples of such substituents are well known to those skilled in the art and are provided herein, such as N-cyclopentylformamide (C(O)NH(cyclopentyl)), (piperidin-1-yl) methyl ketone (C(O)-piperidin-1-yl), and (morpholin-4-yl) methyl ketone (C(O)-morpholin-4-yl).

[0762] In yet another embodiment, the substituent on Ar1 contains nine polyvalent atoms. In this embodiment, one or more hydrogen atoms bonded to the ring atom of Ar1 are replaced by a first polyvalent atom, which in turn is directly or indirectly bonded to each of the second, third, fourth, fifth, sixth, seventh, eighth, and ninth polyvalent atoms, thus providing a substituent formed of nine polyvalent atoms, wherein the open valences on the polyvalent atoms are filled by one or more monovalent atoms. Examples of these substituents are well known to those skilled in the art and are provided herein, such as di-(isopropyl) ester (C(O)O(CH(CH3)2)2, di-(n-propyl) ester (C(O)O(CH2CH2CH3)2), N-cyclohexylformamide (C(O)NH(cyclohexyl)), (4-methylpiperazin-1-yl) methyl ketone (C(O)(4-methylpiperazin-1-yl), 2-(acetylamino)ethylformamide (C(O)NHCH2CH2NHC(O)CH3) and N-phenylformamide (C(O)NH(phenyl)).

[0763] In yet another embodiment, the substituent on Ar1 contains 10 polyvalent atoms. In this embodiment, one or more hydrogen atoms bonded to the ring atoms of Ar1 are replaced by a first polyvalent atom, which in turn is directly or indirectly bonded to each of the second, third, fourth, fifth, sixth, seventh, eighth, ninth, and tenth polyvalent atoms, thus providing a substituent formed of ten polyvalent atoms, wherein the open valences on the polyvalent atoms are filled by one or more monovalent atoms. Examples of such substituents are well known to those skilled in the art and are provided herein, such as N-benzylformamide (C(O)NHCH2(phenyl)).

[0764] In some embodiments, Ar1 is a substituted aryl group, wherein at least one substituent on Ar1 is selected from halogens, hydroxyl groups, thiols, nitro groups, and nitriles.

[0765] In other embodiments, Ar1 is substituted with at least one of the following substituents: halogen, –OH, –CN, –NO2, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 heteroalkyl, C1-C6 cycloalkyl, –OR 0 –CONH2, –C(O)NR 1 R 1' –C(O)(CH2) a NR 1 R 1' –NR 1 R 1' –NR 1 C(O)R 3 –C(O)SR 3 –COR 3 –CO(CH2) a OC(O)R 3 –OC(O)R 3 –C(O)OR 3 –C–O–R 3 , thiols, -R 4 -H、–SOR 1 –S(O)2R 1 –S(O)2NR 1 R 1' –CH2–NR 1 S(O)2R 3 –NR 1 S(O)2R 3 and –C(CH3)=N-(phenyl)-O-CH2-CH; and where:

[0766] R 0 Each time it appears, it is independently selected from C1-C6 alkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C2-C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl.

[0767] R 1 and R 1' Each time it appears, it is independently selected from H, hydroxyl, C1-C6 alkyl, C1-C6 haloalkyl, C1-C 20 Heteroalkyl, C1-C 10 Heteroalkyl –NH2, C2-C6 alkenyl, C2-C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic (e.g., via one or more R... 3Substituent groups), substituted or unsubstituted aryl groups, substituted or unsubstituted arylalkyl groups, and substituted or unsubstituted heteroaryl groups, benzyl groups, –C(=NH)NH2, –CH2CO2R 0 –C(H)(CH) a (COOH))2, -CH2C(O)NHCH2CO2H, -CH2CH2OH, -CH2CH2NHC(O)R 3 –(CH2) a C(O)NH(CH2) a C(O)OR 3 –(CH2) a –CN, spermine, –(CH2) a –NH2、–C(O)R 3 –(CH2) a C(O)OR 3 –(CH2) a OR 3 –C(H)((CH2) a OH) a' –CHCH(OH)(CH2) a OH, –(CH2) a OH, C1-C6–C(O)OH, can be substituted (e.g., via R) 3 (Substituted group) or unsubstituted –(CH2) a - Heterocyclic rings

[0768] , ;and

[0769] Where R 1 and R 1' Together they form substituted or unsubstituted heterocycles, including but not limited to aza-butanes, pyrrolidines, piperidines, piperazines, morpholines, etc.

[0770] or ;

[0771] R 2 Each time it appears, it is independently selected from C2-C6 alkyl, C1-C6 haloalkyl, C1-C6 heteroalkyl, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, and substituted or unsubstituted haloalkoxy.

[0772] R 3Each time it appears, it is independently selected from H, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 heteroalkyl, –C1-C6–OH, –C1-C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine; and

[0773] R 4 Each occurrence is independently selected from alkylene groups interrupted by one or more heteroatoms, wherein the heteroatoms are O, S, NH, or combinations thereof.

[0774] In some embodiments, Ar1 is substituted with at least one substituent selected from the group consisting of: substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, substituted or unsubstituted arylalkyl, substituted or unsubstituted heteroarylalkyl, substituted or unsubstituted haloalkyl and substituted or unsubstituted haloalkoxy.

[0775] In some implementations, Ar1 will include an aromatic moiety as described herein, wherein the aromatic moiety may optionally be substituted, in addition to the substitution described herein, for (Z–Ar2). m Groups (e.g., (triazole-Ar2)) m Other than the group that is substituted.

[0776] In some embodiments, the substituents of Ar1 include, but are not limited to, halides (such as fluorides, chlorides and bromides), alkyl groups having 1 to 6 carbon atoms (such as methyl and ethyl), haloalkyl groups having 1 to 6 carbon atoms (such as trifluoromethyl), cyano, formyl and formamide.

[0777] In other embodiments, the substituents of Ar1 include, but are not limited to: nitro (–NO2); cyano (–CN); carboxylic acid (–COOH or a salt thereof); formamide (–C(O)NH2); C1–C6 alkoxy, such as methoxy; isonicotinic acid ester; isonicotinamide; C1–C6 alkyl, including methyl; C1–C6 haloalkyl, such as trifluoromethyl; C1–C6 heteroalkyl, including amides, such as –NHC(O)(C1–C6 alkyl), –NHC(O)(C1–C6 heteroalkyl), –C(O)NH(C1–C6 alkyl), –C(O)NH(C1–C6 heteroalkyl), –C(O)N(C1–C6 alkyl) C1–C6 alkyl), –C(O)N(C1–C6 alkyl)(C1–C6 heteroalkyl); and –C(O)N(C1–C6 heteroalkyl)(C1–C6 heteroalkyl), including –NHC(O)CH3, C(O)NHCH3, –C(O)N(CH3)2, –NHC(O)CH2CH3, C(O)NHCH2CH3, –C(O)N(CH3)CH2CH3, –C(O)N(CH2CH3)2, –C(O)NH(C1–C6 cycloalkyl) And –NHC(O) (C1–C6 cycloalkyl) (e.g., C(O)NH (cyclopropyl), –NHC(O)–cyclopropyl, C(O)NH (cyclohexyl), NHC(O)–cyclohexyl), C(O)NHCH2CH2CH2CH3, –C(O)NH(C(CH3)3), –C(O)NH(CH2CH2OH); ketones, such as –C(O) (C1–C6 alkyl), including –C(O)CH3, –C(O) (cycloalkyl) This includes –C(O)–cyclohexyl and C(O)–(heterocyclic alkyl), wherein the heterocyclic alkyl can be, for example, morpholino, piperidinyl, piperazinyl, N-methylpiperazinyl; esters, such as –CO2–(C1–C6 alkyl), including –CO2CH3, –O2CH2CH3, –CO2CH2CH2CH3, –CO2CH2(CH3)2; and thioesters, such as C(O)–S–(C1–C6 alkyl), including –C(O)–S–CH3 and –C(O)–S–CH2CH3.

[0778] In some embodiments, the substituents of Ar1 include, but are not limited to, –C(O)–CH3, –C(O)–NH2, –C(O)O–CH2CH3, –CF3, –C(O)O–tert-butyl, –C(O)–N(CH3)2, –C(O)–N(H)CH3, –C(O)–N(H)CH2CH3, –C(O)–N(H) (cycloalkyl), –C(O)–N(H)(CH2)4, –C(O)–N(CH2CH3)2, –C(O)–N(H) (tert-butyl), –C(O)–N(H) (C6–cycloalkyl), –C(O)–N(H)(CH2)3, –C(O)–N(H) (C5–cycloalkyl), –C(O)–N(H) (C5–cycloalkyl), and –C(O)–NH2. –N(H)(C4–cycloalkyl), –C(O)–N(H)(phenyl), –C(O)–N(H)(CH2CH2)N(H)C(O)–CH3, –C(O)–N(H)C(NH)(NH2), –C(O)–N(CH2CH2CH3)2, –C(O)–N–(C(H)(CH3)(CH2CH3))2, –C(O)–N(CH2CH2CH2CH3)2, –C(O)–N(H)(benzyl), –C(O)–N(H)–CH2CH2OH, –C(O)–S–CH2CH3, –C(O)N(CH2)2, –N(H)S(O)(O)–phenyl, –C(O)N(H)CH2CH2CF3, –C(O)N(H)CH3, –C(O)OCH2CH3, –C(O)–N(H)–CH2CH2O–CH2CH2O–CH3, –C(O)–N(H)–(CH2CH2O)8–CH3, –C(O)N(H)–CH2 –CH2–CH2–N(H)–CH2–CH2–CH2–N(H)–CH2–CH2–CH2–NH2, –C(O)N(H)–CH2–CH2–NH2, –C(O)N(H)C(O)CH3, –C(O)N(H)–CH2–CH3, –S(O)(O)N(H) –CH2–CH3, –S(O)(O)N–(CH2–CH3)2, –S(O)(O)N(H)–(C3–cycloalkyl), –S(O)(O)N(H)–(CH2CH2)OH, –S(O)(O)OH, –C(O)N(H)C(H)(CH2OH)2, –C(O)N( CH2CH2OH)2, –C(O)N(H)CH2CH2=CH2, –C(O)CH2N(CH3)(CH2)(COOH), –S(O)(O)(CH2CH3), –S(O)(O)N(H)CH2–CH(OH)(CH2OH), –S(O)(O)–NH2 or –S(O)(O)N(H)CH2–C(O)(O–tert-butyl).

[0779] In one embodiment, Ar1 is substituted with at least one portion selected from: –O–(C 1-6 Alkyl), C 1-6 Alkyl, C 1-6 Halogenated alkyl groups, –CO2–C 1-6 Alkyl, –CONH–C 1-6 Alkyl groups, –CONH2, CN; and –NO2.

[0780] In some embodiments, when n is 1 or 2, the compounds of this disclosure (such as formula (I) and / or (IC)) contain a connector L. In some embodiments, the connector L can be a direct bond. In other embodiments, the connector is not a direct bond, but one or more atoms, particularly atoms selected from carbon, nitrogen, oxygen, and sulfur. In still other embodiments, the connector can be an alkylene group (e.g., a C1–C6 alkylene) or a substituted alkylene. In some embodiments, the connector can be a heteroalkylene connector, which refers to a substituted or unsubstituted alkylene that further contains at least one heteroatom selected from oxygen, nitrogen, or sulfur (e.g., 1, 2, 3, or 4 heteroatoms), said heteroatom being at and / or located at one or more terminal positions of the parent chain. In some embodiments, L is a heteroalkylene with a length of 2 to 10 carbon atoms, wherein one or more carbon atoms are substituted by at least one heteroatom selected from oxygen, nitrogen, and sulfur. In some embodiments, L can be a heteroalkylene linker having at least one N, O, or S heteroatom, wherein the heteroalkylene can be linear or cyclic and optionally substituted, wherein exemplary substituents include oxo, –OH, C 1–4 Alkyl and C 1–4 Alkoxy groups. Non-limiting examples of heteroalkylene linker groups include amide-containing heteroalkylene groups, such as –C(O)NH-alkylene- and –C(O)NH-alkylene-NHC(O)-, wherein the alkylene group is optionally C1–C6 alkylene. Other examples of heteroalkylene groups include ester-containing heteroalkylene groups, such as –C(O)O-alkylene- and –C(O)O-alkylene-OC(O)-, wherein in one embodiment, the alkylene group is an unsubstituted C1–C6 alkylene group, and in another embodiment, the alkylene group is a substituted C1–C6 alkylene group. In some embodiments, the linker is hydrolytically stable, so that it does not decompose or degrade or otherwise break down when PEM is placed in water.

[0781] In some embodiments, the connector L comprises 1 to 25 atoms (excluding hydrogen and halogens in the atom count), wherein, in addition to hydrogen and halogens, the connector may optionally be composed of atoms selected from carbon, nitrogen, oxygen, and sulfur. In some embodiments, the connector has fewer than 25 atoms (excluding hydrogen and halogens), for example, it comprises 1 to 20 atoms, 1 to 15 atoms, 1 to 10 atoms, or 1 to 5 atoms, in each case, excluding hydrogen and halogens from the atom count, wherein the counted atoms may optionally be selected from carbon, oxygen, nitrogen, and sulfur.

[0782] In some embodiments, the "Z" group (e.g., triazole) in the PEM compound of formula (I) may be substituted in addition to being directly bonded to Ar1 and Ar2. Generally, the compounds of this disclosure may optionally be described as comprising the chemical formula Ar1–Z(M)–Ar2 (e.g., Ar1, Ar2, and M are as defined herein. Depending on the specific substituent "Z", Z may be substituted with one or more "M" moieties, such as one "M" moieties, two "M" moieties, three "M" moieties, four "M" moieties, etc. In some embodiments, "Z" is a triazole, and the triazole ring is substituted only with Ar1 and Ar2, i.e., M is hydrogen. In other embodiments, Z is substituted with Ar1, Ar2, and one of methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, or sec-butyl. In still other embodiments, Z is substituted with Ar1, Ar2, and one of fluorine, chlorine, or iodine.

[0783] Therefore, in one embodiment, this disclosure provides a compound having formula (IB):

[0784] (IB),

[0785] Ar1, Ar2, and M are as defined herein. When a compound is substituted with more than one “M” moiety, each “M” moiety is chosen independently each time it appears. However, in one embodiment, M is the same atom each time it appears in the compounds of this disclosure. For example, this disclosure provides compounds in which M is hydrogen each time it appears. In another example, this disclosure provides compounds in which M is an iodide each time it appears. In some embodiments, Z is not a triazole. In some embodiments, Z is not a 1,2,3-triazole or a 1,2,4-triazole.

[0786] In some embodiments, when the compounds of this disclosure contain two or more –Z(M)–Ar2 groups, the Ar2 moiety may optionally have the same chemical structure each time it appears. However, when the PEM compounds of this disclosure contain multiple –Z(M)–Ar2 groups, in one embodiment, these Ar2 moietyes are not necessarily identical to each other; in fact, they may be different. In some embodiments, each Ar2 moiety may differ from each other in terms of Ar2 ring atoms and / or substitutions on Ar2 ring atoms.

[0787] For example, if one Ar2 group is phenyl and the other Ar2 group is pyridyl, then the two Ar2 groups are different in terms of the ring atoms that make up the Ar2 group.

[0788] As another example, if both Ar2 groups are phenyl, but one phenyl group is substituted with a carboxyl group and the other with a methoxy group, for example, 4-(4-(3-(1-(4-methoxyphenyl)-1H-1,2,3-triazol-4-yl)phenyl)-1H-1,2,3-triazol-1-yl)benzoic acid, then the compound is considered to have two distinct Ar2 groups.

[0789] In another example, two Ar2 groups can be positional isomers of each other. For instance, when both Ar2 groups are phenyl and both benzene rings are substituted with hydroxyl and carboxyl groups, but the hydroxyl and / or carboxyl groups are in different positions on the two benzene rings, for example, if on one benzene ring the triazole is located at the 3 position (meta) relative to the carboxyl group, while on the other benzene ring the triazole is located at the 4 position (para) relative to the carboxyl group, then the two Ar2 groups are considered positional isomers and are not the same.

[0790] In some embodiments, the Ar2 ring is identical in all respects each time it appears in the compounds disclosed herein. In some embodiments, the Ar2 ring atoms are identical each time Ar2 appears, but the substitutions on the Ar2 ring are different each time Ar2 appears. In some embodiments, the Ar2 ring atoms are different each time Ar2 appears, and the substitutions on the Ar2 ring may be the same or different.

[0791] In some embodiments, the compound of formula (I) comprises at least one Ar2 moiety. In some embodiments, Ar2 is a monocyclic 6-membered aromatic ring, non-limiting examples of which include phenyl, pyridyl, and pyrazinyl groups, and wherein the Ar2 group optionally includes substituents on the ring atoms. In other embodiments, Ar2 is a 5-membered monocyclic aromatic ring, which may optionally be substituted. In still other embodiments, Ar2 is a 5- or 6-membered aromatic ring, which may optionally be substituted. In yet another embodiment, Ar2 is a 9- or 10-membered fused bicyclic ring comprising two fused 5- and / or 6-membered monocyclic rings, wherein at least one of the two monocyclic rings is an aromatic ring. In still a further embodiment, Ar2 is a 9- or 10-membered fused bicyclic ring comprising two fused 5- and / or 6-membered monocyclic rings, wherein both monocyclic rings are aromatic rings. In a further embodiment, Ar2 is a 13- and 14-member fused tricyclic ring comprising three monocyclic rings together, wherein at least one of the three monocyclic rings is an aromatic ring or a heteroaromatic ring.

[0792] In some implementations, Ar2 can be any of these options, namely, Ar2 is selected from (a) a 5-membered monocyclic aromatic ring, (b) a 6-membered monocyclic aromatic ring, (c) a 9-membered fused bicyclic ring comprising a 5-membered and a 6-membered monocyclic ring fused together, wherein at least one of the two monocyclic rings and optionally the two monocyclic rings are aromatic rings, and (d) a 10-membered fused bicyclic ring comprising two 6-membered monocyclic rings fused together, wherein at least one of the two monocyclic rings and optionally the two monocyclic rings are aromatic rings.

[0793] In compounds having formulas (I), (IA), (IB), (IC), and / or (ID), Ar2 is optionally a 5-membered monocyclic aromatic ring selected from the group consisting of: thiophene, 1,2-thiazole, 1,3-thiazole, furan, 1,2-oxazole, 1,3-oxazole, 1H-pyrrole, 1H-pyrazole, oxadiazole, thiadiazole, 1,2,4-triazole, 1,2,3-triazole, and 1H-imidazole.

[0794] In compounds having formulas (I), (IA), (IB), (IC), and / or (ID), Ar2 is optionally a 6-membered monocyclic aromatic ring selected from the group consisting of benzene, pyridine, pyridazine, pyrimidine, and pyrazine.

[0795] In compounds having formulas (I), (IA), (IB), (IC), and / or (ID), Ar2 is optionally a 9-membered fused bicyclic aromatic ring system selected from the group consisting of: benzofuran, 1,3-benzoxazole, furano[3,2-b]pyridine, furano[3,2-c]pyridine, furano[2,3-c]pyridine, furano[2,3-b]pyridine, indole, 1H-benzimidazole, 1H-pyrrolo[3,2-b]pyridine, 1H-pyrrolo[3,2-b]pyridine, etc. [c]pyridine, 1H-pyrrolo[2,3-c]pyridine, 1H-pyrrolo[2,3-b]pyridine, benzothiophene, 1,3-benzothiazole, thieno[3,2-b]pyridine, thieno[3,2-c]pyridine, thieno[2,3-c]pyridine, benzoxadiazole, benzothiadiazole, benzoisoxazole, benzotriazole and thieno[2,3-b]pyridine.

[0796] In the compounds of formula (I), (IA), (IB), (IC) and / or (ID), Ar2 is optionally a 10-membered fused bicyclic aromatic ring system selected from the group consisting of: naphthylene, quinoline, quinazoline, quinoxaline, 1,5-naphthoidine, 1,6-naphthoidine, 1,7-naphthoidine, 1,8-naphthoidine, isoquinoline, phthalazine, 2,6-naphthoidine and 2,7-naphthoidine.

[0797] As described above, the compounds of this disclosure comprise at least one Ar2 group, wherein the Ar2 group comprises at least one aromatic ring and optionally comprises one or more substituents on the aromatic ring, such as two or more substituents, such as three or more substituents, such as four or more substituents, wherein any of the one or more substituents may be the same or different. In some embodiments, Ar2 comprises at least one, i.e., one or more substituents, such as 1 to 5, 1 to 4, 1 to 3, or 1 to 2 substituents on the aromatic ring. In some embodiments, Ar2 comprises exactly one substituent on the aromatic ring. In other embodiments, Ar2 comprises exactly two substituents on the aromatic ring. In still some embodiments, Ar2 comprises exactly three substituents on the aromatic ring. In a further embodiment, Ar2 comprises exactly four substituents on the aromatic ring. In yet another further embodiment, Ar2 comprises two or more substituents on the aromatic ring.

[0798] In some embodiments, one or more substituents on the ring atoms of Ar2 are selected from substituents optionally named "G", wherein the substituents are selected from halogens, –CN, –NO2, –SO3R. 3 C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–(CH2) aC(O)R 3 , –E–CO2H, –E–CHO, –E–C(O)R 3 , –E–C(O)NH(OH), –E–C(O)NHR 1 –E–C(O)N(H)C(H)(R 1 (R) 1' ), –E–NR 1 R 1' –E–OR 2 –E–C(O)N(R) 1 (CH2) a P(O)(O)OH)2、–E–C(O)N(R 1 (CH2) a SO3H), –E–C(O)NR 1 R 1' –C(O)–heterocyclic alkyl (where the heterocyclic alkyl group may or may not be substituted), –C(O)–N(H)–CH(COOH)((CH2) a – heteroaryl) (wherein the heteroaryl group may or may not be substituted), wherein E is independently selected from direct bond and C1–C6 alkylene group each time it appears;

[0799] Where R 0 Each time it appears, it is independently selected from C1–C6 alkyl, C1–C6 haloalkyl, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl.

[0800] R 1 and R 1' Each time it appears, it is independently selected from H, hydroxyl, C1–C6 alkyl, C1–C6 haloalkyl, C1–C 20 Heteroalkyl, C1–C 10 Heteroalkyl –NH2, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic (e.g., via one or more R... 3 Substituent groups), substituted or unsubstituted aryl groups, substituted or unsubstituted arylalkyl groups, and substituted or unsubstituted heteroaryl groups, benzyl groups, –C(=NH)NH2, –CH2CO2R 0 –C(H)(CH) a (COOH))2, –CH2C(O)NHCH2CO2H, –CH2CH2OH, –CH2CH2NHC(O)R 3 –(CH2) aC(O)NH(CH2) a C(O)OR 3 –(CH2) a –CN, spermine, –(CH2) a –NH2、–C(O)R 3 –(CH2) a C(O)OR 3 –(CH2) a OR 3 –C(H)((CH2) a OH) a' –CHCH(OH)(CH2) a OH, –(CH2) a OH, C1–C6–C(O)OH, can be substituted (e.g., via R...) 3 (Substituted group) or unsubstituted –(CH2) a - Heterocyclic rings

[0801] and ;and

[0802] Where R 1 and R 1' Together they form substituted or unsubstituted heterocycles, including but not limited to aza-butanes, pyrrolidines, piperidines, piperazines, morpholines, etc.

[0803] or ;

[0804] R 2 Each time it appears, it is independently selected from C2–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, and substituted or unsubstituted haloalkoxy; and

[0805] R 3 Each time it appears, it is independently selected from H, C1–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, –C1–C6–OH, –C1–C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine.

[0806] Non-limiting examples of substituents for Ar2 include, but are not limited to, H, –NH2, –C(O)OH, –OH, –OCH3, –C(O)NHOH, –C(O)NH2, –(CH2)3C(O)OH, –CF3, –Cl, –P(O)(OH)2, –F, –C(O)N(H)CH2CH3, –C(O)N(H)CH2C(O)OCH3, –C(O)N(H)CH2CF2CF2CF3, – C(O)N(H)CH2C(O)OH, –C(O)N(H)C(H)(C(O)OOH)CH2C(O)OH, –C(O)N(H)CH2C(O)N(H)CH2C(O)OH, –SO3H, –NO2, –C(O)N(H)CH2PO3H2, –C(O)N(H)CH23O3H, –B(OH)2, –C(OH)(CF3)(CF3), –C(O)–5-membered heterocyclic alkyl –C(O)OH, –C(O)N(H)C(H)(C(O)OH)CH2–imidazolium, –S(O)(O)N(H)–CH2COOH, –C(O)N(H)CH(C(O)OH)(CH2CH2C(O)OH, –C(O)N(H)CH2CH2SO3H, –SO2O – –C(O)N(H)–imidazole–C(O)OH and –C(O)N(CH3)2.

[0807] In some embodiments, the substitution on Ar2 includes an amino group (–NH2). In some embodiments, the substitution on Ar2 includes an alkoxy group, such as a C1–C6 alkoxy group. For example, in one embodiment, the substitution on Ar2 includes a methoxy group. In some embodiments, the substitution on Ar2 includes formic acid or alkylene-formic acid. For example, in some embodiments, the substitution on Ar2 of the PEM compound of formula (I) includes formic acid. In some embodiments, the substitution on Ar2 includes a formate or alkylene-formate. For example, in some embodiments, the substitution on Ar2 of the PEM compound of formula (I) includes –CH2–CO2–CH3. In some embodiments, the substitution on Ar2 includes a haloalkyl group, such as a C1–C6 haloalkyl group. For example, in one embodiment, the substitution on Ar2 of the PEM compound of formula (I) includes a trifluoromethyl group. In some embodiments, the substitution on Ar2 includes a hydroxyl group or a hydroxy-substituted alkyl group, such as a hydroxy-substituted C1–C6 alkyl group. For example, in one embodiment, the substitution of the compound of formula (I) on Ar2 includes a hydroxyl group (–OH).

[0808] In some embodiments, the substitution on Ar2 includes a group selected from formic acid and alkylene-formic acid (e.g., C1-C6 alkylene-formic acid) and another group selected from hydroxyl and hydroxy-substituted alkyl groups (e.g., C1-C6 alkyl substituted with a hydroxyl group). For example, in one embodiment, the substitution on Ar2 is or includes a formic acid or a hydroxyl group.

[0809] In some embodiments, the substitution on Ar2 includes a group selected from formic acid and alkylene-formic acid (e.g., C1-C6 alkylene-formic acid) and a group selected from haloalkyl (e.g., C1-C6 haloalkyl). For example, in one embodiment, the substitution on Ar2 is or includes a formic acid group and a trifluoromethyl group.

[0810] In some embodiments, the substitution on Ar2 includes one group selected from hydroxyl and hydroxy-substituted alkyl groups (e.g., C1-C6 alkyl groups substituted with hydroxyl) and another group selected from haloalkyl groups (e.g., C1-C6 haloalkyl groups). For example, in one embodiment, the substitution on Ar2 is or includes a hydroxyl group and a trifluoromethyl group.

[0811] In some embodiments, the substitution on the Ar2 ring of formula (I) includes at least one of the following: a) formic acid and alkylene-formic acid, such as C1-C6 alkylene-formic acid; b) hydroxyl and hydroxy-substituted alkyl, such as C1-C6 alkyl substituted with one hydroxyl group; and c) haloalkyl, such as C1-C6 haloalkyl. For example, at least one of formic acid, hydroxyl, and trifluoromethyl.

[0812] In some embodiments, the substitution on the Ar2 ring of formula (I) includes at least two of the following: a) formic acid and alkylene-formic acid, such as C1-C6 alkylene-formic acid; b) hydroxyl and hydroxy-substituted alkyl, such as C1-C6 alkyl substituted with one hydroxyl group; and c) haloalkyl, such as C1-C6 haloalkyl. For example, at least two of formic acid, hydroxyl, and trifluoromethyl.

[0813] In some embodiments, the substitutions on the Ar2 ring of formula (I) include all three of the following: a) formic acid and alkylene-formic acid, such as C1-C6 alkylene-formic acid; b) hydroxyl and hydroxy-substituted alkyl, such as C1-C6 alkyl substituted with one hydroxyl group; and c) haloalkyl, such as C1-C6 haloalkyl. That is, Ar2 can be substituted with formic acid, hydroxyl, and trifluoromethyl.

[0814] In some embodiments, the substitution on the Ar2 ring of formula (I) comprises at least one –SO3H moiety or its salt or hydrate. In some embodiments, the substitution on the Ar2 ring of formula (I) comprises at least two –SO3H moiety (at any position on the Ar2 ring relative to each other) or their salt or hydrate.

[0815] In some embodiments, the Ar2 ring of formula (I) comprises at least two substituents, wherein the first substituent of said at least two substituents is a –SO3H moiety or –SO3 - Part; and wherein the second substituent of the at least two substituents is selected from any of the “G” substituents described herein. In some embodiments, the Ar2 ring of formula (I) comprises at least two substituents, wherein the first substituent of the at least two substituents is a –SO3H moiety (or its salt or hydrate); and wherein the second substituent of the at least two substituents is selected from –NO2, –CF3, –F, –Cl, –I, methyl, ethyl, propyl, and butyl. In some embodiments, the Ar2 ring of formula (I) comprises at least three substituents, wherein the first substituent of the at least two substituents is a –SO3H moiety or –SO3 - Partial; and wherein the second and third substituents of the at least three substituents are selected from –NO2, –CF3, –F, –Cl, –I, methyl, ethyl, propyl and butyl.

[0816] In some embodiments, the Ar2 ring of formula (I) comprises at least three substituents, wherein the first substituent of said at least two substituents is a –SO3H moiety or –SO3 - The at least three substituents are selected from –SO3H; and the third substituent is selected from –NO2, –CF3, –F, methyl, ethyl, propyl and butyl.

[0817] In some embodiments, the substitution on the Ar2 ring of formula (I) includes at least one –S(O)(O)N(H)CH2C(O)OH moiety, at least one –C(O)N(H)CH2CH2SO3H moiety, or at least one –C(O)N(H)CH2PO3H2. In other embodiments, the Ar2 moiety is substituted with at least two substituents, wherein the first substituent of said at least two substituents is one of –S(O)(O)N(H)CH2C(O)OH, –C(O)N(H)CH2CH2SO3H, or –C(O)N(H)CH2PO3H2.

[0818] In some embodiments, the substitution on the Ar2 ring of formula (I) includes at least one –SO2O - Partial. In other embodiments, the substitution on the Ar2 ring of formula (I) includes at least one –SO2O - Parts and at least one other part, including any of those parts referred to herein with respect to “G”.

[0819] For example, in some embodiments, the Ar2 group is a substituted phenyl group selected from the following:

[0820]

[0821] In some embodiments, the Ar2 group is a substituted phenyl group, wherein the substituents of the phenyl group are further substituted with G... 2 G 3 G 4 and G 5 The substituted aryl group, and in a more specific embodiment, the substituent of the phenyl group is further substituted with G... 2 G 3 G 4 and G 5 Substituted phenyl groups, such as those further substituted with G... 2 Or via G 2 and G 3 Substituted phenyl groups. For example, the Ar2 group can be via the –SO3H moiety or –SO3 - Replacement, and can be further processed by G 2 G 3 G 4 and G 5 Replacement, such as further via G 2 Or via G 2 and G 3 Substituted phenyl groups. As another example, the Ar2 group can be partially substituted via –C(O)OH, and can be further substituted via G… 2 G 3 G 4 and G 5 , Replacement, such as further via G 2 or G 2 and G 3Substituted phenyl groups. By yet another example, the Ar2 group can be substituted via one of the following moieties: –S(O)(O)N(H)CH2C(O)OH, –C(O)N(H)CH2CH2SO3H, or –C(O)N(H)CH2PO3H2, and can be further substituted via G… 2 G 3 G 4 and G 5 Replacement, such as further via G 2 Or via G 2 and G 3 Substituted phenyl groups. As yet another example, the Ar2 group can be substituted via –SO2O - Partial replacement, and can be further processed by G 2 G 3 G 4 and G 5 , Replacement, such as further via G 2 or G 2 and G 3 Substituted phenyl groups.

[0822] As described above, in one embodiment, the PEM compound of this disclosure may have a hydroxyl group and a formic acid substituted on the Ar2 ring. These two groups may be located at different positions on the Ar2 ring.

[0823] For example, in some embodiments, the PEM compound is described by the following formula:

[0824] .

[0825] In some embodiments, Z is selected from imidazole, pyrazole, pyrimidine, pyridazine, pyrazine, pyrrole, thiophene, triazole (e.g., 1,2,3-triazole; 1,2,4-triazole, thiazole, and isoxazole). In some embodiments, Z is not a triazole. In some embodiments, Z is not 1,2,3-triazole or 1,2,4-triazole. In some embodiments, Ar1 is selected from pyridine, bipyridine, phenyl, and carbazole.In some embodiments, Ar1 may be substituted with the following: –C(O)-CH3, –C(O)-NH2, –C(O)O-CH2CH3, -CF3, –C(O)O-tert-butyl, –C(O)-N(CH3)2, –C(O)-N(H)CH3, –C(O)-N(H)CH2CH3, –C(O)-N(H) (cycloalkyl), –C(O)-N(H)(CH2)4, –C(O)-N(CH2CH3)2, –C(O)-N(H) (tert-butyl), –C(O)-N(H) (C6-cycloalkyl), –C(O)-N(H)(CH2)3, –C(O)-N(H) (C5-cycloalkyl), –C(O)-N(H)(C4- Cycloalkyl), –C(O)-N(H)(phenyl), –C(O)-N(H)(CH2CH2)N(H)C(O)-CH3, –C(O)-N(H)C(NH)(NH2), –C(O)-N(CH2CH2CH3)2, –C(O)-N–(C(H)(CH3)(CH2CH3))2, –C(O)-N(CH2CH2CH2CH3)2, –C(O)-N(H)(benzyl), –C(O)-N(H)-CH2CH2OH, –C(O)–S-CH2CH3, –C(O)N(CH2)2, –N(H)S(O)(O)-phenyl, –C(O)N(H)CH2CH2 CF3, –C(O)N(H)CH3, –C(O)OCH2CH3, –C(O)-N(H)-CH2CH2O-CH2CH2O-CH3, –C(O)-N(H)-(CH2CH2O)8-CH3, –C(O)N(H)-CH2-CH2-CH2-N(H)-CH2-C H2-CH2-N(H)-CH2-CH2-CH2-NH2, –C(O)N(H)-CH2-CH2-NH2, –C(O)N(H)C(O)CH3, –C(O)N(H)-CH2-CH3, –S(O)(O)N(H)-CH2-CH3, –S(O)(O)N-(CH 2-CH3)2, -S(O)(O)N(H)-(C3-cycloalkyl), -S(O)(O)N(H)-(CH2CH2)OH, –S(O)(O)OH, –C(O)N(H)C(H)(CH2OH)2, –C(O)N(CH2CH2OH)2, –C(O)N(H)CH2CH 2=CH2, –C(O)CH2N(CH3)(CH2)(COOH), –S(O)(O)(CH2CH3), –S(O)(O)N(H)CH2-CH(OH)(CH2OH), –S(O)(O)-NH2, –S(O)(O)N(H)CH2-C(O)(O-tert-butyl).

[0826] In one particular embodiment, Z is a triazole, and the compound of formula (I) has the following structure:

[0827] .

[0828] In another embodiment, the PEM compound of this disclosure has a hydroxyl group and a formic acid substitution on Ar2, as provided by the following formula:

[0829] .

[0830] In some embodiments, Z is selected from imidazole, pyrazole, pyrimidine, pyridazine, pyrazine, pyrrole, thiophene, triazole (e.g., 1,2,3-triazole; 1,2,4-triazole, thiazole, and isoxazole). In some embodiments, Z is not a triazole. In some embodiments, Z is not 1,2,3-triazole or 1,2,4-triazole. In some embodiments, Ar1 is selected from pyridine, bipyridine, phenyl, and carbazole.In some embodiments, Ar1 may be substituted with the following: –C(O)-CH3, –C(O)-NH2, –C(O)O-CH2CH3, -CF3, –C(O)O-tert-butyl, –C(O)-N(CH3)2, –C(O)-N(H)CH3, –C(O)-N(H)CH2CH3, –C(O)-N(H) (cycloalkyl), –C(O)-N(H)(CH2)4, –C(O)-N(CH2CH3)2, –C(O)-N(H) (tert-butyl), –C(O)-N(H) (C6-cycloalkyl), –C(O)-N(H)(CH2)3, –C(O)-N(H) (C5-cycloalkyl), –C(O)-N(H)(C4- Cycloalkyl), –C(O)-N(H)(phenyl), –C(O)-N(H)(CH2CH2)N(H)C(O)-CH3, –C(O)-N(H)C(NH)(NH2), –C(O)-N(CH2CH2CH3)2, –C(O)-N–(C(H)(CH3)(CH2CH3))2, –C(O)-N(CH2CH2CH2CH3)2, –C(O)-N(H)(benzyl), –C(O)-N(H)-CH2CH2OH, –C(O)–S-CH2CH3, –C(O)N(CH2)2, –N(H)S(O)(O)-phenyl, –C(O)N(H)CH2CH2 CF3, –C(O)N(H)CH3, –C(O)OCH2CH3, –C(O)-N(H)-CH2CH2O-CH2CH2O-CH3, –C(O)-N(H)-(CH2CH2O)8-CH3, –C(O)N(H)-CH2-CH2-CH2-N(H)-CH2-C H2-CH2-N(H)-CH2-CH2-CH2-NH2, –C(O)N(H)-CH2-CH2-NH2, –C(O)N(H)C(O)CH3, –C(O)N(H)-CH2-CH3, –S(O)(O)N(H)-CH2-CH3, –S(O)(O)N-(CH 2-CH3)2, -S(O)(O)N(H)-(C3-cycloalkyl), -S(O)(O)N(H)-(CH2CH2)OH, –S(O)(O)OH, –C(O)N(H)C(H)(CH2OH)2, –C(O)N(CH2CH2OH)2, –C(O)N(H)CH2CH 2=CH2, –C(O)CH2N(CH3)(CH2)(COOH), –S(O)(O)(CH2CH3), –S(O)(O)N(H)CH2-CH(OH)(CH2OH), –S(O)(O)-NH2, –S(O)(O)N(H)CH2-C(O)(O-tert-butyl).

[0831] In one particular embodiment, Z is a triazole, and the compound of formula (I) has the following structure:

[0832] .

[0833] In yet another embodiment, the PEM compound of this disclosure has a hydroxyl group and a formic acid substitution on Ar2, as shown in the following formula:

[0834] .

[0835] In some embodiments, Z is selected from imidazole, pyrazole, pyrimidine, pyridazine, pyrazine, pyrrole, thiophene, triazole (e.g., 1,2,3-triazole; 1,2,4-triazole, thiazole, and isoxazole). In some embodiments, Z is not a triazole. In some embodiments, Z is not 1,2,3-triazole or 1,2,4-triazole. In some embodiments, Ar1 is selected from pyridine, bipyridine, phenyl, and carbazole.In some embodiments, Ar1 may be substituted with the following: –C(O)-CH3, –C(O)-NH2, –C(O)O-CH2CH3, -CF3, –C(O)O-tert-butyl, –C(O)-N(CH3)2, –C(O)-N(H)CH3, –C(O)-N(H)CH2CH3, –C(O)-N(H) (cycloalkyl), –C(O)-N(H)(CH2)4, –C(O)-N(CH2CH3)2, –C(O)-N(H) (tert-butyl), –C(O)-N(H) (C6-cycloalkyl), –C(O)–N(H)(CH2)3, –C(O)–N(H) (C5-cycloalkyl), –C(O)–N (H)(C4–cycloalkyl), –C(O)–N(H)(phenyl), –C(O)–N(H)(CH2CH2)N(H)C(O)–CH3, –C(O)–N(H)C(NH)(NH2), –C(O)–N(CH2CH2CH3)2, –C(O)–N–(C(H)(CH3)(CH2CH3))2, –C(O)–N(CH2CH2CH2CH3)2, –C(O)–N(H)(benzyl), –C(O)–N(H)–CH2CH2OH, –C(O)–S–CH2CH3, –C(O)N(CH2)2, –N(H)S(O)(O)–phenyl, – C(O)N(H)CH2CH2CF3, –C(O)N(H)CH3, –C(O)OCH2CH3, –C(O)–N(H)–CH2CH2O–CH2CH2O–CH3, –C(O)–N(H)–(CH2CH2O)8–CH3, –C(O)N(H)–CH2– CH2–CH2–N(H)–CH2–CH2–CH2–N(H)–CH2–CH2–CH2–NH2, –C(O)N(H)–CH2–CH2–NH2, –C(O)N(H)C(O)CH3, –C(O)N(H)–CH2–CH3, –S(O)(O)N(H)– CH2–CH3, –S(O)(O)N–(CH2–CH3)2, –S(O)(O)N(H)–(C3–cycloalkyl), –S(O)(O)N(H)–(CH2CH2)OH, –S(O)(O)OH, –C(O)N(H)C(H)(CH2OH)2, –C(O)N( CH2CH2OH)2, –C(O)N(H)CH2CH2=CH2, –C(O)CH2N(CH3)(CH2)(COOH), –S(O)(O)(CH2CH3), –S(O)(O)N(H)CH2–CH(OH)(CH2OH), –S(O)(O)–NH2 Or S(O)(O)N(H)CH2–C(O)(O–tert-butyl).

[0836] In one particular embodiment, Z is a triazole, and the compound of formula (I) has the following structure:

[0837] .

[0838] In one embodiment, the PEM compound of this disclosure has at least hydroxyl and carboxyl substitutions on Ar2, and may have other substitutions on Ar2. For example, Ar2 may be substituted with hydroxyl, formic acid, and alkyl, such as C1–C6 alkyl, to provide compounds of, for example, the following formula:

[0839] .

[0840] In some embodiments, Z is selected from imidazole, pyrazole, pyrimidine, pyridazine, pyrazine, pyrrole, thiophene, triazole (e.g., 1,2,3-triazole; 1,2,4-triazole, thiazole, and isoxazole). In some embodiments, Z is not a triazole. In some embodiments, Z is not 1,2,3-triazole or 1,2,4-triazole. In some embodiments, Ar1 is selected from pyridine, bipyridine, phenyl, and carbazole.In some embodiments, Ar1 may be substituted with the following: –C(O)–CH3, –C(O)–NH2, –C(O)O–CH2CH3, –CF3, –C(O)O–tert-butyl, –C(O)–N(CH3)2, –C(O)–N(H)CH3, –C(O)–N(H)CH2CH3, –C(O)–N(H) (cycloalkyl), –C(O)–N(H) (CH2)4, –C(O)–N(CH2CH3)2, –C(O)–N(H) (tert-butyl), –C(O)–N(H) (C6–cycloalkyl), –C(O)–N(H) (CH2)3, –C(O)–N(H) (C5–cycloalkyl), –C(O)–N (H)(C4–cycloalkyl), –C(O)–N(H)(phenyl), –C(O)–N(H)(CH2CH2)N(H)C(O)–CH3, –C(O)–N(H)C(NH)(NH2), –C(O)–N(CH2CH2CH3)2, –C(O)–N–(C(H)(CH3)(CH2CH3))2, –C(O)–N(CH2CH2CH2CH3)2, –C(O)–N(H)(benzyl), –C(O)–N(H)–CH2CH2OH, –C(O)–S–CH2CH3, –C(O)N(CH2)2, –N(H)S(O)(O)–phenyl, – C(O)N(H)CH2CH2CF3, –C(O)N(H)CH3, –C(O)OCH2CH3, –C(O)–N(H)–CH2CH2O–CH2CH2O–CH3, –C(O)–N(H)–(CH2CH2O)8–CH3, –C(O)N(H)–CH2– CH2–CH2–N(H)–CH2–CH2–CH2–N(H)–CH2–CH2–CH2–NH2, –C(O)N(H)–CH2–CH2–NH2, –C(O)N(H)C(O)CH3, –C(O)N(H)–CH2–CH3, –S(O)(O)N(H)– CH2–CH3, –S(O)(O)N–(CH2–CH3)2, –S(O)(O)N(H)–(C3–cycloalkyl), –S(O)(O)N(H)–(CH2CH2)OH, –S(O)(O)OH, –C(O)N(H)C(H)(CH2OH)2, –C(O)N( CH2CH2OH)2, –C(O)N(H)CH2CH2=CH2, –C(O)CH2N(CH3)(CH2)(COOH), –S(O)(O)(CH2CH3), –S(O)(O)N(H)CH2–CH(OH)(CH2OH), –S(O)(O)–NH2 Or S(O)(O)N(H)CH2–C(O)(O–tert-butyl).

[0841] In one particular embodiment, Z is a triazole, and the compound of formula (I) has the following structure:

[0842] .

[0843] As previously described, in one embodiment, the PEM compound of this disclosure may have a haloalkyl group and a formic acid substituted on Ar2, instead of a hydroxyl group and a formic acid as shown in the structure above.

[0844] In one particular embodiment, the PEM compound disclosed herein can be described by the following formula:

[0845] .

[0846] In some embodiments, Z is selected from imidazole, pyrazole, pyrimidine, pyridazine, pyrazine, pyrrole, thiophene, triazole (e.g., 1,2,3-triazole; 1,2,4-triazole, thiazole, and isoxazole). In some embodiments, Z is not a triazole. In some embodiments, Z is not 1,2,3-triazole or 1,2,4-triazole. In some embodiments, Ar1 is selected from pyridine, bipyridine, phenyl, and carbazole.In some embodiments, Ar1 can be substituted with the following: –C(O)–CH3, –C(O)–NH2, –C(O)O–CH2CH3, –CF3, –C(O)O–tert-butyl, –C(O)–N(CH3)2, –C(O)-N(H)CH3, –C(O)-N(H)CH2CH3, –C(O)-N(H) (cycloalkyl), –C(O)-N(H) (CH2)4, –C(O)-N(CH2CH3)2, –C(O)-N(H) (tert-butyl), –C(O)-N(H) (C6-cycloalkyl), –C(O)-N(H) (CH2)3, –C(O)-N(H) (C5-cycloalkyl), –C(O)-N(H) (C4- Cycloalkyl), –C(O)-N(H)(phenyl), –C(O)-N(H)(CH2CH2)N(H)C(O)-CH3, –C(O)-N(H)C(NH)(NH2), –C(O)-N(CH2CH2CH3)2, –C(O)-N–(C(H)(CH3)(CH2CH3))2, –C(O)-N(CH2CH2CH2CH3)2, –C(O)-N(H)(benzyl), –C(O)-N(H)-CH2CH2OH, –C(O)–S-CH2CH3, –C(O)N(CH2)2, –N(H)S(O)(O)-phenyl, –C(O)N(H)CH2CH2 CF3, –C(O)N(H)CH3, –C(O)OCH2CH3, –C(O)-N(H)-CH2CH2O-CH2CH2O-CH3, –C(O)-N(H)-(CH2CH2O)8-CH3, –C(O)N(H)-CH2-CH2-CH2-N(H)-CH2-C H2-CH2-N(H)-CH2-CH2-CH2-NH2, –C(O)N(H)-CH2-CH2-NH2, –C(O)N(H)C(O)CH3, –C(O)N(H)-CH2-CH3, –S(O)(O)N(H)-CH2-CH3, –S(O)(O)N-(CH 2-CH3)2, -S(O)(O)N(H)-(C3-cycloalkyl), -S(O)(O)N(H)-(CH2CH2)OH, –S(O)(O)OH, –C(O)N(H)C(H)(CH2OH)2, –C(O)N(CH2CH2OH)2, –C(O)N(H)CH2CH 2=CH2, –C(O)CH2N(CH3)(CH2)(COOH), –S(O)(O)(CH2CH3), –S(O)(O)N(H)CH2-CH(OH)(CH2OH), –S(O)(O)-NH2, –S(O)(O)N(H)CH2-C(O)(O-tert-butyl).

[0847] In one particular embodiment, Z is a triazole, and the compound of formula (I) has the following structure:

[0848] .

[0849] In some embodiments, as described herein, the PEM compounds of this disclosure comprise one or more –SO3H moieties or –SO2O. - Partially, and optionally one or more additional substituents on Ar2 (such as G as described herein). 2 G 3 G 4 or G 5 (any of the following). Examples of such PEM compounds include, but are not limited to, those listed below:

[0850]

[0851] Or its salts or hydrates.

[0852] In some embodiments, Z is selected from imidazole, pyrazole, pyrimidine, pyridazine, pyrazine, pyrrole, thiophene, triazole (e.g., 1,2,3-triazole; 1,2,4-triazole, thiazole, and isoxazole). In some embodiments, Z is not a triazole. In some embodiments, Z is not 1,2,3-triazole or 1,2,4-triazole. In some embodiments, Ar1 is selected from pyridine, bipyridine, phenyl, and carbazole. In some embodiments, the Ar1 group may be substituted (such as by any of the substituents described herein) or unsubstituted.For example, Ar1 can be substituted with the following: –C(O)-CH3, –C(O)-NH2, –C(O)O-CH2CH3, -CF3, –C(O)O-tert-butyl, –C(O)-N(CH3)2, –C(O)-N(H)CH3, –C(O)-N(H)CH2CH3, –C(O)-N(H) (cycloalkyl), –C(O)-N(H)(CH2)4, –C(O)-N(CH2CH3)2, –C(O)-N(H) (tert-butyl), –C(O)-N(H) (C6-cycloalkyl), –C(O)–N(H)(CH2)3, –C(O)–N(H) (C5-cycloalkyl), –C(O)–N (H)(C4–cycloalkyl), –C(O)–N(H)(phenyl), –C(O)–N(H)(CH2CH2)N(H)C(O)–CH3, –C(O)–N(H)C(NH)(NH2), –C(O)–N(CH2CH2CH3)2, –C(O)–N–(C(H)(CH3)(CH2CH3))2, –C(O)–N(CH2CH2CH2CH3)2, –C(O)–N(H)(benzyl), –C(O)–N(H)–CH2CH2OH, –C(O)–S–CH2CH3, –C(O)N(CH2)2, –N(H)S(O)(O)–phenyl, – C(O)N(H)CH2CH2CF3, –C(O)N(H)CH3, –C(O)OCH2CH3, –C(O)–N(H)–CH2CH2O–CH2CH2O–CH3, –C(O)–N(H)–(CH2CH2O)8–CH3, –C(O)N(H)–CH2– CH2–CH2–N(H)–CH2–CH2–CH2–N(H)–CH2–CH2–CH2–NH2, –C(O)N(H)–CH2–CH2–NH2, –C(O)N(H)C(O)CH3, –C(O)N(H)–CH2–CH3, –S(O)(O)N(H)– CH2–CH3, –S(O)(O)N–(CH2–CH3)2, –S(O)(O)N(H)–(C3–cycloalkyl), –S(O)(O)N(H)–(CH2CH2)OH, –S(O)(O)OH, –C(O)N(H)C(H)(CH2OH)2, –C(O)N( CH2CH2OH)2, –C(O)N(H)CH2CH2=CH2, –C(O)CH2N(CH3)(CH2)(COOH), –S(O)(O)(CH2CH3), –S(O)(O)N(H)CH2-CH(OH)(CH2OH), –S(O)(O)-NH2 Or S(O)(O)N(H)CH2-C(O)(O-tert-butyl).

[0853] In some embodiments, Z is a triazole, and the compound of formula (I) above has the following structure:

[0854]

[0855] Or its salts or hydrates.

[0856] In some embodiments, Ar1 may be substituted with the following: –C(O)-CH3, –C(O)-NH2, –C(O)O-CH2CH3, -CF3, –C(O)O-tert-butyl, –C(O)-N(CH3)2, –C(O)-N(H)CH3, –C(O)-N(H)CH2CH3, –C(O)-N(H) (cycloalkyl), –C(O)-N(H)(CH2)4, –C(O)-N(CH2CH3)2, –C(O)-N(H) (tert-butyl), –C(O)-N(H) (C6-cycloalkyl), –C(O)-N(H)(CH2)3, –C(O)-N(H) (C5-cycloalkyl), –C(O)-N(H)(C4- Cycloalkyl), –C(O)-N(H)(phenyl), –C(O)-N(H)(CH2CH2)N(H)C(O)-CH3, –C(O)-N(H)C(NH)(NH2), –C(O)-N(CH2CH2CH3)2, –C(O)-N–(C(H)(CH3)(CH2CH3))2, –C(O)-N(CH2CH2CH2CH3)2, –C(O)-N(H)(benzyl), –C(O)-N(H)-CH2CH2OH, –C(O)–S-CH2CH3, –C(O)N(CH2)2, –N(H)S(O)(O)-phenyl, –C(O)N(H)CH2CH2 CF3, –C(O)N(H)CH3, –C(O)OCH2CH3, –C(O)-N(H)-CH2CH2O-CH2CH2O-CH3, –C(O)-N(H)-(CH2CH2O)8-CH3, –C(O)N(H)-CH2-CH2-CH2-N(H)-CH2-C H2-CH2-N(H)-CH2-CH2-CH2-NH2, –C(O)N(H)-CH2-CH2-NH2, –C(O)N(H)C(O)CH3, –C(O)N(H)-CH2-CH3, –S(O)(O)N(H)-CH2-CH3, –S(O)(O)N-(CH 2-CH3)2, -S(O)(O)N(H)-(C3-cycloalkyl), -S(O)(O)N(H)-(CH2CH2)OH, –S(O)(O)OH, –C(O)N(H)C(H)(CH2OH)2, –C(O)N(CH2CH2OH)2, –C(O)N(H)CH2CH 2=CH2, –C(O)CH2N(CH3)(CH2)(COOH), –S(O)(O)(CH2CH3), –S(O)(O)N(H)CH2-CH(OH)(CH2OH), –S(O)(O)-NH2, –S(O)(O)N(H)CH2-C(O)(O-tert-butyl).

[0857] Further examples of PEM compounds comprising an Ar2 moiety substituted with –SO3H (or its salt) according to this disclosure include, but are not limited to, those listed below:

[0858]

[0859] In these embodiments, the –SO3H group (or its salt, i.e., –SO3) - It can be provided at any ring position, such as the para position of group "Z", the meta position of group "Z" or the ortho position of group "Z".

[0860] In some embodiments, Z is selected from imidazole, pyrazole, pyrimidine, pyridazine, pyrazine, pyrrole, thiophene, triazole (e.g., 1,2,3-triazole; 1,2,4-triazole, thiazole, tetraazole, and isoxazole). In some embodiments, Z is not a triazole. In some embodiments, Z is not 1,2,3-triazole or 1,2,4-triazole.

[0861] In some implementations, Ar1 is selected from pyridine, bipyridine, phenyl, and carbazole.

[0862] Further examples of PEM compounds comprising an Ar2 moiety substituted with –SO3H (or its salt) according to this disclosure include, but are not limited to, those listed below:

[0863]

[0864] In some implementations, Ar1 is selected from pyridine, bipyridine, phenyl, and carbazole.

[0865] Further examples of PEM compounds containing substituents on Ar2 include those having the following structures:

[0866]

[0867] Or its salts or hydrates.

[0868] In some embodiments, Z is selected from imidazole, pyrazole, pyrimidine, pyridazine, pyrazine, pyrrole, thiophene, tetrazolium, triazole (e.g., 1,2,3-triazole; 1,2,4-triazole, thiazole, and isoxazole). In some embodiments, Z is not a triazole. In some embodiments, Z is not 1,2,3-triazole or 1,2,4-triazole. In some embodiments, Ar1 is selected from pyridine, bipyridine, phenyl, and carbazole. In some embodiments, the Ar1 group may be substituted (such as by any of the substituents described herein) or unsubstituted.For example, Ar1 can be substituted with the following: –C(O)-CH3, –C(O)-NH2, –C(O)O-CH2CH3, -CF3, –C(O)O-tert-butyl, –C(O)-N(CH3)2, –C(O)-N(H)CH3, –C(O)-N(H)CH2CH3, –C(O)-N(H) (cycloalkyl), –C(O)-N(H)(CH2)4, –C(O)-N(CH2CH3)2, –C(O)-N(H) (tert-butyl), –C(O)-N(H) (C6-cycloalkyl), –C(O)–N(H)(CH2)3, –C(O)–N(H) (C5-cycloalkyl), –C(O)–N (H)(C4–cycloalkyl), –C(O)–N(H)(phenyl), –C(O)–N(H)(CH2CH2)N(H)C(O)–CH3, –C(O)–N(H)C(NH)(NH2), –C(O)–N(CH2CH2CH3)2, –C(O)–N–(C(H)(CH3)(CH2CH3))2, –C(O)–N(CH2CH2CH2CH3)2, –C(O)–N(H)(benzyl), –C(O)–N(H)–CH2CH2OH, –C(O)–S–CH2CH3, –C(O)N(CH2)2, –N(H)S(O)(O)–phenyl, – C(O)N(H)CH2CH2CF3, –C(O)N(H)CH3, –C(O)OCH2CH3, –C(O)–N(H)–CH2CH2O–CH2CH2O–CH3, –C(O)–N(H)–(CH2CH2O)8–CH3, –C(O)N(H)–CH2– CH2–CH2–N(H)–CH2–CH2–CH2–N(H)–CH2–CH2–CH2–NH2, –C(O)N(H)–CH2–CH2–NH2, –C(O)N(H)C(O)CH3, –C(O)N(H)–CH2–CH3, –S(O)(O)N(H)– CH2–CH3, –S(O)(O)N–(CH2–CH3)2, –S(O)(O)N(H)–(C3–cycloalkyl), –S(O)(O)N(H)–(CH2CH2)OH, –S(O)(O)OH, –C(O)N(H)C(H)(CH2OH)2, –C(O)N( CH2CH2OH)2, –C(O)N(H)CH2CH2=CH2, –C(O)CH2N(CH3)(CH2)(COOH), –S(O)(O)(CH2CH3), –S(O)(O)N(H)CH2-CH(OH)(CH2OH), –S(O)(O)-NH2 Or S(O)(O)N(H)CH2-C(O)(O-tert-butyl).

[0869] In some embodiments, the PEM compound of formula (I) includes its solvates, hydrates, chelates, and salts. In some cases, the PEM compound may be amorphous, while in others it may be crystalline. Furthermore, some crystalline forms of the compound may exist as polymorphs, which are also considered herein. Additionally, some compounds may also form solvates with water or other organic solvents. Such solvates are similarly included within the scope of the compounds described herein.

[0870] In some embodiments, the PEM compound of formula (I) may be in the form of a chelate such as a copper chelate. A copper chelate may be formed by combining the PEM compound of this disclosure with copper sulfate. In some embodiments, depending on the substituents on the Ar1 and Ar2 groups, the PEM compound of formula (I) may be in the form of a salt (an acid addition salt or a base addition salt).

[0871] In some embodiments, the PEM structure includes all its stable stereoisomers. Therefore, in some embodiments, the PEM compounds described herein may have one or more chiral (or asymmetric) centers, thus producing enantiomers, diastereomers, and other stereoisomers that can be defined according to absolute stereochemistry as (R)- or (S)-. When the compounds described herein contain an alkene double bond or other geometrically asymmetric centers, unless otherwise stated, it is intended to indicate that these compounds simultaneously contain both E and Z geometric isomers (e.g., cis or trans). Similarly, unless otherwise stated, all possible isomers, as well as their racemic and optically pure forms, and also all tautomers, are intended to be included. Various stereoisomers and mixtures thereof are thus contemplated, including “enantiomers,” which refers to two stereoisomers whose molecules are non-overlapping mirror images of each other. Therefore, compounds may exist in any isomeric form, including racemates, racemic mixtures, and as individual enantiomers or diastereomers.

[0872] In some embodiments, this disclosure provides polymers of the PEM compounds described herein. In some embodiments, the PEM compounds disclosed herein are used as monomers that can be polymerized into polymers (e.g., via cationic polymerization, such as by incubating KCl with PEM). As used herein, the term "polymer" is defined to include homopolymers, copolymers, and oligomers. The term "homogeneous polymer" is defined as a polymer derived from a single monomeric substance (e.g., a single PEM monomeric substance). The term "copolymer" is defined as a polymer derived from more than one monomeric substance, including copolymers obtained by copolymerizing two monomeric substances (such as two different PEM monomeric substances). The term "oligomery" is defined as a low molecular weight polymer wherein the number of repeating units does not exceed twenty. In some embodiments, the polymers of this disclosure include two repeating units, such as three repeating units, such as four repeating units, such as five repeating units, such as six repeating units, etc. In some embodiments, the polymers, copolymers, and / or oligomers are formed through covalent interactions, ionic interactions, or weak interactions.

[0873] In some embodiments, this disclosure provides compositions comprising molecules of any two or more of the compounds disclosed herein (such as two or more compounds having any of formulas (I), (IA), (IB), (IC), and / or (ID)). In some embodiments, the composition comprises molecules of 10 or more of the compounds disclosed herein, such as 20 or more of the compounds disclosed herein, such as 50 or more of the compounds disclosed herein, such as 100 or more of the compounds disclosed herein, such as 200 or more of the compounds disclosed herein, etc.

[0874] In some embodiments, the PEM compounds of this disclosure are generally water-soluble. One measure of water solubility is the logP value of the compound. The logP value can be calculated using commercial software based on the chemical structure of the compound. For example, CHEMDRAW chemical drawing software (Cambridgesoft Limited, a subsidiary of PerkinElmer Holdings) can calculate the logP value of a drawn chemical structure. In one embodiment, the PEM compounds of this disclosure have a logP of at least 4.9.

[0875] In some embodiments, the compounds of this disclosure, for example, PEM compounds of any of formulas (I), (IA), (IB), (IC), and (ID) as described above, can typically be synthesized by reacting a diethynyl compound of formula Ar1(C≡CH)2 with an azide compound of formula Ar2-N3, such as in the presence of a Cu(I) catalyst. See also Crowley JD, McMorran DA (2012) “Click-Triazole” Coordination Chemistry: Exploiting 1,4-Disubstituted-1,2,3-Triazoles as Ligands. See: Košmrlj J. (ed.) Click Triazoles. Topics in Heterocyclic Chemistry, Vol. 28. Springer, Berlin, Heidelberg doi.org / 10.1007 / 7081_2011_67.

[0876] In some embodiments, non-limiting examples of suitable compounds of the formula Ar1(C≡CH)2 include:

[0877] , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , and .

[0878] In other embodiments, the compound of formula Ar1(C≡CH)2 includes:

[0879] , , , , , , , , , , , , , , , , , , , , , , , , and .

[0880] Non-limiting examples of suitable Ar2-N3 compounds include:

[0881] , , , , , , , , , , , , , , , and .

[0882] Specific and similar reactants can also be identified by the index of known chemicals compiled by the American Chemical Society's Chemical Abstracts Service, which is available in most public and university libraries, as well as through online databases (contact the American Chemical Society in Washington, D.C. for more details). Known but not commercially available chemicals can be prepared by custom chemical synthesis companies, many of which offer custom synthesis services to standard chemical suppliers (e.g., those listed above). A reference to the preparation and selection of pharmaceutical salts in this disclosure is PH Stahl & CG Wermuth, “Handbook of Pharmaceutical Salts,” Verlag Helvetica Chimica Acta, Zurich, 2002.

[0883] Compounds of formula Ar1(C≡CH) are commercially available from sources such as TCI America (Portland, Oregon, USA), which sells, for example, 1,3-diethynylbenzene, 1,4-diethynylbenzene, 2,6-diethynylpyridine and 3,6-diethynylcarbazole.

[0884] Generally, ethynyl aromatic compounds can be prepared from aryl aldehydes via a Seyferth-Gilbert homologation reaction using dimethyl (diazomethyl)phosphonate, which is available from Millipore Sigma Corp. (St. Louis, Missouri, USA). Alternatively, dimethyl (diazomethyl)phosphonate can be generated in situ from dimethyl 1-diazo-2-oxopropylphosphonate (Ohira-Bestmann reagent). See, for example, Seyferth et al., J. Org. Chem. 36(10): 1379–1386 (1971). doi:10.1021 / jo00809a014 and Bestman et al., Synlett. 1996 (06):521–522 (1996). doi:10.1055 / s-1996-5474.

[0885] Another route to prepare ethynyl aromatic compounds is by Sonogashira coupling of a haloaromatic compound with (tert-butyldimethylsilyl)acetylene in the presence of a palladium catalyst. The ethynyl aromatic compound is formed upon subsequent deprotection of the silyl group. See, for example, Sonogashira, Organomet. Chem., 653: 46–49 (2002). doi:10.1016 / s0022-328x(02)01158-0.

[0886] The following reactions (I), (II), and (III) illustrate exemplary preparations of diethynyl aromatic compounds. In reaction (I), 2,6-dibromopyridin-4-amine is converted to the corresponding 2,6-diethynylpyridin-4-amine compound. In reaction (II), 2,6-diiodo-4-nitroaniline is converted to the corresponding 2,6-diethynyl-4-nitroaniline. In reaction (III), 2-hydroxy-3,5-diiodobenzoic acid is converted to the corresponding 3,5-diethynyl-2-hydroxybenzoic acid. In each case, the conversion is carried out via the intermediate ditrimethylsilyl (TMS) compound, as shown in the figure.

[0887]

[0888] Each of these reaction products, namely 2,6-diethynylpyridin-4-amine, 2,6-diethynyl-4-nitroaniline, and 3,5-diethynyl-2-hydroxybenzoic acid, can serve as a precursor for the preparation of Ar1 in the PEM of this disclosure. Therefore, each of them represents an Ar1(C≡CH)₂ compound, which can be reacted with an azide compound of formula Ar₂-N₃ in the presence of a Cu(I) catalyst to provide PEM. Reactions (I), (II), and (III) illustrate the preparation of precursors of the substituted Ar1 moiety of this disclosure.

[0889] Compounds of the formula Ar2-N3 are also commercially available, for example, from TCI America (Portland, Oregon, USA), Synthonix (Wake Forest, North Carolina, USA), SigmaAldrich (St. Louis, Missouri, USA), Toronto Research Chemicals (Toronto, Canada), and AnaSpec (Fremont, California, USA). Typically, Ar2-N3 azides can be prepared by nucleophilic substitution of electrophilic compounds such as alkyl, benzyl, or allyl iodide or allyl bromide with sodium azide.

[0890] Generally, the compounds used in the reactions described herein can be prepared from commercially available chemicals and / or from compounds described in chemical literature, according to organic synthesis techniques known to those skilled in the art. "Commercially available chemicals" can be obtained from standard commercial sources including: Across Organics (Pittsburgh, Pennsylvania), Aldrich Chemical (Milwaukee, Wisconsin, including Sigma Chemical and Fluka), Apin Chemicals Ltd (Milton Estate, UK), Avocado Research (Lancashire, UK), BDH Inc. (Toronto, Canada), Bionet (Cornwall, UK), Chemservice Inc. (Westchester, Pennsylvania), Crescent Chemical Co. (Hopog, NY), Eastman Organic Chemicals, Eastman Kodak Company (Rochester, NY), Fisher Scientific Co. (Pittsburgh, Pennsylvania), Fisons Chemicals (Leicestershire, UK), Frontier Scientific (Logan, Utah), ICN Biomedicals, Inc. (Costa Mesa, California), Key Organics (Cornwall, UK), Lancaster Synthesis (Wyndham, New Hampshire), Maybridge Chemical Co. Ltd. (Cornwall, UK), Parish Chemical Co. (Orem, Utah), Pfaltz & Bauer, Inc. (Waterbury, Connecticut), Polyorganix (Houston, Texas), Pierce Chemical Co. (Rockford, Illinois), Riedel de Haen AG (Hanover, Germany), Spectrum Quality Product, Inc. (New Brunswick, New Jersey), TCI America (Portland, Oregon), TransWorld Chemicals, Inc. (Rockville, Maryland), and Wako Chemicals USA, Inc. (Richmond, Virginia).

[0891] In one embodiment, the PEM compound of this disclosure, such as the PEM compound of formula (I), is present in the composition. For example, the PEM compound of this disclosure may be present in a composition that also contains an aqueous buffer. In one embodiment, the PEM compound of this disclosure is present in a composition containing a biomolecule (e.g., a polypeptide and / or a polynucleotide). The polypeptide may be an enzyme, such as a DNA polymerase. The following definitions may help in understanding these compositions and some of their uses.

[0892] As used herein, a “nucleic acid,” also known as a polynucleotide, is a covalently linked chain of nucleotides in which the 3’ position of the pentose sugar of one nucleotide is linked to the 5’ position of the next nucleotide by a phosphodiester group. Nucleic acid molecules can be deoxyribonucleic acid (DNA), ribonucleic acid (RNA), or a combination of both. DNA (deoxyribonucleic acid) and RNA (ribonucleic acid) are biologically existing polynucleotides in which nucleotide residues are linked in a specific sequence by phosphodiester bonds. As used herein, the terms “nucleic acid,” “polynucleotide,” or “oligonucleotide” include any polymeric compound having a linear backbone of nucleotides. Oligonucleotides, also known as oligomers, are typically shorter, chain-like polynucleotides. When targeted for sequencing, nucleic acids are often referred to as “target nucleic acids” or “target sequences.”

[0893] As used herein, the term “template-dependent mode” is intended to refer to a process involving template-dependent extension of primer molecules (e.g., DNA synthesis via DNA polymerase). The term “template-dependent mode” refers to the synthesis of polynucleotides in RNA or DNA, where the sequence of the newly synthesized polynucleotide chain is determined by well-known complementary base pairing rules (see, for example, Watson, JD et al., In: Molecular Biology of the Gene, 4th Ed., WABenjamin, Inc., Menlo Park, Calif. (1987)).

[0894] As used in this article, “nucleic acid polymerase” is an enzyme commonly used to link 3'-OH 5'-triphosphate nucleotides, oligomers and their analogues. Polymerases include, but are not limited to, DNA-dependent DNA polymerase, DNA-dependent RNA polymerase, RNA-dependent DNA polymerase, RNA-dependent RNA polymerase, T7 DNA polymerase, T3 DNA polymerase, T4 DNA polymerase, T7 RNA polymerase, T3 RNA polymerase, SP6 RNA polymerase, DNA polymerase 1, Klenow fragment, Thermophilus aquaticus DNA polymerase, Tth DNA polymerase, VentR® DNA polymerase (New England Biolabs), Deep VentR® DNA polymerase (New England Biolabs), Bst DNA polymerase large fragment, Stoeffel fragment, 9° N DNA polymerase, 9° N DNA polymerase, Pfu DNA polymerase, Tfl DNA polymerase, Tth DNA polymerase, RepliPHI Phi29 polymerase, Tli DNA polymerase, eukaryotic DNA polymerase β, telomerase, and Therminator™. Polymerases (New England Biolabs), KOD HiFi™ DNA polymerase (Novagen), KOD1 DNA polymerase, Q-β replicase, terminal transferase, AMV reverse transcriptase, M-MLV reverse transcriptase, Phi6 reverse transcriptase, HIV-1 reverse transcriptase. The polymerases disclosed herein may be variants, mutants, or chimeric polymerases.

[0895] As used herein, “DPO4-type DNA polymerase” refers to a DNA polymerase or related Y-family DNA polymerase naturally expressed by archaea, sulfur-bearing fungi, or other fungi that typically functions in the replication of damaged DNA through a process called transdamage synthesis (TLS). Y-family DNA polymerases are homologous to DPO4 polymerase; examples include prokaryotic enzymes PolII, PolIV, PolV, archaea Dbh, and eukaryotic enzymes Rev3p, Rev1p, Polƞ, REV3, REV1, PolĮ, and Polĸ DNA polymerases, and their chimeras. Modified recombinant DPO4-type DNA polymerases include one or more mutations relative to naturally occurring wild-type DPO4-type DNA polymerases, such as one or more mutations that increase the ability to utilize large nucleotide analogs as substrates or other polymerase properties, and may include further alterations or modifications to the wild-type DPO4-type DNA polymerase, such as one or more deletions, insertions, and / or fusions of additional peptide or protein sequences (e.g., for immobilizing the polymerase on a surface or otherwise labeling the polymerase). Examples of variant polymerases according to this disclosure are variants of *D. sulfidea* DPO4 described in published PCT patent applications WO2017 / 087281 A1 and PCT patent applications PCTUS2018 / 030972 and PCTUS2018 / 64794, the entire contents of which are incorporated herein by reference.

[0896] As used herein, a “nucleic acid polymerase reaction” refers to an in vitro method for preparing new nucleic acid chains or extending existing nucleic acids (e.g., DNA or RNA) in a template-dependent manner. According to this disclosure, a nucleic acid polymerase reaction includes a primer extension reaction that results in the incorporation of a nucleotide or nucleotide analog into the 3' end of a primer, such that the incorporated nucleotide or nucleotide analog is complementary to the corresponding nucleotide of a target polynucleotide. The primer extension product of the nucleic acid polymerase reaction can be further used for single-molecule sequencing or as a template for the synthesis of additional nucleic acid molecules.

[0897] Primer extension reaction reagents typically include (i) a polymerase; (ii) a buffer; and (iii) one or more extendable nucleotides or nucleotide analogs. Primer extension reactions can be used to measure the length of the resulting nucleic acid products under specific experimental conditions and to determine the effect of various polymerase reaction additives (e.g., PEM) on polymerase activity by comparing the lengths of the extension primer products (e.g., using gel electrophoresis).

[0898] As used in this article, “enhanced nucleic acid polymerase reaction” refers to the ability of an additive (such as PEM) to enable a nucleic acid polymerase to synthesize a primer extension product that is at least one subunit longer than in the absence of PEM.

[0899] As used herein, the rate of a nucleic acid polymerase reaction refers to the average speed at which the nucleic acid polymerase elongates the polymer chain. As used herein, the terms "rate" and "elongation" are used interchangeably. The nucleotide incorporation assay by Hogrefe et al. (Methods in Enzymol. Vol. 334, pp. 91-116 (2001)) can be used to measure the polymerization rate. In short, polymerase activity can be measured by… 32 P-dCTP was incorporated into activated salmon sperm DNA (purchased from Pharmacia; activation protocol see Procedures in Nucl. Acid Res. (edited by Cantoni and Davies), p. 263-276 (1966) at p. 264) for measurement. Reaction buffers could be, for example, 50 mM Tris-HCl (pH 8.0), 5 mM MgCl2, 1 mM dithiothreitol (DTT), 50 µg / ml bovine serum albumin (BSA), and 4% (v / v) glycerol. Large amounts of nucleotide substrate and DNA were used, typically at least 10 times the Km of the polymerase being detected; for example, 200 µM each of dATP, dTTP, and dGTP, 195 µM dCTP plus 5 µM labeled dCTP, and 250 µg / ml activated DNA. The reaction is quenched on ice, and aliquots of the reaction mixture are spotted onto an ion-exchange filter (e.g., Whatman DE81). Undoped nucleotides are washed, and then scintillation counting is performed to measure the incorporated radioactivity.

[0900] As used herein, “increase rate” means an increase of 5% to 10%, 5% to 20%, 10% to 50%, or 50% to 100% or more compared to a polymerization reaction lacking an increase rate as defined herein.

[0901] As used herein, “sustained synthesis capacity” refers to the extent to which a nucleic acid polymerase polymerizes during a single contact between the polymerase and its template; that is, its property of continuing to act on the substrate rather than dissociating from it. The degree of polymerization refers to the amount of nucleotides or nucleotide analogs added by the polymerase during a single contact with its template. Sustained synthesis capacity can depend on the properties of the polymerase, the sequence of the template, the structure of the nucleotide or nucleotide analog substrate, and reaction conditions such as salt concentration, temperature, or the presence of specific additives.

[0902] As used herein, “increased sustained synthetic capacity” means an increase of 5% to 10%, 5% to 20%, 10% to 50%, or 50% to 100% or more compared to a polymerization reaction lacking a PEM with increased sustained synthetic capacity as defined herein. Methods for measuring the sustained synthetic capacity of nucleic acid polymerases are well known in the art, for example, as described in Sambrook et al., 1989, In ​​Molecular Cloning, 2nd Edition, CSH Press, 7.79-7.83 and 13.8, as well as in U.S. Patent Application Publication No. 2002 / 0119467, PCT Application No. WO01 / 92501, and U.S. Patent No. 5,972,603, the entire contents of which are incorporated herein by reference.

[0903] As used herein, the term "fidelity" refers to the accuracy of nucleic acid polymerization by a template-dependent nucleic acid polymerase. DNA polymerase fidelity is measured by the error rate (the frequency of inaccurate nucleotide incorporation, i.e., nucleotides not incorporated in a template-dependent manner). DNA polymerase fidelity or error rate can be measured using assays known in the art (see, for example, Lundburg et al., 1991 Gene, 108:1-6). As used herein, "increased fidelity" means an increase of 5% to 10%, 10% to 50%, or 50% to 100% or more compared to a polymerization reaction lacking an additive that increases fidelity as defined herein.

[0904] As used in this article, the term "multiple" means "at least two".

[0905] “XNTP” is a scalable, 5'-triphosphate-modified nucleotide substrate compatible with template-dependent enzymatic polymerization. XNTP has two distinct functional components: a 5'-triphosphate amino ester nucleobase and a tandem chain linked to each nucleoside triaminophosphate, the position of which allows for controlled expansion via intranucleotide cleavage of the aminophosphate bond. As used herein, XNTP is an exemplary “non-natural, highly substituted nucleotide analog substrate.” Exemplary XNTPs and methods for their preparation are described, for example, in PCT application WO2016 / 081871 published by the applicant, the entire contents of which are incorporated herein by reference.

[0906] The “Xpandomer intermediate” is an intermediate product assembled by XNTPs (also referred to herein as the “daughter strand”), and is formed by template-directed assembly mediated by XNTP polymerase using a target nucleic acid template. The newly synthesized Xpandomer intermediate is a constrained Xpandomer. In a process step where the aminophosphate bonds provided by XNTPs are cleaved, the constrained Xpandomer becomes unconstrained and is the Xpandomer product that extends as the chain lengthens.

[0907] "Xpandomer" or "Xpandomer product" is a synthetic molecular construct generated by constrained Xpandomer amplification, synthesized itself via template-directed assembly of an XNTP substrate. The Xpandomer is elongated relative to the target template from which it originates. It consists of tandem subunits, each subunit being a motif, and each motif being a member of a library, including sequence information, tandem strands, and optionally some or all of the substrate, all derived from a formative substrate construct. The Xpandomer is designed to extend longer than the target template, thereby reducing the linear density of the target template's sequence information along its length. Furthermore, the Xpandomer optionally provides a platform for increasing the size and abundance of the reporter, thereby improving the signal-to-noise ratio of detection. The lower linear information density and stronger signal improve resolution and reduce the sensitivity requirements for detecting and decoding the template strand sequence.

[0908] A “chain” or “chain member” refers to a polymer or molecular construct having a generally linear dimension and end portions at each of its two opposite ends. Chains form XNTPs by being bonded to nucleoside triphosphates at the end portions. Bonds are used to constrain the chain in a “constrained configuration.” Chains have both “constrained configurations” and “extended configurations.” Constrained configurations are found in XNTPs and daughter chains or Xpandomer intermediates. The constrained configuration of a chain is a precursor to the extended configuration, as seen in Xpandomer products. The transition from a constrained configuration to an extended configuration results in the cleavage of selectively cleavable aminophosphate bonds. A chain contains one or more reporter or reporter constructs along its length that can encode sequence information of the substrate. Chains provide a method for extending the length of Xpandomer, thereby reducing the linear density of sequence information.

[0909] A “tethering element” or “tethering segment” is a polymer with a generally linear size and two ends, wherein the ends form terminal bonds for connecting the tethering element. A tethering element is a segment of a tether. Such polymers may include, but are not limited to: polyethylene glycol, polyethylene glycol, polypyridine, polyisocyanate, polyisocyanate, poly(triarylmethyl)methacrylate, polyaldehyde, polypyrrolidone, polyurea, polyethylene glycol phosphate diester, polyacrylate, polymethacrylate, polyacrylamide, polyethylene ester, polystyrene, polyamide, polyurethane, polycarbonate, polybutyrate, polybutadiene, polybutyrolactone, polypyrrolidone, polyphosphonate, polyacetamide, polysaccharide, polyhyaluronic acid ester, polyamide, polyimide, polyester, polyethylene, polypropylene, polystyrene, polycarbonate, polyterephthalate, polysilane, polyurethane, polyether, polyamino acid, polyglycine, polyproline, N-substituted polylysine, peptide, N-substituted peptide with side chain, poly-N-substituted glycine, peptide-like peptide, carboxyl-substituted peptide with side chain, homopeptide, oligonucleotide, ribonucleic acid oligonucleotide, deoxynucleic acid oligonucleotide, modified to prevent Watson-Crick Oligonucleotides with base pairing, oligonucleotide analogs, polycytidylic acid, polyadenylic acid, polyuridine acid, polythymidine, polyphosphate, polynucleotides, polynucleotides, polyethylene glycol-phosphate diesters, peptide polynucleotide analogs, threonine-polynucleotide analogs, ethylene glycol-polynucleotide analogs, morpholino-polynucleotide analogs, locked nucleotide oligomer analogs, polypeptide analogs, branched polymers, comb polymers, star polymers, dendritic polymers, random polymers, gradient and block copolymers, anionic polymers, cationic polymers, stem-ring polymers, rigid segments and flexible segments.

[0910] A reporter consists of one or more reporter elements. Reporter elements are used to analyze the genetic information of a target nucleic acid.

[0911] A reporter construct contains one or more reporters that generate a detectable signal, which typically contains sequence information. This signal information is called the "reporter code," and is subsequently decoded into gene sequence data. The reporter construct may also contain chain segments or other structural components, including polymers, graft copolymers, block copolymers, affinity ligands, oligomers, haptens, aptamers, dendritic polymers, linker groups, or affinity-binding groups (e.g., biotin).

[0912] "Reporter codes" are genetic information derived from the measurement signals of reporter constructs. Reporter codes are decoded to provide sequence-specific genetic information data.

[0913] Therefore, in some embodiments, this disclosure provides a composition comprising PEM as disclosed herein and a buffer. In other embodiments, this disclosure provides a composition comprising PEM as disclosed herein and a plurality of nucleotides and / or nucleotide analogs. In other embodiments, this disclosure provides a composition comprising PEM as disclosed herein and a polynucleotide. In other embodiments, this disclosure provides a composition comprising PEM as disclosed herein and a protein, wherein optionally the protein is a polymerase, including any of the polymerases described above.

[0914] In some embodiments, the composition further comprises one or more of the following: MnCl2, buffer, salt, sugar, single-stranded binding protein (SSB), imidazole, pyrazole, triazole, betaine, molecular crowding agent, dimethyl sulfoxide (DMSO), alkyl glycol, glycerol, N-methyl-2-pyrrolidone (NMP), acetamide, butylated hydroxyanisole (BHA), polyphosphate, urea, and a mixture of nucleotides or nucleotide analogs.

[0915] In some embodiments, the salt is a salt selected from the group consisting of: NaCl, NaBr, NaOAc, NaF, sodium formate, sodium dihydrogen phosphate, disodium hydrogen phosphate, NaSO4, sodium carbonate, sodium bicarbonate, sodium hexanoate, monosodium glutamate, sodium perchlorate, CsCl, LiCl, LiOAc, LiF, lithium carbonate, LiPO4, KCl, KOAc, KF, KSO4, potassium dihydrogen phosphate, dipotassium hydrogen phosphate, potassium carbonate, potassium bicarbonate, potassium glutamate, NH4Cl, NH4F, NH4OAc, NH4SO4, NH4Br, ammonium citrate, ammonium carbonate, ammonium bicarbonate, ammonium sulfite, ammonium glutamate, ammonium dihydrogen phosphate, tetramethylammonium chloride (TMACl), trimethylamine N-oxide (TMAO), tetraethylammonium chloride (TEACl), guanidine chloride, guanidine thiocyanate, guanidine carbonate. In some embodiments, the salt is an inorganic salt. In some embodiments, the salt is NaCl or KCl, such as NaCl.

[0916] In some implementations, the sugar is maltose, trehalose, cellobiose, or sucrose.

[0917] In some implementations, the SSB is selected from the group consisting of: KOD (SEQ ID NO: 6), Gp32 (SEQ ID NO: 7), TTH (SEQ ID NO: 8), SSB1 (SEQ ID NO: 9), RecA (SEQ ID NO: 10), RPA (SEQ ID NO: 11), and NCp7 (SEQ ID NO: 12).

[0918] In some embodiments, imidazole includes (in addition to imidazole itself) imidazole derivatives, such as imidazole chloride, imidazole acetate, 1-methylimidazolium, 2-methylimidazolium, 1-ethylimidazolium, 1-ethyl-3-methylimidazolium chloride, 2-methyl-2-imidazoline, 1-butyl-3-methylimidazolium chloride, 1-methylimidazolium chloride, 1-hexyl-3-methylimidazolium salt, 3-octyl-1-methylimidazolium salt, or 1-decyl-3-methylimidazolium salt.

[0919] In some implementations, the alkanediol is ethylene glycol, propylene glycol, or butanediol, such as propylene glycol.

[0920] In some embodiments, the propylene glycol is 1,2-propanediol or 1,3-propanediol, such as 1,2-propanediol. In some embodiments, the butanediol is 1,2-butanediol, 1,3-butanediol, 1,4-butanediol, 2,3-butanediol, 2,4-butanediol or 3,4-butanediol, such as 1,2-butanediol.

[0921] In some implementations, the molecular crowding agent is PEG, such as PEG4k to PEG25k or PEG4k to PEG10k, such as PEG5k, PEG8k or PEG10k.

[0922] In some implementations, the polyphosphate is tripolyphosphate, tetrapolyphosphate, pentapolyphosphate, hexapolyphosphate, trimetaphosphate (TMP), hexametaphosphate (HMP), or polyphosphate 60.

[0923] In one embodiment, this disclosure provides a composition comprising a PEM compound of the present disclosure (e.g., a PEM compound of any of formulas (I), (IA), (IB), (IC), and (ID)) and a molecular crowding agent. Generally, molecular crowding agents comprise a range of large neutral polymers. Examples of useful molecular crowding agents include, but are not limited to, polyethylene glycol (PEG), sucrose, dextran, or polyvinyl alcohol. Exemplary molecular crowding agents and formulations are described in U.S. Patent No. 7,399,590, which is incorporated herein by reference. In one embodiment, the molecular crowding agent is a polyalkylene glycol, optionally having a number average molecular weight of 4,000-10,000. In one embodiment, the molecular crowding agent is a derivative of a polyalkylene glycol, for example, in which one or both of the terminal hydroxyl groups are in the form of an ester or ether group. In some embodiments, the polyalkylene glycol is polyethylene glycol. In one embodiment, the molecular crowding agent is an inert, water-soluble polymer.

[0924] In one embodiment, this disclosure provides a composition comprising the PEM compound of this disclosure and an aqueous buffer. In one embodiment, the PEM compound has any one of formulas (I), (IA), (IB), (IC), and (ID). In one alternative, the composition has a pH of about 6 to 8.5, and the buffer helps to stabilize the pH of the composition. An exemplary buffer is Tris HCl. Other suitable buffers include those known in the art, such as phosphate buffers, citrate buffers, sodium acetate buffers, sodium carbonate buffers, etc. In some embodiments, the buffer is selected from the group consisting of TrisCl, TrisOAc, NH4OAc, MES, and HEPES.

[0925] In one embodiment, this disclosure provides a composition comprising a PEM compound of the present disclosure (e.g., a PEM compound of any of formulas (I), (IA), (IB), (IC), and (ID)) and a polynucleotide. In one alternative, the polynucleotide is single-stranded, such as single-stranded DNA or single-stranded RNA. When the polynucleotide is intended to be used as a primer, the polynucleotide is a single-stranded DNA molecule. When intended to be used as a primer, the polynucleotide may have a length of about 10-60 mer oligonucleotides, for example, 20-30 oligonucleotides. Alternatively, the polynucleotide may be used as a template, in which case it may be single-stranded DNA or single-stranded RNA and may have a length from 30 bases to kilobases and above, for example, 10 kbps and above.

[0926] In one embodiment, this disclosure provides a composition comprising a PEM compound of the present disclosure (e.g., a PEM compound of any of formulas (I), (IA), (IB), (IC), and (ID)) and a protein. For example, the protein may be an enzyme, a nucleic acid polymerase, or a DNA polymerase. An example of a suitable DNA polymerase is a variant of the DPO4 polymerase discussed herein.

[0927] In one embodiment, this disclosure provides a composition comprising a PEM compound of the present disclosure (e.g., a PEM compound of any of formulas (I), (IA), (IB), (IC), and (ID)) and a mixture of nucleotides or nucleotide analogs, wherein, relative to the same polymerization reaction in the absence of at least one compound, the at least one compound increases the quantity and accuracy of nucleotide analogs incorporated into daughter chains during template-dependent polymerization. Optionally, the mixture of nucleotide analogs comprises nucleoside triaminophosphates, wherein each nucleoside triaminophosphate has a nucleobase selected from adenine, guanine, thymine, and cytosine and a polymer chain moiety, wherein a first end of the polymer chain moiety is attached to a nucleobase, and a second end of the polymer chain moiety is attached to an α-phosphate of the nucleoside triaminophosphate to provide nucleotide analog extension by cleavage of the aminophosphate bond. Optionally, the composition further comprises a buffer containing one or more components selected from MnCl2, buffer solutions, salts, sugars, single-stranded binding proteins (SSBs), imidazoles, pyrazoles, triazoles, betaine, molecular crowding agents, dimethyl sulfoxide (DMSO), alkyl glycols, glycerols, N-methyl-2-pyrrolidone (NMP), acetamides, butylated hydroxyanisole (BHA), polyphosphates, and / or ureas. Optionally, the composition further comprises a single-stranded binding protein. Optionally, the composition comprises urea. Optionally, the mixture of nucleotide analogs comprises nucleotide analogs containing a detectable marker, wherein the detectable marker is optionally one of luminescence, chemiluminescence, fluorescence, fluorescence, chromogenesis, or chromogenesis. In one embodiment, the composition comprises two or more of these options, for example, all of these options.

[0928] In one aspect of this disclosure, the PEM and compositions thereof disclosed herein can be used to enhance nucleic acid polymerization reactions or improve the properties of the resulting nucleic acids, such as the length or accuracy of the reaction products. Polymerization reactions include, for example, primer extension reactions, PCR, mutagenesis, isothermal amplification, DNA sequencing, and probe labeling. Such methods are well known in the art. Enhancement can be provided by stimulating nucleotide incorporation through mechanisms such as increasing the sustained synthetic capacity of polymerase (i.e., reducing polymerase dissociation from the template), increasing the rate of substrate binding or enzyme catalysis, and increasing the accuracy or fidelity of nucleotide incorporation. Furthermore, enhancement can be provided by reducing barriers in the nucleic acid template (e.g., secondary structures and double-stranded DNA). Overcoming or improving these barriers by adding PEM can make the polymerization reaction occur more accurately or efficiently, or allow the use of lower denaturation / extension temperatures or isothermal temperatures.

[0929] In some embodiments, PEM may be used in combination with another class of additives to enhance polymerase reactions. One exemplary class of additives is minor groove binding protein (MGB). In one embodiment, MGB is selected from the group consisting of: tertimigrin A and its synthetic analogues, fusiformin, (+)-CC-1065, pyrrolobenzodiazepines, trabectin and its analogues, Hoechst dye and its derivatives, lexitropsin, thiazotropsin A, diamidine, and polyamides. In some embodiments, the at least one minor groove binding moiety is a Hoechst dye. More information on using MGB to enhance polymerase reactions can be found in the application jointly filed by the applicants entitled “ENHANCEMENT OF NUCLEIC ACID POLYMERIZATION BY MGBS”.

[0930] One exemplary polymerase reaction that can be enhanced by PEM is the polymerization of a non-natural nucleotide analogue called “XNTP,” which forms the basis of the “Extended Sequencing” (SBX) protocol developed by Stratos Genomics (see, for example, Kokoris et al., U.S. Patent No. 7,939,259, “High Throughput Nucleic Acid Sequencing by Expansion”). Generally, SBX uses this biochemical polymerization to transcribe the sequence of a DNA template onto a measurable polymer called “Xpandomer.” The transcribed sequence is encoded along the Xpandomer backbone in high signal-to-noise ratio reporters spaced approximately 10 nm apart and designed for a high signal-to-noise ratio, well-differentiated response. These differences provide a significant enhancement in the read efficiency and accuracy of Xpandomer relative to native DNA. An overview of the SBX process is described in Figures 1A, 1B, 1C, and 1D.

[0931] XNTPs are scalable, 5'-triphosphate-modified nucleotide substrates compatible with template-dependent enzymatic polymerization. A highly simplified XNTP is shown in Figure 1A, highlighting the unique characteristics of these nucleotide analogs: XNTP 100 has two distinct functional regions: a selectively cleavable aminophosphate bond 110 linking a 5'-α-phosphate 115 to a nucleobase 105; and a linker 120 located at certain sites within the nucleoside triaminophosphate, allowing for controlled expansion via intranucleotide cleavage of the aminophosphate bond. The XNTP linker comprises linker arms 125A and 125B separated by the selectively cleavable aminophosphate bond. Each linker is connected to one end of a reporter 130 via a linker group (LG), as disclosed in U.S. Patent No. 8,324,360 to Kokoris et al., which is incorporated herein by reference in its entirety. XNTP 100 is described as having a “restricted configuration,” which is characteristic of both the XNTP substrate and the polymerized daughter chain. The restricted configuration of polymerized XNTP is a precursor to the extended configuration, as seen in the Xpandomer product. The transition from the restricted to the extended configuration occurs when the p–N bonds of the aminophosphate ester within the main backbone of the daughter chain break.

[0932] The synthesis of Xpandomer is summarized in Figures 1B and 1C. During assembly, monomeric XNTP substrates 145 (XATP, XCTP, XGTP, and XTTP) are polymerized at the extendable ends of nascent daughter strands 150 using a template-guided polymerization process guided by a single-stranded template 140. Typically, this process begins with a primer and proceeds along the 5' to 3' direction. Typically, a DNA polymerase or other polymerase is used to form the daughter strand, and conditions are selected to obtain a complementary copy of the template strand. After the daughter strand is synthesized, the coupled ligand contains a constrained Xpandomer that further contains the daughter strand. The ligand in the daughter strand has a “constrained configuration” of the XNTP substrate. The constrained configuration of the ligand is a precursor to the extended configuration, as seen in the Xpandomer product.

[0933] As shown in Figure 1C, the transition from the constrained configuration 160 to the expanded configuration 165 is caused by the selective cleavage of phosphodies (represented by unshaded ellipses for simplicity) within the main chain of the daughter strand. In this embodiment, the tandem strand contains one or more reporter constructs, 130A, 130C, 130G, or 130T, specific to the nucleobases to which they are linked, thereby encoding the sequence information of the template. In this way, the tandem strand provides a means to extend the length of the Xpandomer and reduce the linear density of the parent strand sequence information.

[0934] Figure 1D illustrates the translocation of Xpandomer 165 from cis reservoir 175 to trans reservoir 185 via nanopore 180. After passing through the nanopore, each reporter of the linearized Xpandomer (labeled "G", "C", and "T" in this illustration) generates a unique and repeatable electronic signal (represented by superimposed trace 190) specific to the nucleobase it is attached to.

[0935] Figure 2 illustrates the generalized structure of XNTP in more detail. XNTP 200 comprises a nucleobase triphosphate amino ester 210 with linker arms 220A and 220B, which are separated by selectively cleavable aminophosphate bonds 230. The chain is linked to the nucleoside triphosphate at linker groups 250A and 250B, wherein the first chain terminus is attached to a heterocycle 260 (represented here by cytosine, although the heterocycle may be any of the four standard nucleobases A, C, G, or T), and the second chain terminus is attached to an α-phosphate 270 of the nucleobase backbone. Those skilled in the art will understand that many suitable coupling chemistry known in the art can be used to form the final XNTP substrate product; for example, chain coupling can be accomplished via triazole bonds.

[0936] In this embodiment, tandem 275 comprises several functional elements, including enhancers 280A and 280B, reporter codes 285A and 285B, and translation control elements (TCEs) 290A and 290B. Each of these features performs a unique function during the translocation of Xpandomer through the nanopore and the generation of a unique and reproducible electronic signal. Tandem 275 is designed for translocation control via hybridization (TCH). As shown, the TCE provides a hybridization region that can form a doublet with the complementary oligomer (CO) and is located near the reporter codes. Different reporter code sizes can prevent ions from flowing through the nanopore at different measurable levels. Specific reporter codes can be synthesized efficiently using phosphoramide chemistry commonly used for oligonucleotide synthesis. Reporter codes can be designed by selecting sequences of specific phosphoramides from commercially available libraries. Such libraries include, but are not limited to, polyethylene glycol with a length of 1 to 12 or more ethylene glycol units, aliphatic compounds with a length of 1 to 12 or more carbon units, deoxyadenosine (A), deoxycytosine (C), deoxyguanosine (G), deoxythymidine (T), and baseless compounds (Q). The double-stranded TCE associated with the reporter code also contributes to ion flow blocking; therefore, the combination of the reporter code and the TCE can be referred to as a "reporter." Following the reporter code is the enhancer, which in one embodiment comprises a spermine polymer.

[0937] Figure 3 illustrates one embodiment of Xpandomer cleaved during the translocation of α-hemolysin nanopores. The bio-nanopore is embedded within a lipid bilayer membrane that separates and electrically isolates two electrolyte reservoirs. A typical electrolyte is 1 molar KCl buffered to pH 7.0. When a small voltage (typically 100 mV) is applied across the bilayer, the nanopore restricts the flow of ion currents and is the dominant resistance in the circuit. The Xpandomer reporter is designed to provide a specific level of ion current blocking, and sequence information can be read by measuring the sequence of ion current levels as the reporter sequence translocates through the nanopore.

[0938] α-Hemolysin nanopores are typically oriented, thus translocation occurs by entering the vestibular side and exiting the styloid side. For example... Figure 3 As shown, the nanopores are oriented to capture Xpandomer primarily from the stem side. This orientation is advantageous when using the TCH method because it reduces occlusion artifacts that occur when the Xpandomer first enters the vestibule. Unless otherwise stated, the stem side is initially the assumed translocation orientation. As the Xpandomer translocates, the reporter enters the stem until its duplex, TCE, stops at the stem inlet. The duplex diameter is approximately 2.4 nm, while the inlet diameter is approximately 2.2 nm, thus the reporter is retained in the stem until the complementary strand 395 of the duplex dissociates (releases), and then translocation proceeds to the next reporter. Free complementary strands are highly detrimental to entry into the nanopore because the Xpandomer is still translocating and diffusing out of the pore.

[0939] In one implementation, each member of the reporter code (following the duplex) is formed by an ordered selection of phosphorus amides, selectable from a number of commercial libraries. The phosphorus amide of each component contributes to the net ionic resistance based on its position within the nanopore (following the duplex termination), its displacement, its charge, its interaction with the nanopore, its chemical and thermal environment, and other factors. The charge portion on each phosphorus amide is attributed to phosphate ions with a nominal charge of -1, effectively reduced by counter-ionic shielding. The pulling force on the duplex is due to the local electric field acting on these effective charges along the reporter. Since each reporter molecule can have a different charge distribution, it can exert different forces on the duplex for a given applied voltage. The forces transmitted along the reporter backbone also serve to elongate the reporter to provide a repeatable blocking response.

[0940] The extended sequencing (SBX) method developed by the inventors offers significant performance enhancements in the efficiency and accuracy of Xpandomer sequence reading relative to native DNA. However, the initial transcription of native DNA template sequences into measurable Xpandomers depends on the ability of DNA polymerases to utilize XNTPs as substrates (the general structure of XNTPs is discussed herein with reference to Figures 1A and 2). The inventors have found that most DNA polymerases are inefficient at polymerizing XNTPs. However, the inclusion of suitable additives, such as the PEM disclosed herein, improves the efficiency and accuracy of XNTP polymerization into Xpandomers. Therefore, the PEM disclosed herein can be used in the context of the SBX method to enhance DNA polymerase primer extension reactions using XNTPs as substrates.

[0941] Examples of suitable concentrations of components in a composition, when present, are as follows:

[0942] 0.5 to 50 mM PEM (such as 1 to 50 mM, such as 2 to 50 mM, such as 3 to 50 mM, such as 4 to 50 mM, such as 5 to 50 mM).

[0943] 0.02 to 0.2 µg / µl polymerase,

[0944] 20 to 150 mM nucleotides or nucleotide analogs,

[0945] 0.1 to 1 mM MnCl2,

[0946] 20 to 100 mM buffer,

[0947] 100 to 200 mM salt,

[0948] 0.05% to 0.5% (w / v) sugar,

[0949] 0.05 to 0.5 mM polyphosphate,

[0950] 140 to 300 mM imidazole,

[0951] 50 to 200 mM pyrazole,

[0952] 50 to 200 mM triazole,

[0953] 2% to 8% (w / v) alkanediols,

[0954] 2% to 10% (w / v) acetamide,

[0955] 0.1% to 1% (w / v) glycerin

[0956] 200 to 400 mM betaine,

[0957] 10% to 30% (w / v) PEG,

[0958] 3% to 10% (v / v) NMP,

[0959] 0.5 to 1.5 mM BHA,

[0960] 1% to 5% (v / v) DMSO,

[0961] 0.5 to 2 mM urea,

[0962] 0.02 to 1 µg / µl SSB.

[0963] In some embodiments, the primer extension reaction may comprise an aqueous composition consisting of the following components in water: 0.02 to 0.2 µg / µl polymerase, 0.1 to 1 mM MnCl2, 20 to 150 µM XNTP, 20 to 100 mM buffer, 100 to 200 mM salt, 0.05 to 0.5% (w / v) sugar, 5 to 50 mM PEM, 0.05 to 0.5 mM polyphosphate, and optionally one or more of the following: 140 to 300 mM imidazole, 2% to 8% (w / v) alkyldiol, 2% to 10% (w / v) acetamide, 0.1% to 1% (w / v) glycerol, 200 to 400 mM betaine, 10% to 30% (w / v) PEG, 3% to 10%... (v / v) NMP, 0.5 to 1.5 mM BHA and 1 to 5% (v / v) DMSO, and further optional 0.02 to 1 µg / µl SSB.

[0964] In some embodiments, the primer extension reaction may comprise an aqueous composition consisting of the following components in water: 0.02 to 0.2 µg / µl polymerase, 0.1 to 1 mM MnCl2, 20 to 150 µM XNTP, 20 to 100 mM buffer, 100 to 200 mM salt, 0.05 to 0.5% (w / v) sugar, 5 to 50 mM PEM, 0.05 to 0.5 mM HMP, and optionally one or more of the following: 140 to 300 mM imidazole, 200 to 400 mM betaine, 10% to 30% (w / v) PEG, 3 to 10% (v / v) NMP, 0.5 to 1.5 mM BHA, and 1 to 5% (v / v) DMSO, and further optionally 0.02 to 1 µg / µl SSB.

[0965] In some embodiments, the primer extension reaction may include an aqueous composition consisting of the following components in water: 0.02 to 0.2 µg / µl polymerase, 0.1 to 1 mM MnCl2, 20 to 150 µM XNTP, 20 to 100 mM buffer, 100 to 200 mM salt, 0.05 to 0.5% (w / v) sugar, 5 to 50 mM PEM, 0.05 to 0.5 mM HMP, and optionally one or more of the following: 140 to 300 mM imidazole, 10% to 30% (w / v) PEG and 2% to 8% (w / v) alkyldiol, and further optionally 0.02 to 1 µg / µl SSB.

[0966] In some embodiments, the primer extension reaction may include an aqueous composition consisting of the following components in water: 0.02 to 0.2 µg / µl polymerase, 0.1 to 1 mM MnCl2, 20 to 150 µM XNTP, 20 to 100 mM buffer, 100 to 200 mM salt, 0.05 to 0.5% (w / v) sugar, 5 to 50 mM PEM, 0.05 to 0.5 mM HMP, and optionally one or more of the following: 140 to 300 mM imidazole, 10% to 30% (w / v) PEG and 2% to 10% (w / v) acetamide, and further optionally 0.02 to 1 µg / µl SSB.

[0967] In some embodiments, the primer extension reaction may include an aqueous composition consisting of the following components in water: 0.02 to 0.2 µg / µl polymerase, 0.1 to 1 mM MnCl2, 20 to 150 µM XNTP, 20 to 100 mM buffer, 100 to 200 mM salt, 0.05 to 0.5% (w / v) sugar, 5 to 50 mM PEM, 0.05 to 0.5 mM HMP, and optionally one or more of the following: 140 to 300 mM imidazole, 10% to 30% (w / v) PEG, 0.1% to 1% (w / v) glycerol, 2% to 8% (w / v) alkyldiol and 2% to 10% (w / v) acetamide, and further optionally 0.02 to 1 µg / µl SSB.

[0968] Typically, primer extension reactions may also include primers and templates, such as oligonucleotide templates.

[0969] A representative primer extension reaction may include the following reagents: 2 pmol primers, 2.2 pmol 45 mer oligonucleotide template, 50 pmol of each XNTP (XATP, XCTP, XGTP, and XTTP), 50 mM Tris HCl, pH 6.79, 200 mM NaCl, 20% PEG, 5% NMS, 0.5 nmol polyphosphate 60.1g, 0.3 mM MnCl2, and 0.6 µg purified recombinant DNA polymerase protein. PEM is typically added to this mixture at concentrations ranging from micromolar to millimolecular. Additional additives, such as single-strand binding proteins (SSBs), urea, and NMS, are also included in the reaction. The reaction is run at 23 °C for 1 hour. The reaction product (i.e., the constrained Xpandomer) is treated to cleave the aminophosphodiester bonds, resulting in a linearized Xpandomer. The reaction products were analyzed by gel electrophoresis on 4%–12% acrylamide gels to resolve and visualize Xpandomer products of different lengths.

[0970] Therefore, in one embodiment, this disclosure provides an aqueous (water-containing) composition comprising PEM and a buffer, particularly a buffer suitable for DNA polymerization reactions, wherein Tris HCl is an exemplary buffer of this type. In one embodiment, this disclosure provides a composition comprising PEM and a DNA polymerase protein. In one embodiment, this disclosure provides a composition comprising PEM and a polynucleotide, such as an oligonucleotide of 20-90 mer, 20-60 mer, 30-90 mer, or 30-60 mer. In one embodiment, this disclosure provides a composition comprising each of these components, i.e., an aqueous composition comprising PEM, a buffer, a DNA polymerase protein, and a polynucleotide.

[0971] To investigate ways to enhance the accuracy of XNTP polymerization, the primer extension products can be sequenced using the SBX protocol. In short, the Xpandomer product from constrained XNTP polymerization is cleaved to produce linearized Xpandomer. This is achieved by first quenching the extension reaction with a solution containing 100 mM EDTA, 2 mM THPTA, and 2% Tween-20. The sample is then amine-modified with a DMF solution of 1 M NaHCO3 and 1 M succinic anhydride. Cleavage of the aminophosphate bonds is performed with 37% HCl, and the linearized Xpandomer is purified using a QIAquick column (QIAGEN, Inc.).

[0972] For sequencing, protein nanopores were prepared by inserting α-hemolysin into the DPhPE / hexadecane bilayer member of buffer B1, which contained 2 M NH4Cl and 100 mM HEPES at pH 7.4. Cis-wells were perfused with buffer B2, which contained 0.4 M NH4Cl, 0.6 M GuCl, and 100 mM HEPES at pH 7.4. Xpandomer samples were heated to 70°C for 2 minutes, completely cooled, and then 2 µL of sample was added to the cis-wells. A voltage pulse of 90 mV / 390 mV / 10 µs was then applied, and data were acquired using LabVIEW software.

[0973] Sequence data are analyzed by displaying histograms of sequence read groups from a single SBX reaction. The analysis software aligns each sequence read with the template sequence and trims the sequence range at the ends of reads that are misaligned with the correct template sequence.

[0974] In one embodiment, this disclosure provides a method for improving the accuracy of enhanced XNTP polymerization, wherein the method includes adding a PEM as disclosed herein to a DNA polymerization reaction as described above.

[0975] In some embodiments, this disclosure provides a kit that can be used in the methods described herein. The kit will comprise at least one compound of this disclosure, and one or more of the following: a) a molecular crowding agent, b) an aqueous buffer, c) a protein such as a polymerase, d) a polynucleotide that can be used as, for example, a primer and / or a polynucleotide that can be used as, for example, a template.

[0976] For example, in some embodiments, this disclosure provides a kit for sequencing nucleic acid templates. The kit comprises a mixture of at least one compound of this disclosure and a nucleotide analog. Compared to the same polymerization reaction without at least one compound of this disclosure, the compounds of this disclosure can be used to increase the quantity and accuracy of nucleotide analogs incorporated into daughter chains during template-dependent polymerization. Optionally, the mixture of nucleotide analogs comprises nucleoside triaminophosphates, wherein each nucleoside triaminophosphate comprises a nucleobase selected from the group consisting of adenine, guanine, thymine, and cytosine, and a polymer chain portion, wherein a first end of the polymer chain portion is attached to a nucleobase, and a second end of the polymer chain portion is attached to an α-phosphate of the nucleoside triaminophosphate to provide nucleotide analog expansion through cleavage of the aminophosphate bond. Optionally, the mixture of nucleotide analogs includes nucleotide analogs containing detectable markers, wherein the detectable markers are optically detectable markers selected from the group consisting of luminescent, chemiluminescent, fluorescent, fluorescent, chromogenic, or chromogenic markers. Optionally, the kit includes an aqueous buffer containing one or more components selected from MnCl2, buffer solutions, salts, sugars, single-chain binding proteins (SSBs), imidazoles, pyrazoles, triazoles, betaine, molecular crowding agents, dimethyl sulfoxide (DMSO), alkyl glycols, glycerols, N-methyl-2-pyrrolidone (NMP), acetamide, butylated hydroxyanisole (BHA), polyphosphates, and / or ureas. Optionally, the kit includes a single-chain binding protein. Optionally, the kit includes urea. Optionally, the kit includes two or more of these components, for example, three or four, or all of the specified components.

[0977] The compounds can be prepared by methods known to those skilled in the art, which can be identified from various reference books and databases. Suitable reference books and papers that detail the synthesis of reactants that can be used to prepare the compounds of this disclosure or provide reference to articles describing the preparation include, for example, "Synthetic Organic Chemistry," John Wiley & Sons, Inc., New York; SR Sandler et al., "Organic Functional Group Preparations," 2nd ed., Academic Press, New York, 1983; HO House, "Modern Synthetic Reactions," 2nd ed., WA Benjamin, Inc., Menlo Park, Calif 1972; TL Gilchrist, "Heterocyclic Chemistry," 2nd ed., John Wiley & Sons, New York, 1992; J. March, "Advanced Organic Chemistry: Reactions, Mechanisms and Structure," 4th ed., Wiley-Interscience, New York, 1992. Further details of the synthesis of reactants used to prepare the compounds of this disclosure are provided in the references to other suitable books and papers, or to articles describing the preparation, including, for example, Fuhrhop, J. and Penzlin G., “Organic Synthesis: Concepts, Methods, Starting Materials,” 2nd ed., Revised and Expanded (1994), John Wiley & Sons, ISBN: 3-527-29074-5; Hoffman, RV., “Organic Chemistry, An Intermediate Text” (1996), Oxford University Press, ISBN 0-19-509618-5; Larock, RC., “Comprehensive Organic Transformations: A Guide to Functional Group Preparations,” 2nd ed. (1999), Wiley-VCH, ISBN: 0-471-19031-4; March, J."Advanced Organic Chemistry: Reactions, Mechanisms, and Structure" 4th edition (1992) John Wiley & Sons, ISBN: 0-471-60180-2; Otera, J. (editor) "Modern Carbonyl Chemistry" (2000) Wiley-VCH, ISBN: 3-527-29871-1; Patai, S. "Patai's 1992 Guide to the Chemistry of Functional Groups" (1992) Interscience ISBN: 0-471-93022-9; Quin, LD et al. "AGuide to Organophosphorus Chemistry" (2000) Wiley-Interscience, ISBN: 0-471-31824-8; Solomons, TWG "Organic Chemistry" No. 7 Edition (2000) John Wiley &Sons, ISBN: 0-471-19095-0; Stowell, JC, "Intermediate Organic Chemistry" 2nd edition (1993) Wiley-Interscience, ISBN: 0-471-57456-2; "Industrial OrganicChemicals: Starting Materials and Intermediates: An Ullmann's Encyclopedia" (1999) John Wiley & Sons, ISBN: 3-527-29645-X, in 8 volumes; “OrganicReactions” (1942-2000) John Wiley & Sons, in over 55 volumes; and “Chemistry of Functional Groups” John Wiley & Sons, in 73 volumes. .

[0978] Example

[0979] The compounds shown in Table 1 are synthesized, such as those according to the general examples disclosed herein.

[0980] Materials and Methods. 4-Azide-salicylic acid and 2,6-dibromo-4-pyridinecarboxylic acid were purchased from Toronto Research Chemicals, Inc. (Toronto, Ontario, Canada). 4-Azide-2-(trifluoromethyl)benzoic acid, 3-amino-5-hydroxybenzoic acid, and 4-aminoisophthalic acid were purchased from Matrix Scientific (Columbia, South Carolina, USA). 3-Amino-6-(trifluoromethyl)benzoate, 1-(4-aminophenyl)-2,2,2-trifluoroethyl-1-one, glycylglycine methyl hydrochloride, 3,3,3-trifluoropropyl-1-amine, and diethyl 3-aminopropyl-1-phosphonate were purchased from Enamine LLC (Monmouth, Junction, New Jersey, USA). Tris[(1-benzyl-1H-1,2,3-triazol-4-yl)methyl]amine (TBTA), O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethylurea hexafluorophosphate (HATU), 1,3-diethynylbenzene, 2,6-diethynylpyridine, 3,5-diethynylpyridine, 3,6-diethynylcarbazole, 4-azidobenzoic acid, cyclopropylamine, 6-amino-2-naphthoic acid, 4-amino-o- Phthalic acid, 4-amino-3-hydroxybenzoic acid, 4-amino-2-methylbenzoic acid, 4-amino-2,3,5,6-tetrafluorobenzoic acid, 4-amino-2-nitrobenzoic acid, n-(2-aminoethyl)acetamide, 4-(aminophenyl)phosphonic acid, 2,6-dichloro-9H-purine, dimethyl aspartate hydrochloride, 4-amino-2-(trifluoromethyl)benzonitrile, 3,5-diaminobenzoic acid, and 2,5-dibromofuran were purchased from TCI America (Portland, Oregon, USA). 4-Methoxy-2,6-dibromopyridine, 4-nitro-2,6-dibromopyridine, 2,6-dibromo-4-pyridinecarboxylic acid, and glycine methyl ester hydrochloride were purchased from Chem-Impex International, Inc. (Wooddale, Illinois).Tetra(triphenylphosphine)palladium(0), trimethylsilylacetylene, DMSO, DMF, MeOH, EtOAc, sodium ascorbate, copper sulfate, diisopropylamine, EDTA, morpholine, diethylamine, ammonium hydroxide, ethylamine, ethanol, azacyclobutane, n-ethyl-n-ethylamine, methyl 2,6-dichloropyridine-4-carboxylate, ethyl 2,6-dibromopyridine-4-carboxylate, 4-methyl-2,6-dichloropyridine, 2-chloro-4-cyanopyridine, 4-amino-2-(trifluoromethyl)benzoic acid 2-Bromo-4-cyanopyridine, methyl azidoacetate, 4-azidoaniline hydrochloride, 4-methoxyphenyl azido-4-amino-2-fluorobenzoic acid, n-butylamine, 3-amino-5-(trifluoromethyl)benzoic acid, 3-(4-aminophenyl)propionic acid, 4-(4-aminophenyl)butyric acid, 4-amino-2-methoxybenzoic acid, 2-amino-3-(trifluoromethyl)benzoic acid, aminoethanol, 1,4-diaminobutane, and 2,5-dibromothiophene were purchased from Sigma-Aldrich Corp. (St. Louis, Missouri, USA). Solvents for TLC and rapid chromatography were purchased from Sigma-Aldrich or Thermo Fisher Scientific Inc. (Waltham, Massachusetts, USA).

[0981] Rapid chromatography was performed on a Reveleris preparative purification system purchased from Buchi Corp. (Newcastle, Delaware, USA). The system was equipped with a hand-packed column (2.3 cm diameter x 8 cm height) filled with C18 spherical silica gel (Cat. No. 76646-01) from Sorbent Technologies, Inc. (Nockros, Georgia) and sealed with a polypropylene melt. 1 to 1.5 mL of sample was loaded directly onto the top of the column. The mobile phase was water (A) and acetonitrile (B). The gradient was 0% B to 2% B over 2 min, then 2% B to 100% B over 20 min, at a flow rate of 28 mL / min. UV was monitored at 220 nm, 260 nm, and 280 nm. Fractions were collected at a UV threshold of 0.1 AU. Thin-layer chromatography was performed using an aluminum-backed TLC SilicaGel 60 F254 (Cat. No. 1.05534.0001) from EMD Millipore Corp. (Bill Ricard, MD, USA). ESI mass spectrometry was performed by Numega ResonanceLab (San Diego, CA, USA) using a Perkin Elmer PE-SCIEX API-150 mass spectrometer in both positive and negative modes.

[0982] Table 1

[0983]

[0984] Although the compounds shown in Table 1 include a triazole "Z" group (in respect of formula (I)), other heteroaromatic groups may replace the triazole, wherein the other heteroaromatic groups may contain one, two, three, or four heteroatoms, such as one, two, three, or four heteroatoms selected from O, N, and / or S. For example, each triazole may be substituted with imidazole, pyrazole, pyrimidine, pyridazine, pyrrole, thiophene, thiazole, isoxazole, or tetraazole.

[0985] In some embodiments, the compounds shown in Table 1 can be polymerized (e.g., via cationic polymerization, such as by incubating KCl with PEM). In some embodiments, the composition may comprise molecules of two or more of the compounds shown in Table 1.

[0986] Example 1

[0987] Synthesis of 4,4'-(pyridine-2,6-dimethylbis(1H-1,2,3-triazol-4,1-dimethyl))bis(2-hydroxybenzoic acid)

[0988] (1)

[0989] (A) (B)

[0990] Compound 1 was prepared by mixing 4-azidosalicylic acid B (1.79 mg, 10 µmol) and 2,6-diethynylpyridine A (0.67 mg, 5 µmol) in DMSO (150 µL). This solution was then mixed with a solution of TBTA (5.1 mg, 0.96 µmol) and sodium ascorbate (6.4 mg, 32 µmol) in DMSO (95 µL). A click reaction was initiated by adding 20 M copper sulfate (5 µL) under stirring. The extent of the reaction was analyzed by TLC (94:5:1 ethyl acetate:methanol:acetic acid), and the reaction was completed within 5 minutes based on the consumption of the azide and alkyne. The reaction mixture was brought to a volume of 1 mL with DMSO and 0.5 M EDTA (100 µL). The solids were separated and dissolved in a separate container of DMSO. The combined DMSO solutions were purified by rapid chromatography as described in Materials and Methods above. The product forms a glassy solid during rotary evaporation, with a yield of 50% to 75%. 1 H NMR (300 MHz, DMSO-d6) δppm 3.29 (2H, br.S., ( O (18) H and O (33)H)), 7.17 – 7.29 (4H,m, ( C (7)H, C (11)H, C (28)H, C (32)H)) 7.85 (2H, d,J=8.11 ( C (10) H and C (31)H), 8.06 (3H, s, ( C (15)H, C (16)H, C (17)H)) 9.36 (2H,s, ( C (5) H and C (25)H)).

[0991] Example 2

[0992] Synthesis of 4,4'-(pyridine-3,5-dimethylbis(1H-1,2,3-triazol-4,1-dimethyl))bis(2-hydroxybenzoic acid)

[0993] (2)

[0994] (C)

[0995] Compound 2 was prepared using 4-azidosalicylic acid B and 3,5-diethynylpyridine C, following the method of Example 1. 1 H NMR (300 MHz, DMSO-d6) δ ppm 3.29 (2H, br.S., ( O (18) H and O (33)H))7.17 – 7.29 (4H, m, ( C (7)H, C (11)H, C (28)H, C (31)H)) 7.85 (2H, d, J=8.11 Hz,( C (10) H and C (31)H)) 8.06 (3H, s, ( C (13)H, C (15)H, C (17)H)) 9.36 (2H,s, ( C (5) H and C (25)H)).

[0996] Example 3

[0997] Synthesis of 4,4'-(1,3-phenylenebis(1H-1,2,3-triazol-4,1-diyl))bis(2-hydroxybenzoic acid)

[0998] (3)

[0999] (D)

[1000] Compound 3 was prepared using 4-azidosalicylic acid B and 1,3-diethynylbenzene D, according to the method of Example 1. 1 H NMR (300 MHz, DMSO-d6) δ ppm 3.29 (2H, br.S., ( O (18) H and O (33)H)) 7.16 –7.27 (4H, m, ( C (7)H, C (11)H, C (28)H, C (32)H)) 7.57 – 7.67 (1H,m, ( C (16)H))7.80 – 7.88 (2H,m, ( C (10) H and C(31)H)) 7.94 (2H,d, J=7.63 Hz, ( C (15) H and C (17)H) 8.57(1H,s, ( C (13)H)) 9.35 (2H,s, ( C (5) H and C (25)H).

[1001] Example 4

[1002] Synthesis of 4,4'-((9H-carbazole-3,6-diyl)bis(1H-1,2,3-triazole-4,1-diyl))bis(2-hydroxybenzoic acid)

[1003] (4)

[1004] (E)

[1005] Compound 4 was prepared using 4-azidosalicylic acid B and 3,6-diethynylcarbazole E, according to the method in Example 1. 1 H NMR (300 MHz, DMSO-d6) δ ppm 3.29 (2H, br.S., ( O (25) H and O (40)H))7.16 – 7.27 (4H, m, ( C (7)H, C (11)H, C (35)H, C (39)H)) 7.61 (2H,d, J=8.34 Hz,( C (10) H and C (38)H)) 7.84 (2H,d, J=7.87 Hz, ( C (22)H and C (24)H)) 8.02 (2H,d, J=8.34 Hz, ( C (21) H and C (23)H)) 8.79 (2H,s, ( C (13) H and C (19)H)) 9.28 (2H,s, ( C (5) H and C (32)H)) 11.53 (1H,s, ( N (16)H).

[1006] Example 5

[1007] Synthesis of 4,4'-((9H-carbazole-3,6-diyl)bis(1H-1,2,3-triazole-4,1-diyl))diphenylamine

[1008] (5)

[1009] (F)

[1010] Compound 5 was prepared using 4-azidoaniline hydrochloride F and 3,6-diethynylcarbazole E, according to the method of Example 1.

[1011] Example 6

[1012] Synthesis of 4,4'-((9H-carbazole-3,6-diyl)bis(1H-1,2,3-triazol-4,1-diyl))dibenzoic acid

[1013] (6)

[1014] (G)

[1015] Compound 6 was prepared using 4-azidobenzoic acid G and 3,6-diethynylcarbazole E, according to the method of Example 1.

[1016] Example 7

[1017] Synthesis of 3,6-bis(1-(4-methoxyphenyl)-1H-1,2,3-triazol-4-yl)-9H-carbazole

[1018] (7)

[1019] (H)

[1020] Compound 7 was prepared using 4-azidoanisole H and 3,6-diethynylcarbazole E, according to the method of Example 1.

[1021] Example 8

[1022] Synthesis of dimethyl 2,2'-((9H-carbazole-3,6-diyl)bis(1H-1,2,3-triazole-4,1-diyl))diacetate

[1023] (8)

[1024] (iota)

[1025] Compound 8 was prepared using methyl azidoacetate iota and 3,6-diethynylcarbazole E, according to the method of Example 1.

[1026] Example 9

[1027] Synthesis of 4,4'-((4-methoxypyridin-2,6-diyl)bis(1H-1,2,3-triazol-4,1-diyl))bis(2-hydroxybenzoic acid)

[1028] (9)

[1029] (J)

[1030] The preparation of compound 9 began with the synthesis of 4-methoxy-2,6-diethynylpyridine J from 4-methoxy-2,6-dibromopyridine and trimethylsilylacetylene using the conditions described by Sonogashira (Organomet. Chem., 653:46–49 (2002). doi:10.1016 / s0022-328x(02)01158-0). Compound 9 was synthesized by clicking 4-azidosalicylic acid B and compound J according to the method of Example 1.

[1031] Example 10

[1032] Synthesis of 4,4'-((4-carboxypyridine-2,6-diyl)bis(1H-1,2,3-triazol-4,1-diyl))bis(2-hydroxybenzoic acid)

[1033] (10)

[1034] (K)

[1035] The preparation of compound 10 began with the synthesis of 4-cyano-2,6-diethynylpyridine K from 4-cyano-2,6-dibromopyridine and trimethylsilylacetylene using the conditions described by Sonogashira (Organomet. Chem., 653:46–49 (2002). doi:10.1016 / s0022-328x(02)01158-0). Compound 10 was ...

Claims

1. A compound having formula (I): (I)， Or its solvates, hydrates, tautomers, chelates or salts, in a is 0 or an integer in the range of 1 to 4; a' is 0 or an integer in the range of 1 to 4; m is 1, 2, or 3; m' is 1, 2, or 3; n is 0, 1, or 2; p is 0, 1, or 2; Z is a 5- or 6-membered heteroaromatic ring containing one, two, three or four heteroatoms selected from O, N or S; Y is independently selected from Ar2, –C(O)–Ar2, and –(CH2) each time it appears. a Ar2, –(CH2)3PO(OEt)2 or –CH2CO2Me; L is a linking group; M is independently selected from hydrogen, halogens, and C1-C4 alkyl groups each time it appears; Ar1, each time it appears, is independently selected from optionally substituted phenyl, pyridine, bipyridine, tripyridine, pyrazine, pyridazine, furan, dibenzofuran, thiophene, pyrrole, selenophene, naphthalene, fluorene, phenanthrene, cyclophosphine, phthalazine, quinazoline, quinoxaline, naphthidine, phenanthrene-rholine, purine, and carbazole. The substituents of Ar1 are independently selected each time they appear from halogens, –OH, –CN, –NO2, C1–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, C1–C6 cycloalkyl, –OR 0 –CONH2, –C(O)NR 1 R 1' –C(O)(CH2) a NR 1 R 1' –NR 1 R 1' –NR 1 C(O)R 3 –C(O)SR 3 –COR 3 –CO(CH2) a OC(O)R 3 –OC(O)R 3 –C(O)OR 3 –C–O–R 3 , thiols, -R 4 –H、–SOR 1 –S(O)2R 1 –S(O)2NR 1 R 1' –CH2–NR 1 S(O)2R 3 –NR 1 S(O)2R 3 and –C(CH3)=N–(phenyl)–O–CH2–C–CH; R 0 Each time it appears, it is independently selected from C1–C6 alkyl, C1–C6 haloalkyl, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl. R 1 and R 1' Each time it appears, it is independently selected from H, hydroxyl, C1–C6 alkyl, C1–C6 haloalkyl, C1–C 20 Heteroalkyl, C1–C 10 Heteroalkyl –NH2, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted arylalkyl, and substituted or unsubstituted heteroaryl, benzyl, –C(=NH)NH2, –CH2CO2R 0 –C(H)(CH) a (COOH))2, –CH2C(O)NHCH2CO2H, –CH2(CH2) a OH、–CH2CH2NHC(O)R 3 –(CH2) a C(O)NH(CH2) a C(O)OR 3 –(CH2) a –CN, spermine, –(CH2) a –NH2、–C(O)R 3 –(CH2) a C(O)OR 3 –(CH2) a OR 3 –C(H)((CH2) a OH) a' –CHCH(OH)(CH2) a OH, –(CH2) a OH, C1–C6–C(O)OH, –(CH2) that can be substituted or not substituted a - Heterocyclic rings , ;and Where R 1 and R 1' Together they form substituted or unsubstituted heterocycles, including but not limited to aza-butanes, pyrrolidines, piperidines, piperazines, morpholines, etc. or ; R 2 Each time it appears, it is independently selected from C2–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, and substituted or unsubstituted haloalkoxy. R 3 Each time it appears, it is independently selected from H, C1–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, –C1–C6–OH, –C1–C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine; R 4 Each time it appears, it is independently selected from alkylene groups interrupted by one or more heteroatoms, wherein the heteroatoms are O, S, NH or a combination thereof; Ar2 is independently selected each time it appears from: (i) substituted or unsubstituted 5- and 6-membered monocyclic aromatic or heterocyclic rings; (ii) substituted or unsubstituted 9- and 10-membered fused bicyclic rings comprising two monocyclic rings, wherein at least one of the two monocyclic rings is an aromatic or heterocyclic ring; and (iii) substituted or unsubstituted 13- and 14-membered fused tricyclic rings comprising three monocyclic rings, wherein at least one of the three monocyclic rings is an aromatic or heterocyclic ring. Each Ar2 is independently transmitted via G 1 G 2 G 3 G 4 and G 5 Replace, of which: When Ar2 is monosubstituted, G 1 Each time it appears, it is independently selected from oxo, C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–C(R) 1 (R) 1' (R) 3 –NH2, –NO2, –SO3R 3 SO3O - 、–SO3N(H)(R 1 ), –E–C(O)R 3 , –E–CO2H, –B(OH)2, –C(O)NR 1 R 1' –E–PO(OR) 1 )2, and via G 2 G 3 G 4 and G 5 Substituted aryl; and G 2 G 3 G 4 and G 5 Each time it appears, it is independently selected from groups that are absent or selected from groups containing the following: halogen, –CN, –NO2, –SO3R. 3 C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–(CH2) a C(O)R 3 , –E–CO2H, –E–CHO, –E–C(O)R 3 , –E–C(O)NH(OH), –E–C(O)NHR 1 –E–C(O)N(H)C(H)(R 1 (R) 1' ), –E–C(O)N(R 1 (CH2) a P(O)(O)OH)2、–E–C(O)N(R 1 (CH2) a SO3H), –E–C(O)NR 1 R 1' –E–NR 1 R 1' –E–OR 2 –C(O)–heterocyclic alkyl (where the heterocyclic alkyl is substituted or unsubstituted), –C(O)–N(H)–CH(COOH)((CH2) a – heteroaryl) (wherein the heteroaryl group is substituted or unsubstituted), wherein E is independently selected from direct bond and C1–C6 alkylene group each time it appears; Z is not a triazole.

2. The compound according to claim 1, wherein Z comprises two heteroatoms.

3. The compound according to claim 1, wherein Z is a diazole.

4. The compound of claim 3, wherein the diazole is selected from the group consisting of imidazole, pyrazole, pyrimidine, pyridazine, and pyrazine.

5. The compound according to claim 1, wherein Z comprises three heteroatoms.

6. The compound according to claim 1, wherein Z comprises four heteroatoms.

7. The compound according to claim 1, wherein Z is selected from the group consisting of pyrrole, thiophene, thiazole, isoxazole and tetrazolium.

8. The compound according to any one of claims 1 to 7, wherein Ar1 is phenyl.

9. The compound according to any one of claims 1 to 7, wherein Ar1 is pyridine.

10. The compound according to any one of claims 1 to 7, wherein Ar1 is furan.

11. The compound according to any one of claims 1 to 7, wherein Ar1 is carbazole.

12. The compound according to any one of claims 1 to 7, wherein Ar1 is a naphthyl group.

13. The compound according to any one of claims 1 to 7, wherein Ar1 is a pyridazine.

14. The compound according to any one of claims 1 to 7, wherein Ar1 is thiophene.

15. The compound according to any one of claims 1 to 7, wherein Ar1 is pyrrole.

16. The compound according to any one of claims 1 to 7, wherein Ar1 is dibenzofuran.

17. The compound according to any one of claims 1 to 7, wherein Ar1 is naphthidine.

18. The compound according to any one of claims 1 to 17, wherein each substituent of Ar1 is independently selected from –C(O)–CH3, –C(O)–NH2, –C(O)O–CH2CH3, –CF3, –C(O)O–tert-butyl, –C(O)–N(CH3)2, –C(O)–N(H)CH3, –C(O)–N(H)CH2CH3, –C(O)–N(H) (cycloalkyl), –C(O)–N(H) (CH2)4, –C(O)–N(CH2CH3)2, –C(O)–N(H) (tert-butyl), –C(O)–N(H) (C6–cycloalkyl), –C(O)–N(H) (CH2)3, –C(O)–N(H) (C5–cycloalkyl), –C(O)–N(H) (C5–cycloalkyl), –C(O)–NH2, ... –N(H)(C4–cycloalkyl), –C(O)–N(H)(phenyl), –C(O)–N(H)(CH2CH2)N(H)C(O)–CH3, –C(O)–N(H)C(NH)(NH2), –C(O)–N(CH2CH2CH3)2, –C(O)–N–(C(H)(CH3)(CH2CH3))2, –C(O)–N(CH2CH2CH2CH3)2, –C(O)–N(H)(benzyl), –C(O)–N(H)–CH2CH2OH, –C(O)–S–CH2CH3, –C(O)N(CH2)2, –N(H)S(O)(O)–phenyl , –C(O)N(H)CH2CH2CF3, –C(O)N(H)CH3, –C(O)OCH2CH3, –C(O)–N(H)–CH2CH2O–CH2CH2O–CH3, –C(O)–N(H)–(CH2CH2O)8–CH3, –C(O)N(H)–CH2 –CH2–CH2–N(H)–CH2–CH2–CH2–N(H)–CH2–CH2–CH2–NH2, –C(O)N(H)–CH2–CH2–NH2, –C(O)N(H)C(O)CH3, –C(O)N(H)–CH2–CH3, –S(O)(O)N(H) –CH2–CH3, –S(O)(O)N–(CH2–CH3)2, –S(O)(O)N(H)–(C3–cycloalkyl), –S(O)(O)N(H)–(CH2CH2)OH, –S(O)(O)OH, –C(O)N(H)C(H)(CH2OH)2, –C(O)N( CH2CH2OH)2, –C(O)N(H)CH2CH2=CH2, –C(O)CH2N(CH3)(CH2)(COOH), –S(O)(O)(CH2CH3), –S(O)(O)N(H)CH2–CH(OH)(CH2OH), –S(O)(O)–NH2 and –S(O)(O)N(H)CH2–C(O)(O–tert-butyl).

19. The compound according to any one of claims 1 to 17, wherein Ar1 is via at least one –SO2–N(H)(R 1 ) group substitution.

20. The compound according to claim 19, wherein R 1 It is a C1–C6 alkyl group.

21. The compound according to claim 20, wherein the C1-C6 alkyl group is methyl or ethyl.

22. The compound according to any one of claims 1 to 21, wherein Ar2 is selected from the group consisting of benzene, pyridine, pyridazine, pyrimidine, and pyrazine.

23. The compound according to any one of claims 1 to 22, wherein each Ar2 is partially substituted by one or more of the following groups: –NH2, –C(O)OH, –OH, –OCH3, –C(O)NHOH, –C(O)NH2, –(CH2)3C(O)OH, –CF3, –Cl, –P(O)(OH)2, –F, –C(O)N(H)CH2CH3, –C(O)N(H)CH2C(O)OCH3, –C(O)N(H)CH2CF2CF2CF3 -C(O)N(H)CH2C(O)OH, -C(O)N(H)C(H)(C(O)OOH)CH2C(O)OH, -C(O)N(H)CH2C(O)N(H)CH2C(O)OH, -SO3H, -NO2, -C(O)N(H)CH2PO3H2, -C(O)N(H)CH23O3H, -B(OH)2, -C(OH)(CF3)(CF3), -C(O)-5-membered heterocyclic alkyl-C(O)OH, -C(O)N(H)C(H)(C(O)OH)CH2-imidazole, -S(O)(O)N(H)-CH2COOH, -C(O)N(H)CH(C(O)OH)(CH2CH2C(O)OH), -C(O)N(H)CH2CH2SO3H, -SO2O – –C(O)N(H)–imidazole–C(O)OH and –C(O)N(CH3)2.

24. The compound according to any one of claims 1 to 22, wherein each Ar2 is derived from at least one –SO3H moiety or at least one –SO3 - Partial replacement.

25. The compound according to any one of claims 1 to 22, wherein each Ar2 is derived from at least two –SO3H moieties or at least two –SO3 moieties. - Partial replacement.

26. The compound according to any one of claims 1 to 22, wherein each Ar2 is substituted with at least two moieties, wherein the first moieties of said at least two moieties comprise a –SO3H moiety or a –SO3 moiety. - One of the parts.

27. The compound of claim 26, wherein the second portion of the at least two portions is selected from the group consisting of: –NO2, –CF3, –F, –Cl, –I, methyl, ethyl, propyl, and butyl.

28. The compound according to any one of claims 1 to 21, wherein the compound has any one of formula (IA) or (IB): (IA) or (IB).

29. The compound according to any one of claims 1 to 21, wherein the compound has the following formula: 。 30. The compound according to any one of claims 1 to 21, wherein the compound has any one of the following formulas: , , , , , , , or .

31. The compound according to any one of claims 1 to 21, wherein the compound has any one of the following formulas: , , , , , , , , , , , , , , , , , , , , , , , , , or .

32. A compound having either formula (IA) or (IB): (IA) or (IB), Or its solvates, hydrates, tautomers, chelates or salts, in a is 0 or an integer in the range of 1 to 4; a' is 0 or an integer in the range of 1 to 4; m is 1, 2, or 3; m' is 1, 2, or 3; n is 0, 1, or 2; p is 0, 1, or 2; Z is a 5- or 6-membered heteroaromatic ring containing one, two, three or four heteroatoms selected from O, N or S; Y is independently selected from Ar2, –C(O)–Ar2, and –(CH2) each time it appears. a Ar2, –(CH2)3PO(OEt)2 or –CH2CO2Me; M is independently selected from hydrogen, halogens, and C1-C4 alkyl groups each time it appears; Ar1, each time it appears, is independently selected from optionally substituted phenyl, pyridine, bipyridine, tripyridine, pyrazine, pyridazine, furan, dibenzofuran, thiophene, pyrrole, selenophene, naphthalene, fluorene, phenanthrene, cyclophosphine, phthalazine, quinazoline, quinoxaline, naphthidine, phenanthrene-rholine, purine, and carbazole. Ar1 is converted to –OH and –C(O)NR 1 R 1' –C(O)(CH2) a NR 1 R 1' –CO(CH2) a OC(O)R 3 –OC(O)R 3 –S(O)2R 1 –S(O)2NR 1 R 1' –CH2–NR 1 S(O)2R 3 Or –NR 1 S(O)2R 3 Replace at least one of them; R 0 Each time it appears, it is independently selected from C1–C6 alkyl, C1–C6 haloalkyl, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl. R 1 and R 1' Each time it appears, it is independently selected from H, hydroxyl, C1–C6 alkyl, C1–C6 haloalkyl, C1–C 20 Heteroalkyl, C1–C 10 Heteroalkyl –NH2, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted arylalkyl, and substituted or unsubstituted heteroaryl, benzyl, –C(=NH)NH2, –CH2CO2R 0 –C(H)(CH) a (COOH))2, –CH2C(O)NHCH2CO2H, –CH2(CH2) a OH、–CH2CH2NHC(O)R 3 –(CH2) a C(O)NH(CH2) a C(O)OR 3 –(CH2) a –CN, spermine, –(CH2) a –NH2、–C(O)R 3 –(CH2) a C(O)OR 3 –(CH2) a OR 3 –C(H)((CH2) a OH) a' –CHCH(OH)(CH2) a OH, –(CH2) a OH, C1–C6–C(O)OH, –(CH2) that can be substituted or not substituted a - Heterocyclic rings , ;and Where R 1 and R 1' Together they form substituted or unsubstituted heterocycles, including but not limited to aza-butanes, pyrrolidines, piperidines, piperazines, morpholines, etc. or ; R 2 Each time it appears, it is independently selected from C2–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, and substituted or unsubstituted haloalkoxy. R 3 Each time it appears, it is independently selected from H, C1–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, –C1–C6–OH, –C1–C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine; Ar2 is independently selected each time it appears from: (i) substituted or unsubstituted 5- and 6-membered monocyclic aromatic or heterocyclic rings; (ii) substituted or unsubstituted 9- and 10-membered fused bicyclic rings comprising two monocyclic rings, wherein at least one of the two monocyclic rings is an aromatic or heterocyclic ring; and (iii) substituted or unsubstituted 13- and 14-membered fused tricyclic rings comprising three monocyclic rings, wherein at least one of the three monocyclic rings is an aromatic or heterocyclic ring. Each Ar2 is independently transmitted via G 1 G 2 G 3 G 4 and G 5 Replace, of which: When Ar2 is monosubstituted, G 1 Each time it appears, it is independently selected from oxo, C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–C(R) 1 (R) 1' (R) 3 –NH2, –NO2, –SO3R 3 SO3O - 、–SO3N(H)(R 1 ), –E–C(O)R 3 , –E–CO2H, –B(OH)2, –C(O)NR 1 R 1' –E–PO(OR) 1 )2, and via G 2 G 3 G 4 and G 5 Substituted aryl groups; G 2 G 3 G 4 and G 5 Each time it appears, it is independently selected from groups that are absent or selected from groups containing the following: halogen, –CN, –NO2, –SO3R. 3 C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–(CH2) a C(O)R 3 , –E–CO2H, –E–CHO, –E–C(O)R 3 , –E–C(O)NH(OH), –E–C(O)NHR 1 –E–C(O)N(H)C(H)(R 1 (R) 1' ), –E–C(O)N(R 1 (CH2) a P(O)(O)OH)2、–E–C(O)N(R 1 (CH2) a SO3H), –E–C(O)NR 1 R 1' –E–NR 1 R 1' –E–OR 2 –C(O)–heterocyclic alkyl (where the heterocyclic alkyl is substituted or unsubstituted), –C(O)–N(H)–CH(COOH)((CH2) a – heteroaryl) (wherein the heteroaryl group is substituted or unsubstituted), wherein E is independently selected from direct bond and C1–C6 alkylene group each time it appears.

33. The compound according to claim 32, wherein at least one Ar1 substituent is selected from the group consisting of: –OH, –C(O)NR 1 R 1' –C(O)(CH2) a NR 1 R 1' –CO(CH2) a OC(O)R 3 –OC(O)R 3 –S(O)2R 1 and –S(O)2NR 1 R 1' .

34. The compound according to claim 32, wherein Ar1 is via at least one –C(O)(CH2). a NR 1 R 1' replace.

35. The compound according to claim 34, wherein R 1 For H.

36. The compound according to claim 35, wherein R 1' It is a C1–C6 alkyl group.

37. The compound of claim 36, wherein the C1–C6 alkyl group is branched.

38. The compound of claim 36, wherein the C1–C6 alkyl group comprises at least one substituent.

39. The compound according to claim 37, wherein the at least one substituent is a hydroxyl group.

40. The compound according to claim 34, wherein R 1 and R 1' Together they form cycloalkyl groups or heterocycloalkyl groups.

41. The compound according to claim 32, wherein Ar1 is via at least one –S(O)2NR 1 R 1' replace.

42. The compound according to claim 41, wherein R 1 For H.

43. The compound according to claim 42, wherein R 1' It is a C1–C6 alkyl group.

44. The compound according to claim 43, wherein the C1–C6 alkyl group is branched.

45. The compound of claim 43, wherein the C1–C6 alkyl group comprises at least one substituent.

46. ​​The compound according to claim 45, wherein the at least one substituent is a hydroxyl group.

47. The compound according to claim 41, wherein R 1 and R 1' Together they form cycloalkyl groups or heterocycloalkyl groups.

48. The compound according to any one of claims 32 to 47, wherein each Ar2 is derived from at least one –SO3H moiety or at least one –SO3 - Partial replacement.

49. The compound according to any one of claims 32 to 47, wherein each Ar2 is derived from at least two –SO3H moieties or at least two –SO3 - Partial replacement.

50. The compound according to any one of claims 32 to 47, wherein each Ar2 is substituted with at least two moieties, wherein the first moieties of said at least two moieties comprise a –SO3H moiety or a –SO3 moiety. - One of the parts.

51. The compound of claim 50, wherein the second portion of the at least two portions is selected from the group consisting of: –NO2, –CF3, –F, –Cl, –I, methyl, ethyl, propyl, and butyl.

52. The compound according to any one of claims 32 to 47, wherein the compound has any one of the following formulas: , , , , , , , or .

53. The compound according to any one of claims 32 to 47, wherein the compound has any one of the following formulas: , , , , , , , , , , , , , , , , , , , , , , , , , or .

54. The compound according to claim 32, wherein the compound has the following formula: 。 55. The compound according to claim 32, wherein the compound has the following formula: 。 56. The compound according to any one of claims 32 to 55, wherein Z is a diazole.

57. The compound according to any one of claims 32 to 55, wherein Z is selected from the group consisting of imidazole, pyrazole, pyrimidine, pyridazine, and pyrazine.

58. The compound according to any one of claims 32 to 55, wherein Z is not a triazole.

59. The compound according to any one of claims 32 to 47, wherein the compound has any one of the following formulas: , , , , , and .

60. A compound having any of the following formulas: , , , , , , , , , , , , , , , , , , , and , Or its solvates, hydrates, tautomers, chelates or salts, in Z is a 5- or 6-membered heteroaromatic ring containing one, two, three or four heteroatoms selected from O, N or S; M is independently selected from hydrogen, halogens, and C1–C4 alkyl groups each time it appears; Y is independently selected from Ar2, –C(O)–Ar2, and –(CH2) each time it appears. a Ar2, –(CH2)3PO(OEt)2 or –CH2CO2Me; Ar1 is independently selected each time it appears from optionally substituted phenyl, pyridine, bipyridine, tripyridine, pyrazine, pyridazine, furan, dibenzofuran, thiophene, pyrrole, selenophene, naphthalene, fluorene, phenanthrene, cyclophosphine, phthalazine, quinazoline, quinoxaline, naphthidine, phenanthrene-rholine, purine, and carbazole; Ar2 is independently selected each time it appears from: (i) substituted or unsubstituted 5- and 6-membered monocyclic aromatic or heterocyclic rings; (ii) substituted or unsubstituted 9- and 10-membered fused bicyclic rings comprising two monocyclic rings, wherein at least one of the two monocyclic rings is an aromatic or heterocyclic ring; and (iii) substituted or unsubstituted 13- and 14-membered fused tricyclic rings comprising three monocyclic rings, wherein at least one of the three monocyclic rings is an aromatic or heterocyclic ring. Each Ar2 is independently transmitted via G 1 G 2 G 3 G 4 and G 5 Replace, of which: When Ar2 is monosubstituted, G 1 Each time it appears, it is independently selected from oxo, C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–C(R) 1 (R) 1' (R) 3 –NH2, –NO2, –SO3R 3 SO3O – 、–SO3N(H)(R 1 ), –E–C(O)R 3 , –E–CO2H, –B(OH)2, –C(O)NR 1 R 1' –E–PO(OR) 1 )2, and via G 2 G 3 G 4 and G 5 Substituted aryl groups; G 2 G 3 G 4 and G 5 Each time it appears, it is independently selected from groups that are absent or selected from groups containing the following: halogen, –CN, –NO2, –SO3R. 3 C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–(CH2) a C(O)R 3 , –E–CO2H, –E–CHO, –E–C(O)R 3 , –E–C(O)NH(OH), –E–C(O)NHR 1 –E–C(O)N(H)C(H)(R 1 (R) 1' ), –E–C(O)N(R 1 (CH2) a P(O)(O)OH)2、–E–C(O)N(R 1 (CH2) a SO3H), –E–C(O)NR 1 R 1' –E–NR 1 R 1' –E–OR 2 –C(O)–heterocyclic alkyl (where the heterocyclic alkyl is substituted or unsubstituted), –C(O)–N(H)–CH(COOH)((CH2) a – heteroaryl) (wherein the heteroaryl group is substituted or unsubstituted), wherein E is independently selected from direct bond and C1–C6 alkylene group each time it appears; R 0 Each time it appears, it is independently selected from C1–C6 alkyl, C1–C6 haloalkyl, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl. R 1 and R 1' Each time it appears, it is independently selected from H, hydroxyl, C1–C6 alkyl, C1–C6 haloalkyl, C1–C 20 Heteroalkyl, C1–C 10 Heteroalkyl –NH2, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted arylalkyl, and substituted or unsubstituted heteroaryl, benzyl, –C(=NH)NH2, –CH2CO2R 0 –C(H)(CH) a (COOH))2, –CH2C(O)NHCH2CO2H, –CH2(CH2) a OH、–CH2CH2NHC(O)R 3 –(CH2) a C(O)NH(CH2) a C(O)OR 3 –(CH2) a –CN, spermine, –(CH2) a –NH2、–C(O)R 3 –(CH2) a C(O)OR 3 –(CH2) a OR 3 –C(H)((CH2) a OH) a' –CHCH(OH)(CH2) a OH, –(CH2) a OH, C1–C6–C(O)OH, –(CH2) that can be substituted or not substituted a - Heterocyclic rings , ;and Where R 1 and R 1' Together they form substituted or unsubstituted heterocycles, including but not limited to aza-butanes, pyrrolidines, piperidines, piperazines, morpholines, etc. or ; R 2 Each time it appears, it is independently selected from C2–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, and substituted or unsubstituted haloalkoxy; and R 3 Each time it appears, it is independently selected from H, C1–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, –C1–C6–OH, –C1–C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine.

61. The compound of claim 60, wherein Z comprises two heteroatoms.

62. The compound according to claim 60, wherein Z is a diazole.

63. The compound of claim 62, wherein the diazole is selected from the group consisting of imidazole, pyrazole, pyrimidine, pyridazine, and pyrazine.

64. The compound of claim 60, wherein Z comprises three heteroatoms.

65. The compound of claim 60, wherein Z comprises four heteroatoms.

66. The compound of claim 60, wherein Z is selected from the group consisting of pyrrole, thiophene, thiazole, isoxazole, and tetrazolium.

67. The compound according to claim 60, wherein Z is a triazole.

68. The compound of claim 60, wherein Z is not a triazole.

69. The compound according to any one of claims 60 to 68, wherein Ar1 is phenyl.

70. The compound according to any one of claims 60 to 68, wherein Ar1 is pyridine.

71. The compound according to any one of claims 60 to 68, wherein Ar1 is furan.

72. The compound according to any one of claims 60 to 68, wherein Ar1 is carbazole.

73. The compound according to any one of claims 60 to 68, wherein Ar1 is a naphthyl group.

74. The compound according to any one of claims 60 to 68, wherein Ar1 is a pyridazine.

75. The compound according to any one of claims 60 to 68, wherein Ar1 is thiophene.

76. The compound according to any one of claims 60 to 68, wherein Ar1 is pyrrole.

77. The compound according to any one of claims 60 to 68, wherein Ar1 is a dibenzofuran.

78. The compound according to any one of claims 60 to 68, wherein Ar1 is naphthidine.

79. The compound according to any one of claims 60 to 78, wherein each Ar2 is partially substituted by one or more of the following groups: –NH2, –C(O)OH, –OH, –OCH3, –C(O)NHOH, –C(O)NH2, –(CH2)3C(O)OH, –CF3, –Cl, –P(O)(OH)2, –F, –C(O)N(H)CH2CH3, –C(O)N(H)CH2C(O)OCH3, –C(O)N(H)CH2CF2CF2CF3 -C(O)N(H)CH2C(O)OH, -C(O)N(H)C(H)(C(O)OOH)CH2C(O)OH, -C(O)N(H)CH2C(O)N(H)CH2C(O)OH, -SO3H, -NO2, -C(O)N(H)CH2PO3H2, -C(O)N(H)CH23O3H, -B(OH)2, -C(OH)(CF3)(CF3), -C(O)-5-membered heterocyclic alkyl-C(O)OH, -C(O)N(H)C(H)(C(O)OH)CH2-imidazole, -S(O)(O)N(H)-CH2COOH, -C(O)N(H)CH(C(O)OH)(CH2CH2C(O)OH), -C(O)N(H)CH2CH2SO3H, -SO2O - –C(O)N(H)–imidazole–C(O)OH and –C(O)N(CH3)2.

80. The compound according to any one of claims 60 to 78, wherein each Ar2 is derived from at least one –SO3H moiety or at least one –SO3 - Partial replacement.

81. The compound according to any one of claims 60 to 78, wherein each Ar2 is derived from at least two –SO3H moieties or at least two –SO3 - Partial replacement.

82. The compound according to any one of claims 60 to 78, wherein each Ar2 is substituted with at least two moieties, wherein the first moieties of said at least two moieties comprise a –SO3H moiety or a –SO3 moiety. - One of the parts.

83. The compound of claim 82, wherein the second portion of the at least two portions is selected from the group consisting of: –NO2, –CF3, –F, –Cl, –I, methyl, ethyl, propyl, and butyl.

84. A compound having any of the following formulas: , , , , , , and , Or its solvates, hydrates, tautomers, chelates or salts, in Ar1 is independently selected each time it appears from optionally substituted phenyl, pyridine, bipyridine, tripyridine, pyrazine, pyridazine, furan, dibenzofuran, thiophene, pyrrole, selenophene, naphthalene, fluorene, phenanthrene, cyclophosphine, phthalazine, quinazoline, quinoxaline, naphthidine, phenanthrene-rholine, purine, and carbazole; R 1 and R 1' Each time it appears, it is independently selected from H, hydroxyl, C1–C6 alkyl, C1–C6 haloalkyl, C1–C 20 Heteroalkyl, C1–C 10 Heteroalkyl –NH2, C2–C6 alkenyl, C2–C6 alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted arylalkyl, and substituted or unsubstituted heteroaryl, benzyl, –C(=NH)NH2, –CH2CO2R 0 –C(H)(CH) a (COOH))2, –CH2C(O)NHCH2CO2H, –CH2(CH2) a OH、–CH2CH2NHC(O)R 3 –(CH2) a C(O)NH(CH2) a C(O)OR 3 –(CH2) a –CN, spermine, –(CH2) a –NH2、–C(O)R 3 –(CH2) a C(O)OR 3 –(CH2) a OR 3 –C(H)((CH2) a OH) a' –CHCH(OH)(CH2) a OH, –(CH2) a OH, C1–C6–C(O)OH, –(CH2) that can be substituted or not substituted a - Heterocyclic rings , ;and Where R 1 and R 1' Together they form substituted or unsubstituted heterocycles, including but not limited to aza-butanes, pyrrolidines, piperidines, piperazines, morpholines, etc. or ; R 2 Each time it appears, it is independently selected from C2–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, and substituted or unsubstituted haloalkoxy. R 3 Each time it appears, it is independently selected from H, C1–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, –C1–C6–OH, –C1–C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine; and G 2 and G 3 Each time it appears, it is independently selected from groups that are absent or selected from groups containing the following: halogen, –CN, –NO2, –SO3R. 3 C1–C6 alkyl, C1–C6 haloalkyl, –E–O–R 3 –E–(CH2) a C(O)R 3 , –E–CO2H, –E–CHO, –E–C(O)R 3 , –E–C(O)NH(OH), –E–C(O)NHR 1 –E–C(O)N(H)C(H)(R 1 (R) 1' ), –E–C(O)N(R 1 (CH2) a P(O)(O)OH)2、–E–C(O)N(R 1 (CH2) a SO3H), –E–C(O)NR 1 R 1' –E–NR 1 R 1' –E–OR 2 –C(O)–heterocyclic alkyl (where the heterocyclic alkyl is substituted or unsubstituted), –C(O)–N(H)–CH(COOH)((CH2) a – heteroaryl) (wherein the heteroaryl group is substituted or unsubstituted), wherein E is independently selected from direct bond and C1–C6 alkylene group each time it appears.

85. The compound according to claim 84, wherein the compound has any one of the following formulas: , , , , , , and .

86. The compound of claim 84, wherein the compound has any one of the following formulas: , , , , , , , , , , , , and .

87. The compound according to any one of claims 84 to 85, wherein R 1 For H.

88. The compound according to claim 87, wherein R 1' It is a C1–C6 alkyl group.

89. The compound according to claim 88, wherein the C1–C6 alkyl group is branched.

90. The compound of claim 88, wherein the C1–C6 alkyl group comprises at least one substituent.

91. The compound according to claim 90, wherein the at least one substituent is a hydroxyl group.

92. The compound according to any one of claims 84 to 85, wherein R 1 and R 1' Together they form cycloalkyl groups or heterocycloalkyl groups.

93. The compound according to any one of claims 84 to 92, wherein G 2 and G 3 Each is represented by H.

94. The compound according to any one of claims 84 to 92, wherein G 2 or G 3 One of them is –SO3H or –SO3 - .

95. A compound having the following formula: ， Or its solvates, hydrates, tautomers, chelates or salts, in Z is a 5- or 6-membered heteroaromatic ring containing one, two, three or four heteroatoms selected from O, N or S; M is independently selected from hydrogen, halogens, and C1–C4 alkyl groups each time it appears; Y is independently selected from Ar2, –C(O)–Ar2, and –(CH2) each time it appears. a Ar2, –(CH2)3PO(OEt)2 or –CH2CO2Me; Ar1 is independently selected each time it appears from optionally substituted phenyl, pyridine, bipyridine, tripyridine, pyrazine, pyridazine, furan, dibenzofuran, thiophene, pyrrole, selenophene, naphthalene, fluorene, phenanthrene, cyclophosphine, phthalazine, quinazoline, quinoxaline, naphthidine, phenanthrene-rholine, purine, and carbazole; Ar2 is independently selected each time it appears from: (i) substituted or unsubstituted 5- and 6-membered monocyclic aromatic or heterocyclic rings; (ii) substituted or unsubstituted 9- and 10-membered fused bicyclic rings comprising two monocyclic rings, wherein at least one of the two monocyclic rings is an aromatic or heterocyclic ring; and (iii) substituted or unsubstituted 13- and 14-membered fused tricyclic rings comprising three monocyclic rings, wherein at least one of the three monocyclic rings is an aromatic or heterocyclic ring. Each Ar2 is independently denoted by –C(O)OR 3 –C1–C6 haloalkyl, –NO2 and –SO3H substitution; R 3 Each time it appears, it is independently selected from H, C1–C6 alkyl, C1–C6 haloalkyl, C1–C6 heteroalkyl, –C1–C6–OH, –C1–C6–C(O)OH, or substituted or unsubstituted cycloalkyl, substituted or unsubstituted heterocyclic, substituted or unsubstituted aryl, and substituted or unsubstituted heteroaryl, substituted or unsubstituted haloalkoxy, and guanidine.

96. The compound according to claim 95, wherein the –C1-C6 haloalkyl group is –CF3.

97. The compound according to claim 95, wherein the –C(O)OR 3 It is –C(O)OH.

98. The compound of claim 95, wherein Ar2 comprises two substituents.

99. The compound according to claim 95, wherein Ar2 comprises three substituents.

100. The compound of claim 95, wherein Ar2 comprises four substituents.

101. The compound of claim 95, wherein the compound comprises a –C(O)OR 3 A portion and a –C1-C6 haloalkyl moiety.

102. The compound of claim 95, wherein the compound comprises a –NO2 moiety and a –SO3H moiety.

103. The compound of claim 95, wherein the compound comprises a –SO3H moiety and a –C1-C6 haloalkyl moiety.

104. The compound of claim 95, wherein the compound comprises a –C(O)OR 3 One part and one –NO2 part.

105. The compound of claim 95, wherein the compound comprises at least one –SO3H moiety.

106. A polymer, copolymer, or metal gel comprising any one of the compounds according to claims 1 to 105.

107. A composition comprising at least two molecules of any one of the compounds according to claims 1 to 105.

108. A method for enhancing nucleic acid polymerase reactions, the method comprising: a. Forming a nucleic acid polymerase reaction composition, said nucleic acid polymerase reaction composition comprising: i. Template nucleic acid, ii. Nucleic acid polymerase, iii. A mixture of nucleotides or nucleotide analogs, and iv. at least one of the compounds according to any one of claims 1 to 105; and b. Incubating the nucleic acid polymerase reaction composition under conditions that allow for nucleic acid polymerization, wherein at least one compound according to any one of claims 1 to 105 increases the sustained synthesis capacity, rate, or fidelity of the nucleic acid polymerase reaction.

109. The method according to claim 108, wherein the nucleic acid polymerase is a DNA polymerase.

110. The method of claim 109, wherein the DNA polymerase is DPO4 or a variant thereof.

111. The method of claim 108, wherein the mixture of nucleotides or nucleotide analogs is a mixture of nucleotide analogs comprising nucleoside triaminophosphate, each of wherein each nucleoside triaminophosphate comprises a nucleobase selected from the group consisting of adenine, guanine, thymine, and cytosine, and a polymer chain portion wherein a first end of the polymer chain portion is connected to the nucleobase and a second end of the polymer chain portion is connected to an α-phosphate of the nucleoside triaminophosphate to provide extension of the nucleotide analog by cleavage of the aminophosphate bond.

112. The method of claim 108, wherein the nucleic acid polymerization reaction produces a scalable polymer of a nucleotide analog, wherein the scalable polymer encodes the nucleobase sequence information of the template nucleic acid.

113. The method of claim 108, wherein the conditions allowing the nucleic acid polymerization reaction include a suitable polymerization buffer and oligonucleotide primers.

114. The method of claim 113, wherein the suitable polymerization buffer comprises one or more components selected from MnCl2, buffer, salt, sugar, single-stranded binding protein (SSB), imidazole, pyrazole, triazole, betaine, molecular crowding agent, dimethyl sulfoxide (DMSO), alkyl glycol, glycerol, N-methyl-2-pyrrolidone (NMP), acetamide, butylated hydroxyanisole (BHA), polyphosphate and / or urea.

115. The method of claim 108, wherein the nucleic acid polymerase reaction composition further comprises a single-stranded binding protein.

116. The method of claim 108, wherein the nucleic acid polymerase reaction composition further comprises urea.

117. The method of claim 108, wherein the mixture of nucleotides or nucleotide analogs comprises a nucleotide analog containing a detectable marker.

118. The method of claim 117, wherein the detectable marker is an optically detectable marker selected from the group consisting of luminescent, chemiluminescent, fluorescent, fluorescent, chromogenic, or chromogenic markers.

119. Use of the compound according to any one of claims 1 to 105 in a nucleic acid polymerase reaction.

120. A method for sequencing a DNA or RNA template, the method comprising the following steps: a. Forming a DNA polymerase reaction composition, said DNA polymerase reaction composition comprising: i. DNA or RNA template, ii. Replication primers compounded with the template, iii. DNA polymerase, iv. A mixture of nucleotides or nucleotide analogs, v. At least one of the compounds according to any one of claims 1 to 105; b. Incubating the DNA polymerase reaction composition under conditions that allow DNA polymerization, wherein at least one compound according to any one of claims 1 to 105 increases the rate, fidelity, or sustained synthesis capability of the DNA polymerase reaction; and c. Determine the sequence of the nucleotide or nucleotide analog in the resulting nucleotide or nucleotide analog polymer.

121. The method of claim 120, wherein the mixture of nucleotide analogs comprises nucleoside triaminophosphate, each of the nucleoside triaminophosphates comprising a nucleobase selected from the group consisting of adenine, guanine, thymine, and cytosine, and a polymer chain portion wherein a first end of the polymer chain portion is connected to the nucleobase and a second end of the polymer chain portion is connected to an α-phosphate of the nucleoside triaminophosphate, to provide nucleotide analog extension by cleavage of the aminophosphate bond.

122. The method of claim 120, wherein the DNA polymerase is DPO4 or a variant thereof.

123. The method of claim 120, wherein the resulting nucleotide analog polymer is a scalable polymer.

124. The method according to any one of claims 120 to 123, further comprising the step of contacting the expandable polymer with an aminophosphate cleaving agent to produce an expanded polymer of a nucleotide analog.

125. The method of claim 121, wherein the polymer chain portion of each of the nucleotide analogs comprises a reporter portion characteristic of the nucleobases of the analog.

126. The method of claim 125, wherein each of the reporting portions generates a characteristic electronic signal.

127. The method of claim 120, wherein the step of determining the sequence of the nucleotide analog comprises the step of shifting the expanded polymer of the nucleotide analog through a nanopore.

128. Use of the compound according to any one of claims 1 to 105 in sequencing a DNA or RNA template.

129. A composition comprising: any one of the compounds according to claims 1 to 105; and a molecular crowding agent.

130. The composition of claim 129, wherein the molecular crowding agent is a polyalkylene glycol.

131. A composition comprising: any one of the compounds according to claims 1 to 105; and a buffer solution.

132. The composition of claim 131, wherein the buffer comprises one or more components selected from MnCl2, buffer solution, salt, sugar, single-stranded binding protein (SSB), imidazole, pyrazole, triazole, betaine, molecular crowding agent, dimethyl sulfoxide (DMSO), alkyl glycol, glycerol, N-methyl-2-pyrrolidone (NMP), acetamide, butylated hydroxyanisole (BHA), polyphosphate and / or urea.

133. The composition according to claim 132, wherein the salt is selected from the group consisting of: NaCl, NaBr, NaOAc, NaF, sodium formate, sodium dihydrogen phosphate, disodium hydrogen phosphate, NaSO4, sodium carbonate, sodium bicarbonate, sodium hexanoate, monosodium glutamate, sodium perchlorate, CsCl, LiCl, LiOAc, LiF, lithium carbonate, LiPO4, KCl, KOAc, KF, KSO4, potassium dihydrogen phosphate, dipotassium hydrogen phosphate, potassium carbonate, potassium bicarbonate, potassium glutamate, NH4Cl, NH4F, NH4OAc, NH4SO4, NH4Br, ammonium citrate, ammonium carbonate, ammonium bicarbonate, ammonium sulfite, ammonium glutamate, ammonium dihydrogen phosphate, tetramethylammonium chloride (TMACl), trimethylamine N-oxide (TMAO), tetraethylammonium chloride (TEACl), guanidine chloride, guanidine thiocyanate, and guanidine carbonate.

134. The composition according to claim 132, wherein the salt is an inorganic salt.

135. The composition according to claim 132, wherein the salt is NaCl or KCl.

136. The composition according to claim 132, wherein the sugar is maltose, trehalose, cellobiose, or sucrose.

137. The composition of claim 132, wherein the imidazole is a derivative or analog of an imidazole selected from the group consisting of: imidazole chloride, imidazole acetate, 1-methylimidazolium, 2-methylimidazolium, 1-ethylimidazolium, 1-ethyl-3-methylimidazolium chloride, 2-methyl-2-imidazoline, 1-butyl-3-methylimidazolium chloride, 1-methylimidazolium chloride, 1-hexyl-3-methylimidazolium, 3-octyl-1-methylimidazolium, and 1-decyl-3-methylimidazolium.

138. The composition of claim 132, wherein the alkanediol is ethylene glycol, propylene glycol, or butanediol, such as propylene glycol.

139. The composition of claim 138, wherein the propylene glycol is 1,2-propanediol or 1,3-propanediol, such as 1,2-propanediol. In some embodiments, the butanediol is 1,2-butanediol, 1,3-butanediol, 1,4-butanediol, 2,3-butanediol, 2,4-butanediol or 3,4-butanediol, such as 1,2-butanediol.

140. The composition of claim 132, wherein the molecular crowding agent is PEG, such as PEG4k to PEG25k or PEG4k to PEG10k, such as PEG5k, PEG8k or PEG10k.

141. The composition according to claim 132, wherein the polyphosphate is a tripolyphosphate, tetrapolyphosphate, pentapolyphosphate, hexapolyphosphate, trimetaphosphate (TMP), hexametaphosphate (HMP), or polyphosphate 60.

142. A composition comprising: any one of the compounds according to claims 1 to 105; and a polynucleotide.

143. The composition of claim 142, wherein the polynucleotide is a 20 to 60 mer oligonucleotide.

144. A composition comprising: any one of the compounds according to claims 1 to 105; and a protein.

145. The composition according to claim 144, wherein the protein is a DNA polymerase.

146. A composition comprising: any one of the compounds according to claims 1 to 105; and a mixture of nucleotides or nucleotide analogs.

147. A composition for enhancing the sustained synthesis capacity, fidelity, or rate of a DNA polymerase reaction, said composition comprising: at least one of the compounds according to any one of claims 1 to 105; and a mixture of nucleotide analogs.

148. A composition comprising: at least one of the compounds according to any one of claims 1 to 105; and a mixture of nucleotide analogs, wherein, relative to the same polymerization reaction in the absence of at least one of the compounds according to any one of claims 1 to 105, the at least one compound according to any one of claims 1 to 105 increases the quantity and accuracy of nucleotide analogs incorporated into daughter chains during template-dependent polymerization.

149. The composition of claim 148, wherein the mixture of nucleotide analogs comprises nucleoside triaminophosphate, each of the nucleoside triaminophosphates comprising a nucleobase selected from the group consisting of adenine, guanine, thymine, and cytosine, and a polymer chain portion wherein a first end of the polymer chain portion is attached to the nucleobase and a second end of the polymer chain portion is attached to an α-phosphate of the nucleoside triaminophosphate to provide nucleotide analog extension by cleavage of the aminophosphate bond.

150. The composition of claim 148, further comprising a buffer component selected from at least one of the following: Tris OAc, NH4OAc, PEG, water-miscible organic solvents, polyphosphate 60, NMS, and MnCl2.

151. The composition according to claim 148, further comprising a single-chain binding protein.

152. The composition according to claim 148, further comprising urea.

153. The composition of claim 148, wherein the mixture of nucleotide analogs comprises a nucleotide analog containing a detectable marker.

154. The composition of claim 153, wherein the detectable marker is an optically detectable marker selected from the group consisting of luminescent, chemiluminescent, fluorescent, fluorescent, chromogenic, or chromogenic markers.

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