Methods and compounds for treating Friedreich's ataxia

Abstract

The present disclosure relates to compounds and methods for regulating the expression of fxn and treating diseases and conditions in which fxn plays an active role. The compounds can be transcriptional modulator molecules having a first end, a second end, and an oligomeric backbone, a) the first end comprising a DNA binding moiety capable of non-covalently binding to the trinucleotide repeat sequence GAA, b) the second end comprising a protein binding moiety that binds to a regulatory molecule that regulates the expression of a gene comprising the nucleotide repeat sequence GAA, and c) the oligomeric backbone comprising a linker between the first end and the second end.

Description

[Technical Field]

[0001] cross reference This application claims the benefit of U.S. Patent Application No. 63 / 135,427, filed January 8, 2021, which is incorporated herein by reference in its entirety.

[0002] Disclosed herein are novel chimeric heterocyclic polyamide compounds and compositions and their applications as pharmaceuticals for the treatment of diseases. Methods for modulating the expression of fxn in a human or animal subject are also provided for the treatment of diseases such as Friedreich's ataxia. [Background technology]

[0003] The present disclosure relates to the treatment of inherited genetic diseases characterized by the overproduction of mRNA.

[0004] Friedreich's ataxia ("FA" or "FRDA") is an autosomal recessive neurodegenerative disorder caused by mutations in the fxn gene, which encodes the protein frataxin ("FXN"), an iron-binding mitochondrial protein involved in electron transport and metabolism. In most subjects with FA, a GAA trinucleotide repeat (approximately 66 to over 1000 trinucleotides) is contained in the first intron of fxn, and this overexpansion contributes to the observed pathology. The overexpansion of the GAA repeat results in reduced expression of FXN.

[0005] Friedreich's ataxia is characterized by the progressive degeneration of the nervous system, particularly sensory neurons. Additionally, cardiac myocytes and pancreatic beta cells are susceptible to frataxin depletion. Symptoms usually appear by age 18, but later diagnosis of FA is not uncommon. Patients with FA develop neurodegeneration of large sensory neurons and spinocerebellar tracts, as well as cardiomyopathy and diabetes. Clinical symptoms of FA include ataxia, gait ataxia, muscle weakness, loss of upper body muscle strength, loss of balance, absent reflexes in the lower limbs and tendons, loss of sensation, particularly to vibration, impaired position sense, impaired perception of temperature, touch, and pain, hearing and visual impairment, including distorted color vision and involuntary eye movements, irregular foot shape, including pes cavus and varus, hearing impairment, dysarthria, dysphagia, respiratory problems, scoliosis, diabetes, glucose and carbohydrate intolerance, hypertrophic cardiomyopathy, arrhythmias, myocardial fibrosis, and cardiac dysfunction, including heart failure. Currently, there is no cure for FA, and medical treatment is limited to spinal and cardiac surgical interventions, as well as treatments to assist with balance, coordination, movement, and speech. Summary of the Invention

[0006] The present disclosure utilizes regulatory molecules present in the cell nucleus that control gene expression. Eukaryotic cells provide several mechanisms for controlling gene replication, transcription, and / or translation. Regulatory molecules produced by various biochemical mechanisms within the cell can regulate various processes involved in the conversion of genetic information into cellular components. Some regulatory molecules are known to regulate the production of mRNA, and when directed against fxn, they can regulate the production of fxn mRNA, which causes Friedreich's ataxia, thus reversing the progression of the disease.

[0007] The present disclosure provides compounds and methods for recruiting regulatory molecules into proximity with fxn. The compounds disclosed herein comprise (a) a recruitment moiety that will bind to the regulatory molecule, and (b) a DNA-binding moiety that will selectively bind to fxn. The compounds will counteract the expression of defective fxn in the following ways: (1) The DNA-binding moiety would selectively bind to the characteristic GAA trinucleotide repeat sequence of fxn. (2) Therefore, the DNA-binding site and the associated recruiting moiety would be held in close proximity to fxn. (3) the recruiting moiety now proximal to fxn will recruit regulatory molecules proximal to the gene; and (4) Regulatory molecules may regulate expression and thus counteract the production of defective fxn by direct interaction with the gene.

[0008] The above mechanism would provide an effective treatment for Friedreich's ataxia, which is caused by the expression of defective fxn genes. Therefore, correcting the expression of defective fxn genes represents a promising method for the treatment of Friedreich's ataxia.

[0009] The present disclosure provides recruitment moieties that will bind to regulatory molecules. Small molecule inhibitors of regulatory molecules serve as templates for the design of recruitment moieties, as these inhibitors typically act through non-covalent binding to the regulatory molecule.

[0010] The present disclosure further provides DNA-binding moieties that will selectively bind to one or more copies of the GAA trinucleotide repeat characteristic of a defective fxn gene. The selective binding of the DNA-binding moiety to fxn, made possible by the high GAA number associated with the defective fxn, targets the recruiting moiety to the vicinity of the gene, recruiting regulatory molecules to a position where they upregulate gene transcription.

[0011] The DNA-binding moiety will contain a polyamide segment that will selectively bind to the target GAA sequence. For example, polyamides designed by Dervan (U.S. Patent Nos. 9,630,950 and 8,524,899) can selectively bind to selected DNA sequences. These polyamides reside in the minor groove of double-helical DNA and form hydrogen-bonding interactions with Watson-Crick base pairs. Polyamides that selectively bind to specific DNA sequences can be designed by linking monoamide building blocks according to established chemical rules. One building block is provided for each DNA base pair, and each building block noncovalently and selectively binds to one of the following DNA base pairs: A / T, T / A, G / C, and C / G. Following this guideline, trinucleotides will be linked to the molecule with three amide units, i.e., triamides. Generally, these polyamides are oriented in either direction of the DNA sequence, so that the 5'-GAA-3' trinucleotide repeat sequence of fxn can be targeted by polyamides selectively to either GAA or AAG. Additionally, polyamides that bind to complementary sequences, in this case TTC or CTT, also bind to the trinucleotide repeat sequence of fxn and can be used as well.

[0012] In principle, longer DNA sequences can be targeted with higher specificity and / or higher affinity by combining more monoamide components into longer polyamide chains. Ideally, the binding affinity for a polyamide is simply equal to the sum of each individual monoamide / DNA base pair interaction. However, in practice, due to the fairly strong geometric mismatch between the polyamide and DNA structures, longer polyamide sequences do not bind as tightly to longer DNA sequences as would be expected from simple additive contributions. The geometric mismatch between longer polyamide sequences and longer DNA sequences induces unfavorable geometric distortions that subtract from the otherwise expected binding affinity.

[0013] Thus, the present disclosure provides DNA segments comprising triamides connected by flexible spacers that relieve geometric strain that would otherwise reduce the binding affinity of larger polyamide sequences.

[0014] Disclosed herein are compounds (i.e., transcriptional modulator molecules) comprising polyamides capable of binding to one or more copies of the trinucleotide repeat sequence GAA and regulating the expression of the defective fxn gene. Treating a subject with these compounds can counteract the expression of the defective fxn gene, which can reduce the occurrence, severity, and / or frequency of symptoms associated with Friedreich's ataxia. Certain compounds disclosed herein may provide higher binding affinity and / or selectivity than previously observed for this class of compounds.

[0015] Other objects, features, and advantages of the compounds, methods, and compositions described herein will become apparent from the following detailed description. It should be understood, however, that the detailed description and the specific examples, while indicating specific embodiments of the invention, are given by way of illustration only, since various changes and modifications within the spirit and scope of the disclosure will become apparent to those skilled in the art from this detailed description.

[0016] Incorporation by Reference All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference. DETAILED DESCRIPTION OF THE INVENTION

[0017] The transcriptional modulator molecules described herein represent the interface of chemistry, biology, and precision medicine in that the molecules can be programmed to control the expression of target genes containing the nucleotide repeat GAA. The transcriptional modulator molecules contain a DNA-binding moiety that can selectively bind to one or more copies of the GAA hexanucleotide repeat characteristic of a defective fxn gene. The transcriptional modulator molecules also contain a moiety that binds to a regulatory protein. Selective binding of the target gene brings the regulatory protein into proximity with the target gene, thereby downregulating transcription of the target gene. The molecules and compounds disclosed herein provide higher binding affinity and selectivity than previously observed for this class of compounds and may be more effective in treating diseases associated with defective fxn genes.

[0018] The transcriptional modulator molecules described herein recruit regulatory molecules to regulate the expression of the defective fxn gene, effectively treating and / or alleviating symptoms associated with diseases such as Friedreich's ataxia.

[0019] Transcriptional Modulator Molecules The transcriptional modulator molecules disclosed herein have useful activity for regulating the transcription of target genes having one or more GAA repeats (e.g., fxn) and can be used to treat or prevent diseases or conditions in which the target gene (e.g., fxn) plays an active role. Accordingly, in broad aspects, certain embodiments also provide pharmaceutical compositions comprising one or more compounds disclosed herein together with a pharmaceutically acceptable carrier, as well as methods of making and using the compounds and compositions. Certain embodiments provide methods for modulating the expression of fxn. Other embodiments provide methods for treating a fxn-mediated disorder in a patient in need of such treatment, comprising administering to the patient a therapeutically effective amount of a compound or composition according to the present disclosure. Also provided is the use of certain compounds disclosed herein for use in the manufacture of a medicament for the treatment of a disease or condition alleviated by modulating the expression of fxn.

[0020] Some embodiments relate to a transcription modulator molecule or compound having a first end, a second end, and an oligomeric backbone, wherein a) the first end comprises a DNA-binding moiety capable of non-covalently binding to the nucleotide repeat sequence GAA, b) the second end comprises a protein-binding moiety that binds to a regulatory molecule that regulates expression of a gene comprising the nucleotide repeat sequence GAA, and c) the oligomeric backbone comprises a linker between the first end and the second end. In some embodiments, the second end is a Brd4-binding moiety. In some embodiments, the second end is not a Brd4-binding moiety.

[0021] In certain embodiments, the compound has structural formula (I): XLY Formula (I) or a salt thereof, wherein: X comprises a recruitment moiety capable of non-covalently binding to a regulatory moiety in the nucleus; Y comprises a DNA recognition moiety capable of non-covalently binding to one or more copies of the trinucleotide repeat sequence GAA; L is a linker.

[0022] Certain compounds disclosed herein may have useful activity in regulating the transcription of fxn and may be used in the treatment and / or prevention of diseases or conditions in which fxn plays an active role. Accordingly, in a broad aspect, certain embodiments also provide pharmaceutical compositions comprising one or more compounds disclosed herein together with a pharmaceutically acceptable carrier, as well as methods of making and using the compounds and compositions. Certain embodiments provide methods for modulating the expression of fxn. Other embodiments provide methods for treating a fxn-mediated disorder in a patient in need of such treatment, comprising administering to the patient a therapeutically effective amount of a compound or composition according to the present disclosure. Also provided is the use of certain compounds disclosed herein for use in the manufacture of a medicament for the treatment of a disease or condition alleviated by modulating the expression of fxn.

[0023] In certain embodiments, the regulatory molecule is selected from bromodomain-containing proteins, nucleosome remodeling factors ("NURFs"), bromodomain PHD finger transcription factors ("BPTFs"), ten-eleven translocation enzymes ("TETs"), methylcytosine dioxygenases ("TET1"), DNA demethylases, helicases, acetyltransferases, and histone deacetylases ("HDACs").

[0024] In some embodiments, the first terminus is Y, the second terminus is X, and the oligomer backbone is L.

[0025] In certain embodiments, the compound has structural formula (II): XL-(Y5-Y6-Y7) n -Y0 Formula (II) or a salt thereof, wherein: X comprises a recruitment moiety capable of non-covalently binding to a regulatory molecule in the nucleus; L is a linker, Y5, Y6, and Y7 are internal subunits, each of which is a heterocycle or C 1-6 a moiety selected from straight chain aliphatic segments, each of which is chemically bonded to its two neighbors; Y0 is a terminal subunit comprising a moiety selected from a heterocyclic or a linear aliphatic segment, and is chemically linked to its single adjacent moiety; Each subunit can non-covalently bind to an individual nucleotide within the GAA repeat sequence, n is an integer between 1 and 200, inclusive; (Y5-Y6-Y7) n -Y0 binds to form a DNA recognition moiety that can non-covalently bind to one or more copies of the trinucleotide sequence GAA.

[0026] In certain embodiments, the compound of structural formula (II) comprises a subunit for each individual nucleotide in the GAA repeat sequence.

[0027] In certain embodiments, each internal subunit has an amino (—NH—) group and a carboxy (—CO—) group.

[0028] In certain embodiments, compounds of structural formula (II) contain an amide (—NHCO—) bond between each pair of internal subunits.

[0029] In certain embodiments, compounds of structural formula (II) include an amide (—NHCO—) bond between L and the left-most internal subunit.

[0030] In certain embodiments, the compound of structural formula (II) comprises an amide bond between the rightmost internal subunit and the terminal subunit.

[0031] In certain embodiments, each subunit independently comprises a moiety selected from a heterocycle and an aliphatic chain.

[0032] In certain embodiments, the heterocycle is a monocyclic heterocycle. In certain embodiments, the heterocycle is a monocyclic 5-membered heterocycle. In certain embodiments, each heterocycle contains a heteroatom independently selected from N, O, or S. In certain embodiments, each heterocycle is independently selected from pyrrole, imidazole, thiazole, oxazole, thiophene, and furan.

[0033] In certain embodiments, the aliphatic chain is C 1-6 In certain embodiments, the aliphatic chain has the structural formula -(CH) m -, and m is selected from 1, 2, 3, 4, and 5. In certain embodiments, the aliphatic chain is -CH2CH2-.

[0034] In certain embodiments, each subunit independently comprises:

[0035] [ka]

[0036] [ka] -NH-benzopyrazinylene-CO-, -NH-phenylene-CO-, -NH-pyridinylene-CO-, -NH-piperidinylene-CO-, -NH-pyrimidinylene-CO-, -NH-anthracenylene-CO-, -NH-quinolinylene-CO-, and

[0037] [ka] wherein Z is H, NH, C 1-6 Alkyl, C 1-6 Haloalkyl, or C 1-6 It is alkyl-NH2.

[0038] In some embodiments, Py is

[0039] [ka] and Im is

[0040] [ka] and Hp is

[0041] [ka] and Th is

[0042] [ka] and Pz is

[0043] [ka] and Nt is

[0044] [ka] and Tn is

[0045] [ka] and Nh is

[0046] [ka] and iNt is

[0047] [ka] and iIm is

[0048] [ka] and HpBi is

[0049] [ka] and ImBi is

[0050] [ka] and PyBi is

[0051] [ka] and Dp is

[0052] [ka] and -NH-benzopyrazinylene-CO- is

[0053] [ka] and -NH-phenylene-CO- is

[0054] [ka] and -NH-pyridinylene-CO- is

[0055] [ka] and -NH-piperidinylene-CO- is

[0056] [ka] and -NH-pyrazinylene-CO- is

[0057] [ka] and -NH-anthracenylene-CO- is

[0058] [ka] and -NH-quinolinylene-CO- is

[0059] [ka] In some embodiments, Py is

[0060] [ka] and Im is

[0061] [ka] and Hp is

[0062] [ka] and Th is

[0063] [ka] and Pz is

[0064] [ka] and Nt is

[0065] [ka] and Tn is

[0066] [ka] and Nh is

[0067] [ka] and iNt is

[0068] [ka] and iIm is

[0069] [ka] is.

[0070] In certain embodiments, n is between 1 and 100, inclusive. In certain embodiments, n is between 1 and 50, inclusive. In certain embodiments, n is between 1 and 20, inclusive. In certain embodiments, n is between 1 and 10, inclusive. In certain embodiments, n is between 1 and 5, inclusive. In certain embodiments, n is an integer between 1 and 3, inclusive. In certain embodiments, n is selected from 1 and 2. In certain embodiments, n is 1.

[0071] In certain embodiments, n is an integer from 1 to 5, inclusive.

[0072] In certain embodiments, n is an integer from 1 to 3, inclusive.

[0073] In certain embodiments, n is an integer from 1 to 2, inclusive.

[0074] In certain embodiments, n is 1.

[0075] In certain embodiments, L is C 1-6 Contains a straight chain aliphatic segment.

[0076] In certain embodiments, L is (CH2OCH2) m wherein m is an integer from 1 to 20, inclusive. In certain further embodiments, m is an integer from 1 to 10, inclusive. In certain further embodiments, m is an integer from 1 to 5, inclusive.

[0077] In certain embodiments, the compound has structural formula (III): XL-(Y5-Y6-Y7)-(W-Y5-Y6-Y7) n -Y0 Formula (III) or a salt thereof, wherein: X comprises a recruitment moiety capable of non-covalently binding to a regulatory molecule in the nucleus; L is a linker, Y5, Y6, and Y7 are internal subunits, each of which is a heterocycle or C 1-6 a moiety selected from straight chain aliphatic segments, each of which is chemically bonded to its two neighbors; Y0 is a terminal subunit comprising a moiety selected from a heterocyclic or a linear aliphatic segment, and is chemically linked to its single adjacent moiety; Each subunit can non-covalently bind to an individual nucleotide within the GAA repeat sequence, W is a spacer, n is an integer between 1 and 200, inclusive; (Y5-Y6-Y7)-(W-Y5-Y6-Y7) n-Y0 binds to form a DNA recognition moiety that can non-covalently bind to one or more copies of the trinucleotide repeat sequence GAA.

[0078] In certain embodiments, Y5-Y6-Y7 is

[0079] [ka] is.

[0080] In certain embodiments, Y5-Y6-Y7 is

[0081] [ka] is.

[0082] In certain embodiments, Y5-Y6-Y7 is Im-Py-β.

[0083] In certain embodiments, Y5-Y6-Y7 is Im-Im-β.

[0084] In certain embodiments, each Y5-Y6-Y7 is independently selected from β-Py-Im and β-Im-Im.

[0085] In certain embodiments, at most one Y5-Y6-Y7 is β-Im-Im.

[0086] In certain embodiments of compounds of structural formula (III), n is between 1 and 100, inclusive. In certain embodiments of compounds of structural formula (III), n is between 1 and 50, inclusive. In certain embodiments of compounds of structural formula (III), n is between 1 and 20, inclusive. In certain embodiments of compounds of structural formula (III), n is between 1 and 10, inclusive. In certain embodiments of compounds of structural formula (III), n is between 1 and 5, inclusive. In certain embodiments of compounds of structural formula (III), n is selected from 1 and 2. In certain embodiments of compounds of structural formula (III), n is 1.

[0087] In certain embodiments, the compound has structural formula (IV): XL-(Y5-Y6-Y7)-V-(Y8-Y9-Y 10 )-Y0 Formula (IV) or a salt thereof, wherein: X comprises a recruitment moiety capable of non-covalently binding to a regulatory molecule in the nucleus; Y5, Y6, Y7, Y8, Y9, and Y 10 are internal subunits, each of which is a heterocycle or C 1-6 a moiety selected from straight chain aliphatic segments, each of which is chemically bonded to its two neighbors; Y0 is a terminal subunit comprising a moiety selected from a heterocyclic or a linear aliphatic segment, and is chemically linked to its single adjacent moiety; Each subunit can non-covalently bind to an individual nucleotide within the GAA repeat sequence, L is a linker, V is a curve component for forming a hairpin curve, n is an integer between 1 and 200, inclusive; (Y5-Y6-Y7)-V-(Y8-Y9-Y 10 )-Y0 combine to form a DNA recognition moiety that can non-covalently bind to one or more copies of the trinucleotide repeat sequence GAA.

[0088] In certain embodiments of compounds of structural formula (IV), n is between 1 and 100, inclusive. In certain embodiments of compounds of structural formula (IV), n is between 1 and 50, inclusive. In certain embodiments of compounds of structural formula (IV), n is between 1 and 20, inclusive. In certain embodiments of compounds of structural formula (IV), n is between 1 and 10, inclusive. In certain embodiments of compounds of structural formula (IV), n is between 1 and 5, inclusive. In certain embodiments of compounds of structural formula (IV), n is selected from 1 and 2. In certain embodiments of compounds of structural formula (IV), n is 1.

[0089] In certain embodiments, V is —HN—CH 2 CH 2 CH 2 —CO—.

[0090] In certain embodiments, the compound has structural formula (V): XC(=O)-CH2CH2-(Y5-Y6-Y7) n -NH-Y0 Formula (V) or a salt thereof, wherein: X comprises a recruitment moiety capable of non-covalently binding to a regulatory molecule in the nucleus; each Y5-Y6-Y7 is independently selected from β-Py-Im and β-Im-Im; Y0 is a terminal subunit comprising a moiety selected from a heterocyclic or a linear aliphatic segment, and is chemically linked to its single adjacent moiety; n is an integer from 1 to 200, inclusive.

[0091] In certain embodiments of compounds of structural formula (V), at most one Y5-Y6-Y7 is β-Im-Im.

[0092] In certain embodiments of compounds of structural formula (V), Y5-Y6-Y7 is β-Py-Im.

[0093] In certain embodiments of compounds of Structural Formula (V), n is between 1 and 100, inclusive. In certain embodiments of compounds of Structural Formula (V), n is between 1 and 50, inclusive. In certain embodiments of compounds of Structural Formula (V), n is between 1 and 20, inclusive. In certain embodiments of compounds of Structural Formula (V), n is between 1 and 10, inclusive. In certain embodiments of compounds of Structural Formula (V), n is between 1 and 5, inclusive. In certain embodiments of compounds of Structural Formula (V), n is selected from 1 and 2. In certain embodiments of compounds of Structural Formula (V), n is 1.

[0094] In certain embodiments, the compound has structural formula (VI):

[0095] [ka] Formula (VI) or a salt thereof, wherein: X comprises a recruitment moiety capable of non-covalently binding to a regulatory molecule in the nucleus; Y0 is a terminal subunit comprising a moiety selected from a heterocyclic or a linear aliphatic segment, and is chemically linked to its single adjacent moiety; n is an integer from 1 to 200, inclusive.

[0096] In certain embodiments of compounds of structural formula (VI), n is between 1 and 100, inclusive. In certain embodiments of compounds of structural formula (VI), n is between 1 and 50, inclusive. In certain embodiments of compounds of structural formula (VI), n is between 1 and 20, inclusive. In certain embodiments of compounds of structural formula (VI), n is between 1 and 10, inclusive. In certain embodiments of compounds of structural formula (VI), n is between 1 and 5, inclusive. In certain embodiments of compounds of structural formula (VI), n is selected from 1 and 2. In certain embodiments of compounds of structural formula (VI), n is 1.

[0097] In certain embodiments, the compound has structural formula (VII):

[0098] [ka] Formula (VII) or a salt thereof, wherein: X comprises a recruitment moiety capable of non-covalently binding to a regulatory molecule in the nucleus; W is a spacer, Y0 is a terminal subunit comprising a moiety selected from a heterocyclic or a linear aliphatic segment, and is chemically linked to its single adjacent moiety; n is an integer from 1 to 200, inclusive.

[0099] In certain embodiments of compounds of structural formula (VII), n is between 1 and 100, inclusive. In certain embodiments of compounds of structural formula (VII), n is between 1 and 50, inclusive. In certain embodiments of compounds of structural formula (VII), n is between 1 and 20, inclusive. In certain embodiments of compounds of structural formula (VII), n is between 1 and 10, inclusive. In certain embodiments of compounds of structural formula (VII), n is between 1 and 5, inclusive. In certain embodiments of compounds of structural formula (VII), n is selected from 1 and 2. In certain embodiments of compounds of structural formula (VII), n is 1.

[0100] In certain embodiments of compounds of structural formula (VII), W is —NHCH—(CHOCH) p -CH2CO-, and p is an integer from 1 to 4, inclusive.

[0101] In some embodiments, (V) is —(CH) a -NR 1 -(CH2) b -, -(CH2) a -, -(CH2) a -O-(CH2) b -, -(CH2) a- CH(NHR 1 )-, -(CH2) a -CH(NHR 1 )-, -(CR 2 R 3 ) a - or -(CH2) a -CH(NR 1 3) + -(CH2) b wherein each a is independently an integer from 2 to 4; 1 is H, optionally substituted C 1-6 Alkyl, optionally substituted C 3-10 Cycloalkyl, optionally substituted C 6-10 aryl, optionally substituted 4- to 10-membered heterocyclyl, or optionally substituted 5- to 10-membered heteroaryl, and each R 2 and R 3are independently H, halogen, OH, NHAc, or C 1-4 In some embodiments, R 1 is H. In some embodiments, R 1 is a C optionally substituted by 1 to 3 substituents selected from -C(O)-phenyl 1-6 In some embodiments, (V) is -(CR 2 R 3 )-(CH2)a- or -(CH2)a-(CR 2 R 3 )-(CH2) b wherein each a is independently 1 to 3, b is 0 to 3, and each R 2 and R 3 are independently H, halogen, OH, NHAc, or C 1-4 In some embodiments, (V) is —(CH)—CH(NH) + -(CH2)- or -(CH2)-CH2CH(NH3) + -It is.

[0102] In one aspect, compounds of the present disclosure bind to the GAA of fxn and recruit regulatory moieties to the vicinity of fxn, which, due to their proximity to the gene, will likely regulate the expression of fxn.

[0103] Also provided are embodiments in which any of the compounds disclosed above, including compounds of Formulae (I)-(VIII), are singly, partially, or fully deuterated. Methods for exchanging deuterium for hydrogen are known in the art.

[0104] Also provided are embodiments in which any of the above embodiments may be combined with any one or more of these embodiments, provided that the combinations are not mutually exclusive.

[0105] As used herein, two embodiments are "mutually exclusive" if one is defined as being different from the other. For example, an embodiment in which two groups are bonded to form a cycloalkyl is mutually exclusive from an embodiment in which one group is ethyl and the other group is hydrogen. Similarly, an embodiment in which one group is CH2 is mutually exclusive from an embodiment in which the same group is NH.

[0106] In one aspect, compounds of the present disclosure bind to the GAA of fxn and recruit regulatory moieties to the vicinity of fxn, which, due to their proximity to the gene, will likely regulate the expression of fxn.

[0107] In one embodiment, the compounds of the present disclosure provide a polyamide sequence for interaction of a single polyamide subunit with each base pair in the GAA repeat sequence. In one embodiment, the compounds of the present disclosure provide a curve component V to enable hairpin binding of the compound to GAA, with each nucleotide pair interacting with two subunits of the polyamide.

[0108] In one embodiment, a compound of the present disclosure provides more than one copy of a polyamide sequence for non-covalent attachment to fxn, and the individual polyamide sequences in the compound are linked by a spacer W, as defined above. The spacer W allows the compound to adjust its geometry as needed to relieve geometric strain that would otherwise affect non-covalent attachment of longer polyamide sequences.

[0109] First terminal DNA binding site The first end interacts with and binds to the minor groove of the gene, particularly the GAA sequence. In one embodiment, the compound of the present disclosure provides a polyamide sequence for the interaction of a single polyamide subunit with each base pair in the GAA repeat sequence. In one embodiment, the compound of the present disclosure provides a curved component (e.g., an aliphatic amino acid portion) to enable the compound to bind to GAA in a hairpin, and each nucleotide pair interacts with two subunits of the polyamide.

[0110] In one aspect, the molecules of the present disclosure are more likely to bind to the repeated GAA of fxn than to GAA elsewhere in the DNA of a subject due to the higher number of GAA repeats associated with fxn.

[0111] In one embodiment, the compounds of the present disclosure provide more than one copy of the polyamide sequence for non-covalent binding to GAA. In one embodiment, the compounds of the present disclosure bind to fxn with higher affinity than corresponding compounds containing a single polyamide sequence.

[0112] In one aspect, a compound of the present disclosure provides more than one copy of a polyamide sequence for non-covalent attachment to GAA, and the individual polyamide sequences in the compound are linked by a spacer W, as defined above. The spacer W allows the compound to adjust its geometry as needed to relieve geometric strain that would otherwise affect non-covalent attachment of longer polyamide sequences.

[0113] In certain embodiments, the DNA recognition or binding moiety binds to the minor groove of DNA.

[0114] In certain embodiments, the DNA recognition or binding moiety comprises a polymeric sequence of monomers, each monomer in the polymer selectively binding to a specific DNA base pair.

[0115] In certain embodiments, the DNA recognition or binding moiety comprises a polyamide moiety.

[0116] In certain embodiments, the DNA recognition or binding moiety comprises a polyamide portion comprising heteroaromatic monomers, each non-covalently bound to a specific nucleotide, and each heteroaromatic monomer is linked to its neighbor or neighbors via amide bonds.

[0117] In certain embodiments, the DNA recognition moiety binds to a sequence comprising at least 1000 pentanucleotide repeats. In certain embodiments, the DNA recognition moiety binds to a sequence comprising at least 500 trinucleotide repeats. In certain embodiments, the DNA recognition moiety binds to a sequence comprising at least 200 trinucleotide repeats. In certain embodiments, the DNA recognition moiety binds to a sequence comprising at least 100 trinucleotide repeats. In certain embodiments, the DNA recognition moiety binds to a sequence comprising at least 50 trinucleotide repeats. In certain embodiments, the DNA recognition moiety binds to a sequence comprising at least 20 trinucleotide repeats.

[0118] In certain embodiments, the compound comprises a cell-permeable ligand moiety.

[0119] In certain embodiments, the cell-permeable ligand moiety is a polypeptide.

[0120] In certain embodiments, the cell-permeable ligand moiety is a polypeptide comprising fewer than 30 amino acid residues.

[0121] In certain embodiments, the polypeptide is selected from any one of SEQ ID NO: 1 to SEQ ID NO: 37, inclusive.

[0122] The form of the polyamide selected can vary based on the target gene. The first end can comprise a polyamide selected from the group consisting of linear polyamides, hairpin polyamides, H-pin polyamides, overlapping polyamides, slipped polyamides, cyclic polyamides, tandem polyamides, and extended polyamides. In some embodiments, the first end comprises a linear polyamide. In some embodiments, the first end comprises a hairpin polyamide.

[0123] The binding affinity between a polyamide and a target gene can be adjusted based on the composition of the polyamide. In some embodiments, a polyamide can bind to DNA with an affinity of less than about 600 nM, about 500 nM, about 400 nM, about 300 nM, about 250 nM, about 200 nM, about 150 nM, about 100 nM, or about 50 nM. In some embodiments, a polyamide can bind to DNA with an affinity of less than about 300 nM. In some embodiments, a polyamide can bind to DNA with an affinity of less than about 200 nM. In some embodiments, a polyamide can bind to DNA with an affinity of greater than about 200 nM, about 150 nM, about 100 nM, about 50 nM, about 10 nM, or about 1 nM. In some embodiments, the polyamide can bind to DNA with an affinity in the range of about 1-600 nM, 10-500 nM, 20-500 nM, 50-400 nM, or 100-300 nM.

[0124] The binding affinity between polyamides and target DNA can be determined using quantitative footprinting experiments. The experiments measure the dissociation constant, K, of polyamides for the target sequence at either 24°C or 37°C using either standard polyamide assay solution conditions or approximate intracellular solution conditions. d This involves measuring

[0125] The binding affinity between the control protein and the ligand on the second end can be determined using an assay suitable for the particular protein. The experiment can be performed to determine the dissociation constant, K, of the ligand for the protein. d and involves using either standard protein assay solution conditions or approximating intracellular solution conditions.

[0126] In some embodiments, the first terminus comprises -NH-QC(O)-, where Q is an optionally substituted C 6-10 In some embodiments, Q is an optionally substituted C 6-10In some embodiments, Q is an optionally substituted 5- to 10-membered heteroarylene group. In some embodiments, Q is an optionally substituted 5- to 10-membered heteroarylene group. In some embodiments, the 5- to 10-membered heteroarylene group is selected from the group consisting of H, OH, halogen, C 1-10 Alkyl, NO2, CN, NR′R″, C 1-6 Haloalkyl, C 1-6 Alkoxyl, C 1-6 Haloalkoxy, (C 1-6 Alkoxy)C 1-6 Alkyl, C 2-10 Alkenyl, C 2-10 Alkynyl, C 3-7 Carbocyclyl, 4-10 membered heterocyclyl, C 6-10 Aryl, 5- to 10-membered heteroaryl, (C 3-7 Carbocyclyl)C 1-6 Alkyl, (4-10 membered heterocyclyl)C 1-6 Alkyl, (C 6-10 Aryl)C 1-6 Alkyl, (C 6-10 Aryl)C 1-6 Alkoxy, (5-10 membered heteroaryl)C 1-6 Alkyl, (C 3-7 (carbocyclyl)-amines, (4- to 10-membered heterocyclyl)amines, (C 6-10 (aryl)amine, (5-10 membered heteroaryl)amine, acyl, C-carboxy, O-carboxy, C-amido, N-amido, S-sulfonamido, N-sulfonamido, -SR ’ , COOH, or CONR′R″, wherein each R′ and R is independently H, C 1-10 Alkyl, C 1-10 haloalkyl haloalkyl, C 1-10 alkoxyl.

[0127] In some embodiments, the first end comprises at least three aromatic carboxamide moieties selected to correspond to the nucleotide repeat sequence GAA and at least one aliphatic amino acid residue selected from the group consisting of glycine, β-alanine, γ-aminobutyric acid, 2,4-diaminobutyric acid, and 5-aminovaleric acid, hi some embodiments, the first end comprises at least one β-alanine subunit.

[0128] In some embodiments, the monomeric elements are independently selected from the group consisting of optionally substituted pyrrole carboxamide monomers, optionally substituted imidazole carboxamide monomers, optionally substituted C-C linked heteromonocyclic / heterobicyclic moieties, and β-alanine.

[0129] In some embodiments, the first end comprises the structure of formula (A-1), or a pharmaceutically acceptable salt thereof: -L 1a -[AM] p -E1 (A-1) During the ceremony, each [AM] occurs p times, where p is an integer ranging from 1 to 10; L 1a is a bond, C 1-6 Alkylene, -NR a -C 1-6 Alkylene-C(O)-, -NR a C(O)-, -NR a -C 1-6 Alkylene, -O-, or -OC 1-6 is alkylene, Each A is a bond, C 1-10 Alkylene, optionally substituted C 6-10 arylene group, optionally substituted 4- to 10-membered heterocyclene, optionally substituted 5- to 10-membered heteroarylene group, -C 1-10 Alkylene-C(O)-, -C 1-10 Alkylene-NR a -, -CO-, -NR a -,-CONR a -,-CONR a C 1-4Alkylene-, -NR a CO-C 1-4 Alkylene-, -C(O)O-, -O-, -S-, -S(O)-, -S(O)2-, -C(=S)-NH-, -C(O)-NH-NH-, -C(O)-N=N-, -C(O)-CH=CH-, -(CH2) 0-4 -CH=CH-(CH2) 0-4- , -N(CH3)-C 1-6 alkylene-,

[0130] [ka] , -NH-C 1-6 Alkylene-NH-, -OC 1-6 selected from the group consisting of alkylene-O-, -NH-N=N-, -NH-C(O)-NH-, and any combination thereof, wherein at least one A is -CONH-; Each M is an optionally substituted C 6-10 an arylene group, an optionally substituted 4- to 10-membered heterocyclene, an optionally substituted 5- to 10-membered heteroarylene group, or an optionally substituted alkylene; E1 is H or -A E -G, A E is absent or -NHCO-, G is an optionally substituted C 6-10 Aryl, optionally substituted 4- to 10-membered heterocyclyl, optionally substituted 5- to 10-membered heteroaryl, optionally substituted C 1-6 Alkyl, C 0-4 Alkylene-NHC(=NH)NH, -CN, -C 0-4 Alkylene-C(=NH)(NR a R b ), -C 0-4 Alkylene-C(=N + H2)(NR a R b )C 1-5 Alkylene-NR a R b , C 0-4 Alkylene-NHC(=NH)R aand optionally substituted amines; Each R a and R b are independently H, optionally substituted C 1-6 Alkyl, optionally substituted C 3-10 Cycloalkyl, optionally substituted C 6-10 It is selected from the group consisting of aryl, optionally substituted 4- to 10-membered heterocyclyl, and optionally substituted 5- to 10-membered heteroaryl.

[0131] In some embodiments, the first end comprises a polyamide having a structure of Formula (A-2), or a pharmaceutically acceptable salt thereof:

[0132] [ka] Formula (A-2) During the ceremony, m1 is 1 to 4, n1 is 0 to 2, Each Y 1 , Y 2 , Y 3 , and Y 4 are independently CH or N; each Z 1 , Z 2 , Z 3 , and Z 4 are independently O, S, or NR 1D and Each L 3 is an optionally substituted C1-C6 alkylene, C3-C7 cycloalkylene, 3- to 7-membered heterocyclene, or 5- to 6-membered heteroarylene; Each R 30 is hydrogen or C1-C6 alkyl, or Each R 30 and L 3 are bonded together with the atoms to which they are attached to form a 4- to 7-membered heterocyclic ring, W1 is hydrogen, optionally substituted C1-C6 alkyl, -NR 1E -C(O)-NR 1E R 1F , -C(O)-NR1E R 1F , or (AA) 1-10 and W2 is hydrogen, optionally substituted C1-C6 alkyl, -C(O)-NR 1E R 1F , or (AA) 1-10 and Each R 1D and R 1E are independently hydrogen, optionally substituted C-C 50 Alkyl, C1-C 50 Heteroalkyl, or PEG 1-50 and R 1F is hydrogen, optionally substituted C1-C 20 Alkyl, C1-C 20 Heteroalkyl, PEG 1-20 , or one or more AA, Each AA is independently a naturally occurring amino acid.

[0133] In some embodiments, each L 3 is an optionally substituted C1-C6 alkylene. In some embodiments, L 3 is a C2, C3, C4, or C5 alkylene optionally substituted with one or more hydrogen, halogen, C1-C6 alkyl, C1-C6 heteroalkyl, C1-C6 haloalkyl, C1-C6 hydroxyalkyl, C3-C6 cycloalkyl, or a 4- to 7-membered heterocycloalkyl ring. 3 is a C2 or C3 alkylene optionally substituted with one or more hydrogen, halogen, C1-C6 alkyl, C1-C6 heteroalkyl, C3-C6 cycloalkyl, or 4-7 membered heterocycloalkyl ring. 3 is a C2 alkylene optionally substituted with one or two hydrogens, C1-C6 alkyl, C1-C6 heteroalkyl, C3-C6 cycloalkyl, or a 4- to 7-membered heterocycloalkyl ring.

[0134] In some embodiments, each L 3is independently C-C cycloalkylene. In some embodiments, L 3 is a cyclobutylene, cyclopentylene ring, cyclohexylene, or cycloheptylene ring. 3 is cyclobutylene. In some embodiments, L 3 is cyclopentylene. In some embodiments, L 3 is cyclohexylene.

[0135] In some embodiments, each L 3 is a 3- to 7-membered heterocyclene. In some embodiments, L 3 is a 4-, 5-, or 6-membered heterocyclene.

[0136] In some embodiments, each R 30 is independently hydrogen. In some embodiments, each R 30 is independently C1-C6 alkyl.

[0137] In some embodiments, L 3 and R 30 are joined together with the atoms to which they are attached to form a 4- to 7-membered heterocyclic ring. In some embodiments, the ring is a 4-membered heterocyclic ring. In some embodiments, the ring is a 5-membered heterocyclic ring. In some embodiments, the ring is a 6-membered heterocyclic ring. In some embodiments, the ring is a 7-membered heterocyclic ring.

[0138] In some embodiments, the first end comprises a polyamide having a structure of Formula (A-3), or a pharmaceutically acceptable salt thereof:

[0139] [ka] Formula (A-3) During the ceremony, m1 is 1 to 4, n1 is 0 to 2, Each Y 1 , Y 2 , Y 3, and Y 4 are independently CH or N; each Z 1 , Z 2 , Z 3 , and Z 4 are independently O, S, or NR 1D and W1 is hydrogen, optionally substituted C1-C6 alkyl, -NR 1E -C(O)-NR 1E R 1F , -C(O)-NR 1E R 1F , or (AA) 1-10 and W2 is hydrogen, optionally substituted C1-C6 alkyl, -C(O)-NR 1E R 1F , or (AA) 1-10 and Each R 1D and R 1E are independently hydrogen, optionally substituted C-C 50 Alkyl, C1-C 50 Heteroalkyl, or PEG 1-50 and R 1F is hydrogen, optionally substituted C1-C 20 Alkyl, C1-C 20 Heteroalkyl, PEG 1-20 , or one or more AA, Each AA is independently an amino acid residue selected from β-alanine, lysine, and arginine.

[0140] In some embodiments, the linker moiety is connected to the DNA-binding moiety (i.e., polyamide) at W2. In some embodiments, W2 is optionally substituted C1-C6 alkyl, -C(O)-NR 1E R 1F , or (AA) 1-10 In some embodiments, W 2 is hydrogen.

[0141] In some embodiments, W 2 is (AA) 1-10In some embodiments, AA is β-alanine. In some embodiments, AA is one β-alanine. In some embodiments, AA is two β-alanines.

[0142] In some embodiments, the first end is represented by formula (A-4):

[0143] [ka] Formula (A-4) or a pharmaceutically acceptable salt thereof.

[0144] In some embodiments, each R 1D and R 1E are independently hydrogen, optionally substituted C-C 20 Alkyl, C1-C 20 Heteroalkyl, or PEG 1-20 In some embodiments, each R 1D and R 1E are independently hydrogen, optionally substituted C-C 10 Alkyl, C1-C 10 Heteroalkyl, or PEG 1-20 is.

[0145] In some embodiments, each R 1D are independently optionally substituted C-C 20 Alkyl, C1-C 20 Heteroalkyl, or PEG 1-20 and each of which is optionally substituted with amido, alkyl, alkynyl, azido, amino, halogen, haloalkyl, hydroxy, nitro, oxo (=O), phosphorus hydroxide, or PEG. In some embodiments, each R 1D are optionally substituted C-C groups optionally substituted with -CN, -NH, -N, -OH, CF, -OP(=O)(OH), -OP(=O)(OCH), -OP(=O)(OCH)(OH), or -OP(=O)OH; 20 In some embodiments, each R1D is independently PEG 1-50 In some embodiments, each R 1D are independently -C(O)-NR 2A R 2B or -NR 2A R 2B and each R 2A and R 2B are independently hydrogen, C1-C 50 Alkyl, or PEG 1-50 is.

[0146] In some embodiments, each Z 1 , Z 2 , Z 3 , and Z 4 are independently O or S.

[0147] In some embodiments, each Z 1 , Z 2 , Z 3 , and Z 4 is independently, NR 1D and R 1D is an optionally substituted C1-C 20 Alkyl or C1-C 20 It is heteroalkyl.

[0148] In some embodiments, each Z 1 , Z 2 , Z 3 , and Z 4 are independently NCH3.

[0149] In some embodiments, each Z 1 , Z 2 , Z 3 , and Z 4 are independently NH.

[0150] In some embodiments, the first end is represented by formula (A-5):

[0151] [ka] Formula (A-5) or a pharmaceutically acceptable salt thereof.

[0152] In some embodiments, each Y 1 and Y 3 is N, and each Y 2 and Y 4 is CH or N. In some embodiments, each Y 2 and Y 4 is independently CH. In some embodiments, each Y 2 and Y 4 is independently N. In some embodiments, Y 2 is CH and Y 4 is N. In some embodiments, Y 2 is N and Y 4 is CH.

[0153] In some embodiments, each unit m1 and n1 is different or the same. In some embodiments, each unit m1 is different. In some embodiments, each unit m1 is the same. In some embodiments, each unit n1 is different. In some embodiments, each unit n1 is the same.

[0154] In some embodiments, m1 is 2 or 3 and n1 is 0 or 1.

[0155] In some embodiments, m1 is 2. In some embodiments, m1 is 1.

[0156] In some embodiments, n1 is 0. In some embodiments, n1 is 1.

[0157] In some embodiments, the linker moiety is connected to the DNA-binding moiety via W. In some embodiments, W is optionally substituted C-C alkyl, or —C(O)—NR 1E R 1F In some embodiments, W is —C(O)—NR 1E R 1F and R1E is hydrogen and R 1F is hydrogen, optionally substituted C1-C 10 Alkyl, or PEG 1-20 is.

[0158] In some embodiments, W1 is hydrogen.

[0159] In some embodiments, the first end comprises a polyamide having a structure of Formula (A-6):

[0160] [ka] Formula (A-6) During the ceremony, Each R 1H , R 1J , R 1K , and R 1L are independently hydrogen, halogen, C1-C6 alkyl, C1-C6 heteroalkyl, C1-C6 haloalkyl, or C1-C6 hydroxyalkyl; R 1H and R 1J or R 1L and R 1K are joined together with the atoms to which they are attached to form a C3-C6 cycloalkyl or 4- to 7-membered heterocycloalkyl ring.

[0161] In some embodiments, each R 1H , R 1J , R 1K , and R 1L is independently hydrogen, halogen, C-C alkyl, C-C heteroalkyl, C-C haloalkyl, or C-C hydroxyalkyl. 1H , R 1J , R 1K , and R 1L is independently hydrogen, halogen, or C-C alkyl. In some embodiments, each R 1H , R 1J , R 1K , and R1L is independently halogen. In some embodiments, each R 1H , R 1J , R 1K , and R 1L is independently C-C alkyl. In some embodiments, each R 1H , R 1J , R 1K , and R 1L are independently hydrogen.

[0162] In some embodiments, R 1H and R 1J or R 1L and R 1K are joined together with the atoms to which they are attached to form a C3-C6 cycloalkyl or 4- to 7-membered heterocycloalkyl ring. In some embodiments, R 1H and R 1J or R 1L and R 1K are joined together with the atoms to which they are attached to form a C-C cycloalkyl. In some embodiments, R 1H and R 1J or R 1L and R 1K are joined together with the atoms to which they are attached to form a 4- to 7-membered heterocycloalkyl ring.

[0163] In some embodiments, the first end comprises a polyamide having a structure of Formula (A-7):

[0164] [ka] Formula (A-7) In the formula, each v1 and v2 is independently 1 to 3.

[0165] In some embodiments, each v1 is independently 1. In some embodiments, each v1 is independently 2. In some embodiments, each v1 is independently 3. In some embodiments, each v2 is independently 1. In some embodiments, each v2 is independently 2. In some embodiments, each v2 is independently 3.

[0166] In some embodiments, the first end is represented by formula (A-8):

[0167] [ka] Formula (A-8) or a pharmaceutically acceptable salt thereof.

[0168] In some embodiments, the first end is represented by formula (A-9):

[0169] [ka] Formula (A-9) or a pharmaceutically acceptable salt thereof.

[0170] In some embodiments, the first end is represented by formula (A-10):

[0171] [ka] Formula (A-10) or a pharmaceutically acceptable salt thereof.

[0172] In some embodiments, the first end comprises a polyamide having a structure of Formula (A-11), or a pharmaceutically acceptable salt thereof:

[0173] [ka] Formula (A-11) During the ceremony, Each R 1H , R 1J , R 1K , and R 1L are independently hydrogen, halogen, C1-C6 alkyl, C1-C6 heteroalkyl, C1-C6 haloalkyl, or C1-C6 hydroxyalkyl; R 1H and R 1J or R 1L and R 1K are joined together with the atoms to which they are attached to form a C3-C6 cycloalkyl or 4- to 7-membered heterocycloalkyl ring; R 1H and R 1J or R 1L and R 1K are joined together with the atoms to which they are attached to form a C3-C6 cycloalkyl or 4- to 7-membered heterocycloalkyl ring.

[0174] In some embodiments, the first end comprises a polyamide having a structure of formula (A-12), or a pharmaceutically acceptable salt thereof:

[0175] [ka] Formula (A-12) During the ceremony, Each A 1 is -NH- or -NH-(CH2) m -CH2-C(O)-NH-, Each M is an optionally substituted C 6-10 an arylene group, an optionally substituted 4- to 10-membered heterocyclene, an optionally substituted 5- to 10-membered heteroarylene group, or an optionally substituted alkylene; m is an integer from 1 to 10, n is an integer of 1 to 6.

[0176] In some embodiments, the formula (A-6) [A 1 -M 1 ] for each M 1 is C 6-10an arylene group, a 4- to 10-membered heterocyclene, an optionally substituted 5- to 10-membered heteroarylene group, or C 1-6 alkylene, each of which is H, OH, halogen, or C 1-10 Alkyl, NO2, CN, NR′R″, C 1-6 Haloalkyl, -C 1-6 Alkoxyl, C 1-6 Haloalkoxy, (C 1-6 Alkoxy)C 1-6 Alkyl, C 2-10 Alkenyl, C 2-10 Alkynyl, C 3-7 Carbocyclyl, 4- to 10-membered heterocyclyl, 4- to 10-membered heterocyclyl, C 6-10 Aryl, 5-10 membered heteroaryl, -(C 3-7 Carbocyclyl)C 1-6 Alkyl, (4-10 membered heterocyclyl)C 1-6 Alkyl, (C 6-10 Aryl)C 1-6 Alkyl, (C 6-10 Aryl)C 1-6 Alkoxy, (5-10 membered heteroaryl)C 1-6 Alkyl, -(C 3-7 (carbocyclyl)-amines, (4- to 10-membered heterocyclyl)amines, (C 6-10 (aryl)amine, (5-10 membered heteroaryl)amine, acyl, C-carboxy, O-carboxy, C-amido, N-amido, S-sulfonamido, N-sulfonamido, -SR ’ , COOH, or CONR′R″, wherein each R′ and R″ is independently H, C 1-10 Alkyl, C 1-10 Haloalkyl, -C 1-10 In some embodiments, the [A 1 -R 1 ] each R 1 is a 5- to 10-membered heteroarylene containing at least one heteroatom selected from O, S, and N, or C 1-6 alkylene, heteroarylene or C 1-6 Alkylene is a group that contains OH, halogen, and C 1-10Alkyl, NO2, CN, NR′R″, C 1-6 Haloalkyl, -C 1-6 Alkoxyl, C 1-6 Haloalkoxy, C 3-7 Carbocyclyl, 4-10 membered heterocyclyl, C 6-10 Aryl, 5-10 membered heteroaryl, -SR ’ , COOH, or CONR′R″, wherein each R′ and R″ is independently H, C 1-10 Alkyl, C 1-10 Haloalkyl, -C 1-10 In some embodiments, the [A 1 -R 1 ] each R 1 is a 5- to 10-membered heteroarylene containing at least one heteroatom selected from O, S, and N, and the heteroarylene is selected from OH, C 1-6 Alkyl, halogen, and C 1-6 It is optionally substituted with 1 to 3 substituents selected from alkoxyl.

[0177] The DNA recognition, binding moiety, or polyamide may be

[0178] [ka]

[0179] [ka] -NH-benzopyrazinylene-CO-, -NH-phenylene-CO-, -NH-pyridinylene-CO-, -NH-piperidinylene-CO-, -NH-pyrimidinylene-CO-, -NH-anthracenylene-CO-, -NH-quinolinylene-CO-, and

[0180] [ka] wherein Z is H, NH, C 1-6Alkyl or C 1-6 It is alkylNH2.

[0181] In some embodiments, Py is

[0182] [ka] and Im is

[0183] [ka] and Hp is

[0184] [ka] and Th is

[0185] [ka] and Pz is

[0186] [ka] and Nt is

[0187] [ka] and Tn is

[0188] [ka] and Nh is

[0189] [ka] and iNt is

[0190] [ka] and iIm is

[0191] [ka] and HpBi is

[0192] [ka] and ImBi is

[0193] [ka] and PyBi is

[0194] [ka] and Dp is

[0195] [ka] and -NH-benzopyrazinylene-CO- is

[0196] [ka] and -NH-phenylene-CO- is

[0197] [ka] and -NH-pyridinylene-CO- is

[0198] [ka] and -NH-piperidinylene-CO- is

[0199] [ka] and -NH-pyrazinylene-CO- is

[0200] [ka] and -NH-anthracenylene-CO- is

[0201] [ka] and -NH-quinolinylene-CO- is

[0202] [ka] is.

[0203] In some embodiments, the first terminus comprises one or more subunits selected from the group consisting of optionally substituted N-methylpyrrole, optionally substituted N-methylimidazole, and β-alanine.

[0204] The first end of the molecules described herein has a high binding affinity for sequences having multiple repeats of GAA and binds to the target nucleotide repeat preferentially over other nucleotide repeats or nucleotide sequences. In some embodiments, the first end has a higher binding affinity for sequences having multiple repeats of GAA than for sequences having CGG repeats. In some embodiments, the first end has a higher binding affinity for sequences having multiple repeats of GAA than for sequences having CCG repeats. In some embodiments, the first end has a higher binding affinity for sequences having multiple repeats of GAA than for sequences having CCTG repeats. In some embodiments, the first end has a higher binding affinity for sequences having multiple repeats of GAA than for sequences having TGGAA repeats. In some embodiments, the first end has a higher binding affinity for sequences having multiple repeats of GAA than for sequences having GGGGCC repeats. In some embodiments, the first end has a higher binding affinity for sequences having multiple repeats of GAA than for sequences having CAG repeats. In some embodiments, the first end has a higher binding affinity for sequences with multiple repeats of GAA than for sequences with repeats of CTG.

[0205] Preferential binding between the first end and the target nucleotide repeat causes the transcription modulator molecules described herein to localize around regions with multiple repeats of GAA. In some embodiments, the localized concentration of a first end or molecule described herein is higher near a sequence with multiple repeats of GAA than near a sequence with CGG repeats. In some embodiments, the localized concentration of a first end or molecule described herein is higher near a sequence with multiple repeats of GAA than near a sequence with CCG repeats. In some embodiments, the localized concentration of a first end or molecule described herein is higher near a sequence with multiple repeats of GAA than near a sequence with CCTG repeats. In some embodiments, the localized concentration of a first end or molecule described herein is higher near a sequence with multiple repeats of GAA than near a sequence with TGGAA repeats. In some embodiments, the localized concentration of a first end or molecule described herein is higher near a sequence with multiple repeats of GAA than near a sequence with GGGGCC repeats. In some embodiments, the localized concentration of a first terminus or molecule described herein is higher near a sequence having multiple repeats of GAA than near a sequence having repeats of CTG, hi some embodiments, the localized concentration of a first terminus or molecule described herein is higher near a sequence having multiple repeats of GAA than near a sequence having repeats of CAG.

[0206] The first end is localized to a sequence having multiple repeats of GAA and binds preferentially to the target nucleotide repeat over other nucleotide repeats. In some embodiments, the sequence has at least 2, 3, 4, 5, 8, 10, 12, 15, 20, 25, 30, 40, 50, 100, 200, 300, 400, or 500 repeats of GAA. In certain embodiments, the sequence comprises at least 1,000 nucleotide repeats of GAA. In certain embodiments, the sequence comprises at least 500 nucleotide repeats of GAA. In certain embodiments, the sequence comprises at least 200 nucleotide repeats of GAA. In certain embodiments, the sequence comprises at least 100 nucleotide repeats of GAA. In certain embodiments, the sequence comprises at least 50 nucleotide repeats of GAA. In certain embodiments, the sequence comprises at least 20 nucleotide repeats of GAA.

[0207] In one aspect, compounds of the present disclosure are capable of binding to the repeat GAA of fxn in preference to GAA elsewhere in the DNA of a subject.

[0208] Polyamides composed of preselected combinations of subunits can selectively bind to DNA in the minor groove. In their hairpin structure, antiparallel juxtapositions of two aromatic amino acids bind to the DNA sequence, with the polyamide rings specifically packed for each DNA base. N-methylpyrrole (Py) prefers T, A, and C bases, excluding G; N-methylimidazole (Im) is a G-reading factor; and 3-hydroxyl-N-methylpyrrole (Hp) is specific for thymine bases. Nucleotide base pairs can be recognized using different pairings of amino acid subunits, using the pairing principles shown in Tables 1A and 1B below. For example, Im / Py pairing symmetry reads G·C, Py / Im pairing reads C·G, Hp / Py pairing can distinguish T·A from A·T, G·C, and C·G, and Py / Py pairing nonspecifically distinguishes both A·T and T·A from G·C and C·G.

[0209] In some embodiments, the first end comprises Im corresponding to the nucleotide G, Py or beta corresponding to the nucleotide A, Py corresponding to the nucleotide A, where Im is N-alkylimidazole, Py is N-alkylpyrrole, and beta is β-alanine. In some embodiments, the first end comprises Im / Py corresponding to the nucleotide pair G / C, Py / beta or Py / Py corresponding to the nucleotide pair A / T, where Im is N-alkylimidazole (e.g., N-methylimidazole), Py is N-alkylpyrrole (e.g., N-methylpyrrole), and beta is β-alanine.

[0210] [Table 1-1]

[0211] [Table 1-2]

[0212] [Table 1-3]

[0213] [Table 2-1]

[0214] [Table 2-2]

[0215] The monomer subunits of the polyamide can be linked together based on the pairing principles shown in Tables 1A and 1B. The monomer subunits of the polyamide can be linked together based on the pairing principles shown in Tables 1C and 1D.

[0216] Table 1C shows examples of monomer subunits that can be linked to specific nucleotides. The first end can include a polyamide, where several monomer subunits are linked together, with the monomer subunits selected from each column. For example, a polyamide can include Im-β-Py linked to GAA, where Im is selected from the first G column, β is selected from the A column, and Py is selected from the second A column. The polyamide can be any combination of subunits selected from each column of Table 1C that link to the GAA subunits, with the subunits linked together in GAA order.

[0217] Additionally, polyamides can also contain partial or multiple sets of five subunits, for example, 1.5, 2, 2.5, 3, 3.5, or 4 sets of three subunits. Polyamides can contain 3, 4, 5, 6, 7, 8, 9, 10, 12, 14, and 16 monomer subunits. Multiple sets can be linked together by W. In addition to five or ten subunits, polyamides can also contain 1 to 4 additional subunits that can link multiple sets of five subunits.

[0218] A polyamide can include monomer subunits that link 2, 3, 4, or 5 nucleotides of GAA. For example, a polyamide can link GA, AA, GAA, AAG, AGA, GAAG, AAGA, GAAGA, or GAAGAA. A polyamide can include monomer subunits that link 3, 4, 5, 6, 7, 8, 9, or 10 nucleotides of GAA repeats. The nucleotides can be linked by W.

[0219] When a monomer subunit is positioned as a terminal unit, it does not have a terminal amine group, carbonyl group, or carboxylic acid group. The terminal amine or carboxylic acid group is replaced with a hydrogen atom. For example, when Py is used as a terminal unit,

[0220] [ka] (for example,

[0221] [ka] ) and when Im is located as a terminal unit,

[0222] [ka] (for example,

[0223] [ka] In addition, when Py or Im is used as a terminal, Py and Im are each represented by the formula: PyT

[0224] [ka] (for example,

[0225] [ka] ) and ImT

[0226] [ka] (for example,

[0227] [ka] ) can be replaced with

[0228] Linear polyamides can have non-limiting examples including, but not limited to, β-Py-Im, Im-Py-β-Im-Py-β-Im-Py, Im-Py-β-Im-Py-Py-Im-β, Im-Py-Py-Im-Py-β-Im-β, and any combination thereof.

[0229] [Table 3]

[0230] The DNA-binding moiety can also include a hairpin polyamide having subunits that link together based on the pairing principle shown in Table 1B. Table 1D shows some examples of monomer subunit pairs that selectively bind to nucleotide pairs. The hairpin polyamide can include 2n monomer subunits, where n is an integer ranging from 2 to 8, and the polyamide further includes a W at the center of the 2n monomer subunits. W is -(CH2) a -NR 1 -(CH2) b -, -(CH2) a -, -(CH2) a -O-(CH2) b -, -(CH2) a -CH(NHR 1 )-, -(CH2)a-CH(NHR 1 )-, -(CR 2 R 3 ) a - or -(CH2) a -CH(NR 1 3) + -(CH2) b wherein each a is independently an integer from 2 to 4; 1 is H, optionally substituted C 1-6 Alkyl, optionally substituted C 3-10 Cycloalkyl, optionally substituted C 6-10 aryl, optionally substituted 4- to 10-membered heterocyclyl, or optionally substituted 5- to 10-membered heteroaryl, and each R 2 and R 3 are independently H, halogen, OH, NHAc, or C1-4 In some embodiments, W is —(CH)—CH(NH) + -(CH2)- or -(CH2)-CH2CH(NH3) + In some embodiments, R 1 is H. In some embodiments, R 1 is a C optionally substituted by 1 to 3 substituents selected from -C(O)-phenyl 1-6 In some embodiments, W is -(CR 2 R 3 )-(CH2)a- or -(CH2) a -(CR 2 R 3 )-(CH2) b wherein each a is independently 1 to 3, b is 0 to 3, and each R 2 and R 3 are independently H, halogen, OH, NHAc, or C 1-4 W is an alkyl. W can be an aliphatic amino acid residue shown in Table 4, such as gAB.

[0231] Because the target gene can contain multiple repeats of GAA, the subunits can be linked together to bind to at least 2, 3, 4, 5, 6, 7, 8, 9, or 10 nucleotides in one or more GAA repeats (e.g., GAAGAAGAAGAA). For example, the polyamide can bind to the GAA repeat by binding to a partial copy, a full copy, or multiple repeats of GAA, such as GA, AA, GAA, AAG, AGA, GAAG, AAGA, GAAGA, or GAAGAA. For example, the polyamide can include Im-Py-β-W-Py-β-Py that binds to GAA and its complementary nucleotide on double-stranded DNA, where the Im / Py pair binds to G·C, the Py / β pair binds to A·T, and the β / Py pair binds to G·A. In another example, Im-Py-β-Im-W-β-Py-β-Py binds to GAAG and its complementary nucleotide on double-stranded DNA, where the Im / Py pair binds to G·C, the Py / β pair binds to A·T, the β / Py pair binds to G·A, and the Im / β pair binds to G·C. W can be an aliphatic amino acid residue such as gAB or other suitable spacer, as shown in Table 4. In another example, Im-Py-β-Im-gAB-Im-Py binds to a portion of the complementary nucleotide (ACG) on double-stranded DNA, where Im binds to G and Py binds to A. □ / Py binds to A·T, and Im / Im binds to G·C.

[0232] Additional examples of polyamides include, but are not limited to, Im-Py-Py-Im-gAB-Py-Im-Im-Py, Im-Py-Py-Im-gAB-Py-Im-Im-PyT, Im-Py-Py-Im-gAB-Py-Im-Im-β, Im-Py-Py-Im-gAB-Py-Im-Im-β-G, Im-β-β-Py-Im-gAB-Py-Im-Im-β, Im-β-Py-Im-gAB-Py-Im-Im-β-G, Im-β-Py-Im-gAB-Py-Im-Im-Py, Im-β-Py-Im-gAB-Py-Im-Im-PyT, Py-Py-Im-β-gAB-Im-Py-Im-Im, Py-Py-Im-β-gAB-Im-Py-Im-ImT, Py-Py-Im-Py-gAB-Im-Py-Im-Im, Py-Py-Im-Py-gAB-Im-Py-Im-ImT, Py-Py-Im-β-gAB-Im-β-Im-Im, Py-Py-Im-β-gAB-Im-β-Im-ImT, Py-Py-Im-Py-gAB-Im-β-Im-Im, Py-Py-Im-Py-gAB-Im-β-Im-ImT, Im-β-Py-gAB-Im-Im-Py, Im-β-Py-gAB-Im-Im-PyT, Im-β-Py-gAB-Im-Im-β, Im-β-Py-gAB-Im-Im-β-G, Im-Py-Py-gAB-Im-Im-β, Im-Py-Py-gAB-Im-Im-β-G, Im-Py-Py-gAB-Im-Im-Py, Im-Py-Py-gAB-Im-Im-PyT, Im-p-Py-gAB-Im-Im-Py, and Im-β-Py-gAB-Im-Im-PyT, where G can be hydrogen, alkyl, alkenyl, alkynyl, or -C(O)-R B can be, and R B can be hydrogen, a C1-C6 alkyl, C1-C6 alkenyl, or C1-C6 alkynyl group. In some embodiments, the hairpin polyamide has the structure of Im-Py-β-Im-gAB-Im-Py, Im-Py-β-Im-gAB-Im-Py-β-Im, Py-β-Im-gAB-Im-Py-β-Im, or β-Im-gAB-Im-Py-β-Im.

[0233]

Table 4

[0234] Recognition of nucleotide repeats or DNA sequences by two antiparallel polyamide strands relies on the encoding of parallel aromatic amino acid pairs in the minor groove, typically oriented N to C relative to the 5' to 3' direction of the DNA helix. Enhanced affinity and specificity of polyamide-nucleotide binding is achieved by covalently linking the antiparallel strands. A "hairpin motif" connects the N- and C-termini of the two strands with W (e.g., gamma-aminobutyric acid units (gamma turns)) to form a folded linear chain. An "H-pin motif" connects the antiparallel strands across a central or near-central ring / ring pair by a short, flexible bridge.

[0235] The DNA-binding moiety can also include H-pin polyamides, which have subunits that link together based on the pairing principles shown in Table 1A and / or Table 1B. Table 1C shows some examples of monomer subunits that selectively bind to nucleotides, and Table 1D shows some examples of monomer subunit pairs that selectively bind to nucleotide pairs. The h-pin polyamide can include two strands, each of which can have several monomer subunits (each strand can contain 2 to 8 monomer subunits). The polyamide also includes a bridge L1 at or near the center of each strand to connect the two strands. At least one or two of the monomer subunits on each strand pair with the corresponding monomer subunit on the other strand according to the pairing principles of Table 1D, preferring binding of either G·C or C·G, A·T, or T·A pairs. These monomer subunit pairs are often located in the center, near the central region, or at or near the bridge connecting the two strands. In some cases, the H-pin polyamide can have all of the monomer subunits pair with corresponding monomer subunits on the antiparallel strand based on the pairing principles of Tables 1B and 1D to bind to nucleotide pairs on double-stranded DNA. In some cases, the H-pin polyamide can have a portion (2, 3, 4, 5, or 6) of the monomer subunits pair with corresponding monomer subunits on the antiparallel strand based on the binding principles of Tables 1B and 1D to bind to nucleotide pairs on double-stranded DNA, while the remainder of the monomer subunits bind to nucleotides based on the binding principles of Tables 1A and 1C but do not pair with monomer subunits on the antiparallel strand. The h-pin polyamide can have one or more overhanging monomer subunits that bind to nucleotides but do not pair with monomer subunits on the antiparallel strand.

[0236] Another polyamide structure derived from the h-pin structure connects two antiparallel strands at the ends via a bridge, and only the two monomer subunits connected by the bridge form pairs that bind with the nucleotide pair G·C or C·G based on the binding principles in Table 1B / 1D, while the remaining monomer subunits on the strand form overhangs that bind with nucleotides based on the binding principles in Table 1A and / or 1C and do not pair with monomer subunits on the other strand.

[0237] The bridge is

[0238] [ka] a C 1-10 Alkylene, -NH-C 0-6 Alkylene -C(O)-, -N(CH3)-C 0-6 alkylene, and

[0239] [ka] , -(CH2) a -NR 1 -(CH2) b -, -(CH2) a -, -(CH2) a -O-(CH2) b -, -(CH2) a -CH(NHR 1 )-, -(CH2) a -CH(NHR 1 )-, -(CR 2 R 3 ) a - or -(CH2) a -CH(NR 1 3) + -(CH2) b -, m is an integer ranging from 0 to 10, n is an integer ranging from 0 to 10, each a is independently an integer from 2 to 4, and R 1 is H, optionally substituted C 1-6 Alkyl, optionally substituted C 3-10 Cycloalkyl, optionally substituted C6-10 aryl, optionally substituted 4- to 10-membered heterocyclyl, or optionally substituted 5- to 10-membered heteroaryl, and each R 2 and R 3 are independently H, halogen, OH, NHAc, or C 1-4 In some embodiments, W is —(CH)—CH(NH) + -(CH2)- or -(CH2)-CH2CH(NH3) + In some embodiments, R 1 is H. In some embodiments, R 1 is a C optionally substituted by 1 to 3 substituents selected from -C(O)-phenyl 1-6 In some embodiments, L is -(CR 2 R 3 )-(CH2) a -or-(CH2) a -(CR 2 R 3 )-(CH2) b wherein each a is independently 1 to 3, b is 0 to 3, and each R 2 and R 3 are independently H, halogen, OH, NHAc, or C 1-4 L1 is C 2-9 Alkylene or (PEG) 2-8 It could be.

[0240] Additional examples of polyamides include, but are not limited to, Im-Py-Py-Im (central bond - either position 2 or 3) to Py-Py-Py-Py, Im-Py-Py-Im (central bond - py and Py at position 3) to Im-Py-β-Py-Py, Im-Py-β-Im (bond at bold position) Im-Py, Im-Py-β-Im (central bond, either position 2 or 3) Im-Py-β-Im, Py-β-Im (bond at bold central position) Im-Py-β-Im, or β-Im (bond at bold position) Im-Py-β-Im.

[0241] Second end - regulatory binding site In certain embodiments, the regulatory molecule is selected from nucleosome remodeling factors ("NURFs"), bromodomain PHD finger transcription factors ("BPTFs"), ten-eleven translocation enzymes ("TETs"), methylcytosine dioxygenases ("TET1"), DNA demethylases, helicases, acetyltransferases, and histone deacetylases ("HDACs").

[0242] In some embodiments, the protein binding moiety is selected from the group consisting of CREB-binding protein (CBP), P300, O-linked β-N-acetylglucosamine transferase (OGT), P300-CBP associated factor (PCAF), histone methyltransferase, histone demethylase, chromodomain, cyclin-dependent kinase-9 (CDK9), nucleosome remodeling factor (NURF), bromodomain PHD finger transcription factor (BPTF), ten-eleven The gene binds to a regulatory molecule selected from the group consisting of translocation enzymes (TET-), methylcytosine dioxygenases (TET1-), histone acetyltransferases (HATs), histone deacetylases (HDACs), host cell factor 1 (HCF1-), octamer-binding transcription factors (OCT1-), P-TEFb-, cyclin-T1-, PRC2-, DNA-demethylases, helicases, acetyltransferases, histone deacetylases, and methylated histone lysine proteins.

[0243] In some embodiments, the second end comprises a moiety that binds to O-linked β-N-acetylglucosamine transferase (OGT) or CREB binding protein (CBP). In some embodiments, the protein-binding moiety is a residue of a compound that binds to O-linked β-N-acetylglucosamine transferase (OGT) or CREB binding protein (CBP).

[0244] In some embodiments, the second end comprises a bromodomain-binding moiety. In some embodiments, the bromodomain-binding moiety is a BRD2, BRD3, BRD4, or BRDT-binding moiety. In some embodiments, the bromodomain-binding moiety is a BRD4-binding moiety.

[0245] In some embodiments, the regulatory molecule is a bromodomain-containing protein selected from BRD2, BRD3, BRD4, and BRDT.

[0246] In some embodiments, the regulatory molecule is BRD4. In certain embodiments, the recruitment moiety is a BRD4 activator.

[0247] In certain embodiments, the regulatory molecule regulates histone rearrangement.

[0248] In certain embodiments, the regulatory molecule modulates glycosylation, phosphorylation, alkylation, or acylation of histones.

[0249] In certain embodiments, the regulatory molecule is a transcription factor.

[0250] In certain embodiments, the regulatory molecule is an RNA polymerase.

[0251] In certain embodiments, the regulatory molecule is a moiety that regulates the activity of an RNA polymerase.

[0252] In certain embodiments, the recruitment moiety binds to a regulatory molecule but does not inhibit the activity of the regulatory molecule. In certain embodiments, the recruitment moiety binds to a regulatory molecule and inhibits the activity of the regulatory molecule. In certain embodiments, the recruitment moiety binds to a regulatory molecule and increases the activity of the regulatory molecule.

[0253] In certain embodiments, the recruitment moiety binds to an active site of a regulatory molecule. In certain embodiments, the recruitment moiety binds to a regulatory site of a regulatory molecule.

[0254] The binding affinity between the regulatory protein and the second end can be adjusted based on the composition of the molecule or the type of protein. In some embodiments, the second end binds to the regulatory molecule with an affinity of less than about 600 nM, about 500 nM, about 400 nM, about 300 nM, about 250 nM, about 200 nM, about 150 nM, about 100 nM, or about 50 nM. In some embodiments, the second end binds to the regulatory molecule with an affinity of less than about 300 nM. In some embodiments, the second end binds to the regulatory molecule with an affinity of less than about 200 nM. In some embodiments, the polyamide can bind to DNA with an affinity of greater than about 200 nM, about 150 nM, about 100 nM, about 50 nM, about 10 nM, or about 1 nM. In some embodiments, the polyamide can bind to DNA with an affinity in the range of about 1-600 nM, 10-500 nM, 20-500 nM, 50-400 nM, 100-300 nM, or 50-200 nM.

[0255] In some embodiments, the second terminus comprises a diazine or diazepine ring, the diazine or diazepine ring comprising one or more heteroatoms selected from S, N, and O. 6-10 The second terminus is fused to an aryl or 5- to 10-membered heteroaryl ring. In some embodiments, the second terminus comprises an optionally substituted bicyclic or tricyclic structure. In some embodiments, the optionally substituted bicyclic or tricyclic structure comprises a diazepine ring fused to a thiophene ring. In some embodiments, the second terminus comprises an optionally substituted bicyclic structure, wherein the bicyclic structure comprises a diazepine ring fused to a thiophene ring.

[0256] In some embodiments, the second end comprises a compound having the structure of Formula (9-A):

[0257] [ka] Formula (9-A) During the ceremony, Ring A is absent or an optionally substituted 6-membered monocyclic aryl or heteroaryl; Y is —NH— or —O—; R 8 is hydrogen or C 1-6 is alkyl, R 9 , R 10 , and R 11 each independently represents hydrogen, optionally substituted C 1-6 Alkyl, C 1-6 Haloalkyl, or C 1-6 hydroxyalkyl; R 12 is hydrogen, halogen, -NO2, -CN, optionally substituted aryl, optionally substituted C 1-20 Alkyl, C 1-20 Heteroalkyl, C 1-6 Haloalkyl or C 1-6 hydroxyalkyl; or R 12 is -NR A R B and R A and R B each independently represents hydrogen, optionally substituted C 1-20 Alkyl or C 1-20 is heteroalkyl, x1 is an integer from 1 to 6.

[0258] In some embodiments, ring A is an optionally substituted 6-membered monocyclic aryl or heteroaryl, each of which is optionally substituted with alkyl, amino, halogen, hydroxy, hydroxyalkyl, or PEG. In some embodiments, ring A is phenyl. In some embodiments, ring A is a 6-membered monocyclic heteroaryl. In some embodiments, ring A is pyridine or pyrimidine.

[0259] In some embodiments, ring A is absent.

[0260] In some embodiments, Y is -NH-. In some embodiments, Y is -O-.

[0261] In some embodiments, R 8 is hydrogen.

[0262] In some embodiments, R 9 , R 10 , and R 11 each independently represents an optionally substituted C 1-6 Alkyl, C 1-6 Haloalkyl, or C 1-6 In some embodiments, R 9 , R 10 , and R 11 each independently represents an optionally substituted C 1-6 In some embodiments, R 9 , R 10 , and R 11 are each independently methyl, ethyl, or propyl. 9 , R 10 , and R 11 are each independently methyl.

[0263] In some embodiments, R 12 is hydrogen, halogen, optionally substituted C 1-6 Alkyl, C 1-6 Haloalkyl, or C 1-6 In some embodiments, R 12 is bromo, chloro, or fluoro.

[0264] In some embodiments, R 12 is -NR A R B and R A and R B each independently represents hydrogen, optionally substituted C 1-6 It is alkyl.

[0265] In some embodiments, x1 is an integer from 1 to 5, 1 to 4, 1 to 3, or 1 to 2. In some embodiments, x1 is 1. In some embodiments, x1 is 2.

[0266] In some embodiments, the second end is represented by formula (9-B):

[0267] [ka] Formula (9-B) or a pharmaceutically acceptable salt thereof.

[0268] In some embodiments, the second end comprises a compound of Formula (10-A), or a pharmaceutically acceptable salt thereof:

[0269] [ka] Formula (10-A) During the ceremony, Ring B is absent or an optionally substituted 5- to 6-membered monocyclic aryl or heteroaryl, or a 4- to 8-membered heterocycle; Y is —NH— or —O—; R 13 is selected from hydrogen or optionally substituted C1-C6 alkyl; R 14 and R 15 each independently represents hydrogen, optionally substituted C 1-6 Alkyl, C 1-6 Haloalkyl, or C 1-6 hydroxyalkyl; R 16 is hydrogen, halogen, -NO2, -CN, optionally substituted aryl, optionally substituted C 1-20 Alkyl, C 1-20 Heteroalkyl, C 1-6 Haloalkyl or C 1-6 hydroxyalkyl; or R 16 is -NR A RB and R A and R B each independently represents hydrogen, optionally substituted C 1-20 Alkyl or C 1-20 is heteroalkyl, x2 is an integer from 1 to 6.

[0270] In some embodiments, Ring B is an optionally substituted 6-membered monocyclic aryl or heteroaryl, each of which is optionally substituted with alkyl, amino, halogen, hydroxy, hydroxyalkyl, or PEG. In some embodiments, Ring B is phenyl. In some embodiments, Ring B is a 6-membered monocyclic heteroaryl. In some embodiments, Ring B is pyridine or pyrimidine.

[0271] In some embodiments, ring B is absent.

[0272] In some embodiments, Y is -NH-. In some embodiments, Y is -O-.

[0273] In some embodiments, R 13 is hydrogen.

[0274] In some embodiments, R 14 and R 15 each independently represents an optionally substituted C 1-6 Alkyl, C 1-6 Haloalkyl, or C 1-6 In some embodiments, R 14 and R 15 each independently represents an optionally substituted C 1-6 In some embodiments, R 14 and R 15 are each independently methyl, ethyl, or propyl. 14 and R 15are each independently methyl.

[0275] In some embodiments, R 16 is hydrogen, halogen, optionally substituted C 1-6 Alkyl, C 1-6 Haloalkyl, or C 1-6 In some embodiments, R 16 is bromo, chloro, or fluoro.

[0276] In some embodiments, R 16 is -NR A R B and R A and R B each independently represents hydrogen, optionally substituted C 1-6 It is alkyl.

[0277] In some embodiments, x2 is an integer from 1 to 5, 1 to 4, 1 to 3, or 1 to 2. In some embodiments, x2 is 1. In some embodiments, x2 is 2.

[0278] In some embodiments, the second end is represented by formula (10-B):

[0279] [ka] Formula (10-B) or a pharmaceutically acceptable salt thereof.

[0280] In some embodiments, the second end comprises a compound having the structure of Formula (11), or a pharmaceutically acceptable salt thereof:

[0281] [ka] Formula (11) During the ceremony, Ring E is absent or an optionally substituted 5- to 6-membered monocyclic aryl or heteroaryl, or a 4- to 8-membered heterocycle; Y is —NH— or —O—; R 17 is hydrogen or C1-C6 alkyl, R 31 is hydrogen or a substituted monocyclic aryl or heteroaryl.

[0282] In some embodiments, the second end comprises a compound having the structure of Formula (11-A), or a pharmaceutically acceptable salt thereof:

[0283] [ka] Formula (11-A) During the ceremony, Ring E is absent or an optionally substituted 5- to 6-membered monocyclic aryl or heteroaryl, or a 4- to 8-membered heterocycle; Y is —NH— or —O—; R 17 is hydrogen or C1-C6 alkyl, R 18 and R 19 are each independently hydrogen, halogen, —CN, —NO, optionally substituted —C1-C6 alkyl, C1-C6 haloalkyl, or C1-C6 hydroxyalkyl; or R 18 is -NR A R B and R A and R B each independently represents hydrogen, optionally substituted C 1-6 Alkyl or C 1-6 is heteroalkyl, R 25 is an arbitrarily substituted C 1-6 Alkyl, C 1-6 Heteroalkyl, C 1-6 Alkenyl, C 1-6 Alkynyl, C 1-6 Hydroxyalkyl, or -NHSO2R A and R 32 is hydrogen or optionally substituted C1-6 is alkyl, y1 is an integer from 1 to 3.

[0284] In some embodiments, Ring E is an optionally substituted 5- or 6-membered monocyclic aryl or heteroaryl, each of which is optionally substituted, each of which is optionally substituted with alkyl, amino, halogen, hydroxy, hydroxyalkyl, or PEG.

[0285] In some embodiments, Ring E is phenyl. In some embodiments, Ring E is a 6-membered heteroaryl. In some embodiments, Ring E is pyridine, pyrazine, or triazine. In some embodiments, Ring E is pyridine. In some embodiments, Ring E is pyrazine. In some embodiments, Ring E is triazine. In some embodiments, Ring E is a 5-membered heteroaryl. In some embodiments, Ring E is pyrazole. In some embodiments, Ring E is triazole, pyrrole, imidazole, oxazole, oxadiazole, thiazole, or thiadiazole. In some embodiments, Ring E is triazole. In some embodiments, Ring E is imidazole or pyrrole. In some embodiments, oxazole or oxadiazole. In some embodiments, Ring E is thiazole or thiadiazole.

[0286] In some embodiments, ring E is absent.

[0287] In some embodiments, R 17 is hydrogen. In some embodiments, R 17 is C1-C6 alkyl. In some embodiments, R 17 is methyl, ethyl, or propyl. In some embodiments, R 17 is methyl.

[0288] In some embodiments, R 18 and R 19are each independently hydrogen, —CN, or —NO. In some embodiments, R 18 and R 19 are each independently halogen or optionally substituted -C1-C6 alkyl. In some embodiments, R 18 and R 19 are each independently bromo, chloro, fluoro, methyl, or ethyl. 18 and R 19 are each independently fluoro or methyl.

[0289] In some embodiments, R 25 is an arbitrarily substituted C 1-6 Alkyl, C 1-6 Heteroalkyl, C 1-6 Alkenyl, C 1-6 Alkynyl, or C 1-6 and hydroxyalkyl, each of which is optionally substituted with amido, alkyl, alkynyl, azido, amino, halogen, haloalkyl, hydroxy, nitro, oxo (=O), phosphorus hydroxide, or PEG.

[0290] In some embodiments, R 25 is an arbitrarily substituted C 1-6 Alkyl, C 1-6 Heteroalkyl, or C 1- In some embodiments, R 25 is C 1-6 Alkyl or C 1-6 and heteroalkyl, each or they are optionally substituted with -CN, -NH2, -N3, -OH, CF3, or -OP(=O)(OH)2.

[0291] In some embodiments, R 25 is -NHSO2R A In some embodiments, R 25 is -NHSOEt. In some embodiments, R 25 is -NHSO2Me.

[0292] In some embodiments, R 32 is optionally substituted with haloalkyl, phosphorus hydroxide, 1-6 In some embodiments, R 32 is C optionally substituted with -OP(=O)(OH)2 1-6 In some embodiments, R 32 is the unsubstituted C 1-6 In some embodiments, R 32 is methyl, ethyl, or tributyl. In some embodiments, R 32 is hydrogen.

[0293] In some embodiments, y1 is 1. In some embodiments, y1 is 2. In some embodiments, y1 is 3.

[0294] In some embodiments, the second end is represented by formula (11-B):

[0295] [ka] Formula (11-B) or a pharmaceutically acceptable salt thereof.

[0296] In some embodiments, the second end is represented by formula (11-C):

[0297] [ka] Formula (11-C) or a pharmaceutically acceptable salt thereof.

[0298] In some embodiments, the second end is represented by formula (11-D):

[0299] [ka] Formula (11-D) or a pharmaceutically acceptable salt thereof.

[0300] In some embodiments, the second end is represented by formula (11-E):

[0301] [ka] Formula (11-E) or a pharmaceutically acceptable salt thereof.

[0302] In some embodiments, the second end is represented by formula (11-F):

[0303] [ka] Formula (11-F) or a pharmaceutically acceptable salt thereof.

[0304] In some embodiments, the second end is represented by formula (11-G):

[0305] [ka] Formula (11-G) or a pharmaceutically acceptable salt thereof.

[0306] In some embodiments, the second end comprises a compound having the structure of Formula (12-A):

[0307] [ka] Formula (12-A) During the ceremony, Each R 20 , R 21 , and R 22 are independently hydrogen, halogen, optionally substituted C1-C6 alkyl, C1-C6 haloalkyl, or C1-C6 hydroxyalkyl; Each z1, z2, and z3 is independently 1 to 4.

[0308] In some embodiments, each R 20 , R 21 , and R 22 is independently an optionally substituted C-C alkyl, C-C haloalkyl, or C-C hydroxyalkyl. In some embodiments, each R 20 , R 21 , and R 22 is independently methyl, ethyl, or propyl. In some embodiments, each R 20 , R 21 , and R 22 is independently halogen. In some embodiments, each R 20 , R 21 , and R 22 are independently hydrogen.

[0309] In some embodiments, each z1, z2, and z3 is independently an integer from 1 to 3 or 1 to 2. In some embodiments, each z1, z2, and z3 is independently 1. In some embodiments, each z1, z2, and z3 is independently 2. In some embodiments, each z1, z2, and z3 is independently 3.

[0310] In some embodiments, the second end is represented by formula (12-B):

[0311] [ka] Formula (12-B) or a pharmaceutically acceptable salt thereof.

[0312] In some embodiments, the second end is a group represented by formula (12-C):

[0313] [ka] Formula (12-C) or a pharmaceutically acceptable salt thereof.

[0314] In some embodiments, the second end comprises a compound having the structure of Formula (13-A):

[0315] [ka] Formula (13-A) During the ceremony, Ring C is absent or an optionally substituted monocyclic 6-membered aryl or heteroaryl; X 1 is CH or N, L 2 is -NR D -or-CR D H- and R 23 is C1-C6 alkyl or C3-C6 cycloalkyl, R 24 is halogen, alkyl, hydroxyalkyl, haloalkyl, optionally substituted C1-C6 alkyl, C1-C6 haloalkyl, or C1-C6 hydroxyalkyl; R D is hydrogen or C 1-3 It is alkyl.

[0316] In some embodiments, Ring C is an optionally substituted 6-membered monocyclic aryl or heteroaryl, each of which is optionally substituted, each of which is optionally substituted with alkyl, amino, halogen, hydroxy, hydroxyalkyl, or PEG. In some embodiments, Ring C is phenyl. In some embodiments, Ring C is a 6-membered monocyclic heteroaryl. In some embodiments, Ring C is pyridine or pyrimidine.

[0317] In some embodiments, ring C is

[0318] [ka] is.

[0319] In some embodiments, ring C is absent.

[0320] In some embodiments, X 1 is CH. In some embodiments, X 1 is N.

[0321] In some embodiments, L 2 is -NR D In some embodiments, L 2 is —NH—. In some embodiments, L 2 -CR D H. In some embodiments, L 2 is -CH2-.

[0322] In some embodiments, R 23 is methyl, ethyl, or propyl. In some embodiments, R 23 is methyl. In some embodiments, R 23 is ethyl. In some embodiments, R 23 is propyl. In some embodiments, R 23 is cyclopropyl.

[0323] In some embodiments, R 24 is alkyl, hydroxyalkyl, haloalkyl, optionally substituted C-C alkyl, C-C haloalkyl, or C-C hydroxyalkyl. In some embodiments, R 24 is hydroxyalkyl. In some embodiments, R 24 is halogen. In some embodiments, R 24 is bromo, chloro, or fluoro.

[0324] In some embodiments, the second end is represented by formula (13-B):

[0325] [ka] Formula (13-B) or a pharmaceutically acceptable salt thereof.

[0326] In some embodiments, the second end is represented by formula (13-C):

[0327] [ka] Formula (13-C) or a pharmaceutically acceptable salt thereof.

[0328] In some embodiments, the second end is

[0329] [ka]

[0330] [ka] or a pharmaceutically acceptable salt thereof.

[0331] In some embodiments, the second end is

[0332] [ka]

[0333] [ka] or a pharmaceutically acceptable salt thereof.

[0334] In some embodiments, the protein binding moiety is

[0335] [ka] or a pharmaceutically acceptable salt thereof.

[0336] Linker The oligomeric backbone includes a linker connecting the first and second ends, bringing the regulatory molecule into proximity with the target gene to regulate gene expression.

[0337] The length of the linker also depends on the type of regulatory protein and the target gene. In some embodiments, the linker has a length of less than about 50 angstroms. In some embodiments, the linker has a length of about 20-30 angstroms.

[0338] In some embodiments, the linker comprises between 5 and 50 chain atoms.

[0339] In some embodiments, the linker comprises a multimer having 2 to 50 spacing moieties, The spacing part can be independently -((CR 3a R 3b ) x -O) y -,-((CR 3a R 3b ) x -NR 4a ) y -,-((CR 3a R 3b ) x -CH=CH-(CR 3a R 3b ) x -O) y -, optionally substituted -C 1-12 Alkyl, optionally substituted C 2-10 Alkenyl, optionally substituted C 2-10 Alkynyl, optionally substituted C 6-10 Arylene, optionally substituted C 3-7 Cycloalkylene, optionally substituted 5- to 10-membered heteroarylene, optionally substituted 4- to 10-membered heterocycloalkylene, amino acid residue, -O-, -C(O)NR 4a -, -NR 4a C(O)-, -C(O)-, -NR1 -, -C(O)O-, -O-, -S-, -S(O)-, -SO2-, -SO2NR 4a -, -NR 4a selected from the group consisting of SO2-, and -P(O)OH-, and any combination thereof; each x is independently 2 to 4; each y is independently 1 to 10; Each R 3a and R 3b are independently selected from hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted alkoxy, optionally substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acylamino, aminoacyl, optionally substituted alkylamido, sulfonyl, optionally substituted thioalkoxy, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted cycloalkyl, and optionally substituted heterocyclyl; Each R 4a are independently hydrogen or optionally substituted C 1-6 It is alkyl.

[0340] In some embodiments, the oligomer backbone is -(T 1 -V 1 ) a -(T 2 -V 2 ) b -(T 3 -V 3 ) c -(T 4 -V 4 ) d -(T 5 -V 5 ) e -including In the formula, a, b, c, d, and e each independently represent 0 or 1, and the sum of a, b, c, d, and e is 1 to 5; T 1 , T 2 , T 3 , T 4 , and T 5 are each independently optionally substituted (C-C 12) alkylene, optionally substituted alkenylene, optionally substituted alkynylene, (EA) w , (EDA) m , (PEG) n , (modified PEG) n , (AA) p , -(CR 2a OH) h -, optionally substituted (C6-C 10 ) arylene, optionally substituted C 3-7 selected from cycloalkylene, optionally substituted 5- to 10-membered heteroarylene, optionally substituted 4- to 10-membered heterocycloalkylene, acetal groups, disulfides, hydrazines, carbohydrates, beta-lactams, and esters; w is an integer from 1 to 20, m is an integer from 1 to 20; n is an integer from 1 to 30, p is an integer from 1 to 20; h is an integer from 1 to 12; The EA has the following structure:

[0341] [ka] EDA has the following structure:

[0342] [ka] In the formula, each q is independently an integer of 1 to 6, each x is independently an integer of 1 to 4, and each r is independently 0 or 1; (PEG) n is -(CR 2a R 2b -CR 2a R 2b -O) n -CR 2a R 2b -having the structure (Modified PEG) n is (PEG) n At least one of -(CR 2a R 2b -CR 2aR 2b -O)-(CH2-CR 2a =CR 2a -CH2-O)- or -(CR 2a R 2b -CR 2a R 2b -S)-, AA is an amino acid residue, V 1 , V 2 , V 3 , V 4 , and V 5 each independently represents a bond, CO—, or —NR 1a -,-CONR 1a -, -NR 1a CO-, -CONR 1a C 1-4 Alkyl-, -NR 1a CO-C 1-4 Alkyl-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR 1a -, -NR 1a selected from the group consisting of SO2-, and -P(O)OH-; Each R 1a are independently hydrogen or optionally substituted C 1-6 is alkyl, Each R 2a and R 2b is independently selected from hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, halogen, alkoxy, substituted alkoxy, amino, substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acylamino, aminoacyl, alkylamido, substituted alkylamido, sulfonyl, thioalkoxy, substituted thioalkoxy, aryl, substituted aryl, heteroaryl, substituted heteroaryl, cycloalkyl, substituted cycloalkyl, heterocyclyl, and substituted heterocyclyl.

[0343] In some embodiments, a, b, c, d, and e are each independently 0 or 1, and the sum of a, b, c, d, and e is 1. In some embodiments, a, b, c, d, and e are each independently 0 or 1, and the sum of a, b, c, d, and e is 2. In some embodiments, a, b, c, d, and e are each independently 0 or 1, and the sum of a, b, c, d, and e is 3. In some embodiments, a, b, c, d, and e are each independently 0 or 1, and the sum of a, b, c, d, and e is 4. In some embodiments, a, b, c, d, and e are each independently 0 or 1, and the sum of a, b, c, d, and e is 5.

[0344] In some embodiments, n is 3 to 9. In some embodiments, n is 4 to 8. In some embodiments, n is 5 or 6.

[0345] In some embodiments, T 1 , T 2 , T 3 , and T 4 , and T 5 are each independently (C1-C 12 ) alkyl, substituted (C1-C 12 ) alkyl, (EA) w , (EDA) m , (PEG) n , (modified PEG) n , (AA) p , -(CR 2a OH) h -, phenyl, substituted phenyl, piperidine-4-amino (P4A), para-amino-benzyloxycarbonyl (PABC), meta-amino-benzyloxycarbonyl (MABC), para-amino-benzyloxy (PABO), meta-amino-benzyloxy (MABO), para-aminobenzyl, acetal group, disulfide, hydrazine, carbohydrate, beta-lactam, ester, (AA) p -MABC-(AA) p , (AA) p -MABO-(AA) p , (AA) p-PABO-(AA) p , and (AA) p -PABC-(AA) p In some embodiments, piperidine-4-amino (P4A) is selected from:

[0346] [ka] where R 1a is H or C 1-6 It is alkyl.

[0347] In some embodiments, T 1 , T 2 , T 3 , T 4 , and T 5 are each independently (C1-C 12 ) alkyl, substituted (C1-C 12 ) alkyl, (EA) w , (EDA) m , (PEG) n , (modified PEG) n , (AA) p , -(CR 2a OH) h -, optionally substituted (C6-C 10 ) arylene, 4- to 10-membered heterocycloalkene, and optionally substituted 5- to 10-membered heteroarylene. In some embodiments, EA has the structure:

[0348] [ka] is selected from EDA has the following structure:

[0349] [ka]

[0350] In some embodiments, for EA and EDA, x is 2 to 3 and q is 1 to 3. In some embodiments, R 1a is H or C1-6 It is alkyl.

[0351] In some embodiments, T 4 or T 5 is optionally substituted (C6-C 10 ) arylene.

[0352] In some embodiments, T 4 or T 5 is phenylene or substituted phenylene. In some embodiments, T 4 or T 5 is phenylene or -C 1-6 In some embodiments, T is phenylene substituted with 1 to 3 substituents selected from alkyl, halogen, OH, or amine. 4 or T 5 is 5-10 membered heteroarylene or substituted heteroarylene. In some embodiments, T 4 or T 5 is a 4- to 10-membered heterosilene or substituted heterocyclylene. In some embodiments, T 4 or T 5 -C 1-6 It is a heteroarylene or heterosylene optionally substituted with 1 to 3 substituents selected from alkyl, halogen, OH, or amine.

[0353] In some embodiments, T 1 , T 2 , T 3 , T 4 , and T 5 , and V 1 , V 2 , V 3 , V 4 , and V 5 is selected from Table 2 below.

[0354] [Table 5-1]

[0355] [Table 5-2]

[0356] In some embodiments, the linker is

[0357] [ka] or any combination thereof, wherein r is an integer of 1 to 10, preferably 3 to 7, and X is O, S, or NR 1a In some embodiments, X is O or NR 1a In some embodiments, X is O.

[0358] In some embodiments, the linker is

[0359] [ka] or any combination thereof, wherein at least one -(CH2-CH2-O)- is -((CR 1a R 1b ) x -CH=CH-(CR 1a R 1b ) x -O)-, or any combination thereof, W' is absent, (CH) 1-5 , -(CH2) 1-5 O, (CH2) 1-5- C(O)NH-(CH2) 1-5 -O, (CH2) 1-5- C(O)NH-(CH2) 1-5 , -(CH2) 1-5 NHC(O)-(CH2) 1-5 -O, or -(CH2) 1-5- NHC(O)-(CH2) 1-5 - and E 3 is an arbitrarily substituted C 6-10 an arylene group, an optionally substituted 4- to 10-membered heterocycloalkylene, or an optionally substituted 5- to 10-membered heteroarylene, wherein X is O, S, or NH; and each R 1a and R1b are independently H or C 1-6 alkyl, r is an integer from 1 to 10, and x is an integer from 1 to 15. In some embodiments, X is O. In some embodiments, X is NH. In some embodiments, E 3 -C 1-6 C optionally substituted with 1 to 3 substituents selected from alkyl, halogen, OH, or amine 6-10 It is an arylene group.

[0360] In some embodiments, E 3 is phenylene or substituted phenylene.

[0361] In some embodiments, the linker is

[0362] [ka] Includes.

[0363] In some embodiments, the linker is -X(CH) m (CH2CH2O) n wherein X is —O—, —NH—, or —S—, and wherein m is 0 or greater and n is at least 1.

[0364] In some embodiments, the linker is followed at the second end by

[0365] [ka] Contains R c is a bond, -N(R 1a )-, -O-, and -S-; R d is -N(R 1a )-, -O-, and -S-; R e are independently hydrogen and optionally substituted C 1-6 alkyl.

[0366] In some embodiments, the linker is

[0367] [ka] -C 1-12 Alkyl, arylene, cycloalkylene, heteroarylene, heterocycloalkylene, -O-, -C(O)NR 1a -, -C(O)-, -NR 1a -, -(CH2CH2CH2O) y - and -(CH2CH2CH2NR 1a ) y -, wherein each d and y is independently 1 to 10; and each R 1a are independently hydrogen or C 1-6 In some embodiments, d is 4 to 8.

[0368] In some embodiments, the linker is

[0369] [ka] wherein each d is independently 3 to 7. In some embodiments, d is 4 to 6.

[0370] In some embodiments, the linker is -N(R 1a )(CH2) x N(R 1b )(CH2) x N-containing, wherein R 1a and R 1b are each independently selected from hydrogen or optionally substituted C1-C6 alkyl, and each x is independently an integer ranging from 1 to 6.

[0371] In some embodiments, the linker is -(CH2-C(O)N(R'')-(CH2) q -N(R')-(CH2) q -N(R'')C(O)-(CH2) x -C(O)N(R'')-A 2 -, -(CH2)x -C(O)N(R'')-(CH2CH2O) y (CH2) x -C(O)N(R'')-A 2 -, -C(O)N(R'')-(CH2) q -N(R')-(CH2) q -N(R'')C(O)-(CH2) x -A 2 -, -(CH2) x -O-(CH2CH2O) y -(CH2) x -N(R'')C(O)-(CH2) x -A 2 - or -N(R'')C(O)-(CH2)-C(O)N(R'')-(CH2) x -O(CH2CH2O) y (CH2) x -A 2 wherein R' is methyl, R'' is hydrogen, each x and y is independently an integer from 1 to 10, each q is independently an integer from 2 to 10, and each A 2 are independently a bond, an optionally substituted C 1-12 Alkyl, optionally substituted C 6-10 Arylene, optionally substituted C 3-7 It is selected from cycloalkylene, optionally substituted 5- to 10-membered heteroarylene, and optionally substituted 4- to 10-membered heterocycloalkylene.

[0372] In some embodiments, the linker is -(CH2CH2-O) x1 -or-(CH2CH2-O) x2 -A 2 -(CH2CH2-O) x3 -, wherein A 2 is an optionally substituted 4-10 membered heterocycloalkylene or spirocyclene, and each x1, x2, and x3 is independently an integer from 1 to 15.

[0373] In some embodiments, A 2 teeth,

[0374] [ka] In some embodiments, A 2 teeth,

[0375] [ka] In some embodiments, A 2 teeth,

[0376] [ka] In some embodiments, A 2 teeth,

[0377] [ka] is.

[0378] In some embodiments, A 2 comprises a moiety having the structure:

[0379] [ka] During the ceremony, X 2 is absent or -C(O)-, R 26 is an arbitrarily substituted C 1-50 Alkyl or C 1-50 It is heteroalkyl.

[0380] In some embodiments, X 2 is —C(O)—. In some embodiments, X 2 is non-existent.

[0381] In some embodiments, R 26 is C 1-50 In some embodiments, R 26 is C 1-40In some embodiments, R 26 is C 1-30 In some embodiments, R 26 is C 1-20 In some embodiments, R 26 is C 1-10 In some embodiments, R 26 is C 1-50 In some embodiments, R 26 is C 1-40 In some embodiments, R 26 is C 1-30 In some embodiments, R 26 is C 1-20 In some embodiments, R 26 is C 1-10 In some embodiments, the heteroalkyl is polyethylene glycol (PEG).

[0382] In some embodiments, the linker is -CO-, -NR 1a -, C 1-12 Alkyl, -CONR 1a - and -NR 1a CO—, wherein each R 1a are independently hydrogen or optionally substituted C 1-6 alkyl, or optionally substituted -C 1-12 Alkylene, optionally substituted C 2-10 Alkenylene, optionally substituted C 2-10 Alkynylene, optionally substituted C 6-10 Arylene, optionally substituted C 3-7 cycloalkylene, optionally substituted 5- to 10-membered heteroarylene, and optionally substituted 4- to 10-membered heterocycloalkylene.

[0383] In some embodiments, the linker is -CO-, -NR 1a -,-CONR 1a -, -NR 1aCO-, -CONR 1a C 1-4 Alkyl-, -NR 1a CO-C 1-4 Alkyl-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR 1a -, -NR 1 SO2-, -P(O)OH-, -((CH2) x -O)-, -((CH2) y -NR 1a )-, optionally substituted -C 1-12 Alkylene, optionally substituted C 2-10 Alkenylene, optionally substituted C 2-10 Alkynylene, optionally substituted C 6-10 Arylene, optionally substituted C 3-7 attached to the first terminus by a group selected from cycloalkylene, optionally substituted 5- to 10-membered heteroarylene, and optionally substituted 4- to 10-membered heterocycloalkylene; each x is independently 1 to 4; each y is independently 1 to 4; and each R 1a are independently hydrogen or optionally substituted C 1-6 It is alkyl.

[0384] In some embodiments, the linker is -CO-, -NR 1a -, C 1-12 Alkyl, -CONR 1a - and -NR 1a and attached to the first end by a group selected from CO—.

[0385] In some embodiments, the linker is -CO-, -NR 1a -,-CONR 1a -, -NR 1a CO-, -CONR 1a C 1-4 Alkyl-, -NR 1a CO-C 1-4 Alkyl-, -C(O)O-, -OC(O)-, -O-, -S-, -S(O)-, -SO2-, -SO2NR 1a -, -NR 1 SO2-, -P(O)OH-, -((CH2) x-O)-, -((CH2) y -NR 1a )-, optionally substituted -C 1-12 Alkylene, optionally substituted C 2-10 Alkenylene, optionally substituted C 2-10 Alkynylene, optionally substituted C 6-10 Arylene, optionally substituted C 3-7 attached to the second end by a group selected from cycloalkylene, optionally substituted 5- to 10-membered heteroarylene, and optionally substituted 4- to 10-membered heterocycloalkylene; each x is independently 1 to 4; each y is independently 1 to 4; and each R 1a are independently hydrogen or optionally substituted C 1-6 It is alkyl.

[0386] In some embodiments, the linker is -CO-, -NR 1a -,-CONR 1a -, -NR 1a CO-, -((CH2) x -O)-, -((CH2) y -NR 1a )-, -O-, optionally substituted -C 1-12 Alkyl, optionally substituted C 6-10 Arylene, optionally substituted C 3-7 attached to the second end by a group selected from cycloalkylene, optionally substituted 5- to 10-membered heteroarylene, and optionally substituted 4- to 10-membered heterocycloalkylene; each x is independently 1 to 4; each y is independently 1 to 4; and each R 1 are independently hydrogen or optionally substituted C 1-6 In some embodiments, the linker is -CO-, -NR 1a -, C 1-12 Alkyl, -CONR 1a - and -NR 1a and attached to the second end by a group selected from CO-.

[0387] In some embodiments, the linker is attached at its first or second terminus to a group selected from optionally substituted 4- to 10-membered heterocycloalkylene. In some embodiments, the linker is attached at its second terminus to a group selected from optionally substituted 4- to 10-membered heterocycloalkylene.

[0388] In some embodiments, the linker is attached to the second end by a moiety comprising the structure of formula (C-1), or a pharmaceutically acceptable salt thereof:

[0389] [ka] Formula (C-1) During the ceremony, Ring D is absent, arylene, or heterocycloacrylene; L 1 is absent or is an optionally substituted alkylene or alkenylene; each X 3 and X 4 are independently CH or N, p1 is 0 to 3, ** indicates the bond to the second end.

[0390] In some embodiments, Ring D is absent. In some embodiments, Ring D is C4-C7 heterocycloalkylene.

[0391] In some embodiments, X 3 is N. In some embodiments, X 3 is CH.

[0392] In some embodiments, X 4 is N. In some embodiments, X 4 is CH.

[0393] In some embodiments, the linker is attached to the second end by a moiety comprising the structure of formula (C-2), or a pharmaceutically acceptable salt thereof:

[0394] [ka] Formula (C-2) During the ceremony, each X 5 and X 6 is independently N or CH.

[0395] In some embodiments, X 4 and X 5 are each independently N or CH, and X 6 is N.

[0396] In some embodiments, L 1 is non-existent.

[0397] In some embodiments, L 1 is -(C R1G R 1G ) x -(Alkylene)2-(CR 1G R 1G ) y wherein x and y are each independently 0 or 1; and each R 1G is hydrogen or C1-C3 alkyl.

[0398] In some embodiments, L 1 is C1-C3 alkylene or C1-C3 alkenylene.

[0399] In some embodiments, L 1 is —CH—, —CHCH—, —C≡C—, or —C≡CC≡C—. In some embodiments, L 1 is —CH— or —CHCH—. In some embodiments, L 1 is -C≡C-. In some embodiments, L 1 is -C≡CC≡C-.

[0400] In some embodiments, the linker is attached to the second end by a moiety comprising the structure of formula (C-3), or a pharmaceutically acceptable salt thereof:

[0401] [ka] Formula (C-3) During the ceremony, p1 is 0 to 3, r1 is 1 to 3, R 27 is an arbitrarily substituted C 1-50 Alkyl, C 1-50 Heteroalkyl, -C(O)(C 1-50 alkyl), or -C(O)(C 1-50 heteroalkyl), wherein each alkyl and heteroalkyl is optionally substituted; Each R 1G are independently hydrogen or C1-C3 alkyl; ** indicates the bond to the second end.

[0402] In some embodiments, R 27 is an arbitrarily substituted C 1-50 Alkyl or C 1-50 In some embodiments, R 27 is -C(O)(C 1-50 alkyl) or -C(O)(C 1-50 heteroalkyl), wherein each alkyl and heteroalkyl is optionally substituted.

[0403] In some embodiments, R 27 is C 1-50 In some embodiments, R 27 is C 1-40 In some embodiments, R 27 is C 1-30 In some embodiments, R 27 is C 1-20 In some embodiments, R 27 is C 1-10 In some embodiments, R 27 is C 1-50 In some embodiments, R 27 is C1-40 In some embodiments, R 27 is C 1-30 In some embodiments, R 27 is C 1-20 In some embodiments, R 27 is C 1-10 In some embodiments, the heteroalkyl is polyethylene glycol (PEG).

[0404] In some embodiments, each R 1G is independently hydrogen. In some embodiments, each R 1G is independently C1-C3 alkyl. In some embodiments, C1-C3 alkyl is methyl, ethyl, or propyl. In some embodiments, each R 1G is independently methyl.

[0405] In some embodiments, p1 is 0, 1, or 2. In some embodiments, p1 is 0. In some embodiments, p1 is 1. In some embodiments, p1 is 2.

[0406] In some embodiments, r1 is 1 or 2. In some embodiments, r1 is 1. In some embodiments, r1 is 2.

[0407] In some embodiments, the linker is

[0408] [ka] and ** indicates a bond to the first and / or second end.

[0409] In some embodiments, the linker is

[0410] [ka] and ** indicates a bond to the first and / or second end.

[0411] In some embodiments, the linker is independently attached to the first and second termini using one of the groups described above. In some embodiments, the linker is attached to the first terminus using any of the groups described above. In some embodiments, the linker is attached to the second terminus using any of the groups described above.

[0412] Cell-permeable ligands In certain embodiments, the compound comprises a cell-permeable ligand moiety.

[0413] In certain embodiments, the cell-permeable ligand moiety is a polypeptide.

[0414] In certain embodiments, the cell-permeable ligand moiety is a polypeptide comprising fewer than 30 amino acid residues.

[0415] In certain embodiments, the polypeptide is selected from any one of SEQ ID NO: 1 to SEQ ID NO: 37, inclusive.

[0416] Also provided are embodiments in which any of the above embodiments may be combined with any one or more of these embodiments, provided that the combinations are not mutually exclusive.

[0417] As used herein, two embodiments are "mutually exclusive" if one is defined as being different from the other. For example, an embodiment in which two groups are bonded to form a cycloalkyl is mutually exclusive from an embodiment in which one group is ethyl and the other group is hydrogen. Similarly, an embodiment in which one group is CH2 is mutually exclusive from an embodiment in which the same group is NH.

[0418] In some embodiments, non-limiting examples of transcription modulator compounds described herein, or pharmaceutically acceptable salts thereof, are shown in Table 3.

[0419] [Table 6-1]

[0420] [Table 6-2]

[0421] [Table 6-3]

[0422] [Table 6-4]

[0423] [Table 6-5]

[0424] [Table 6-6]

[0425] [Table 6-7]

[0426] [Table 6-8]

[0427] [Table 6-9]

[0428] [Table 6-10]

[0429]

Table 6-11

[0430]

Table 6-12

[0431]

Table 6-13

[0432]

Table 6-14

[0433]

Table 6-15

[0434]

Table 6-16

[0435]

Table 6-17

[0436]

Table 6-18

[0437]

Table 6-19

[0438]

Table 6-20

[0439]

Table 6-21

[0440]

Table 6-22

[0441]

Table 6-23

[0442]

Table 6-24

[0443]

Table 6-25

[0444]

Table 6-26

[0445]

Table 6-27

[0446]

Table 6-28

[0447]

Table 6-29

[0448]

Table 6-30

[0449]

Table 6-31

[0450]

Table 6-32

[0451]

Table 6-33

[0452]

Table 6-34

[0453]

Table 6-35

[0454]

Table 6-36

[0455]

Table 6-37

[0456]

Table 6-38

[0457]

Table 6-39

[0458]

Table 6-40

[0459]

Table 6-41

[0460]

Table 6-42

[0461]

Table 6-43

[0462]

Table 6-44

[0463]

Table 6-45

[0464]

Table 6-46

[0465]

Table 6-47

[0466]

Table 6-48

[0467]

Table 6-49

[0468]

Table 6-50

[0469]

Table 6-51

[0470]

Table 6-52

[0471]

Table 6-53

[0472]

Table 6-54

[0473]

Table 6-55

[0474]

Table 6-56

[0475]

Table 6-57

[0476]

Table 6-58

[0477]

Table 6-59

[0478]

Table 6-60

[0479]

Table 6-61

[0480]

Table 6-62

[0481]

Table 6-63

[0482]

Table 6-64

[0483]

Table 6-65

[0484]

Table 6-66

[0485]

Table 6-67

[0486]

Table 6-68

[0487]

Table 6-69

[0488]

Table 6-70

[0489]

Table 6-71

[0490]

Table 6-72

[0491]

Table 6-73

[0492]

Table 6-74

[0493]

Table 6-75

[0494]

Table 6-76

[0495]

Table 6-77

[0496]

Table 6-78

[0497]

Table 6-79

[0498]

Table 6-80

[0499]

Table 6-81

[0500] [Table 6-82]

[0501] [Table 6-83]

[0502] [Table 6-84]

[0503] [Table 6-85]

[0504] [Table 6-86]

[0505] [Table 6-87]

[0506] How to use The present disclosure also relates to methods of modulating the transcription of fxn, comprising contacting fxn with a compound described herein. Changes in cell phenotype, cell proliferation, transcription of fxn, production of mRNA from transcription of fxn, translation of fxn, changes in biochemical output produced by a protein encoded by fxn, or non-covalent binding of a protein encoded by fxn to a natural binding partner can be monitored. Such methods can be disease treatment modalities, biological assays, cellular assays, biochemical assays, etc.

[0507] Also provided herein is a method for treating a disease mediated by the transcription of fxn, comprising administering to a patient in need thereof a therapeutically effective amount of a transcription modulator molecule disclosed herein or a salt thereof.

[0508] In certain embodiments, the disease is Friedreich's ataxia (FA).

[0509] Also provided is the use of the transcriptional modulator molecules disclosed herein as agents for the treatment of diseases mediated by the transcription of fxn.

[0510] Also provided herein are methods of modulating the transcription of fxn, comprising contacting fxn with a transcription modulator molecule disclosed herein, or a salt thereof.

[0511] Also provided herein is a method of treating Friedreich's ataxia in a subject in need thereof, comprising administering to the patient a transcription modulator molecule described herein.

[0512] Also provided herein is a method for achieving an effect in a patient comprising administering to the patient a therapeutically effective amount of a compound disclosed herein or a salt thereof, wherein the effect is selected from improved neurosensation, improved vision, improved balance, improved gait, reduced sensitivity to glucose, and reduced sensitivity to carbohydrates.

[0513] In some embodiments, the method comprises reducing one or more of muscle atrophy, ataxia, fasciculation, or dementia.

[0514] Certain compounds of the present disclosure may be effective in treating subjects whose genotype has 5 or more repeats of GAA. Certain compounds of the present disclosure may be effective in treating subjects whose genotype has 10 or more repeats of GAA. Certain compounds of the present disclosure may be effective in treating subjects whose genotype has 20 or more repeats of GAA. Certain compounds of the present disclosure may be effective in treating subjects whose genotype has 50 or more repeats of GAA. Certain compounds of the present disclosure may be effective in treating subjects whose genotype has 100 or more repeats of GAA. Certain compounds of the present disclosure may be effective in treating subjects whose genotype has 200 or more repeats of GAA. Certain compounds of the present disclosure may be effective in treating subjects whose genotype has 500 or more repeats of GAA.

[0515] Also provided are methods for modulating fxn-mediated functions in a subject, comprising administering a therapeutically effective amount of a compound disclosed herein.

[0516] Also provided are pharmaceutical compositions comprising a compound disclosed herein together with a pharmaceutically acceptable carrier.

[0517] In certain embodiments, the pharmaceutical composition is formulated for oral administration.

[0518] In certain embodiments, the pharmaceutical composition is formulated for intravenous injection and / or infusion.

[0519] In certain embodiments, the oral pharmaceutical composition is selected from a tablet and a capsule.

[0520] In certain embodiments, an ex vivo treatment method is provided. Ex vivo methods typically involve cells, organs, and / or tissues removed from a subject. The cells, organs, and / or tissues can be, for example, incubated with a drug under appropriate conditions. The contacted cells, organs, and / or tissues are typically returned to the donor, placed in a recipient, or stored for future use. Therefore, the compound is typically present in a pharmaceutically acceptable carrier.

[0521] In certain embodiments, administration of the pharmaceutical composition modulates expression of fxn within 6 hours of treatment. In certain embodiments, administration of the pharmaceutical composition modulates expression of fxn within 24 hours of treatment. In certain embodiments, administration of the pharmaceutical composition modulates expression of fxn within 72 hours of treatment.

[0522] In certain embodiments, administration of the pharmaceutical composition causes a 2-fold increase in expression of FXN. In certain embodiments, administration of the pharmaceutical composition causes a 5-fold increase in expression of FXN. In certain embodiments, administration of the pharmaceutical composition causes a 10-fold increase in expression of FXN. In certain embodiments, administration of the pharmaceutical composition causes a 20-fold increase in expression of FXN.

[0523] In certain embodiments, administration of the pharmaceutical composition causes a 20% decrease in the expression of FXN. In certain embodiments, administration of the pharmaceutical composition causes a 50% decrease in the expression of FXN. In certain embodiments, administration of the pharmaceutical composition causes an 80% decrease in the expression of FXN. In certain embodiments, administration of the pharmaceutical composition causes a 90% decrease in the expression of FXN. In certain embodiments, administration of the pharmaceutical composition causes a 95% decrease in the expression of FXN. In certain embodiments, administration of the pharmaceutical composition causes a 99% decrease in the expression of FXN.

[0524] In certain embodiments, administration of the pharmaceutical composition reduces the expression of FXN to within 25% of the expression level observed in healthy individuals. In certain embodiments, administration of the pharmaceutical composition reduces the expression of FXN to within 50% of the expression level observed in healthy individuals. In certain embodiments, administration of the pharmaceutical composition reduces the expression of FXN to within 75% of the expression level observed in healthy individuals. In certain embodiments, administration of the pharmaceutical composition reduces the expression of FXN to within 90% of the expression level observed in healthy individuals.

[0525] Pharmaceutical Compositions and Administration Also provided are methods for modulating fxn-mediated functions in a subject, comprising administering a therapeutically effective amount of a compound disclosed herein.

[0526] Also provided are pharmaceutical compositions comprising a compound disclosed herein together with a pharmaceutically acceptable carrier.

[0527] In certain embodiments, the pharmaceutical composition is formulated for oral administration.

[0528] In certain embodiments, the pharmaceutical composition is formulated for intravenous injection or infusion.

[0529] In certain embodiments, the oral pharmaceutical composition is selected from a tablet and a capsule.

[0530] In certain embodiments, an ex vivo treatment method is provided. Ex vivo methods typically involve cells, organs, or tissues removed from a subject. The cells, organs, or tissues can be, for example, incubated with a drug under appropriate conditions. The contacted cells, organs, or tissues are typically returned to the donor, placed in a recipient, or stored for future use. Therefore, the compound is typically present in a pharmaceutically acceptable carrier.

[0531] In certain embodiments, the compound is effective at a concentration of less than about 5 μM. In certain embodiments, the compound is effective at a concentration of less than about 1 μM. In certain embodiments, the compound is effective at a concentration of less than about 400 nM. In certain embodiments, the compound is effective at a concentration of less than about 200 nM. In certain embodiments, the compound is effective at a concentration of less than about 100 nM. In certain embodiments, the compound is effective at a concentration of less than about 50 nM. In certain embodiments, the compound is effective at a concentration of less than about 20 nM. In certain embodiments, the compound is effective at a concentration of less than about 10 nM.

[0532] Abbreviations and definitions As used herein, the following terms have the meanings indicated.

[0533] It is understood that a particular radical naming convention can include either a monoradical or a diradical, depending on the context. For example, if a substituent requires two points of attachment to the rest of the molecule, the substituent is understood to be a diradical. For example, a substituent identified as alkyl, which requires two points of attachment, includes diradicals such as -CH-, -CHCH-, -CHCH(CH)CH-, etc. Other radical naming conventions explicitly indicate that the radical is a diradical, such as "alkylene," "alkenylene," "arylene," "heteroarylene," etc.

[0534] When two R groups are said to form a ring "together with the atoms to which they are attached" (e.g., a carbocyclyl, heterocyclyl, aryl, or heteroaryl ring), it means that the collective unit of the atoms and two R groups is the recited ring. Otherwise, the ring is not limited by the definition of each R group when taken individually. For example, if the following substructure is present:

[0535] [ka] R 1 and R 2 is defined as selected from the group consisting of hydrogen and alkyl, or R 1 and R 2 together with the nitrogen to which they are attached form a heterocyclyl, which is represented by R 1 and R 2 can be selected from hydrogen or alkyl, or the moiety has the structure:

[0536] [ka] wherein ring A is the nitrogen-containing heteroaryl ring as shown.

[0537] Similarly, when two "adjacent" R groups are said to form a ring "together with the atoms to which they are attached," it is meant that the collective unit of the atoms, intervening bonds, and two R groups is the recited ring. For example, if the following substructure is present:

[0538] [ka] R 1 and R 2 is defined as selected from the group consisting of hydrogen and alkyl, or R 1 and R 2 together with the atom to which they are attached form an aryl or carbocyclyl, which is 1 and R 2 can be selected from hydrogen or alkyl, or the moiety has the structure:

[0539] [ka] wherein A is an aryl ring or carbocyclyl containing the double bond shown.

[0540] When a substituent is depicted as a diradical (i.e., having two points of attachment to the rest of the molecule), it is understood that the substituent may be attached in any orientation unless otherwise indicated. Thus, for example, -AE-, or

[0541] [ka] Substituents depicted as include those where A is attached to the rightmost point of attachment of the molecule as well as those oriented so that A is attached to the leftmost point of attachment of the molecule.

[0542] When a range of values ​​is disclosed and the notation "n1... to n2" or "between n1... to n2" is used, n1 and n2 are numbers, and unless otherwise specified, this notation is intended to include the numbers themselves and the range therebetween. The range may be an integer or a continuous value between them, and may be inclusive of the end value. As an example, carbon is an integer unit, so the range "2 to 6 carbons" is intended to include 2, 3, 4, 5, and 6 carbons. By comparison, as an example, the range "1 to 3 μM (micromolar)" is intended to include 1 μM, 3 μM, and all increments therebetween to any significant figure (e.g., 1.255 μM, 2.1 μM, 2.9999 μM, etc.).

[0543] As used herein, the term "about" is intended to modify the numerical value that it modifies, expressing such value as variable within a margin of error. When a specific error, such as a standard deviation to the average value given in a chart or table of data, is not recited, the term "about" should be understood to mean a range that encompasses the recited value, and also by rounding up or down to that numerical value in light of significant values.

[0544] The term "polyamide" refers to a polymer of linkable units chemically linked by amide (i.e., CONH) bonds, and optionally, the polyamide includes a chemical probe attached thereto. Polyamides can be synthesized by the stepwise condensation of carboxylic acids (COOH) with amines (RR'NH) using methods known in the art. Alternatively, polyamides can be formed using in vitro enzymatic reactions or by microbial fermentation.

[0545] The term "linkable unit" refers to methylimidazole, methylpyrrole, and linear and branched chain aliphatic functional groups (e.g., methylene, ethylene, propylene, butylene, etc.) optionally containing nitrogen substituents, and chemical derivatives thereof. The aliphatic functional groups of the linkable unit can be provided, for example, by condensation of β-alanine or dimethylaminopropylamine during the synthesis of polyamides by methods well known in the art.

[0546] The term "linker" refers to a chain of at least 10 contiguous atoms. In certain embodiments, a linker comprises 20 or fewer non-hydrogen atoms. In certain embodiments, a linker comprises 40 or fewer non-hydrogen atoms. In certain embodiments, a linker comprises 60 or fewer non-hydrogen atoms. In certain embodiments, a linker comprises an atom selected from C, H, N, O, and S. In certain embodiments, all non-hydrogen atoms are chemically bonded to either two adjacent atoms in the linker, or one adjacent atom in the linker and the terminus of the linker. In certain embodiments, a linker forms an amide bond with at least one of the two other groups to which it is attached. In certain embodiments, a linker forms an ester or ether bond with at least one of the two other groups to which it is attached. In certain embodiments, a linker forms a thioester or thioether bond with at least one of the two other groups to which it is attached. In certain embodiments, a linker forms a direct carbon-carbon bond with at least one of the two other groups to which it is attached. In certain embodiments, the linker forms an amine or amide bond with at least one of the two other groups to which it is attached. In certain embodiments, the linker comprises a -(CH2OCH2)- unit. In certain embodiments, the linker comprises a -(CH(CH3)OCH2)- unit. In certain embodiments, the linker comprises a -(CH(CH3)OCH2)- unit. In certain embodiments, the linker comprises a -(CH(CH3)OCH2)- unit. N =C 1-4 For alkyl, -(CHNR N CH2) units. In certain embodiments, the linker comprises an arylene, cycloalkylene, or heterocycloalkylene moiety.

[0547] The term "spacer" refers to a chain of at least 5 consecutive atoms. In certain embodiments, a spacer comprises 10 or fewer non-hydrogen atoms. In certain embodiments, a spacer comprises an atom selected from C, H, N, O, and S. In certain embodiments, a spacer forms an amide bond with the two other groups to which it is attached. In certain embodiments, a spacer comprises a -(CH2OCH2)- unit. In certain embodiments, a spacer is a group selected from R N =C 1-4 For alkyl, -(CHNR N In certain embodiments, the spacer comprises at least one positive charge at physiological pH.

[0548] The term "curve element" refers to a chain of about 4-10 contiguous atoms. In certain embodiments, the curve element comprises an atom selected from C, H, N, O, and S. In certain embodiments, the curve element forms an amide bond with the two other groups to which it is attached. In certain embodiments, the curve element comprises at least one positive charge at physiological pH.

[0549] The terms "nucleic acid" and "nucleotide" refer to ribonucleotides and deoxyribonucleotides and their analogs as known in the art.

[0550] The term "oligonucleotide sequence" refers to a plurality of nucleic acids having a defined sequence and length (e.g., 2, 3, 4, 5, 6, or more nucleotides). The term "oligonucleotide repeat" refers to a contiguous stretch of an oligonucleotide sequence.

[0551] The term "transcription" is well known in the art and refers to the synthesis of RNA (i.e., ribonucleic acid) by DNA-directed RNA polymerase. The term "modulating transcription" refers to a change in transcription levels, which can be measured by methods well known in the art, for example, by assaying the transcription product, mRNA. In certain embodiments, modulation is an increase in transcription. In other embodiments, modulation is a decrease in transcription.

[0552] The term "contacting" refers to bringing a compound (e.g., a transcriptional molecule of the present disclosure) into proximity with a desired target gene. Contacting may result in binding to the target moiety or may result in a conformational change in the target moiety.

[0553] The term "acyl," as used herein, alone or in combination, refers to a carbonyl bound to an alkenyl, alkyl, aryl, cycloalkyl, heteroaryl, heterocycle, or any other moiety where the atom bound to the carbonyl is carbon. An "acetyl" group refers to a -C(O)CH group. An "alkylcarbonyl" or "alkanoyl" group refers to an alkyl group bound to the parent molecular moiety through a carbonyl group. Examples of such groups include methylcarbonyl and ethylcarbonyl. Examples of acyl groups include formyl, alkanoyl, and aroyl.

[0554] As used herein, the term "alkenyl," alone or in combination, refers to a straight- or branched-chain hydrocarbon radical having one or more double bonds and containing 2 to 20 carbon atoms. In certain embodiments, the alkenyl will contain 2 to 6 carbon atoms. The term "alkenylene" refers to a carbon-carbon double bond system attached at two or more positions, such as ethenylene [(-CH=CH-), (-C::C-)]. Examples of suitable alkenyl radicals include ethenyl, propenyl, 2-methylpropenyl, 1,4-butadienyl, and the like. Unless otherwise specified, the term "alkenyl" can include "alkenylene" groups.

[0555] The term "alkoxy," as used herein, alone or in combination, refers to an alkyl ether radical, where the term alkyl is as defined below. Examples of suitable alkyl ether radicals include methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, iso-butoxy, sec-butoxy, tert-butoxy, and the like.

[0556] As used herein, the term "alkyl," alone or in combination, refers to a straight- or branched-chain alkyl radical containing 1 to 20 carbon atoms. In certain embodiments, the alkyl will contain 1 to 10 carbon atoms. In further embodiments, the alkyl will contain 1 to 8 carbon atoms. The alkyl group can be optionally substituted as defined herein. Examples of alkyl radicals include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, pentyl, iso-amyl, hexyl, octyl, noyl, and the like. As used herein, the term "alkylene," alone or in combination, refers to a saturated aliphatic group derived from a straight- or branched-chain saturated hydrocarbon bonded at two or more positions, such as methylene (-CH-). Unless otherwise specified, the term "alkyl" can include an "alkylene" group.

[0557] The term "alkylamino," as used herein, alone or in combination, refers to an alkyl group attached to the parent molecular moiety through an amino group. Suitable alkylamino groups can be mono- or di-alkylated to form, for example, N-methylamino, N-ethylamino, N,N-dimethylamino, N,N-ethylmethylamino, and the like groups.

[0558] The term "alkylidene," as used herein, alone or in combination, refers to an alkenyl group in which one carbon atom of the carbon-carbon double bond belongs to the moiety to which the alkenyl group is attached.

[0559] As used herein, the term "alkylthio," alone or in combination, refers to an alkyl thioether (RS-) radical, where the term alkyl is as defined above and the sulfur may be singly or doubly oxidized. Examples of suitable alkyl thioether radicals include methylthio, ethylthio, n-propylthio, isopropylthio, n-butylthio, iso-butylthio, sec-butylthio, tert-butylthio, methanesulfonyl, ethanesulfinyl, and the like.

[0560] As used herein, the term "alkynyl," alone or in combination, refers to a straight- or branched-chain hydrocarbon radical having one or more triple bonds and containing 2 to 20 carbon atoms. In certain embodiments, the alkynyl contains 2 to 6 carbon atoms. In further embodiments, the alkynyl contains 2 to 4 carbon atoms. The term "alkynylene" refers to a carbon-carbon triple bond attached at two positions, such as ethynylene [(-C:::C-, -C≡C-)]. Examples of alkynyl radicals include ethynyl, propynyl, hydroxypropynyl, butyn-1-yl, butyn-2-yl, pentyn-1-yl, 3-methylbutyn-1-yl, hexyn-2-yl, and the like. Unless otherwise specified, the term "alkynyl" can include "alkynylene" groups.

[0561] As used herein, the terms "amido" and "carbamoyl," alone or in combination, refer to an amino group, as described below, attached to the parent molecular moiety through a carbonyl group, or vice versa. As used herein, the term "C-amido," alone or in combination, refers to a -C(O)N(RR') group, where R and R' are as defined herein or as defined by the specifically enumerated "R" group designated. As used herein, the term "N-amido," alone or in combination, refers to a RC(O)N(R')- group, where R and R' are as defined herein or as defined by the specifically enumerated "R" group designated. As used herein, the term "acylamino," alone or in combination, encompasses an acyl group attached to the parent moiety through an amino group. An example of an "acylamino" group is acetylamino (CHC(O)NH-).

[0562] As used herein, the term "amide," alone or in combination, refers to -C(O)NRR', where R and R' are independently selected from hydrogen, alkyl, acyl, heteroalkyl, aryl, cycloalkyl, heteroaryl, and heterocycloalkyl, any of which may themselves be optionally substituted. Additionally, R and R' may together form a heterocycloalkyl, any of which may itself be optionally substituted. Amides may be formed by direct condensation of a carboxylic acid with an amine or by using an acid chloride. Additionally, coupling reagents are known in the art, including carbodiimide-based compounds such as DCC and EDCI.

[0563] As used herein, the term "amino", alone or in combination, refers to -NRR ’wherein R and R' are independently selected from hydrogen, alkyl, acyl, heteroalkyl, aryl, cycloalkyl, heteroaryl, and heterocycloalkyl, any of which may themselves be optionally substituted. Additionally, R and R' may together form a heterocycloalkyl, any of which may be optionally substituted.

[0564] As used herein, the term "aryl," alone or in combination, refers to a carbocyclic aromatic system containing one, two, or three rings, with polycyclic ring systems being fused together. The term "aryl" encompasses aromatic groups such as phenyl, naphthyl, anthracenyl, and phenanthryl. The term "arylene" encompasses aromatic groups such as phenylene, naphthylene, anthracenylene, and phenanthrylene.

[0565] The terms "arylalkenyl" or "aralkenyl," as used herein, alone or in combination, refer to an aryl group attached to the parent molecular moiety through an alkenyl group.

[0566] The terms "arylalkoxy" or "aralkoxy," as used herein, alone or in combination, refer to an aryl group attached to the parent molecular moiety through an alkoxy group.

[0567] The terms "arylalkyl" or "aralkyl," as used herein, alone or in combination, refer to an aryl group attached to the parent molecular moiety through an alkyl group.

[0568] The terms "arylalkynyl" or "aralkynyl," as used herein, alone or in combination, refer to an aryl group attached to the parent molecular moiety through an alkynyl group.

[0569] As used herein, the terms "arylalkanoyl" or "aralkanoyl" or "aroyl," alone or in combination, refer to an acyl radical derived from an aryl-substituted alkanecarboxylic acid, such as benzoyl, naphthoyl, phenylacetyl, 3-phenylpropionyl(hydrocinnamoyl), 4-phenylbutyryl, (2-naphthyl)acetyl, 4-chlorohydrocinnamoyl, and the like.

[0570] The term aryloxy, as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an oxy.

[0571] As used herein, the terms "benzo" and "benz," alone or in combination, refer to the divalent radical CH= derived from benzene. Examples include benzothiophene and benzimidazole.

[0572] The term "carbamate," as used herein, alone or in combination, refers to an ester of carbamic acid (-NHCOO-), which can be attached to the parent molecular moiety through either the nitrogen or the acid terminus, and can be optionally substituted as defined herein.

[0573] The term "O-carbamyl," as used herein, alone or in combination, refers to an --OC(O)NRR' group, with R and R' as defined herein.

[0574] The term "N-carbamyl," as used herein, alone or in combination, refers to an ROC(O)NR'- group, with R and R' as defined herein.

[0575] As used herein, the term "carbonyl" when alone includes formyl [-C(O)H] and in combination is a -C(O)- group.

[0576] As used herein, the terms "carboxyl" or "carboxy" refer to -C(O)OH or the corresponding "carboxylic acid" anion, such as in a carboxylate salt. An "O-carboxy" group refers to a -RC(O)O- group, where R is as defined herein. A "C-carboxy" group refers to a -C(O)OR group, where R is as defined herein.

[0577] The term "cyano," as used herein, alone or in combination, refers to --CN.

[0578] As used herein, "cycloalkyl," or alternatively, "carbocycle," alone or in combination, refers to a saturated or partially saturated monocyclic, bicyclic, or tricyclic alkyl group, each cyclic moiety containing 3 to 12 carbon atom ring members, and may optionally be a benzo-fused ring system, optionally substituted as defined herein. In certain embodiments, the cycloalkyl will contain 5 to 7 carbon atoms. Examples of such cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, tetrahydronaphthyl, indanyl, octahydronaphthyl, 2,3-dihydro-1H-indenyl, adamantyl, and the like. As used herein, "bicyclic" and "tricyclic" are intended to include fused ring systems, e.g., decahydronaphthalene, octahydronaphthalene, and polycyclic (multi-center) saturated or partially unsaturated types. The latter type of isomer is commonly exemplified by bicyclo[1,1,1]pentane, camphor, adamantane, and bicyclo[3,2,1]octane.

[0579] The term "ester," as used herein, alone or in combination, refers to a carboxy group bridging two moieties joined at a carbon atom.

[0580] The term "ether," as used herein, alone or in combination, refers to an oxy group bridging two moieties joined at a carbon atom.

[0581] The terms "halo" or "halogen," as used herein, alone or in combination, refer to fluorine, chlorine, bromine, or iodine.

[0582] The term "haloalkoxy," as used herein, alone or in combination, refers to a haloalkyl group attached to the parent molecular moiety through an oxygen atom.

[0583] As used herein, the term "haloalkyl," alone or in combination, refers to an alkyl radical having the meaning defined above, in which one or more hydrogen atoms are replaced by halogen. Specifically, monohaloalkyl, dihaloalkyl, and polyhaloalkyl radicals are encompassed. For example, a monohaloalkyl radical may have an iodo, bromo, chloro, or fluoro atom within the radical. Dihalo and polyhaloalkyl radicals may have two or more of the same halo atoms or a combination of different halo radicals. Examples of haloalkyl radicals include fluoromethyl, difluoromethyl, trifluoromethyl, chloromethyl, dichloromethyl, trichloromethyl, pentafluoroethyl, heptafluoropropyl, difluorochloromethyl, dichlorofluoromethyl, difluoroethyl, difluoropropyl, dichloroethyl, and dichloropropyl. "Haloalkylene" refers to a haloalkyl group attached at two or more positions. Examples include fluoromethylene (-CFH-), difluoromethylene (-CF2-), chloromethylene (-CHCl-), and the like.

[0584] As used herein, the term "heteroalkyl," alone or in combination, refers to a stable, linear or branched chain, or combination thereof, which may be fully saturated or contain one to three degrees of unsaturation, consisting of the specified number of carbon atoms and one to three heteroatoms selected from N, O, and S, wherein the N and S atoms may be optionally oxidized, and the N heteroatom may be optionally quaternized. The heteroatom may be placed at any interior position of the heteroalkyl group. Up to two heteroatoms may be consecutive, for example, -CH-NH-OCH.

[0585] As used herein, the term "heteroaryl," alone or in combination, refers to a 3- to 15-membered unsaturated heteromonocycle or fused monocyclic, bicyclic, or tricyclic ring system in which at least one of the fused rings is aromatic and contains at least one atom selected from N, O, and S. In certain embodiments, the heteroaryl will contain 1 to 4 heteroatoms as ring members. In further embodiments, the heteroaryl will contain 1 to 2 heteroatoms as ring members. In certain embodiments, the heteroaryl will contain 5 to 7 atoms. The term also encompasses fused polycyclic groups in which a heterocycle is fused to an aryl ring, a heteroaryl ring is fused to another heteroaryl ring, a heteroaryl ring is fused to a heterocycloalkyl ring, or a heteroaryl ring is fused to a cycloalkyl ring. Examples of heteroaryl groups include pyrrolyl, pyrrolinyl, imidazolyl, pyrazolyl, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl, triazolyl, pyranyl, furyl, thienyl, oxazolyl, isoxazolyl, oxadiazolyl, thiazolyl, thiadiazolyl, isothiazolyl, indolyl, isoindolyl, indolizinyl, benzimidazolyl, quinolyl, isoquinolyl, quinoxalinyl, quinazolinyl, indazolyl, benzotriazolyl, benzodioxolyl, benzopyranyl, benzoxazolyl, benzoxadiazolyl, benzothiazolyl, benzothiadiazolyl, benzofuryl, benzothienyl, chromonyl, coumarinyl, benzopyranyl, tetrahydroquinolinyl, tetrazolopyridazinyl, tetrahydroisoquinolinyl, thienopyridinyl, furopyridinyl, pyrrolopyridinyl, and the like. Exemplary tricyclic heterocyclic groups include carbazolyl, benzidolyl, phenanthrolinyl, dibenzofuranyl, acridinyl, phenanthridinyl, xanthenyl, and the like.

[0586] As used herein, the terms "heterocycloalkyl," and interchangeably, "heterocycle," alone or in combination, refer to a saturated, partially unsaturated, or fully unsaturated (but non-aromatic), monocyclic, bicyclic, or tricyclic heterocyclic group, respectively, containing at least one heteroatom as a ring member, where each heteroatom may be independently selected from nitrogen, oxygen, and sulfur. In certain embodiments, the heterocycloalkyl will contain 1 to 4 heteroatoms as ring members. In further embodiments, the heterocycloalkyl will contain 1 to 2 heteroatoms as ring members. In certain embodiments, the heterocycloalkyl will contain 3 to 8 ring members in each ring. In further embodiments, the heterocycloalkyl will contain 3 to 7 ring members in each ring. In still further embodiments, the heterocycloalkyl will contain 5 to 6 ring members in each ring. "Heterocycloalkyl" and "heterocycle" are intended to include sulfone, sulfoxide, N-oxide of tertiary nitrogen ring members, and fused carbocyclic and benzofused ring systems; additionally, both terms include systems in which a heterocycle is fused to an aryl group, as defined herein, or an additional heterocyclic group. Examples of heterocyclic groups include tetrahydroisoquinoline, aziridinyl, azetidinyl, 1,3-benzodioxolyl, dihydroisoindolyl, dihydroisoquinolinyl, dihydrocinnolinyl, dihydrobenzodioxinyl, dihydro[1,3]oxazolo[4,5-b]pyridinyl, benzothiazolyl, dihydroindolyl, dihydropyridinyl, 1,3-dioxanyl, 1,4-dioxanyl, 1,3-dioxolanyl, isoindolinyl, morpholinyl, piperazinyl, pyrrolidinyl, tetrahydropyridinyl, piperidinyl, thiomorpholinyl, and the like. The heterocycle groups may be optionally substituted unless specifically prohibited.

[0587] The term "hydrazinyl," as used herein, alone or in combination, refers to two amino groups joined by a single bond, ie, --NN--.

[0588] The term "hydroxy," as used herein, alone or in combination, refers to --OH.

[0589] The term "hydroxyalkyl," as used herein, alone or in combination, refers to a hydroxy group attached to the parent molecular moiety through an alkyl group.

[0590] The term "imino," as used herein, alone or in combination, refers to =N-.

[0591] The term "iminohydroxy," as used herein, alone or in combination, refers to ═N(OH) and ═NO—.

[0592] The phrase "in the backbone" refers to the longest contiguous or adjacent chain of carbon atoms beginning at the point of attachment of the group to a compound or molecule of any one of the formulas disclosed herein.

[0593] The term "isocyanato" refers to an --NCO group.

[0594] The term "isothiocyanato" refers to the group -NCS.

[0595] The phrase "linear chain of atoms" refers to the longest linear chain of atoms independently selected from carbon, nitrogen, oxygen, and sulfur.

[0596] As used herein, the term "lower", alone or in combination, means containing 1 to 6 carbon atoms (ie, C1-C6 alkyl), unless otherwise defined.

[0597] As used herein, the term "lower aryl," alone or in combination, means phenyl or naphthyl, either of which, if provided, may be optionally substituted.

[0598] The term "lower heteroaryl," as used herein, alone or in combination, means either 1) a monocyclic heteroaryl containing 5 or 6 ring members, of which 1 to 4 of the ring members can be heteroatoms selected from N, O, and S, or 2) a bicyclic heteroaryl in which each of the fused rings contains 5 or 6 ring members and between them 1 to 4 heteroatoms selected from N, O, and S.

[0599] As used herein, "lower cycloalkyl," alone or in combination, refers to a monocyclic cycloalkyl having 3 to 6 ring members (i.e., C3-C6 cycloalkyl). Lower cycloalkyls can be unsaturated. Examples of lower cycloalkyls include cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl.

[0600] As used herein, the term "lower heterocycloalkyl," alone or in combination, means a monocyclic heterocycloalkyl having 3 to 6 ring members, 1 to 4 of which may be heteroatoms selected from N, O, and S (i.e., C3-C6 heterocycloalkyl). Examples of lower heterocycloalkyl include pyrrolidinyl, imidazolidinyl, pyrazolidinyl, piperidinyl, piperazinyl, and morpholinyl. Lower heterocycloalkyl may be unsaturated.

[0601] As used herein, the term "lower amino," alone or in combination, refers to --NRR', where R and R' are independently selected from hydrogen and lower alkyl, either of which may be optionally substituted.

[0602] The term "mercaptyl," as used herein, alone or in combination, refers to an RS-group, where R is as defined herein.

[0603] The term "nitro," as used herein, alone or in combination, refers to --NO.sub.2.

[0604] The terms "oxy" or "oxa," as used herein, alone or in combination, refer to --O--.

[0605] The term "oxo," as used herein, alone or in combination, refers to =O.

[0606] The term "perhaloalkoxy" refers to an alkoxy group in which all hydrogen atoms have been replaced with halogen atoms.

[0607] The term "perhaloalkyl," as used herein, alone or in combination, refers to an alkyl group in which all of the hydrogen atoms have been replaced with halogen atoms.

[0608] As used herein, the terms "sulfonate," "sulfonic acid," and "sulfonic," alone or in combination, refer to the -SO3H group and its anion when sulfonic acid is used in salt formation.

[0609] The term "sulfanyl," as used herein, alone or in combination, refers to --S--.

[0610] The term "sulfinyl" as used herein, alone or in combination, refers to -S(O)-.

[0611] The term "sulfonyl," as used herein, alone or in combination, refers to -S(O)2-.

[0612] The term "N-sulfonamido" refers to an RS(=O)2NR'- group, with R and R' as defined herein.

[0613] The term "S-sulfonamido" refers to a -S(=O)2NRR' group, where R and R' are as defined herein.

[0614] The terms "thia" and "thio," as used herein, alone or in combination, refer to an -S- group or ether where the oxygen is replaced by sulfur. The oxidized derivatives of the thio group, i.e., sulfinyl and sulfonyl, are included in the definition of thia and thio.

[0615] The term "thiol," as used herein, alone or in combination, refers to a --SH group.

[0616] As used herein, the term "thiocarbonyl" when alone includes thioformyl-C(S)H and in combination is a -C(S)- group.

[0617] The term "N-thiocarbamyl" refers to the group ROC(S)NR', ​​where R and R' are as defined herein.

[0618] The term "O-thiocarbamyl" refers to an OC(S)NRR' group, where R and R' are as defined herein.

[0619] "The term "thiocyanato" refers to a CNS group.

[0620] The term "trihalomethanesulfonamide" refers to a X3CS(O)2NR group where X is a halogen and R is as defined herein.

[0621] The term "trihalomethanesulfonyl" refers to a X3CS(O)2 group where X is a halogen.

[0622] The term "trihalomethoxy" refers to a X3CO group where X is a halogen.

[0623] As used herein, the term "trisubstituted silyl," alone or in combination, refers to a silicone group substituted at its three free valences with a group listed herein under the definition of substituted amino. Examples include trimethylsilyl, tert-butyldimethylsilyl, triphenylsilyl, etc.

[0624] Any definition herein can be used in combination with any other definition to describe a composite structural group. By convention, the element following such a definition is the one that is attached to the parent moiety. For example, the composite group alkylamido represents an alkyl group attached to the parent molecule via an amide group, and the term alkoxyalkyl represents an alkoxy group attached to the parent molecule via an alkyl group.

[0625] The term "optionally substituted" means that the preceding group may be substituted or unsubstituted. When substituted, the substituents of the "optionally substituted" group may include, but are not limited to, one or more substituents independently selected from the following groups or the set of specific specified groups, alone or in combination: lower alkyl, lower alkenyl, lower alkynyl, lower alkanoyl, lower heteroalkyl, lower heterocycloalkyl, lower haloalkyl, lower haloalkenyl, lower haloalkynyl, lower perhaloalkyl, lower perhaloalkoxy, lower cycloalkyl, phenyl, aryl, aryloxy, lower alkoxy, lower haloa alkoxy, oxo, lower acyloxy, carbonyl, carboxyl, lower alkylcarbonyl, lower carboxyester, lower carboxamide, cyano, hydrogen, halogen, hydroxy, amino, lower alkylamino, arylamino, amido, nitro, thiol, lower alkylthio, lower haloalkylthio, lower perhaloalkylthio, arylthio, sulfonate, sulfonic acid, trisubstituted silyl, N3, SH, SCH3, C(O)CH3, CO2CH3, CO2H, pyridinyl, thiophene, furanyl, lower carbamate, and lower urea. Where structurally feasible, two substituents may be joined together to form a fused 5-, 6-, or 7-membered carbocyclic or heterocyclic ring consisting of 0 to 3 heteroatoms, for example, methylenedioxy or ethylenedioxy. Optionally substituted groups can be unsubstituted (e.g., -CH2CH3), fully substituted (e.g., -CF2CF3), monosubstituted (e.g., -CH2CH2F), or substituted at a level between fully and monosubstituted (e.g., -CH2CF3). When substituents are listed without qualification as to substitution, both substituted and unsubstituted forms are encompassed. When a substituent is modified as "substituted," the substituted form is specifically intended. Additionally, different sets of optional substituents for a particular moiety may be defined as needed; in these cases, the optional substitution is often as defined immediately following the phrase "optionally substituted with."

[0626] As used herein, a substituent is derived from an unsubstituted parent group in which one or more hydrogen atoms have been replaced with another atom or group. Unless otherwise indicated, when a group is considered to be "substituted," the group is independently selected from C-C alkyl, C-C alkenyl, C-C alkynyl, C-C heteroalkyl, C-C carbocyclyl (optionally substituted with halo, C-C alkyl, C-C alkoxy, C-C haloalkyl, and C-C haloalkoxy), C-C-carbocyclyl-C-C-alkyl (optionally substituted with halo, C-C alkyl, C-C alkoxy, C-C haloalkyl, and C-C haloalkoxy). substituted), 3-10 membered heterocyclyl (optionally substituted with halo, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, and C1-C6 haloalkoxy), 3-10 membered heterocyclyl-C1-C6-alkyl (optionally substituted with halo, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, and C1-C6 haloalkoxy), aryl (optionally substituted with halo, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, and C1-C6 haloalkoxy), aryl ( C1-C6)alkyl (optionally substituted with halo, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, and C1-C6 haloalkoxy), 5-10 membered heteroaryl (optionally substituted with halo, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, and C1-C6 haloalkoxy), 5-10 membered heteroaryl(C1-C6)alkyl (optionally substituted with halo, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, and C1-C6 haloalkoxy), halo, cyclohexane ... Ano, hydroxy, C1-C6 alkoxy, C1-C6 alkoxy(C1-C6)alkyl (i.e., ether), aryloxy, sulfhydryl (mercapto), halo(C1-C6)alkyl (e.g., -CF3), halo(C1-C6)alkoxy (e.g., -OCF3), C1-C6 alkylthio, arylthio, amino, amino(C1-C6)alkyl, nitro, O-carbamyl, N-carbamyl, O-thiocarbamyl, N-thiocarbamyl, C-amido, N-amido, S-sulfonamido, N-sulfonamido,It means substituted with one or more substituents selected from C-carboxy, O-carboxy, acyl, cyanato, isocyanato, thiocyanato, isothiocyanato, sulfinyl, sulfonyl, and oxo (=O). Whenever a group is described as "optionally substituted," that group can be substituted with the above substituents.

[0627] The term R or R', unless otherwise defined, appears alone and without a number designation and refers to a moiety selected from hydrogen, alkyl, cycloalkyl, heteroalkyl, aryl, heteroaryl, and heterocycloalkyl, any of which may be optionally substituted. Such R and R' groups may be optionally substituted as defined herein. Regardless of whether an R group has a number designation, R, R', and R n All R groups, including those in formulas where n = (1, 2, 3, ... n), all substituents, and all terms should be understood to be independent of all others in terms of selection from groups. When any variable, substituent, or term (e.g., aryl, heterocycle, R, etc.) occurs more than once within a formula or generic structure, its definition at each occurrence is independent of its definition at every other occurrence. Those skilled in the art will further recognize that certain groups may be attached to the parent molecule or may occupy a position within the chain of elements from either end as written. For example, an unsymmetrical group such as -C(O)N(R)- can be attached to the parent moiety at either the carbon or the nitrogen.

[0628] Asymmetric centers exist in the compounds or molecules disclosed herein. These centers are designated by the symbols "R" or "S," depending on the configuration of substituents around the chiral carbon atom. It is understood that the present disclosure encompasses all stereochemically isomeric forms, including diastereomeric, enantiomeric, and epimeric forms, as well as d- and l-isomers, and mixtures thereof. Individual stereoisomers of compounds or molecules can be prepared synthetically from commercially available starting materials containing chiral centers, or by preparation of an enantiomeric product mixture followed by conversion to a diastereomeric mixture, followed by separation or recrystallization, chromatographic techniques, direct separation of enantiomers on a chiral chromatographic column, or any other suitable method known in the art. Starting compounds or molecules of particular stereochemistry are commercially available or can be made and resolved by techniques known in the art. Additionally, compounds or molecules disclosed herein can exist as geometric isomers. The present disclosure includes all cis, trans, syn, anti, entgegen (E), and zusammen (Z) isomers, as well as the appropriate mixtures thereof. Additionally, compounds or molecules may exist as tautomers, with all tautomeric isomers being provided by the present disclosure. Additionally, compounds or molecules disclosed herein may exist in unsolvated as well as solvated forms with pharmaceutically acceptable solvents, such as water, ethanol, and the like. In general, solvated forms are considered equivalent to unsolvated forms.

[0629] The term "bond" refers to a covalent bond between two atoms, or two moieties when the atoms connected by the bond are considered to be part of a larger substructure. Unless otherwise specified, the bond may be a single, double, or triple bond. A dashed line between two atoms in a drawing of a molecule indicates that an additional bond may or may not be present at that position.

[0630] As used herein, the term "disease" is generally intended to be synonymous with, and used interchangeably with, the terms "disorder," "syndrome," and "condition" (as in medical condition), all of which reflect an abnormal condition of the human and animal body, or parts thereof, that impairs normal functioning, is typically manifested by characteristic signs and symptoms, and reduces the duration or quality of a human or animal's life.

[0631] The term "combination therapy" refers to the administration of two or more therapeutic agents to treat a therapeutic condition or disorder described in this disclosure. Such administration encompasses the co-administration of these therapeutic agents in a substantially simultaneous manner, such as in a single capsule having a fixed ratio of active ingredients, or in multiple separate capsules for each active ingredient. In addition, such administration also encompasses the use of each type of therapeutic agent in a sequential manner. In either case, the treatment regimen will provide the beneficial effects of the drug combination in treating the condition or disorder described herein.

[0632] The phrase "therapeutically effective" is intended to qualify the amount of active ingredient used in the treatment of a disease or disorder or for the effect of a clinical endpoint.

[0633] The term "therapeutically acceptable" refers to those compounds or molecules (or salts, prodrugs, tautomers, zwitterionic forms, etc.) that are suitable for use in contact with the tissues of patients without undue toxicity, irritation, or allergic response, are commensurate with a reasonable benefit / risk ratio, and are effective for their intended use.

[0634] As used herein, reference to "treatment" of a patient is intended to include prevention. Treatment may also be preemptive in nature, i.e., include prevention of disease. Prevention of disease may include complete protection from disease, as in the case of, for example, prevention of infection by a pathogen, or may include prevention of disease progression. For example, prevention of disease may not mean completely eliminating the effects associated with the disease at any level, but instead may mean preventing disease symptoms to a clinically significant or detectable level. Prevention of disease may also mean preventing the progression of disease to a later stage of disease.

[0635] The term "patient" is generally synonymous with the term "subject" and includes all mammals, including humans. Examples of patients include humans, livestock such as cows, goats, sheep, pigs, and rabbits, and companion animals such as dogs, cats, rabbits, and hares. Preferably, the patient is a human.

[0636] The term "prodrug" refers to a compound or molecule that is more active in vivo. Certain compounds or molecules disclosed herein may also exist as prodrugs, as described in Hydrolysis in Drug and Prodrug Metabolism: Chemistry, Biochemistry, and Enzymology (Testa, Bernard and Mayer, Joachim M. Wiley-VHCA, Zurich, Switzerland 2003). Prodrugs of the compounds described herein are structurally modified forms of the compounds that readily undergo chemical changes under physiological conditions to provide the compounds. Additionally, prodrugs can be converted to the compounds by chemical or biochemical methods in an ex vivo environment. For example, prodrugs can be slowly converted to the compounds when placed in a transdermal patch reservoir with a suitable enzyme or chemical reagent. Prodrugs are often useful because, in some cases, they may be easier to administer than the compound or parent drug. They may be bioavailable, for example, by oral administration, whereas the parent drug is not. Prodrugs may also have improved solubility in pharmaceutical compositions over the parent drug. A wide variety of prodrug derivatives are known in the art, including those that rely on hydrolytic cleavage or oxidative activation of the prodrug. Examples of prodrugs include, but are not limited to, compounds that are administered as an ester (the "prodrug"), but are subsequently metabolically hydrolyzed to the carboxylic acid, the active agent. Additional examples include peptidyl derivatives of the compounds.

[0637] The compounds or molecules disclosed herein can exist as therapeutically acceptable salts. The present disclosure includes the compounds or molecules listed above in the form of salts, including acid addition salts. Suitable salts include those formed with both organic and inorganic acids. Such acid addition salts are generally pharmaceutically acceptable. However, salts of non-pharmaceutically acceptable salts may also be useful in the preparation and purification of the compound or molecule in question. Basic addition salts are also formed and are pharmaceutically acceptable. For a more complete discussion of salt preparation and selection, see Pharmaceutical Salts: Properties, Selection, and Use (Stahl, P. Heinrich, Wiley-VCHA, Zurich, Switzerland, 2002).

[0638] Base addition salts can be prepared during the final isolation and purification of a compound or molecule by reacting a carboxyl group with a suitable base, such as a hydroxide, carbonate, or bicarbonate of a metal cation, or with ammonia, or with an organic primary, secondary, or tertiary amine. Therapeutically acceptable salt cations include lithium, sodium, potassium, calcium, magnesium, and aluminum, as well as non-toxic quaternary amine cations such as ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, diethylamine, ethylamine, tributylamine, pyridine, N,N-dimethylaniline, N-methylpiperidine, N-methylmorpholine, dicyclohexylamine, procaine, dibenzylamine, N,N-dibenzylphenethylamine, 1-ephenamine, and N,N'-dibenzylethylenediamine. Other representative organic amines useful for the formation of base addition salts include ethylenediamine, ethanolamine, diethanolamine, piperidine, and piperazine.

[0639] Other carrier materials and modes of administration known in the pharmaceutical art may also be used. The pharmaceutical compositions of the present disclosure may be prepared by any of the well-known techniques of pharmacy, including effective formulation and administration procedures. Preferred unit dosage formulations are those containing an effective dose, as recited herein below, or an appropriate fraction thereof, of the active ingredient.

[0640] It will be understood that in addition to the ingredients particularly mentioned above, the above formulations may include other agents conventional in the art having regard to the type of formulation in question; for example, those suitable for oral administration may include flavoring agents.

[0641] The amount of active ingredient that can be combined with the carrier materials to produce a single dosage form will vary depending upon the host treated and the particular mode of administration.

[0642] The compound or molecule can be administered in various ways, for example, orally, topically, or by injection. The exact amount of compound administered to a patient is the responsibility of the attending physician. The specific dose level for any particular patient will depend on various factors, including the activity of the specific compound used, age, body weight, general health, sex, diet, administration time, administration route, excretion rate, drug combination, the exact disease being treated, and the severity of the indication or condition being treated. In addition, the administration route may vary depending on the symptoms and their severity. The above considerations regarding effective formulations and administration procedures are well known in the art and are described in standard textbooks.

[0643] Combination and Combined Therapies In certain cases, it may be appropriate to administer at least one of the compounds described herein (or a pharmaceutically acceptable salt thereof) in combination with another therapeutic agent. By way of example only, if one of the side effects experienced by a patient when receiving one of the compounds described herein is hypertension, it may be appropriate to administer an antihypertensive drug in combination with the first therapeutic agent. Or, by way of example only, the therapeutic effectiveness of one of the compounds described herein may be enhanced by the administration of an adjuvant (i.e., the adjuvant itself may have minimal therapeutic benefit, but in combination with another therapeutic agent, the overall therapeutic benefit to the patient is enhanced). Or, by way of example only, the benefit experienced by a patient may be increased by administering one of the compounds described herein with another therapeutic agent (including a treatment regimen) that has therapeutic benefit. By way of example only, in the treatment of diabetes involving the administration of one of the compounds described herein, providing the patient with another diabetes medication may result in an increased therapeutic benefit. In either case, regardless of the disease, disorder, or condition being treated, the overall benefit experienced by the patient may simply be the addition of the two therapeutic agents, or the patient may experience a synergistic benefit.

[0644] Specific non-limiting examples of possible combination therapies include the use of certain compounds of the present disclosure together with ACE inhibitors.

[0645] In either case, the multiple therapeutic agents (at least one of which is a compound disclosed herein) may be administered in any order, or simultaneously. If simultaneously, the multiple therapeutic agents may be provided in a single, integrated form, or in multiple forms (by way of example only, as a single pill or as two separate pills). One of the therapeutic agents may be administered in multiple doses, or both may be administered in multiple doses. If not simultaneously, the timing between the multiple administrations may be any period ranging from a few minutes to four weeks.

[0646] Accordingly, in another aspect, certain embodiments provide a method for treating a fxn-mediated disorder in a human or animal subject in need of such treatment, comprising administering to the subject an amount of a compound disclosed herein in combination with at least one additional agent known in the art for treating the disorder effective to alleviate or prevent the disorder. In a related aspect, certain embodiments provide a therapeutic composition comprising at least one compound disclosed herein in combination with one or more additional agents for treating fxn-mediated.

[0647] In addition to being useful for human treatment, certain compounds and formulations disclosed herein may also be useful for the veterinary treatment of companion animals, exotic animals, and farm animals, including mammals, rodents, etc. More preferred animals include horses, dogs, and cats.

[0648] Compound synthesis The compounds of the present disclosure can be prepared using the methods exemplified in the general synthetic schemes and experimental procedures detailed below. The general synthetic schemes and experimental procedures are illustrative and are not intended to be limiting. The starting materials used to prepare the compounds of the present disclosure are commercially available or can be prepared using routine methods readily known in the art.

[0649] List of abbreviations Ac2O = acetic anhydride; AcCl = acetyl chloride; AcOH = acetic acid; AIBN = azobisisobutyronitrile; aq. = aqueous; Bu3SnH = tributyltin hydride; CD3OD = deuterated methanol; CDCl3 = deuterated chloroform; CDI = 1,1'-carbonyldiimidazole; DBU = 1,8-diazabicyclo[5.4.0]undec-7-ene; DCM = dichloromethane; DEAD = diethyl azodicarboxylate; DIBAL-H = di-isobutylaluminum hydride; DIEA = DIPEA = N,N-diisopropylethylamine; DMAP = 4-dimethylaminopyridine; DMF = N,N-dimethylformamide; DMSO-d6 = deuterated dimethyl sulfoxide; DMSO = dimethyl sulfoxide; DPPA = diphenylphosphoryl azide; EDC.HCl = EDCI.HCl = 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride; Et2O = diethyl ether; EtOAc = ethyl acetate; EtOH = ethanol; h = hour; HATU = 2-(1H-7-azabenzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate methanamine; HMDS = hexamethyldisilazane; HOBT = 1-hydroxybenzotriazole; i-PrOH = isopropanol; LAH = lithium aluminum hydride; LiHMDS = lithium bis(trimethylsilyl)amide; MeCN = acetonitrile; MeOH = methanol; MP-carbonate resin = macroporous triethylammonium methyl polystyrene carbonate resin; MsCl = mesyl chloride; MTBE = Methyl tertiary butyl ether; MW = microwave irradiation; n-BuLi = n-butyllithium; NaHMDS = sodium bis(trimethylsilyl)amide; NaOMe = sodium methoxide; NaOtBu = sodium t-butoxide; NBS = N-bromosuccinimide; NCS = N-chlorosuccinimide; NMP = N-methyl-2-pyrrolidone; Pd(Ph3)4 = tetrakis(triphenylphosphine)palladium(0); Pd2(dba)3 = tris(dibenzylideneacetone)dipalladium(0); PdCl2(PPh3)2 = bis(triphenylphosphine)palladium(II) dichloride; PG = protecting group; Prep-HPLC = preparative high-performance liquid chromatography; PyBop = (benzotriazol-1-yloxy)-tripyrrolidinophosphonium Hexafluorophosphate; Pyr = pyridine; RT = room temperature; RuPhos = 2-dicyclohexylphosphino-2',6'-diisopropoxybiphenyl; sat. = saturated; ss = saturated solution; t-BuOH = tert-butanol; T3P = propylphosphonic anhydride; TBS = TBDMS = tert-butyldimethylsilyl; TBSCl = TBDMSCl = tert-butyldimethylchlorosilane; TEA = Et3N = triethylamine; TFA = trifluoroacetic acid; TFAA = trifluoroacetic anhydride; THF = tetrahydrofuran; Tol = toluene; TsCl = tosyl chloride; XPhos = 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl.

[0650] General synthetic methods for preparing compounds In general, the polyamides of the present disclosure can be synthesized by solid-support synthesis methods using compounds such as Boc-protected linear aliphatic and heteroaromatic amino acids and their alkylated derivatives, which are cleaved from the support by aminolysis, deprotected (e.g., with sodium thiophenoxide), and purified by reverse-phase HPLC, as is well known in the art. The identity and purity of the polyamides can be determined by 1 This can be verified using any of a variety of analytical techniques available to those skilled in the art, such as 1 H-NMR, analytical HPLC, or mass spectrometry.

[0651] The following scheme can be used to practice the present disclosure:

[0652] [ka]

[0653] The compounds disclosed herein can be synthesized using Scheme I. For clarity and brevity, the scheme depicts the synthesis of a diamide containing subunits "C" and "D," both of which are depicted as unspecified five-membered rings with amino and carboxy moieties. The amino group of subunit "D" is protected with a protecting group "PG," such as Boc or CBz carbamate, to yield 101. The (free) carboxylic acid is then reacted with a solid support using a coupling reagent, such as EDC, to yield supported compound 103. Removal of PG under acidic conditions yields free amine 104, which is coupled with nitrogen-protected carboxylic acid 105 to yield amide 106. Removal of PG under acidic conditions yields free amine 107. In this example, the free amine is reacted with acetic anhydride to form an acetamide (not shown). The molecule is then cleaved from the solid support under basic conditions to yield carboxylic acid 108. Methods for attachment of the linker L and recruiting moiety X are disclosed below.

[0654] Those skilled in the art will appreciate that many variations of the above scheme are available to provide a wide range of compounds: 1) The sequence 104-106-107 can be repeated as often as desired to form longer polyamine sequences. 2) A variety of aminoheterocyclic carboxylic acids can be used to form the different subunits. Table 4 provides, but is not intended to be limiting, some heterocyclic amino acids contemplated for the synthesis of compounds of the present disclosure. The carbamate protecting group PG can be incorporated using techniques well established in the art.

[0655] [Table 7-1]

[0656] [Table 7-2]

[0657] 3) Hydroxy-containing heterocyclic amino acids can be incorporated into Scheme I as their TBS ethers. While not intended to be limiting, Scheme B provides the synthesis of TBS-protected heterocyclic amino acids contemplated for the synthesis of molecules in this disclosure.

[0658] [ka]

[0659] 4) Aliphatic amino acids can be used in the above synthesis to form the spacer unit "W" and subunits for recognition of DNA nucleotides. Table 5 provides, but is not intended to be limiting, some aliphatic amino acids contemplated for synthesis of the molecules in this disclosure.

[0660] [Table 8-1]

[0661] [Table 8-2]

[0662] [ka]

[0663] Attachment of linker L and recruiting moiety X can be achieved by the method disclosed in Scheme C, which uses a triethylene glycol moiety in linker L. The mono-TBS ether of triethylene glycol 301 is converted to the bromo compound 302 under Mitsunobu conditions. Recruiting moiety X is attached by displacement of the bromine with the hydroxyl moiety to give ether 303. The TBS group is then removed by treatment with fluoride to give alcohol 304, which may be suitable for coupling to a polyamide moiety. Other methods for incorporating alternative linkers L, including, but not limited to, propylene glycol or polyamine linkers, or alternative attachment points for recruiting moiety X, including, but not limited to, the use of amines and thiols, will be apparent to those skilled in the art.

[0664] [ka]

[0665] The synthesis of the XLY molecule can be completed as shown in Scheme D. Carboxylic acid 108 is converted to acid chloride 401. Reaction with the alcohol functionality of 301 under basic conditions affords coupled product 402. Other methods for carrying out the coupling procedure will be apparent to those skilled in the art, including, but not limited to, the use of carbodiimide reagents. For example, amide coupling reagents include, but are not limited to, carbodiimides such as dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIC), and ethyl-(N',N'-dimethylamino)propylcarbodiimide hydrochloride (EDC), in combination with reagents such as 1-hydroxybenzotriazole (HOBt), 4-(N,N-dimethylamino)pyridine (DMAP), and diisopropylethylamine (DIEA).Other reagents, depending on the actual coupling reaction, include (benzotriazol-1-yloxy)tris(dimethylamino)phosphonium hexafluorophosphate (BOP), (benzotriazol-1-yloxy)tripyrrolidinophosphonium hexafluorophosphate (PyBOP), (7-azabenzotriazol-1-yloxy)tripyrrolidinophosphonium hexafluorophosphate (PyAOP), bromotripyrrolidinophosphonium hexafluorophosphate (PyBrOP), bis(2-oxo-3-oxazolidinyl)phosphinic chloride (BOP-Cl), O-(benzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HBTU), O- (Benzotriazol-1-yl)-N,N,N',N'-tetramethyluronium tetrafluoroborate (TBTU), O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU), O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium tetrafluoroborate (TATU), O-(6-chlorobenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HCTU), carbonyldiimidazole (CDI), and N,N,N',N'-tetramethylchloroformamidinium hexafluorophosphate (TCFH) are also often used.

[0666] Conjugating protein-binding molecules to an oligomeric backbone Generally, the oligomer backbone is functionalized to accommodate the types of chemical reactions that can be performed to link the oligomer to a binding position within the protein binding site. Suitable reaction types include, but are not limited to, amide coupling reactions, ether-forming reactions (O-alkylation reactions), amine-forming reactions (N-alkylation reactions), and, in some cases, carbon-carbon coupling reactions. Typical reactions used to link oligomers and protein-binding agents are shown in Schemes (E-G) below. The compounds and structures shown in Table 2 can be attached to the oligomer backbone described herein at any chemically feasible position that does not interfere with hydrogen bonding between the compound and the control protein.

[0667] [ka]

[0668] Either the oligomer or the protein binding agent can be functionalized with a carboxylic acid, and the other coupling counterpart with an amino group, allowing the moieties to be conjugated with amide coupling reagents, including, but not limited to, carbodiimides such as dicyclohexylcarbodiimide (DCC), diisopropylcarbodiimide (DIC), and ethyl-(N',N'-dimethylamino)propylcarbodiimide hydrochloride (EDC), in combination with reagents such as 1-hydroxybenzotriazole (HOBt), 4-(N,N-dimethylamino)pyridine (DMAP), and diisopropylethylamine (DIEA). Other reagents, depending on the actual coupling reaction, include (benzotriazol-1-yloxy)tris(dimethylamino)phosphonium hexafluorophosphate (BOP), (benzotriazol-1-yloxy)tripyrrolidinophosphonium hexafluorophosphate (PyBOP), (7-azabenzotriazol-1-yloxy)tripyrrolidinophosphonium hexafluorophosphate (PyAOP), bromotripyrrolidinophosphonium hexafluorophosphate (PyBrOP), bis(2-oxo-3-oxazolidinyl)phosphinic chloride (BOP-Cl), O-(benzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HBTU), O- (Benzotriazol-1-yl)-N,N,N',N'-tetramethyluronium tetrafluoroborate (TBTU), O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU), O-(7-azabenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium tetrafluoroborate (TATU), O-(6-chlorobenzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HCTU), carbonyldiimidazole (CDI), and N,N,N',N'-tetramethylchloroformamidinium hexafluorophosphate (TCFH) are also often used.

[0669] [ka]

[0670] In the ether-forming reaction, either the oligomer or the protein-binding agent can be functionalized to have a hydroxyl group (phenol or alcohol), while other coupling counterparts are functionalized with leaving groups such as halides, tosylates, and mesylates, allowing the moieties to be conjugated together via a base or catalyst. The base can be selected from, but is not limited to, sodium hydride, potassium hydride, sodium hydroxide, potassium hydroxide, sodium carbonate, and potassium carbonate. The catalyst can be selected from silver oxide, phase transfer reagents, iodide salts, and crown ethers.

[0671] [ka]

[0672] In N-alkylation reactions, either the oligomer or the protein-binding agent can be functionalized to have an amino group (arylamine or alkylamine), while other coupling counterparts are functionalized with leaving groups such as halides, tosylates, and mesylates, and the moieties can be conjugated together directly or in conjunction with a base or catalyst. The base can be selected from, but is not limited to, sodium hydride, potassium hydride, sodium hydroxide, potassium hydroxide, sodium carbonate, and potassium carbonate. The catalyst can be selected from silver oxide, phase transfer reagents, iodide salts, and crown ethers. Alkylation of amines can also be achieved by reductive amination reactions, where either the oligomer or the protein-binding agent can be functionalized to have an amino group (arylamine or alkylamine), while other coupling counterparts are functionalized with aldehyde or ketone groups, and the moieties can be conjugated together directly by treatment with a reducing reagent (hydride source) or in combination with a dehydrating agent. The reducing reagent can be selected from, but is not limited to, NaBH4, NaHB(OAc)3, NaBH3CN, and the dehydrating agent is typically Ti(iPrO)4, Ti(OEt)4, Al(iPrO)3, orthoformates, and activated molecular sieves.

[0673] Cell-permeable ligands In one aspect, the molecules of the present disclosure include a cell-permeable ligand moiety. The cell-permeable ligand moiety serves to facilitate transport of a compound across a cell membrane. In certain embodiments, the cell-permeable ligand moiety is a polypeptide. Some peptide sequences include polycationic sequences such as polyR, (RXR) n (X=6-aminohexanoic acid) and (RXRRBR) n These peptides facilitate passage into cells, including arginine-rich sequences interspersed with spacers such as (B = beta-alanine), sequences derived from penetratin peptides, and sequences derived from PNA / PMO internalization peptides (Pips). The Pip5 series is characterized by the sequence ILFQY.

[0674] In certain embodiments, the cell-penetrating polypeptide comprises an N-terminal cationic sequence H2N-(R) n -CO-, where n=5 to 10, inclusive. In certain embodiments, the N-terminal cationic sequence independently comprises one, two, or three substitutions of R for an amino acid residue selected from β-alanine and 6-aminohexanoic acid.

[0675] In certain embodiments, the cell-penetrating polypeptide comprises an ILFQY sequence. In certain embodiments, the cell-penetrating polypeptide comprises a QFLY sequence. In certain embodiments, the cell-penetrating polypeptide comprises a QFL sequence.

[0676] In certain embodiments, the cell-penetrating polypeptide comprises a C-terminal cationic sequence -HN-(R) n and -COOH, where n=5-10, inclusive. In certain embodiments, the C-terminal cationic sequence comprises one, two, or three substitutions of R for an amino acid residue independently selected from beta-alanine and 6-aminohexanoic acid. In certain embodiments, the C-terminal cationic sequence is substituted at every other position with an amino acid residue independently selected from beta-alanine and 6-aminohexanoic acid. In certain embodiments, the C-terminal cationic sequence is -HN-RXRBRXRB-COOH.

[0677] [Table 9-1]

[0678] [Table 9-2] [Example]

[0679] The following examples are presented for the purpose of illustrating various embodiments of the present invention and are not intended to limit the invention in any manner. The examples, along with the methods described herein, are presently representative of preferred embodiments and are illustrative and do not limit the scope of the invention. Modifications and other uses which are encompassed within the spirit of the invention as defined by the claims will occur to those skilled in the art.

[0680] While preferred embodiments of the present invention have been shown and described herein, it will be apparent to those skilled in the art that such embodiments are provided by way of example only. Numerous variations, changes, and substitutions will now occur to those skilled in the art without departing from the invention. It should be understood that various alternatives to the embodiments described herein may be employed. The following claims define the scope of the invention, and it is intended that methods and structures within the scope of these claims and their equivalents be covered thereby.

[0681] Synthesis of representative polyamides

[0682] Example 1. Synthesis of 3-([1-methyl-4-[3-([1-methyl-4-[1-methyl-4-(3-[[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido]propanamido)imidazol-2-amido]pyrrol-2-yl]formamido)propanamido]imidazol-2-yl]formamido)propanoic acid (PA-001)

[0683] [ka]

[0684] Step 1: Synthesis of ethyl 4-amino-1-methylimidazole-2-carboxylate

[0685] To a solution of ethyl 1-methyl-4-nitroimidazole-2-carboxylate (30.00 g, 150.63 mmol, 1.00 equiv) in EtOH (120.00 mL) and EA (120.00 mL) was added Pd / C (8.01 g, 27% w / w). The reaction was then stirred at room temperature under an H atmosphere for 17.0 h. The solid was filtered and the filtrate was concentrated to give ethyl 4-amino-1-methylimidazole-2-carboxylate (22.30 g, 75.20%) as a yellow solid. LC / MS: C7H 11 Calculated mass for N3O2: 169.09, Measured: 170.10 [M+H] + . 1 H NMR(400MHz,DMSO-d6)δ:7.37(s,1H), 4.29-4.34(m,2H), 3.94(s,3H), 1.31(t,J=7.2Hz,3H).

[0686] Step 2: Synthesis of ethyl 4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylate

[0687] A 500 mL flask was charged with 3-[(tert-butoxycarbonyl)amino]propanoic acid (22.45 g, 118.65 mmol, 0.90 equiv.) and DMF (180.00 mL). The mixture was cooled to 0° C., and then HATU (75.18 g, 197.71 mmol, 1.50 equiv.) and DIEA (51.11 g, 395.43 mmol, 3.00 equiv.) were added. The mixture was stirred for 10.0 minutes, and then ethyl 4-amino-1-methylimidazole-2-carboxylate (22.30 g, 131.81 mmol, 1.00 equiv.) was added in portions. The reaction was stirred at room temperature for 1.0 hour. The reaction was quenched with ice water (600 mL), and the solution was stirred for 15.0 minutes. The precipitated solid was collected by filtration, washed with water (3 x 50 mL), and dried under vacuum to give ethyl 4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylate (34.50 g, 76.90%) as a pale yellow solid. LC / MS: C 15 H 24Calculated mass for N4O5: 340.17, Measured: 341.20 [M+H] + . 1 H NMR(400MHz,DMSO-d6)δ:10.63(s,1H), 7.52(s,1H), 6.80(t,J=5.6Hz,1H), 4.23-4.28(m,2H) ), 3.90(s,3H), 3.15-3.20(m,2H), 2.42(t,J=7.2Hz,2H), 1.37(s,9H), 1.29(t,J=7.2Hz,3H).

[0688] Step 3: Synthesis of 4-[3-[(Tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylic acid

[0689] To a stirred solution of ethyl 4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylate (34.50 g, 101.36 mmol, 1.00 equiv) in MeOH (200.00 mL) was added dropwise LiOH solution (2 M, 202.00 mL, 4.00 equiv) at room temperature. The resulting mixture was stirred at 45 °C for 2.0 h. The resulting mixture was concentrated under reduced pressure. The residue was dissolved in HO (50 mL). The mixture was acidified to pH 3-5 with 2 M HCl. The precipitated solid was collected by filtration, washed with HO (3 × 30 mL), and dried under vacuum. 4-[3-[(Tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylic acid (30.00 g, 94.77%) was obtained as a white solid. LC / MS:C 13 H 20 Calculated mass for N4O5: 312.14, Measured: 313.15 [M+H] + . 1 H NMR(300MHz,DMSO-d6)δ:10.53(s,1H), 7.48(s,1H), 6.79(t,J=5.4Hz,1H), 3.89(s,3H), 3.15-3.22(m,2H), 2.43(t,J=7.2Hz,2H), 1.37(s,9H).

[0690] Step 4: Synthesis of methyl 4-(4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazol-2-amido)-1-methylpyrrole-2-carboxylate

[0691] To a stirred solution of 4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylic acid (16.00 g, 51.23 mmol, 1.00 equiv.) in CHCN (150.00 mL), TCFH (21.56 g, 76.84 mmol, 1.50 equiv.), NMI (12.62 g, 153.69 mmol, 3.00 equiv.), and methyl 4-amino-1-methylpyrrole-2-carboxylate hydrochloride (10.74 g, 56.34 mmol, 1.10 equiv.) were added in small portions at 0 °C. The resulting mixture was stirred at room temperature for 2.0 h. The precipitated solid was collected by filtration, washed with CHCN (3 × 20 mL), and dried under vacuum. Methyl 4-(4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazol-2-amido)-1-methylpyrrole-2-carboxylate (19.00 g, 82.70%) was obtained as a white solid. LC / MS: C 20 H 28 Mass calculation value for N6O6: 448.21, actual value: 449.25[M+H]+.1H NMR(300MHz,DMSO-d6)δ:10.24(s,1H), 10.11(s,1H), 7.52(s,1H), 7.33(s,1H), 6.99(s,1H), 6.82(t,J=5 .1Hz,1H), 3.94(s,3H), 3.85(s,3H), 3.74(s,3H), 3.16-3.23(m,2H), 2.47(t,J=6.9Hz,2H), 1.38(s,9H).

[0692] Step 5: Synthesis of methyl 4-[4-(3-aminopropanamido)-1-methylimidazol-2-amido]-1-methylpyrrole-2-carboxylate hydrochloride

[0693] A solution of methyl 4-(4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazol-2-amido)-1-methylpyrrole-2-carboxylate (19.00 g, 42.37 mmol, 1.00 equiv) in HCl / 1,4-dioxane (4 M, 200.00 mL) was stirred at room temperature for 2.0 h. The resulting mixture was concentrated in vacuo to give methyl 4-[4-(3-aminopropanamido)-1-methylimidazol-2-amido]-1-methylpyrrole-2-carboxylate (19.00 g crude) as a yellow solid. LC / MS: Calculated mass value for C15H21ClN6O4: 348.15, Actual value: 349.05[M+H]+.1H NMR(300MHz,CD3OD)δ:7.37(s,2H), 6.91(s,1H), 4.03(s,3H), 3.88(s,3H), 3.79(s,3H), 3.09(t,J=6.6Hz,2H), 2.64(t,J=6.6Hz,2H).

[0694] Step 6: Synthesis of methyl 3-[(4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazol-2-yl)formamido]propanoate

[0695] A 1000 mL flask was charged with 4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylic acid (11.00 g, 35.22 mmol, 1.00 equiv.) and DMF (300.00 mL). The mixture was cooled to 0 °C. HATU (20.09 g, 52.83 mmol, 1.50 equiv.) and DIEA (18.21 g, 140.88 mmol, 4.00 equiv.) were then added dropwise. The mixture was stirred for 10 minutes, and methyl 3-aminopropanoate (3.63 g, 35.22 mmol, 1.00 equiv.) was added portionwise. The reaction was stirred at room temperature for 1.0 hour. The reaction mixture was poured into water / ice (600 mL), and the solid was filtered and dried under vacuum. The aqueous phase was extracted with EA (3 x 200 mL), and the combined organic phases were washed with HO (1 x 200 mL) and NaCl (1 x 200 mL), and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified on a silica gel column eluted with pure EA. The combined fractions were concentrated. Methyl 3-[(4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazol-2-yl)formamido]propanoate (13.00 g, 87.95%) was obtained as a yellow solid. LC / MS: C 17 H 27 Calculated mass for N5O6: 397.20, Found: 398.20 [M+H] + . 1 H NMR(400MHz,DMSO-d6)δ:10.28(s,1H), 7.92(t,J=6.0Hz,1H), 7.37(s,1H), 6.77(t,J=6.0Hz,1H), 3.88(s,3H) , 3.59(s,3H), 3.42-3.47(m,2H), 3.13-3.18(m,2H), 2.56(t,J=6.0Hz,2H), 2.42(t,J=6.0Hz,2H), 1.35(s,9H).

[0696] Step 7: Synthesis of methyl 3-[[4-(3-aminopropanamido)-1-methylimidazol-2-yl]formamido]propanoate hydrochloride

[0697] A solution of methyl 3-[(4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazol-2-yl)formamido]propanoate (11.00 g, 27.678 mmol, 1.00 equiv) in HCl / 1,4-dioxane (4 M, 110.00 mL) was stirred at room temperature for 1.0 h. The resulting mixture was concentrated in vacuo to give methyl 3-[[4-(3-aminopropanamido)-1-methylimidazol-2-yl]formamido]propanoate hydrochloride (11.00 g, crude) as a yellow oil. LC / MS: C 12 H 19 Calculated mass for N5O4: 297.14, Found: 298.20 [M+H] + . 1 H NMR(400MHz,DMSO-d6)δ:10.57(s,1H), 7.92(t,J=6.0Hz,1H), 7.37(s,1H), 3.89(s,3H), 3. 59(s,3H), 3.43-3.47(m,2H), 2.97-3.05(m,2H), 2.57-2.71(m,2H), 2.56(t,J=6.0Hz,2H).

[0698] Step 8: Synthesis of methyl 1-methyl-4-(1-methylimidazol-2-amido)pyrrole-2-carboxylate

[0699] To a stirred solution of 1-methylimidazole-2-carboxylic acid (10.00 g, 79.29 mmol, 7.00 equiv) in DMF (150.00 mL) was added TBTU (38.19 g, 118.94 mmol, 1.50 equiv), methyl 4-amino-1-methylpyrrole-2-carboxylate hydrochloride (16.63 g, 87.24 mmol, 1.10 equiv), and DIEA (30.74 g, 237.88 mmol, 3.00 equiv) in small portions at 0 °C. The resulting mixture was stirred at room temperature for 17.0 h. The reaction was poured into water / ice (450 mL). The precipitated solid was collected by filtration, washed with HO (3 x 50 mL), and dried under vacuum. Methyl 1-methyl-4-(1-methylimidazol-2-amido)pyrrole-2-carboxylate (16.50 g, 78.37%) was obtained as a white solid. LC / MS: C12 H 14 Calculated mass for N4O3: 262.11, Measured mass: 263.15 [M+H] + . 1 H NMR(300MHz,DMSO-d6)δ:10.54(s,1H), 7.54(s,1H), 7.40(s,1H), 7.04(s,2H), 3.99(s,3H), 3.85(s,3H), 3.74(s,3H).

[0700] Step 9: Synthesis of 1-methyl-4-(1-methylimidazol-2-amido)pyrrole-2-carboxylic acid

[0701] To a stirred solution of methyl 1-methyl-4-(1-methylimidazol-2-amido)pyrrole-2-carboxylate (16.50 g, 62.91 mmol, 1.00 equiv) in MeOH (100.00 mL) was added LiOH solution (2 M, 158.00 mL, 5.00 equiv) dropwise at room temperature. The resulting mixture was stirred at 45 °C for 2.0 h. The resulting mixture was concentrated under reduced pressure. The residue was dissolved in HO (50 mL). The mixture was acidified to pH 3-5 with 2 M HCl. The precipitated solid was collected by filtration, washed with HO (3 x 30 mL), and dried under vacuum. 1-Methyl-4-(1-methylimidazol-2-amido)pyrrole-2-carboxylic acid (12.00 g, 76.84%) was obtained as a white solid. LC / MS: C 11 H 12 Calculated mass for N4O3: 248.09, Measured: 249.10 [M+H] + . 1 H NMR(300MHz,DMSO-d6)δ:10.52(s,1H), 7.48(s,1H), 7.41(s,1H), 7.06(s,1H), 6.99(s,1H), 3.99(s,3H), 3.82(s,3H).

[0702] Step 10: Synthesis of methyl 1-methyl-4-[1-methyl-4-(3-[[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido]propanamido)imidazol-2-amido]pyrrole-2-carboxylate

[0703] To a stirred solution of 1-methyl-4-(1-methylimidazol-2-amido)pyrrole-2-carboxylic acid (9.00 g, 36.255 mmol, 1.00 equiv) in DMF (150.00 mL) was added HATU (20.68 g, 54.38 mmol, 1.50 equiv), DIEA (14.06 g, 108.77 mmol, 3.00 equiv), and methyl 4-[4-(3-aminopropanamido)-1-methylimidazol-2-amido]-1-methylpyrrole-2-carboxylate (13.89 g, 39.872 mmol, 1.10 equiv) in portions at 0 °C. The resulting mixture was stirred at room temperature for 17.0 h. The reaction was poured into water / ice (450 mL) at 0 °C. The precipitated solid was collected by filtration, washed with HO (3 x 50 mL), and dried under vacuum to give methyl 1-methyl-4-[1-methyl-4-(3-[[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido]propanamido)imidazol-2-amido]pyrrole-2-carboxylate (14.00 g, 63.54%) as a yellow solid. LC / MS: C 26 H 30 N 10 Calculated mass for O6: 578.23, Measured: 579.10 [M+H] + .1H NMR(300MHz,DMSO-d6)δ:10.53(s,1H), 10.29(s,1H), 10.11(s,1H), 8.10(t,J=5.4Hz,1H), 7.52(s,1H), 7.47(s,2H), 7.25(s,1H), 7.17(s, 1H), 6.99(s,1H), 6.97(s,1H), 3.99(s,3H), 3.95(s,3H), 3.84(s,3H), 3.82(s,3H), 3.69(s,3H), 3.42-3.49(m,2H), 2.60(t,J=7.2Hz,2H).

[0704] Step 11: Synthesis of 1-methyl-4-[1-methyl-4-(3-[[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido]propanamido)imidazole-2-afoldamido]pyrrole-2-carboxylic acid

[0705] To a solution of methyl 1-methyl-4-[1-methyl-4-(3-[[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide]propanamido)imidazol-2-amido]pyrrol-2-yl]formamidecarboxylate (14.00 g, 24.20 mmol, 1.00 equiv) in MeOH (70.00 mL) was added LiOH solution (2 M, 72.00 mL, 6.00 equiv). The mixture was stirred at 45 °C for 2.0 h. The resulting mixture was concentrated under reduced pressure. The residue was dissolved in HO (50 mL). The mixture was acidified to pH 3-5 with 2 M HCl. The precipitated solid was collected by filtration, washed with HO (3 x 20 mL), and dried under vacuum. 1-Methyl-4-[1-methyl-4-(3-[[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido]propanamido)imidazole-2-afoldamido]pyrrole-2-carboxylic acid (12.00 g, 81.49%) was obtained as a yellow solid. LC / MS: C 25 H 28 N 10 Calculated mass for O6: 564.22, Measured mass: 565.15 [M+H] + . 1 H NMR(300MHz,DMSO-d6)δ:10.72(s,1H), 10.32(s,1H), 10.08(s,1H), 8.14(t,J=6.0Hz,1H), 7.51(s,1H), 7.47(s,2H), 7.27(s, 1H), 7.23(s,1H), 6.98(s,1H), 6.94(s,1H), 4.00(s,3H), 3.95(s,3H), 3.82(s,6H), 3.44-3.46(m,2H), 2.60(t,J=6.6Hz,2H).

[0706] Step 12: Synthesis of methyl 3-([1-methyl-4-[3-([1-methyl-4-[1-methyl-4-(3-[[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido]propanamido)imidazol-2-amido]pyrrol-2-yl]formamido)propanamido]imidazol-2-yl]formamido)propanoate

[0707] To a stirred solution of 1-methyl-4-[1-methyl-4-(3-[[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido]propanamido)imidazole-2-amido]pyrrole-2-carboxylic acid (12.00 g, 21.26 mmol, 1.00 equiv) in DMF (100.00 mL), HATU (12.12 g, 31.88 mmol, 1.50 equiv), DIEA (8.24 g, 63.77 mmol, 3.00 equiv), and methyl 3-[[4-(3-aminopropanamido)-1-methylimidazol-2-yl]formamido]propanoate (6.95 g, 23.38 mmol, 1.10 equiv) were added in small portions at 0° C. The resulting mixture was stirred at room temperature for 2.0 h. The reaction was poured into water / ice (300 mL) at 0 °C. The precipitated solid was collected by filtration, washed with HO (3 x 30 mL), and dried under vacuum. Methyl 3-([1-methyl-4-[3-([1-methyl-4-[1-methyl-4-(3-[[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido]propanamido)imidazol-2-amido]pyrrol-2-yl]formamido)propanamido]imidazol-2-yl]formamido)propanoate (13.00 g, 64.77%) was obtained as a yellow solid. LC / MS: C 37 H 45 N 15 Calculated mass for O9: 843.35, Found: 844.55 [M+H] + .1H NMR(300MHz,DMSO-d6)δ:10.41(s,1H), 10.37(s,1H), 10.32(s,1H), 9.96(s,1H), 8.08(s,2H), 7.96(s,1H), 7.46(s,1H), 7.42(s,1H), 7.38(s, 1H), 7.24(s,2H), 7.03(s,1H), 6.98(s,1H), 6.93(s,1H), 4.13(s,3H), 3.98(s,3H), 3.95(s,3H), 3.81(s,9H), 3.60(s,6H), 2.57-2.69(m,6H).

[0708] Step 13: Synthesis of 3-([1-methyl-4-[3-([1-methyl-4-[1-methyl-4-(3-[[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido]propanamido)imidazol-2-amido]pyrrol-2-yl]formamido)propanamido]imidazol-2-yl]formamido)propanoic acid (PA-001)

[0709] To a solution of methyl 3-([1-methyl-4-[3-([1-methyl-4-[1-methyl-4-(3-[[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide]propanamido)imidazol-2-amido]pyrrol-2-yl]formamide)propanamido]imidazol-2-yl]formamide)propanoate (10.00 g, 10.59 mmol, 1.00 equiv) in MeOH (60.00 mL), 2 M LiOH (21.20 mL, 42.40 mmol, 4.00 equiv) was added, and the resulting mixture was stirred at 45 °C for 2.0 h. The resulting mixture was concentrated under reduced pressure. The resulting mixture was diluted with water (60 mL). The mixture was acidified to pH 3-5 with 2 M HCl. The precipitated solid was collected by filtration and washed with water (3 × 20 mL). The solid was dried under vacuum to give 3-([1-methyl-4-[3-([1-methyl-4-[1-methyl-4-(3-[[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido]propanamido)imidazol-2-amido]pyrrol-2-yl]formamido)propanamido]imidazol-2-yl]formamido)propanoic acid (8.70 g, 84.14%) as a brown solid. LC / MS: C 36 H 43 N 15Calculated mass for O9: 829.34, Found: 830.25 [M+H] + .1H NMR(300MHz,DMSO-d6)δ:10.46(s,1H), 10.39(s,1H), 10.31(s,1H), 9.93( s,1H), 8.05-8.10(m,2H), 7.87(t,J=6.0Hz,1H), 7.42-7.46(m,3H), 7.20- 7.23(m,2H), 7.07(s,1H), 6.90-6.95(m,2H), 3.95(s,3H), 3.92(s,3H), 3. 89(s,3H), 3.79(s,3H), 3.78(s,3H), 3.38-3.41(m,6H), 2.44-2.59(m,6H).

[0710] Example 2. Synthesis of 1-methyl-4-(1-methyl-4-{1-methyl-4-[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-amido]pyrrol-2-amido}imidazol-2-amido)pyrrole-2-carboxylic acid (PA-047)

[0711] [ka]

[0712] Step 1: Synthesis of ethyl 4-[4-[(tert-butoxycarbonyl)amino]-1-methylpyrrol-2-amido]-1-methylimidazole-2-carboxylate

[0713] To a stirred solution of 4-[(tert-butoxycarbonyl)amino]-1-methylpyrrole-2-carboxylic acid (11.50 g, 47.87 mmol, 1.00 equiv.) in DMF (200.00 mL) was added EDCI (22.94 g, 119.66 mmol, 2.50 equiv.), ethyl 4-amino-1-methylimidazole-2-carboxylate (8.10 g, 47.87 mmol, 1.00 equiv.), and DMAP (14.62 g, 119.66 mmol, 2.50 equiv.) at 0° C. The resulting mixture was stirred at 35° C. for 17.0 h. After the reaction, the reaction mixture was poured into 500 mL of ice / water. The precipitated solid was collected by filtration, washed with water (3×50 mL), and dried under vacuum. This gave ethyl 4-{4-[(tert-butoxycarbonyl)amino]-1-methylpyrrol-2-amido}-1-methylimidazole-2-carboxylate (16.00 g, 85.48% yield) as a pale yellow solid. LC / MS: C 18 H 25 Calculated mass for N5O5: 391.19, Found: 392.30 [M+H] + .

[0714] Step 2: Synthesis of 4-[4-[(tert-butoxycarbonyl)amino]-1-methylpyrrol-2-amido]-1-methylimidazole-2-carboxylic acid

[0715] The procedure was the same as for 4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylic acid (Example 1, Step 1), except that the reaction temperature was room temperature and the reaction time was 1.0 h. Using 970.00 mg of ethyl 4-[4-[(tert-butoxycarbonyl)amino]-1-methylpyrrol-2-amido]-1-methylimidazole-2-carboxylate, 638.00 mg of 4-[4-[(tert-butoxycarbonyl)amino]-1-methylpyrrol-2-amido]-1-methylimidazole-2-carboxylic acid was obtained as a yellow solid (64.36% yield). LC / MS: C 16 H 21 Calculated mass for N5O5: 363.15, Measured: 364.15 [M+H]+ .

[0716] Step 3: Synthesis of methyl 4-(4-{4-[(tert-butoxycarbonyl)amino]-1-methylpyrrole-2-amido}-1-methylimidazol-2-amido)-1-methylpyrrole-2-carboxylate

[0717] 4-{4-[(tert-butoxycarbonyl)amino]-1-methylpyrrole-2-amide}-1-methylimidazole-2-carboxylic acid (6.00 g, 16.51 mmol, 1.00 equiv.) was dissolved in DMF (60.00 mL). PyBOP (8.59 g, 16.51 mmol, 1.00 equiv.), methyl 4-amino-1-methylpyrrole-2-carboxylate (2.55 g, 16.51 mmol, 1.00 equiv.), and DIEA (6.40 g, 49.536 mmol, 3.00 equiv.) were added sequentially to the solution at 0 °C. The mixture was warmed to room temperature and stirred for 1.0 h. Upon completion of the reaction, the mixture was added dropwise to ice water (150 mL). A solid formed, which was filtered, washed with water (2x15 mL) and dried under vacuum to give methyl 4-(4-{4-[(tert-butoxycarbonyl)amino]-1-methylpyrrole-2-amido}-1-methylimidazol-2-amido)-1-methylpyrrole-2-carboxylate (7.10 g, 86.08%) as a reddish-brown solid. LC / MS: C 23 H 29 Calculated mass for N7O6: 499.21, Found: 500.15 [M+H] + .

[0718] Step 4: Synthesis of methyl 4-[4-(4-amino-1-methylpyrrol-2-amido)-1-methylimidazol-2-amido]-1-methylpyrrole-2-carboxylate

[0719] To a stirred solution of methyl 4-(4-{4-[(tert-butoxycarbonyl)amino]-1-methylpyrrole-2-amido}-1-methylimidazole-2-amido)-1-methylpyrrole-2-carboxylate (250.00 mg, 0.500 mmol, 1.00 equiv) in DCM (2.50 mL) was added TFA (0.50 mL) dropwise at room temperature. The resulting mixture was stirred at room temperature for 1.0 h. The resulting mixture was concentrated in vacuo to afford methyl 4-[4-(4-amino-1-methylpyrrole-2-amido)-1-methylimidazole-2-amido]-1-methylpyrrole-2-carboxylate (250.00 mg, crude) as a tan oil. LC / MS: C 18 H 21 Calculated mass for N7O4: 399.17, Found: 400.35 [M+H] + .

[0720] Step 5: Synthesis of methyl 1-methyl-4-(1-methyl-4-{1-methyl-4-[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-amido]pyrrol-2-amido}imidazol-2-amido)pyrrole-2-carboxylate

[0721] To a stirred solution of 1-methyl-4-(1-methylimidazol-2-amido)pyrrole-2-carboxylic acid (156.62 mg, 0.63 mmol, 0.90 equiv) in DMF (2.00 mL), PyBOP (361.16 mg, 0.69 mmol, 1.00 equiv), methyl 4-[4-(4-amino-1-methylpyrrol-2-amido)-1-methylimidazol-2-amido]-1-methylpyrrole-2-carboxylate (280.00 mg, 0.70 mmol, 1.00 equiv), and DIEA (453.02 mg, 3.51 mmol, 5.00 equiv) were added in small portions at 0° C. The resulting mixture was stirred at room temperature for 1.0 h. The reaction mixture was directly purified by reverse-phase column chromatography using the following conditions: column, C18 column; mobile phase, ACN (0.05% TFA) in water, 5% to 70% gradient over 50 min; detector, UV 254 nm. Fractions were combined and concentrated. Methyl 1-methyl-4-(1-methyl-4-{1-methyl-4-[1-methyl-4-(1-methylimidazole-2-amido)pyrrol-2-amido]pyrrol-2-amido}imidazole-2-amido)pyrrole-2-carboxylate (240.00 mg, 51.65% yield) was obtained as a white solid. LC / MS: C 29 H 31 N 11 Calculated mass for O6: 629.25, Measured mass: 630.25 [M+H] + .

[0722] Step 6: Synthesis of 1-methyl-4-(1-methyl-4-{1-methyl-4-[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-amido]pyrrol-2-amido}imidazol-2-amido)pyrrole-2-carboxylic acid (PA-047)

[0723] The procedure was the same as for 4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylic acid (Example 1, Step 3). 240.00 mg of methyl 1-methyl-4-(1-methyl-4-{1-methyl-4-[1-methyl-4-(1-methylimidazole-2-amido)pyrrol-2-amido]pyrrol-2-amido}imidazole-2-amido)pyrrole-2-carboxylate was used to obtain 178.00 mg of 1-methyl-4-(1-methyl-4-{1-methyl-4-[1-methyl-4-(1-methylimidazole-2-amido)pyrrol-2-amido]pyrrol-2-amido}imidazole-2-amido)pyrrole-2-carboxylic acid as a white solid (62.96% yield). LC / MS: C 28 H 29 N 11 Calculated mass for O6: 615.23, Measured mass: 616.25 [M+H] + .

[0724] Example 3. Synthesis of 1-methyl-4-(1-methyl-4-{1-methyl-4-[2-(1-methylimidazol-2-yl)-3H-1,3-benzodiazol-5-amido]pyrrol-2-amido}imidazole-2-amido)pyrrole-2-carboxylic acid (PA-047-P2BNZ)

[0725] [ka]

[0726] Step 1: Synthesis of 2-(1-methylimidazol-2-yl)-3H-1,3-benzodiazole-5-carboxylic acid

[0727] To a stirred solution of 3,4-diaminobenzoic acid (1.00 g, 6.57 mmol, 1.00 equiv) in DMF (15.00 mL) was added 1-methylimidazole-2-carbaldehyde (723.00 mg, 6.57 mmol, 1.00 equiv). The resulting mixture was stirred at 80 °C for 1.0 h. FeCl3.6H2O (53.00 mg, 0.20 mmol, 0.03 equiv) was added in small portions. The resulting mixture was stirred at 120 °C under air for 1.0 h. The resulting mixture was concentrated in vacuo. The residue was purified by silica gel column chromatography (0-10% EA / MeOH) to give 2-(1-methylimidazol-2-yl)-3H-1,3-benzodiazole-5-carboxylic acid (130.00 mg, 8.17% yield) as a yellow solid. LC / MS: C 12 H 10 Calculated mass for N4O2: 242.08, Measured: 243.10 [M+H] + .

[0728] Step 2: Synthesis of methyl 1-methyl-4-(1-methyl-4-{1-methyl-4-[2-(1-methylimidazol-2-yl)-3H-1,3-benzodiazol-5-amido]pyrrol-2-amido}imidazole-2-amido)pyrrole-2-carboxylate

[0729] The procedure was the same as for methyl 4-(4-{4-[(tert-butoxycarbonyl)amino]-1-methylpyrrol-2-amido}-1-methylimidazole-2-amido)-1-methylpyrrole-2-carboxylate (Example 2, Step 3). Using 120.00 mg of 2-(1-methylimidazol-2-yl)-3H-1,3-benzodiazole-5-carboxylic acid, 290.00 mg of methyl 1-methyl-4-(1-methyl-4-{1-methyl-4-[2-(1-methylimidazol-2-yl)-3H-1,3-benzodiazole-5-amido]pyrrol-2-amido}imidazole-2-amido)pyrrole-2-carboxylate was obtained as a yellow solid (93.87% yield). LC / MS: C 30 H 29 N 11Calculated mass for O5: 623.23, Measured: 624.50 [M+H] + .

[0730] Step 3: Synthesis of 1-methyl-4-(1-methyl-4-{1-methyl-4-[2-(1-methylimidazol-2-yl)-3H-1,3-benzodiazol-5-amido]pyrrole-2-amido}imidazole-2-amido)pyrrole-2-carboxylic acid

[0731] The procedure was the same as for 4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylic acid (Example 1, Step 3), except that the reaction time was 3.0 hours. Using 280.00 mg of methyl 1-methyl-4-(1-methyl-4-{1-methyl-4-[2-(1-methylimidazol-2-yl)-3H-1,3-benzodiazol-5-amido]pyrrol-2-amido}imidazole-2-amido)pyrrole-2-carboxylate, 210.00 mg of 1-methyl-4-(1-methyl-4-{1-methyl-4-[2-(1-methylimidazol-2-yl)-3H-1,3-benzodiazol-5-amido]pyrrol-2-amido}imidazole-2-amido)pyrrole-2-carboxylic acid was obtained as a yellow solid (76.73% yield). LC / MS:C 29 H 27 N 11 Calculated mass for O5: 609.21, Measured: 610.45 [M+H] + .

[0732] Example 4. Synthesis of 1-methyl-4-[3-({1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrol-2-yl}formamido)propanamido]imidazole-2-carboxylic acid (PA-048)

[0733] [ka]

[0734] Step 1: Synthesis of 2-(1-methylimidazol-2-yl)-3H-1,3-benzodiazole-5-carboxylic acid

[0735] The procedure was the same as (Example 1, Step 7), but the reaction time was 1.0 h. Using 2.00 g of ethyl 4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylate, 2.00 g of crude ethyl 4-(3-aminopropanamido)-1-methyl-1H-imidazole-2-carboxylate was obtained as an off-white solid. LC / MS: C 10 H 16 Calculated mass for N4O3: 240.12, Measured mass: 241.10 [M+H] + .

[0736] Step 2: Ethyl 1-methyl-4-[3-({1-methyl-4-[1-methyl-4-(3-{[1-

[0737] Synthesis of methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrol-2-yl}formamido)propanamido]imidazole-2-carboxylate

[0738] The procedure was the same as for methyl 4-(4-{4-[(tert-butoxycarbonyl)amino]-1-methylpyrrol-2-amido}-1-methylimidazol-2-amido)-1-methylpyrrole-2-carboxylate (Example 2, step 3). Using 270.00 mg of 1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamide)imidazole-2-amido]pyrrole-2-carboxylic acid, 460.00 mg of ethyl 1-methyl-4-[3-({1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamide)imidazole-2-amido]pyrrol-2-yl}formamide)propanamide]imidazole-2-carboxylate was obtained as an off-white solid (96.45% yield). LC / MS: C 35 H 42 N 14 Calculated mass for O8: 786.33, Found: 809.60 [M+Na] + .

[0739] Step 3: Synthesis of 1-methyl-4-[3-({1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamide)imidazol-2-amido]pyrrol-2-yl}formamide)propanamide]imidazole-2-carboxylic acid

[0740] The procedure was the same as for 4-[3-[(Tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylic acid (Example 1, Step 3). Using 470.00 mg of ethyl 1-methyl-4-[3-({1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamide)imidazol-2-amido]pyrrol-2-yl}formamide)propanamide]imidazole-2-carboxylate, 400.00 mg of 1-methyl-4-[3-({1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamide)imidazol-2-amido]pyrrol-2-yl}formamide)propanamide]imidazole-2-carboxylic acid was obtained as an off-white solid (74.41% yield). LC / MS: C 33 H 38 N 14 Calculated mass for O8: 758.30, Found: 759.55 [M+H] + .

[0741] Example 5. Synthesis of 1-methyl-4-[2-(1-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrol-2-amido}cyclopropyl)acetamido]imidazole-2-carboxylic acid (PA-048-P6CP)

[0742] [ka]

[0743] Step 1: Synthesis of ethyl 4-(2-{1-[(tert-butoxycarbonyl)amino]cyclopropyl}acetamido)-1-methylimidazole-2-carboxylate

[0744] The procedure was the same as for ethyl 3-[(4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazol-2-yl)formamido]propanoate. 260.00 mg of {1-[(tert-butoxycarbonyl)amino]cyclopropyl}acetic acid was used to obtain 320.00 mg of ethyl 4-(2-{1-[(tert-butoxycarbonyl)amino]cyclopropyl}acetamido)-1-methylimidazole-2-carboxylate as a white solid (68.69% yield). LC / MS: C 17 H 26 Calculated mass for N4O5: 366.19, Measured: 367.10 [M+H] + .

[0745] Step 2: Synthesis of ethyl 4-[2-(1-aminocyclopropyl)acetamido]-1-methylimidazole-2-carboxylate

[0746] The procedure was the same as for methyl 4-[4-(4-amino-1-methylpyrrol-2-amido)-1-methylimidazole-2-amido]-1-methylpyrrole-2-carboxylate (Example 2, Step 4). 260.00 mg of ethyl 4-(2-{1-[(tert-butoxycarbonyl)amino]cyclopropyl}acetamido)-1-methylimidazole-2-carboxylate was used to obtain 260.00 mg of crude ethyl 4-[2-(1-aminocyclopropyl)acetamido]-1-methylimidazole-2-carboxylate as a yellow oil. LC / MS: C 12 H 18 Calculated mass for N4O3: 266.14, Measured mass: 267.05 [M+H] + .

[0747] Step 3: Synthesis of ethyl 1-methyl-4-[2-(1-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrol-2-amido}cyclopropyl)acetamido]imidazole-2-carboxylate

[0748] The procedure was the same as for methyl 4-(4-{4-[(tert-butoxycarbonyl)amino]-1-methylpyrrol-2-amido}-1-methylimidazole-2-amido)-1-methylpyrrole-2-carboxylate (Example 2, Step 3). Using 150.00 mg of ethyl 4-[2-(1-aminocyclopropyl)acetamido]-1-methylimidazole-2-carboxylate, 220.00 mg of ethyl 1-methyl-4-[2-(1-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-amido]pyrrol-2-amido}cyclopropyl)acetamido]imidazole-2-carboxylate was obtained as a yellow solid (37.48% yield). LC / MS: C 37 H 44 N 14 Calculated mass for O8: 812.35, Measured mass: 813.35 [M+H] + .

[0749] Step 4: Synthesis of 1-methyl-4-[2-(1-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrol-2-amido}cyclopropyl)acetamido]imidazole-2-carboxylic acid

[0750] The procedure was the same as for 4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylic acid (Example 1, step 3). Using 240.00 mg of ethyl 1-methyl-4-[2-(1-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrol-2-amido}cyclopropyl)acetamido]imidazole-2-carboxylate, 155.00 mg of 1-methyl-4-[2-(1-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrol-2-amido}cyclopropyl)acetamido]imidazole-2-carboxylic acid was obtained as a white solid (54.85% yield). LC / MS: C 35 H 40 N 14 Calculated mass for O8: 784.32, Found: 785.55 [M+H] + .

[0751] Example 6. Synthesis of 1-methyl-4-[3-methyl-3-({1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrol-2-yl}formamido)butanamido]imidazole-2-carboxylic acid (PA-048-P6GM)

[0752] [ka]

[0753] Step 1: Synthesis of ethyl 4-{3-[(tert-butoxycarbonyl)amino]-3-methylbutanamido}-1-methylimidazole-2-carboxylate

[0754] The procedure was the same as for methyl 3-[(4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazol-2-yl)formamido]propanoate, but the reaction time was 17.0 hours and the crude product was not purified by silica gel column. 385.00 mg of 3-[(tert-butoxycarbonyl)amino]-3-methylbutanoic acid was used to give 540.00 mg of crude ethyl 4-{3-[(tert-butoxycarbonyl)amino]-3-methylbutanamido}-1-methylimidazole-2-carboxylate as a white solid. LC / MS: C 17 H 28 Calculated mass for N4O5: 368.21, Measured: 369.15 [M+H] + .

[0755] Step 2: Synthesis of ethyl 4-(3-amino-3-methylbutanamido)-1-methylimidazole-2-carboxylate

[0756] The procedure was the same as for methyl 4-[4-(4-amino-1-methylpyrrol-2-amido)-1-methylimidazole-2-amido]-1-methylpyrrole-2-carboxylate (Example 2, Step 3). 540.00 mg of ethyl 4-{3-[(tert-butoxycarbonyl)amino]-3-methylbutanamido}-1-methylimidazole-2-carboxylate was used to obtain 540.00 mg of crude ethyl 4-(3-amino-3-methylbutanamido)-1-methylimidazole-2-carboxylate as a yellow oil. LC / MS: C 12 H 20 Calculated mass for N4O3: 268.15, Measured mass: 269.30 [M+H] + .

[0757] Step 3: Synthesis of ethyl 1-methyl-4-[3-methyl-3-({1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrol-2-yl}formamido)butanamido]imidazole-2-carboxylate

[0758] To a stirred solution of 1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-amido]pyrrole-2-carboxylic acid (600.00 mg, 1.06 mmol, 1.00 equiv) in DMF (5.00 mL) was added DIEA (412.07 mg, 3.19 mmol, 3.00 equiv), ethyl 4-(3-amino-3-methylbutanamido)-1-methylimidazole-2-carboxylate (313.67 mg, 1.17 mmol, 1.10 equiv), and PyBOP (1106.11 mg, 2.13 mmol, 2.00 equiv) in small portions at 0° C. The resulting mixture was stirred at room temperature for 17.0 h. The reaction was poured into ice / water (15 mL). The precipitated solid was collected by filtration, washed with HO (3 x 2 mL), and dried under vacuum. The reaction mixture was purified by silica gel column chromatography, eluting with DCM / MeOH (5:1) to give ethyl-methyl-4-[3-methyl-3-({1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamide)imidazol-2-amido]pyrrol-2-yl}formamide)butanamide]imidazole-2-carboxylate (800.00 mg, 77.49% yield) as a yellow oil. LC / MS: C 37 H 46 N 14 Calculated mass for O8: 814.36, Measured: 408.50 [M / 2 / +H]+.

[0759] Step 4: Synthesis of 1-methyl-4-[3-methyl-3-({1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamido)imidazol-2-amido]pyrrol-2-yl}formamide)butanamido]imidazole-2-carboxylic acid

[0760] To a stirred solution of ethyl 1-methyl-4-(3-methyl-3-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamide)imidazol-2-amido]pyrrole-2-carbonyloxy}butanamide)imidazole-2-carboxylate (725.00 mg, 0.89 mmol, 1.00 equiv) in MeOH (8.00 mL) was added LiOH solution (2 M in HO, 1.80 mL, 4.00 equiv) at room temperature. The resulting mixture was stirred at 45 °C for 2.0 h. The resulting mixture was concentrated under reduced pressure. The residue was dissolved in HO (9 mL). The mixture was acidified to pH 3-5 with 2 M HCl. The mixture was then concentrated, and the residue was directly purified by reverse-phase column chromatography using the following conditions: column, C18 silica gel; mobile phase, ACN (0.05% TFA) in water, 10% to 50% gradient over 50 minutes; detector, UV 254 nm. The fractions were combined and concentrated. 1-Methyl-4-[3-methyl-3-({1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamide)imidazol-2-amido]pyrrol-2-yl}formamide)butanamide]imidazole-2-carboxylic acid (700.00 mg, 95.88% yield) was obtained as a yellow solid. LC / MS: C 35 H 42 N 14 Calculated mass for O8: 786.33, Measured mass: 394.40 [M / 2+H] + .

[0761] Example 7. Synthesis of 1-methyl-4-[(1r,3r)-3-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrol-2-amido}cyclobutanamido]imidazole-2-carboxylic acid (PA-048-P6CB)

[0762] [ka]

[0763] Step 1: Synthesis of ethyl 1-methyl-4-[(1r,3r)-3-[(tert-butoxycarbonyl)amino]cyclobutanamido]imidazole-2-carboxylate

[0764] The procedure was the same as for methyl 3-[(4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazol-2-yl)formamido]propanoate. Using 200.00 mg of (1r,3r)-3-[(tert-butoxycarbonyl)amino]cyclobutane-1-carboxylic acid, 330.00 mg of ethyl 1-methyl-4-[(1r,3r)-3-[(tert-butoxycarbonyl)amino]cyclobutanamido]imidazole-2-carboxylate was obtained as an orange solid (96.93% yield). LC / MS: C 17 H 26 Calculated mass for N4O5: 366.19, Measured: 367.25 [M+H] + .

[0765] Step 2: Synthesis of ethyl 1-methyl-4-[(1r,3r)-3-aminocyclobutanamido]imidazole-2-carboxylate

[0766] The procedure was the same as for methyl 4-[4-(4-amino-1-methylpyrrol-2-amido)-1-methylimidazole-2-amido]-1-methylpyrrole-2-carboxylate (Example 2, Step 3). 145.00 mg of ethyl 1-methyl-4-[(1r,3r)-3-[(tert-butoxycarbonyl)amino]cyclobutanamido]imidazole-2-carboxylate was used to obtain 145.00 mg of crude ethyl 1-methyl-4-[(1r,3r)-3-aminocyclobutanamido]imidazole-2-carboxylate as a yellow oil. LC / MS: C 12 H 18 Calculated mass for N4O3: 266.14, Measured mass: 267.10 [M+H] + .

[0767] Step 3: Synthesis of ethyl 1-methyl-4-[(1r,3r)-3-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrol-2-amido}cyclobutanamido]imidazole-2-carboxylate

[0768] The procedure was the same as for methyl 4-(4-{4-[(tert-butoxycarbonyl)amino]-1-methylpyrrol-2-amido}-1-methylimidazol-2-amido)-1-methylpyrrole-2-carboxylate (Example 2, Step 3). Using 105.00 mg of ethyl 1-methyl-4-[(1r,3r)-3-aminocyclobutanamido]imidazole-2-carboxylate, 250.00 mg of ethyl 1-methyl-4-[(1r,3r)-3-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrol-2-amido}cyclobutanamido]imidazole-2-carboxylate was obtained as a pale yellow solid (78.15% yield). LC / MS:C 37 H 44 N 14Calculated mass for O8: 812.35, Measured mass: 813.50 [M+H] + .

[0769] Step 4: Synthesis of 1-methyl-4-[(1r,3r)-3-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrol-2-amido}cyclobutanamido]imidazole-2-carboxylic acid

[0770] The procedure was the same as for 4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylic acid (Example 1, step 3). Using 250.00 mg of ethyl 1-methyl-4-[(1r,3r)-3-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-amido]pyrrol-2-amido}cyclobutanamido]imidazole-2-carboxylate, 210.00 mg of 1-methyl-4-[(1r,3r)-3-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-amido]pyrrol-2-amido}cyclobutanamido]imidazole-2-carboxylic acid was obtained as a pale yellow solid (87.00% yield). LC / MS: C 35 H 40 N 14 Calculated mass for O8: 784.32, Found: 785.40 [M+H] + .

[0771] Example 8. Synthesis of 1-methyl-4-(1-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrole-2-carbonyl}azetidin-3-amido)imidazole-2-carboxylic acid (PA-048-P6AZ)

[0772] [ka]

[0773] Step 1: Synthesis of ethyl 4-[1-(tert-butoxycarbonyl)azetidin-3-amido]-1-methylimidazole-2-carboxylate

[0774] The procedure was the same as for methyl 3-[(4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazol-2-yl)formamido]propanoate (Example 1, Step 6). 300.00 mg of ethyl 4-amino-1-methylimidazole-2-carboxylate was used to obtain 450.00 mg of ethyl 4-[1-(tert-butoxycarbonyl)azetidin-3-amido]-1-methylimidazole-2-carboxylate as a white solid (72.02% yield). LC / MS: C 16 H 24 Calculated mass for N4O5: 352.17, Measured: 353.10 [M+H] + .

[0775] Step 2: Synthesis of ethyl 4-(azetidin-3-amido)-1-methylimidazole-2-carboxylate

[0776] The procedure was the same as for methyl 4-[4-(4-amino-1-methylpyrrol-2-amido)-1-methylimidazole-2-amido]-1-methylpyrrole-2-carboxylate (Example 2, Step 4). 140.00 mg of ethyl 4-[1-(tert-butoxycarbonyl)azetidin-3-amido]-1-methylimidazole-2-carboxylate was used to obtain 140.00 mg of crude ethyl 4-(azetidin-3-amido)-1-methylimidazole-2-carboxylate as a yellow oil. LC / MS: C 11 H 16 Calculated mass for N4O3: 252.12, Measured: 253.10 [M+H] + .

[0777] Step 3: Synthesis of ethyl 1-methyl-4-(1-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrole-2-carbonyl}azetidin-3-amido)imidazole-2-carboxylate

[0778] The procedure was the same as for methyl 4-(4-{4-[(tert-butoxycarbonyl)amino]-1-methylpyrrol-2-amido}-1-methylimidazole-2-amido)-1-methylpyrrole-2-carboxylate (Example 2, Step 3). Using 140.00 mg of ethyl 4-(azetidin-3-amido)-1-methylimidazole-2-carboxylate, 260.00 mg of ethyl 1-methyl-4-(1-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazole-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-amido]pyrrole-2-carbonyl}azetidin-3-amido)imidazole-2-carboxylate was obtained as a white solid (53.96% yield). LC / MS: C 36 H 42 N 14 Calculated mass for O8: 798.33, Found: 799.45. [M+H] + .

[0779] Step 4: Synthesis of 1-methyl-4-(1-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamido)imidazol-2-amido]pyrrole-2-carbonyl}azetidin-3-amido)imidazole-2-carboxylic acid

[0780] The procedure was the same as for 4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylic acid (Example 1, step 3). Using 250.00 mg of ethyl 1-methyl-4-(1-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrole-2-carbonyl}azetidin-3-amido)imidazole-2-carboxylate, 160.00 mg of 1-methyl-4-(1-{1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrole-2-carbonyl}azetidin-3-amido)imidazole-2-carboxylic acid was obtained as a yellow solid (62.08% yield). LC / MS: C 34 H 38 N 14 Calculated mass for O8: 770.30, Found: 771.60 [M+H] + .

[0781] Example 9. Synthesis of 3-({1-methyl-4-[3-({4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]-1-[14-(2,5,8,11-tetraoxatrican-13-yl)-2,5,8,11-tetraoxa-14-azanonadecane-19-yl]pyrrol-2-yl}formamido)propanamido]imidazol-2-yl}formamido)propanoic acid (PA-035-Bis(mPEG4)

[0782] [ka]

[0783] Step 1: Synthesis of ethyl 1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-carboxylate

[0784] The procedure was the same as for methyl 4-(4-{4-[(tert-butoxycarbonyl)amino]-1-methylpyrrol-2-amido}-1-methylimidazol-2-amido)-1-methylpyrrole-2-carboxylate (Example 2, Step 3). 2.00 g of 1-methyl-4-(1-methylimidazol-2-amido)pyrrole-2-carboxylic acid was used to give 4.00 g of ethyl 1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-carboxylate as an off-white solid. LC / MS: C 21 H 26 Calculated mass for N8O5: 470.20, Found: 471.15 [M+H] + .

[0785] Step 2: Synthesis of ethyl 1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-carboxylate

[0786] The procedure was the same as for 4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylic acid (Example 1, Step 3), except that the reaction solvent was MeOH / THF (2:1), the reaction temperature was room temperature, and the reaction time was 1.0 h. Using 4.00 g of ethyl 1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-carboxylate, 2.85 g of 1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-carboxylic acid was obtained as an off-white solid. LC / MS: C19 H 22 Calculated mass for N8O5: 442.17, Measured mass: 443.30 [M+H] + .

[0787] Step 3: Synthesis of ethyl 4-[(tert-butoxycarbonyl)amino]-1H-pyrrole-2-carboxylate

[0788] To a solution of ethyl 4-nitro-1H-pyrrole-2-carboxylate (5.00 g, 27.15 mmol, 1.00 equiv) in EtOH (50.00 mL) and EA (50.00 mL) was added Pd / C (1.00 g, 20% w / w) and (Boc)2O (11.85 g, 54.304 mmol, 2.00 equiv). The reaction was then stirred under an H2 atmosphere at room temperature for 17.0 h. The mixture was filtered, and the filtrate was concentrated to give ethyl 4-[(tert-butoxycarbonyl)amino]-1H-pyrrole-2-carboxylate (5.00 g, 71.54% yield) as a white solid. LC / MS: C 12 H 18 Calculated mass for N2O4: 254.13, Found: 255.25 [M+H] + .

[0789] Step 4: Synthesis of ethyl 1-(5-bromopentyl)-4-[(tert-butoxycarbonyl)amino]pyrrole-2-carboxylate

[0790] To a solution of ethyl 4-[(tert-butoxycarbonyl)amino]pyrrole-2-carboxylate (4.00 g, 15.73 mmol, 1.00 equiv) in MeCN (70.00 mL) was added KCO (6.52 g, 47.19 mmol, 3.00 equiv), and 1,5-dibromopentane (36.17 g, 157.30 mmol, 10.00 equiv). The reaction was then stirred at 70 °C for 36.0 h. The reaction mixture was filtered, and the filtrate was concentrated. The residue was purified by silica gel column chromatography eluting with PE / EA (5:1) to give ethyl 1-(5-bromopentyl)-4-[(tert-butoxycarbonyl)amino]pyrrole-2-carboxylate (4.82 g, 67.43% yield) as an orange solid. LC / MS: C 17 H 27 Calculated mass for BrN2O4: 402.12, Found: 403.00 [M+H] + .

[0791] Step 5: Synthesis of ethyl 4-[(tert-butoxycarbonyl)amino]-1-[14-(2,5,8,11-tetraoxatrican-13-yl)-2,5,8,11-tetraoxa-14-azanonadecane-19-yl]pyrrole-2-carboxylate

[0792] The procedure was the same as for ethyl 1-(5-bromopentyl)-4-[(tert-butoxycarbonyl)amino]pyrrole-2-carboxylate (Example 9, Step 4), except that the reaction time was 17.0 hours. The crude product was purified by reverse-phase column chromatography. Using 1.00 g of ethyl 1-(5-bromopentyl)-4-[(tert-butoxycarbonyl)amino]pyrrole-2-carboxylate, 1.00 g of ethyl 4-[(tert-butoxycarbonyl)amino]-1-[14-(2,5,8,11-tetraoxatrican-13-yl)-2,5,8,11-tetraoxa-14-azanonadecane-19-yl]pyrrole-2-carboxylate was obtained as a tan oil (61.29% yield). LC / MS: C 35 H 65 N3O 12Calculated mass: 719.46, Measured mass: 720.45 [M+H] + .

[0793] Step 6: Synthesis of ethyl 4-amino-1-[14-(2,5,8,11-tetraoxatrican-13-yl)-2,5,8,11-tetraoxa-14-azanonadecane-19-yl]pyrrole-2-carboxylate

[0794] The procedure was the same as for methyl 4-[4-(4-amino-1-methylpyrrol-2-amido)-1-methylimidazol-2-amido]-1-methylpyrrole-2-carboxylate (Example 2, Step 4). 350.00 mg of ethyl 4-[(tert-butoxycarbonyl)amino]-1-[14-(2,5,8,11-tetraoxatrican-13-yl)-2,5,8,11-tetraoxa-14-azanonadecane-19-yl]pyrrole-2-carboxylate was used to obtain 350.00 mg of crude ethyl 4-amino-1-[14-(2,5,8,11-tetraoxatrican-13-yl)-2,5,8,11-tetraoxa-14-azanonadecane-19-yl]pyrrole-2-carboxylate as a yellow oil. LC / MS: C 30 H 57 N3O 10 Calculated mass: 619.40, Measured mass: 620.60 [M+H] + .

[0795] Step 7: Synthesis of ethyl 4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]-1-[14-(2,5,8,11-tetraoxatrican-13-yl)-2,5,8,11-tetraoxa-14-azanonadecane-19-yl]pyrrole-2-carboxylate

[0796] The procedure was the same as for methyl 1-methyl-4-(1-methyl-4-{1-methyl-4-[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-amido]pyrrol-2-amido}imidazol-2-amido)pyrrole-2-carboxylate (INT-450-11). Using 220.00 mg of 1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-carboxylic acid, 460.00 mg of ethyl 4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]-1-[14-(2,5,8,11-tetraoxatrican-13-yl)-2,5,8,11-tetraoxa-14-azanonadecane-19-yl]pyrrole-2-carboxylate was obtained as a yellow oil (88.59% yield). LC / MS: C 49 H 77 N 11 O 14 Calculated mass: 1043.57, Measured mass: 1044.60 [M+H] + .

[0797] Step 8: Synthesis of 4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]-1-[14-(2,5,8,11-tetraoxatrican-13-yl)-2,5,8,11-tetraoxa-14-azanonadecane-19-yl]pyrrole-2-carboxylic acid

[0798] The procedure was the same as for 1-methyl-4-[3-methyl-3-({1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamide)imidazol-2-amido]pyrrol-2-yl}formamide)butanamide]imidazole-2-carboxylic acid, but the reaction time was 17.0 h. 450.00 mg of ethyl 4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]-1-[14-(2,5,8,11-tetraoxatrican-13-yl)-2,5,8,11-tetraoxa-14-azanonadecane-19-yl]pyrrole-2-carboxylate was used to prepare 400. Obtained 0.00 mg of 4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]-1-[14-(2,5,8,11-tetraoxatrican-13-yl)-2,5,8,11-tetraoxa-14-azanonadecane-19-yl]pyrrole-2-carboxylic acid as a yellow oil (91.34% yield). LC / MS: C 47 H 73 N 11 O 14 Calculated mass: 1015.53, Measured mass: 1039.05 [M+Na] + .

[0799] Step 9: Synthesis of ethyl 3-({1-methyl-4-[3-({4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]-1-[14-(2,5,8,11-tetraoxatrican-13-yl)-2,5,8,11-tetraoxa-14-azanonadecane-19-yl]pyrrol-2-yl}formamido)propanamido]imidazol-2-yl}formamido)propanoate

[0800] The procedure was the same as for methyl 1-methyl-4-(1-methyl-4-{1-methyl-4-[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-amido]pyrrol-2-amido}imidazol-2-amido)pyrrole-2-carboxylate (INT-450-11). 390.00 mg of 4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]-1-[14-(2,5,8,11-tetraoxatrican-13-yl)-2,5,8,11-tetraoxa-14-azanonadecane-19-yl]pyrrole-2-carboxylic acid was used to prepare 180.00 mg of ethyl 3-({1-methyl-4-[3-({4-[1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]-1-[14-(2,5,8,11-tetraoxatrican-13-yl)-2,5,8,11-tetraoxa-14-azanonadecane-19-yl]pyrrole-2-carboxylic acid. -4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]-1-[14-(2,5,8,11-tetraoxatrican-13-yl)-2,5,8,11-tetraoxa-14-azanonadecane-19-yl]pyrrol-2-yl}formamido)propanamido]imidazol-2-yl}formamido)propanoate was obtained as a yellow oil (35.82% yield). LC / MS: C 60 H 92 N 16 O 17 Calculated mass: 1308.68, Measured mass: 1309.90 [M+H] + .

[0801] Step 10: Synthesis of 3-({1-methyl-4-[3-({4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]-1-[14-(2,5,8,11-tetraoxatrican-13-yl)-2,5,8,11-tetraoxa-14-azanonadecane-19-yl]pyrrol-2-yl}formamido)propanamido]imidazol-2-yl}formamido)propanoic acid (PA-035-Bis(mPEG4))

[0802] The procedure was the same as for 1-methyl-4-[3-methyl-3-({1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamide)imidazol-2-amido]pyrrol-2-yl}formamide)butanamide]imidazole-2-carboxylic acid, but the reaction solvent was MeOH / THF (1:1), and the reaction time was 17.0 h. 170.00 mg of ethyl 3-({1-methyl-4-[3-({4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]-1-[14-(2,5,8,11-tetraoxatrican-13-yl)-2,5,8,11-tetraoxa-14-azanonadecane-19-yl]pyrrol-2-yl}formamido)propanamido]imidazol-2-yl}formamido)propanoate was used to prepare 80.0 Obtained 0 mg of 3-({1-methyl-4-[3-({4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]-1-[14-(2,5,8,11-tetraoxatrican-13-yl)-2,5,8,11-tetraoxa-14-azanonadecane-19-yl]pyrrol-2-yl}formamido)propanamido]imidazol-2-yl}formamido)propanoic acid as a yellow oil (48.09% yield). LC / MS: C 58 H 88 N 16 O 17 Calculated mass: 1280.65, measured mass: 641.90 [M / 2+H] + .

[0803] Example 10. Synthesis of 3-{[4-(3-{[1-(6-azidohexyl)-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamido)imidazol-2-amido]pyrrol-2-yl]formamide}propanamido)-1-methylimidazol-2-yl]formamide}propanoic acid (PA-001-P5C6N3)

[0804] [ka]

[0805] Step 1: Synthesis of ethyl 1-(6-bromohexyl)-4-[(tert-butoxycarbonyl)amino]pyrrole-2-carboxylate

[0806] The procedure was the same as for ethyl 1-(5-bromopentyl)-4-[(tert-butoxycarbonyl)amino]pyrrole-2-carboxylate (Example 9, Step 4), but the reaction time was 17.0 hours. Using 8.10 g of ethyl 4-[(tert-butoxycarbonyl)amino]-1H-pyrrole-2-carboxylate, 10.70 g of ethyl 1-(6-bromohexyl)-4-[(tert-butoxycarbonyl)amino]pyrrole-2-carboxylate was obtained as a white solid (80.49% yield). LC / MS: C 18 H 29 Calculated mass for BrN2O4: 416.13, Found: 417.20, 419.20 [M+H, M+2+H] + .

[0807] Step 2: Synthesis of ethyl 1-(6-azidohexyl)-4-[(tert-butoxycarbonyl)amino]pyrrole-2-carboxylate

[0808] To a 250 mL flask was added ethyl 1-(6-bromohexyl)-4-[(tert-butoxycarbonyl)amino]pyrrole-2-carboxylate (10.48 g, 25.11 mmol, 1.00 equiv.), DMF (100.00 mL), and NaN (2.15 g, 33.15 mmol, 1.32 equiv.), and the reaction was stirred at room temperature for 1.0 h. The reaction was quenched by the addition of saturated NH Cl (aqueous) (100 mL) at 0 °C. The resulting mixture was extracted with EtOAc (3 × 100 mL). The combined organic layers were washed with water (1 × 100 mL), NaCl solution (1 × 100 mL), and dried over anhydrous Na SO . After filtration, the filtrate was concentrated under reduced pressure. This gave ethyl 1-(6-azidohexyl)-4-[(tert-butoxycarbonyl)amino]pyrrole-2-carboxylate (9.00 g, crude) as a yellow oil. LC / MS: C 18 H 29 Calculated mass for N5O4: 379.22, Found: 402.40 [M+Na] + .

[0809] Step 3: Synthesis of ethyl 4-amino-1-(6-azidohexyl)pyrrole-2-carboxylate

[0810] The procedure was the same as for methyl 4-[4-(4-amino-1-methylpyrrol-2-amido)-1-methylimidazol-2-amido]-1-methylpyrrole-2-carboxylate (Example 2, Step 4). 300.00 mg of ethyl 1-(6-azidohexyl)-4-[(tert-butoxycarbonyl)amino]pyrrole-2-carboxylate was used to obtain 300.00 mg of crude ethyl 4-amino-1-(6-azidohexyl)pyrrole-2-carboxylate as a yellow oil. LC / MS: C 13 H 21 Calculated mass for N5O2: 279.17, Found: 280.35 [M+H] + .

[0811] Step 4: Synthesis of ethyl 1-(6-azidohexyl)-4-(4-{3-[(tert-butoxycarbonyl)amino]propanamido}-1-methylimidazol-2-amido)pyrrole-2-carboxylate

[0812] The procedure was the same as for ethyl 4-(1-methylimidazol-2-amido)-1H-pyrrole-2-carboxylate (INT-459-2). 300.00 mg of ethyl 4-amino-1-(6-azidohexyl)pyrrole-2-carboxylate was used to obtain 345.00 mg of ethyl 1-(6-azidohexyl)-4-(4-{3-[(tert-butoxycarbonyl)amino]propanamido}-1-methylimidazol-2-amido)pyrrole-2-carboxylate as a pink solid (56.00% yield). LC / MS: C 26 H 39 Calculated mass for N9O6: Exact mass: 573.30, Measured mass: 574.55 [M+H] + .

[0813] Step 5: Synthesis of ethyl 1-(6-azidohexyl)-4-(4-{3-[(tert-butoxycarbonyl)amino]propanamido}-1-methylimidazol-2-amido)pyrrole-2-carboxylate

[0814] The procedure was the same as for methyl 4-[4-(4-amino-1-methylpyrrol-2-amido)-1-methylimidazol-2-amido]-1-methylpyrrole-2-carboxylate (Example 2, Step 4). 340.00 mg of ethyl 1-(6-azidohexyl)-4-(4-{3-[(tert-butoxycarbonyl)amino]propanamido}-1-methylimidazol-2-amido)pyrrole-2-carboxylate was used to obtain 340.00 mg of crude ethyl 4-[4-(3-aminopropanamido)-1-methylimidazol-2-amido]-1-(6-azidohexyl)pyrrole-2-carboxylate as a yellow oil. (56.00% yield). LC / MS: C 21 H 31Calculated mass for N9O4: Exact mass: 473.25, Measured mass: 474.40 [M+H] + .

[0815] Step 6: Synthesis of ethyl 1-(6-azidohexyl)-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrole-2-carboxylate

[0816] The procedure was the same as for ethyl 1-methyl-4-[3-methyl-3-({1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamide)imidazol-2-amido]pyrrol-2-yl}formamide)butanamide]imidazole-2-carboxylate (Example 6, Step 3). Using 140.00 mg of 1-methyl-4-(1-methylimidazol-2-amido)pyrrole-2-carboxylic acid, 320.00 mg of ethyl 1-(6-azidohexyl)-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrole-2-carboxylate was obtained as a yellow oil (80.62% yield). LC / MS: C 32 H 41 N 13 Calculated mass for O6: 703.33, Measured mass: 704.55 [M+H] + .

[0817] Step 7: Synthesis of 1-(6-azidohexyl)-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrole-2-carboxylic acid

[0818] The procedure was the same as 4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylic acid (Example 1, Step 3), but the reaction temperature was 50° C. and the reaction time was 17.0 hours. Using 310.00 mg of ethyl 1-(6-azidohexyl)-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-amido]pyrrole-2-carboxylate, 270.00 mg of 1-(6-azidohexyl)-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-amido]pyrrole-2-carboxylic acid was obtained as a yellow solid (90.90% yield). LC / MS: C 30 H 37 N 13 Calculated mass for O6: Exact mass: 675.30, Measured mass: 676.60 [M+H] + .

[0819] Step 8: Synthesis of ethyl 3-{[4-(3-{[1-(6-azidohexyl)-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrol-2-yl]formamido}propanamido)-1-methylimidazol-2-yl]formamido}propanoate

[0820] The procedure was the same as for methyl 4-(4-{4-[(tert-butoxycarbonyl)amino]-1-methylpyrrol-2-amido}-1-methylimidazol-2-amido)-1-methylpyrrole-2-carboxylate (Example 2, step 3). Using 200.00 mg of 1-(6-azidohexyl)-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamido)imidazol-2-amido]pyrrole-2-carboxylic acid, 290.00 mg of crude ethyl 3-{[4-(3-{[1-(6-azidohexyl)-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamido)imidazol-2-amido]pyrrol-2-yl]formamide}propanamido)-1-methylimidazol-2-yl]formamide}propanoate was obtained as a white solid. LC / MS: C 43 H 56 N 18 Calculated mass for O9: 968.45, Found: 969.30 [M+H] + .

[0821] Step 9: Synthesis of 3-{[4-(3-{[1-(6-azidohexyl)-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamido)imidazol-2-amido]pyrrol-2-yl]formamide}propanamido)-1-methylimidazol-2-yl]formamide}propanoic acid

[0822] The procedure was the same as in 4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylic acid (Example 1, Step 3), except that the reaction solvent was MeOH / THF (1:1), the reaction temperature was room temperature, and the reaction time was 1.0 h. 280.00 mg of ethyl 3-{[4-(3-{[1-(6-azidohexyl)-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamido)imidazol-2-amido]pyrrol-2-yl]formamide}propanamido)-1-methylimidazol-2-yl]formamide}propanoate was used to obtain 240.0 mg of ethyl 3-{[4-(3-{[1-(6-azidohexyl)-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamido)-1-methylimidazol-2-yl]formamide}propanoate. 0 mg of 3-{[4-(3-{[1-(6-azidohexyl)-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamido)imidazol-2-amido]pyrrol-2-yl]formamide}propanamido)-1-methylimidazol-2-yl]formamide}propanoic acid was obtained as a white solid (88.27% yield). LC / MS: C 41 H 52 N 18 Calculated mass for O9: 940.42, Found: 941.20 [M+H] +

[0823] Synthesis of representative ligands

[0824] Example 11: Synthesis of 4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylic acid

[0825] [ka]

[0826] Step 1: Synthesis of 4-bromo-7-methoxy-1-(4-methylbenzenesulfonyl)pyrrolo[2,3-c]pyridine

[0827] To a solution of 4-bromo-7-methoxy-1H-pyrrolo[2,3-c]pyridine (5.00 g, 22.12 mmol, 1.00 equiv) in DMF (20.00 mL) was added NaH (60%, 796.46 mg, 33.19 mmol, 1.50 equiv) in small portions at 0 °C. The reaction was then stirred for 15 min, followed by the addition of TsCl (6.30 g, 33.19 mmol, 1.50 equiv) at 0 °C. The resulting mixture was stirred at room temperature for an additional 2.0 h. The mixture was poured into ice water (60 mL). The solid was filtered, washed with HO (10 mL), and dried to give 4-bromo-7-methoxy-1-(4-methylbenzenesulfonyl)pyrrolo[2,3-c]pyridine (7.50 g, 83.98% yield) as a white solid. LCMS:C 15 H 13 Calculated mass for BrN2O3S: 379.98, Found: 380.95, 382.95 [M+H, M+2+H] + .

[0828] Step 2: Synthesis of ethyl 4-bromo-7-methoxy-1-tosyl-1H-pyrrolo[2,3-c]pyridine-2-carboxylate

[0829] To a solution of 4-bromo-7-methoxy-1-tosyl-1H-pyrrolo[2,3-c]pyridine (6.30 g, 16.58 mmol, 1.00 equiv) in THF (80.00 mL), LDA (2 M in THF, 12.50 mL, 24.87 mmol, 1.50 equiv) was added dropwise at -78 °C. The mixture was stirred at -78 °C to -50 °C for 1.0 h, followed by the dropwise addition of ClCOOEt (2.69 g, 24.87 mmol, 1.50 equiv). After 2.0 h, the reaction mixture was quenched with saturated NH4Cl (aqueous), and the residue was extracted with EA (3 × 300 mL). The organic phases were combined, dried over Na2SO4, filtered, and concentrated. The residue was purified by silica gel column chromatography eluting with PE / EA=10:1 to give ethyl 4-bromo-7-methoxy-1-tosyl-1H-pyrrolo[2,3-c]pyridine-2-carboxylate (7.00 g, 90.11% yield) as a white solid. LCMS: C 18 H 17Calculated mass for BrN2O5S: 452.00, Found: 453.00, 455.00 [M+H, M+2+H] + .

[0830] Step 3: Synthesis of ethyl 4-bromo-7-oxo-1-tosyl-6,7-dihydro-1H-pyrrolo[2,3-c]pyridine-2-carboxylate

[0831] To a stirred solution of ethyl 4-bromo-7-methoxy-1-tosyl-1H-pyrrolo[2,3-c]pyridine-2-carboxylate (4.00 g, 8.850 mmol, 1.00 equiv) in CHCN (80.00 mL) was added TMSCl (1.45 g, 13.28 mmol, 1.50 equiv) and NaI (2.00 g, 13.28 mmol, 1.50 equiv) in small portions at room temperature under a N atmosphere. The mixture was stirred at room temperature for 1.0 h, and then HO (238.95 mg, 13.28 mmol, 1.50 equiv) was added dropwise at 65 °C. The mixture was stirred at 65 °C for 2.0 h. The reaction mixture was cooled to room temperature. The precipitate was filtered, washed with water (50 mL), and dried in vacuo. Ethyl 4-bromo-7-oxo-1-tosyl-6,7-dihydro-1H-pyrrolo[2,3-c]pyridine-2-carboxylate (4.30 g, crude) was obtained as a brown solid. LCMS: C 17 H 15 Calculated mass for BrN2O5S: 437.99, Found: 438.95, 440.95 [M+H, M+2+H] + .

[0832] Step 4: Synthesis of ethyl 4-bromo-6-methyl-1-(4-methylbenzenesulfonyl)-7-oxopyrrolo[2,3-c]pyridine-2-carboxylate

[0833] To a solution of ethyl 4-bromo-7-oxo-1-tosyl-6,7-dihydro-1H-pyrrolo[2,3-c]pyridine-2-carboxylate (4.30 g, 9.82 mmol, 1.00 equiv.) in DMF (20.00 mL) was added CsCO (3.83 g, 11.78 mmol, 1.20 equiv.), and MeI (1.67 g, 11.78 mmol, 1.20 equiv.) was added dropwise to the reaction. The reaction mixture was stirred at room temperature under a N atmosphere for 17.0 hours. The mixture was poured into ice water (60 mL). The solid was filtered, washed with HO (10 mL) and dried to give ethyl 4-bromo-6-methyl-1-(4-methylbenzenesulfonyl)-7-oxopyrrolo[2,3-c]pyridine-2-carboxylate (4.30 g, crude) as a brown solid. LCMS: C 18 H 17 Calculated mass for BrN2O5S: 452.00, Found: 453.15, 455.15 [M+H, M+2+H] + .

[0834] Step 5: Synthesis of ethyl 6-methyl-1-(4-methylbenzenesulfonyl)-7-oxo-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[2,3-c]pyridine-2-carboxylate

[0835] To a solution of ethyl 4-bromo-6-methyl-1-(4-methylbenzenesulfonyl)-7-oxopyrrolo[2,3-c]pyridine-2-carboxylate (1.00 g, 2.21 mmol, 1.00 equiv.) in THF (30.00 mL), bis(pinacolato)diboron (1.12 g, 4.41 mmol, 2.00 equiv.), KOAc (650.00 mg, 6.62 mmol, 3.00 equiv.), X-Phos Pd G2 (175.00 mg, 0.22 mmol, 0.10 equiv.), and X-Phos (106.00 mg, 0.22 mmol, 0.10 equiv.) were added under a N2 atmosphere at room temperature. The resulting mixture was stirred at 75 °C for 17 h under a N2 atmosphere. The mixture was concentrated, and 40 mL of H2O was added to the residue. The mixture was then extracted with EA (3 x 40 mL). The combined organic phases were washed with NaCl solution (40 mL) and dried over Na2SO4. The solid was filtered, and the filtrate was concentrated. Ethyl 6-methyl-1-(4-methylbenzenesulfonyl)-7-oxo-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[2,3-c]pyridine-2-carboxylate (2.20 g, crude) was obtained as a yellow solid. The crude product was used directly in the next step. LCMS: C 21 H 25 Calculated mass for BN2O5S: 500.18, Measured mass: 501.10 [M+H] + .

[0836] Step 6: Synthesis of 2-(4-fluoro-2,6-dimethylphenyl)-4,4,5,5-tetramethyl-1,3,2-dioxaborolane

[0837] To a solution of 2-bromo-5-fluoro-1,3-dimethylbenzene (5.00 g, 24.62 mmol, 1.00 equiv) in THF (15.00 mL) was added n-BuLi (2.5 M, 14.77 mL, 36.94 mmol, 1.50 equiv) dropwise at −78° C. under a N atmosphere. The mixture was stirred at −78° C. for 3.0 hours, and then 2-isopropoxy-4,4,5,5-tetramethyl-1,3,2-dioxaborolane (6.87 g, 36.94 mmol, 1.50 equiv) was added dropwise at −78° C. The resulting mixture was allowed to warm to room temperature and stirred for 3.0 hours. After the reaction, the reaction was quenched with water (20 mL) at 0° C. The resulting mixture was extracted with EtOAc (3×20 mL). The combined organic layers were dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure to give 2-(4-fluoro-2,6-dimethylphenyl)-4,4,5,5-tetramethyl-1,3,2-dioxaborolane (6.70 g, crude) as a pale yellow oil. The crude product was used directly in the next step without further purification. LC / MS: C 14 H 20 Calculated mass for BFO2: 250.15, Measured mass: 251.30 [M+1] + .

[0838] Step 7: Synthesis of 4-fluoro-2,6-dimethylphenol

[0839] To a stirred solution of 2-(4-fluoro-2,6-dimethylphenyl)-4,4,5,5-tetramethyl-1,3,2-dioxaborolane (6.70 g, 26.79 mmol, 1.00 equiv.) in THF (20.00 mL), NaOH (1.61 g, 40.25 mmol, 1.50 equiv.) and HO (9.99 mL, 428.59 mmol, 16.00 equiv.) were added dropwise at −10° C. under a N atmosphere. The resulting mixture was stirred at room temperature for 17.0 h. After the reaction, the mixture was acidified to pH = 1 with HCl (aqueous 2 M). The resulting mixture was extracted with EtOAc (3 × 20 mL). The combined organic layers were washed with saturated NaHCO (aqueous) (1 × 10 mL) and saturated NaSO (aqueous) (1 × 10 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with hexane / EtOAc (12:1) to give 4-fluoro-2,6-dimethylphenol (2.70 g, 66.88% yield) as a white solid. 1 HNMR(400MHz,DMSO)δ:8.12(s,1H), 6.73(d,J=9.3Hz,2H), 2.16(s,6H).

[0840] Step 8: Synthesis of methyl 3-bromo-4-(4-fluoro-2,6-dimethylphenoxy)benzoate

[0841] To a stirred solution of 4-fluoro-2,6-dimethylphenol (2.70 g, 19.26 mmol, 1.00 equiv.) and methyl 3-bromo-4-fluorobenzoate (4.94 g, 21.20 mmol, 1.10 equiv.) in DMSO (20.00 mL) was added CsCO (9.41 g, 28.90 mmol, 1.50 equiv.) at room temperature. The resulting mixture was stirred at 80 °C for 2.0 h. After the reaction, the reaction was quenched by adding water (30 mL) at room temperature. The resulting mixture was extracted with EtOAc (3 × 20 mL). The combined organic layers were dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluting with hexane / EtOAc (12:1) to give methyl 3-bromo-4-(4-fluoro-2,6-dimethylphenoxy)benzoate (6.80 g, 94.95% yield) as a white solid. LC / MS: C 16 H 14 Calculated mass for BrFO3: 352.01, Found: 353.15 [M+H] + .

[0842] Step 9: Synthesis of 2-[3-bromo-4-(4-fluoro-2,6-dimethylphenoxy)phenyl]propan-2-ol

[0843] To a stirred solution of methyl 3-bromo-4-(4-fluoro-2,6-dimethylphenoxy)benzoate (2.00 g, 5.66 mmol, 1.00 equiv) in THF (10.00 mL) was added bromo(methyl)magnesium (3.00 M in 2-Me-THF, 11.33 mL, 33.98 mmol, 6.00 equiv) at 0° C. under a N atmosphere. The resulting mixture was stirred at 0° C. for 1.0 h under a N atmosphere. After the reaction, the reaction was quenched by adding saturated NH Cl (aqueous) (10 mL) at 0° C. The resulting mixture was extracted with EtOAc (3×10 mL). The combined organic layers were combined and dried over anhydrous Na SO . After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluting with hexane / EtOAc (10:1) to give 2-[3-bromo-4-(4-fluoro-2,6-dimethylphenoxy)phenyl]propan-2-ol (1.80 g, 77.40% yield) as a white solid. LC / MS: C 17 H 18 Calculated mass for BrFO2: 352.05, Measured mass: 335.00 [M-OH] + .

[0844] Step 10: Synthesis of ethyl 4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-1-(4-methylbenzenesulfonyl)-7-oxopyrrolo[2,3-c]pyridine-2-carboxylate

[0845] 2-[3-bromo-4-(4-fluoro-2,6-dimethylphenoxy)phenyl]propan-2-ol (500.00 mg, 1.42 mmol, 1.00 equiv.) and ethyl 6-methyl-1-(4-methylbenzenesulfonyl)-7-oxo-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[2,3-c]pyridine-2-carboxamide in dioxane (16.00 mL) and HO (4.00 mL). To a stirred solution of Pd(dba)(2,3-dimethyl-2,4-trioxa-6-phenyl-6-phosphaadamantane) (1.42 g, 2.83 mmol, 2.00 equiv.) was added Pd(dba)(2,3-dimethyl-2,4-trioxa-6-phenyl-6-phosphaadamantane) (129.62 mg, 0.14 mmol, 0.10 equiv.), KPO (901.39 mg, 4.25 mmol, 3.00 equiv.), and 1,3,5,7-tetramethyl-2,4,8-trioxa-6-phenyl-6-phosphaadamantane (82.00 mg, 0.28 mmol, 0.20 equiv.) under a N atmosphere at room temperature. The resulting mixture was stirred at 75 °C under a N atmosphere for 1.0 h. The reaction was quenched with water at room temperature. The resulting mixture was extracted with EtOAc (3 × 20 mL). The combined organic layers were dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluting with hexane / EtOAc (1:1) to give ethyl 4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-1-(4-methylbenzenesulfonyl)-7-oxopyrrolo[2,3-c]pyridine-2-carboxylate (620.00 mg, 48.76% yield) as a white solid. LC / MS: C 35 H 35 Calculated mass for FN2O7S: 646.21, Found: 647.20 [M+H] + .

[0846] Step 11: Synthesis of 4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylic acid

[0847] To a stirred solution / mixture of ethyl 4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-1-(4-methylbenzenesulfonyl)-7-oxopyrrolo[2,3-c]pyridine-2-carboxylate (600.00 mg, 0.93 mmol, 1.00 equiv.) in MeOH (15.00 mL) was added KOH (2 M, 3.71 mL, 7.42 mmol, 8.00 equiv.) at room temperature. The resulting mixture was stirred at 40 °C for 4.0 h. After the reaction, the resulting mixture was concentrated in vacuo. The residue was then dissolved in water (10 mL) and acidified to pH 3 with HCl (2 M aqueous). The precipitated solid was collected by filtration and washed with water (3 × 10 mL). The solid was concentrated under vacuum to give 4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylic acid (390.00 mg, 64.26% yield) as a white solid. LC / MS: C 26 H 25 Calculated mass for FN2O5: 464.17, Measured mass: 465.15 [M+H] + .

[0848] Example 12. Synthesis of 1-ethyl-4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxopyrrolo[2,3-c]pyridine-2-carboxylic acid

[0849] [ka]

[0850] Step 1: Synthesis of ethyl 4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylate

[0851] To a stirred solution of ethyl 4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-1-(4-methylbenzenesulfonyl)-7-oxopyrrolo[2,3-c]pyridine-2-carboxylate (3.40 g, 5.26 mmol, 1.00 equiv.) in ethyl alcohol (50.00 mL) was added sodium ethoxide (894.40 mg, 13.14 mmol, 2.50 equiv.) at room temperature. The resulting mixture was stirred at room temperature for 2.0 hours. After the reaction, the reaction mixture was poured into citric acid solution (3.32 g, 3.00 equiv., 125 mL). The resulting mixture was then extracted with EtOAc (3 × 150 mL). The combined organic layers were washed with water (2 × 50 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure. The residue was washed with diethyl ether (3x10 mL) to give ethyl 4-[2-(4-fluoro-2,6-)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylate (2.00 g, 71.78% yield) as a white solid. LCMS: C 28 H 29 Calculated mass for FN2O5: 492.21, Found: 493.40 [M+H] + .

[0852] Step 2: Synthesis of ethyl 1-ethyl-4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxopyrrolo[2,3-c]pyridine-2-carboxylate

[0853] The procedure was the same as for ethyl 1-(5-bromopentyl)-4-[(tert-butoxycarbonyl)amino]pyrrole-2-carboxylate, but the reaction time was 2.0 hours. Using 500.00 mg of ethyl 4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylate, 500.00 mg of ethyl 1-ethyl-4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxopyrrolo[2,3-c]pyridine-2-carboxylate was obtained as a white solid (94.61% yield). LC / MS: C 30 H 33 Calculated mass for FN2O5: 520.24, Found: 521.35 [M+H] + .

[0854] Step 3: Synthesis of 1-ethyl-4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxopyrrolo[2,3-c]pyridine-2-carboxylic acid

[0855] The procedure was the same as for 4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylic acid, except that the reaction solvent was MeOH / THF (1:5). 500.00 mg of ethyl 1-ethyl-4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxopyrrolo[2,3-c]pyridine-2-carboxylate was used to obtain 514.00 mg of crude 1-ethyl-4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxopyrrolo[2,3-c]pyridine-2-carboxylic acid as a pale yellow solid. LC / MS: C 28 H 29 Calculated mass for FN2O5: 492.21, Found: 493.15 [M+H] + .

[0856] Example 13: Synthesis of 4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-methanesulfonylphenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylic acid

[0857] [ka]

[0858] Step 1: Synthesis of ethyl 4-bromo-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylate

[0859] To a stirred solution of ethyl 6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylate (10.00 g, 45.41 mmol, 1.00 equiv.) in tetrahydrofuran (150.00 mL), NBS (8.08 g, 45.41 mmol, 1.00 equiv.) and p-TsOH (3.91 g, 22.70 mmol, 0.50 equiv.) were added. The resulting mixture was stirred at room temperature for 1.0 h. The resulting mixture was concentrated in vacuo. The residue was purified by silica gel column chromatography (0-10% MeOH / DCM) to give ethyl 4-bromo-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylate (13.00 g, 95.71% yield) as a yellow solid. LC / MS: C 11 H 11 Calculated mass for BrN2O3: 298.00, Found: 299.00, 301.00 [M+H, M+2+H] + .

[0860] Step 2: Synthesis of ethyl 6-methyl-7-oxo-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrrolo[2,3-c]pyridine-2-carboxylate

[0861] To a stirred solution of ethyl 4-bromo-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylate (13.00 g, 43.46 mmol, 1.00 equiv.) in dioxane (150.00 mL) was added bis(pinacolato)diboron (22.07 g, 86.92 mmol, 2.00 equiv.), Pd(dba).CHCl (4.00 g, 4.36 mmol, 0.10 equiv.), and AcOK (8.53 g, 86.92 mmol, 2.00 equiv.). The final reaction mixture was microwaved at 120 °C for 1.0 h. The reaction was run on a 1.0 g scale and repeated 13 times. The reaction mixtures were then combined and processed together. 150 mL of HO was added, and the resulting mixture was extracted with EA (3 × 150 mL). The organic layers were combined, dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (50-70% EA / PE) to give ethyl 6-methyl-7-oxo-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrrolo[2,3-c]pyridine-2-carboxylate (10.00 g, 66.47% yield) as a yellow solid. LC / MS: C 17 H 23 Calculated mass for BN2O5: 346.17, Found: 347.20 [M+H] + .

[0862] Step 3: Synthesis of 5-fluoro-2-(4-methanesulfonyl-2-nitrophenoxy)-1,3-dimethylbenzene

[0863] The procedure was the same as for methyl 3-bromo-4-(4-fluoro-2,6-dimethylphenoxy)benzoate (SM15-52), except that the reaction temperature was 120° C. and the reaction time was 1.0 h. Using 2.00 g of 4-fluoro-2,6-dimethylphenol, 4.60 g of the desired product was obtained as an off-white solid (94.05% yield). 1H NMR(300MHz,DMSO-d6)δ:8.60(s,1H), 8.07(d,J=9.0Hz,1H), 7.16(d,J=9.0Hz,2H), 6.88(d,J=9.0Hz,1H), 3.31(s,3H), 2.09(s,6H).

[0864] Step 4: Synthesis of 2-(2,4-difluorophenoxy)-5-methanesulfonylaniline

[0865] To a stirred solution of 1-(2,4-difluorophenoxy)-4-methanesulfonyl-2-nitrobenzene (500.00 mg, 1.52 mmol, 1.00 equiv) in THF (10.00 mL) was added Pd / C (100.00 mg, 20% w / w). The mixture was hydrogenated at room temperature under a H atmosphere using a hydrogen balloon for 17.0 h. The resulting mixture was filtered, and the filter cake was washed with EA (3×20 mL). The filtrate was concentrated under reduced pressure to give 2-(2,4-difluorophenoxy)-5-methanesulfonylaniline (450.00 mg, crude) as a pale yellow oil. The crude product was used directly in the next step without further purification. LC / MS: C 15 H 16 Calculated mass for FNO3S: 309.08 Measured mass: 310.10 [M+H] + .

[0866] Step 5: Synthesis of 5-fluoro-2-(2-iodo-4-methanesulfonylphenoxy)-1,3-dimethylbenzene

[0867] To a stirred solution of 2-(4-fluoro-2,6-dimethylphenoxy)-5-methanesulfonylaniline (500.00 mg, 1.62 mmol, 1.00 equiv) in dioxane (5.00 mL) was added concentrated hydrogen chloride (1.00 mL) dropwise at 0° C. The resulting mixture was stirred at 0° C. for 10.0 minutes. To the above mixture was added sodium nitrite (133.81 mg, 1.94 mmol, 1.20 equiv) at 0° C. The resulting mixture was stirred at 0° C. for an additional 1.0 hour. To the above mixture was added KI (536.60 mg, 3.23 mmol, 2.00 equiv) at 0° C. The resulting mixture was stirred at 40° C. for an additional 17.0 hours. After the reaction, the reaction was quenched with water (5 mL) at room temperature. The resulting mixture was extracted with EtOAc (3×10 mL). The combined organic layers were washed with water (1x10 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by preparative TLC (PE / EA 5:1) to give 5-fluoro-2-(2-iodo-4-methanesulfonylphenoxy)-1,3-dimethylbenzene (250.00 mg, 31.65% yield) as a pale yellow oil. LC / MS: C 15 H 14 Calculated mass for FiO3S: 419.97, Measured: 442.95 [M+Na] + .

[0868] Step 6: Synthesis of ethyl 4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-methanesulfonylphenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylate

[0869] To a stirred solution of 5-fluoro-2-(2-iodo-4-methanesulfonylphenoxy)-1,3-dimethylbenzene (380.00 mg, 0.90 mmol, 1.00 equiv) in toluene (6.00 mL) and water (1.50 mL), ethyl 6-methyl-7-oxo-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-pyrrolo[2,3-c]pyridine-2-carboxylate (469.56 mg, 1.36 mmol, 1.50 equiv), KPO (383.88 mg, 1.81 mmol, 2.00 equiv), and Pd(dtbpf)Cl (58.93 mg, 0.09 mmol, 0.10 equiv) were added under a N atmosphere at room temperature. The resulting mixture was stirred at 70°C under a N2 atmosphere for 2.0 hours. After the reaction, the reaction was quenched with water (10 mL) at room temperature. The resulting mixture was extracted with EtOAc (3 x 10 mL). The combined organic layers were washed with water (1 x 5 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography eluting with PE / EA (0-100%) to give ethyl 4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-methanesulfonylphenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylate (250.00 mg, 52.86% yield) as a dark yellow solid. LC / MS: C 26 H 25 Calculated mass for FN2O6S: 512.14, Measured mass: 513.30 [M+H] + .

[0870] Step 7: Synthesis of 4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-methanesulfonylphenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylic acid

[0871] To a stirred solution of ethyl 4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-methanesulfonylphenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylate (240.00 mg, 0.47 mmol, 1.00 equiv.) in tetrahydrofuran (1.00 mL) and water (5.00 mL), sodium hydroxide (74.91 mg, 1.87 mmol, 4.00 equiv.) was added at room temperature. The resulting mixture was stirred at 70°C for 2.0 hours. After the reaction, the resulting mixture was concentrated under reduced pressure. The residue was dissolved in water (5 mL). The mixture was acidified to pH = 4 with HCl (aqueous 2 M). The precipitated solid was collected by filtration, washed with water (3 x 5 mL), and dried under vacuum. This gave 4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-methanesulfonylphenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylic acid (170.00 mg, 73.44% yield) as a pale yellow solid. LC / MS: C 24 H 21 Calculated mass for FN2O6S: 484.11, Found: 485.10 [M+H] + .

[0872] Example 14: Synthesis of 4-[5-(ethanesulfonyl)-2-(4-fluoro-2,6-dimethylphenoxy)phenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylic acid

[0873] [ka]

[0874] Step 1: Synthesis of 2-bromo-4-(ethanesulfonyl)-1-fluorobenzene

[0875] To a stirred solution of fluoresone (1.00 g, 5.31 mmol, 1.00 equiv) in H2SO4 (6.00 mL) was added NBS (1.04 g, 5.84 mmol, 1.10 equiv). The resulting mixture was stirred at room temperature for 16.0 h. The resulting mixture was poured into ice water (20 mL). The precipitated solid was collected by filtration, washed with PE (50 mL), and dried to give 2-bromo-4-(ethanesulfonyl)-1-fluorobenzene (890.00 mg, 62.71% yield) as a yellow solid. LC / MS: Mass calculated for C8H8BrFO2S: 265.94, 267.05, 268.95 [M+H, M+H +2 ].

[0876] Step 2: Synthesis of 2-[2-bromo-4-(ethanesulfonyl)phenoxy]-5-fluoro-1,3-dimethylbenzene

[0877] The procedure was the same as for methyl 3-bromo-4-(4-fluoro-2,6-dimethylphenoxy)benzoate (SM15-52), except that the reaction temperature was 110° C. and the reaction time was 1.0 h. The crude product was used in the next step without purification. Using 870.00 mg of 1,3-dibromo-5-(ethanesulfonyl)-2-fluorobenzene, 950.00 mg of 2-[2-bromo-4-(ethanesulfonyl)phenoxy]-5-fluoro-1,3-dimethylbenzene was obtained as a yellow solid (97.56% yield). LC / MS: C 16 H 16 Calculated mass for BrFO3S: 386.00, Found: 387.05, 389.05 [M+H, M+2+H] + .

[0878] Step 3: Synthesis of 2-[2-bromo-4-(ethanesulfonyl)phenoxy]-5-fluoro-1,3-dimethylbenzene

[0879] The procedure was the same as for ethyl 4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-methanesulfonylphenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylate (INT-444-4), except the reaction temperature was 75 °C and the reaction time was 1.0 h. Using 950.00 mg of 2-[2-bromo-4-(ethanesulfonyl)phenoxy]-5-fluoro-1,3-dimethylbenzene, 870.00 mg of ethyl 4-[5-(ethanesulfonyl)-2-(4-fluoro-2,6-dimethylphenoxy)phenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylate was obtained as a yellow solid (67.35% yield). LC / MS: C 27 H 27 Calculated mass for FN2O6S: 526.15, Found: 527.35 [M+H] + .

[0880] Step 4: Synthesis of 4-[5-(ethanesulfonyl)-2-(4-fluoro-2,6-dimethylphenoxy)phenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylic acid

[0881] The procedure was the same as for 4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylic acid. Using 860.00 mg of ethyl 4-[5-(ethanesulfonyl)-2-(4-fluoro-2,6-dimethylphenoxy)phenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylate, 590.00 mg of 4-[5-(ethanesulfonyl)-2-(4-fluoro-2,6-dimethylphenoxy)phenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylic acid was obtained as a yellow solid (72.46% yield). LC / MS: C 25 H 23 Calculated for FN2O6S: 498.12, Found: 499.25 [M+H] + .

[0882] Example 15. Synthesis of (R)-2-(4-(4-chlorophenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetic acid

[0883] [ka]

[0884] The procedure was the same as for methyl 4-[4-(4-amino-1-methylpyrrol-2-amido)-1-methylimidazol-2-amido]-1-methylpyrrole-2-carboxylate (Example 2, Step 3). 500.00 mg of tert-butyl (R)-2-(4-(4-chlorophenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate was used to obtain 500.00 mg of the crude desired product as a yellow oil. LC / MS: C 19 H 17 Calculated for ClN4O2S: 400.08, Found: 400.95 [M+H] + .

[0885] Example 16 Synthesis of (S)-2-(4-(4-(16-((2-(1H-indol-3-yl)ethyl)(((9H-fluoren-9-yl)methoxy)carbonyl)amino)hexadecanamido)phenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetic acid

[0886] [ka]

[0887] Step 1: Synthesis of tert-butyl (S)-2-(4-(4-((diphenylmethylene)amino)phenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate

[0888] To a solution of tert-butyl (S)-2-(4-(4-chlorophenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate (1000.00 mg, 2.19 mmol, 1.00 equiv) in toluene (30.00 mL) was added diphenylmethaneimide. CsCO3 (594.89 mg, 3.28 mmol, 1.50 equiv.), CsCO3 (2138.91 mg, 6.56 mmol, 3.00 equiv.), Pd(dba)3.CHCl3 (200.38 mg, 0.22 mmol, 0.10 equiv.), and S-Phos (89.83 mg, 0.22 mmol, 0.10 equiv.) were added at room temperature under a N2 atmosphere. The resulting mixture was stirred at 110 °C for 17.0 h under a N2 atmosphere. After cooling to room temperature, the reaction solvent was removed under reduced pressure. The resulting residue was purified by silica gel chromatography (40-50% EA / PE) to give the desired product (1180.00 mg, 80.65% yield) as a yellow solid. LC / MS: C 36 H 35 Calculated mass for N5O2S: 601.25, Measured: 602.35 [M+H] + .

[0889] Step 2: Synthesis of tert-butyl (S)-2-(4-(4-aminophenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate

[0890] To a stirred solution of tert-butyl (S)-2-(4-(4-((diphenylmethylene)amino)phenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate (1180.00 mg, 1.96 mmol, 1.00 equiv) in THF (10.00 mL) was added 1 M HCl (2.00 mL) dropwise at room temperature. The resulting mixture was stirred at room temperature for 2.0 h. The mixture was diluted with EA (100 mL), and the organic layer was separated. The organic layer was concentrated and purified by preparative HPLC under the following conditions: Column: CHIRALPAK IH-3, 3.0*50 mm, 3 μm; Mobile phase, MeOH (0.1% DEA; Flow rate: 2 mL / min; Gradient: 10% B; 220 nm. The fractions were combined and directly lyophilized. Tert-butyl (S)-2-(4-(4-aminophenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate (600.00 mg, 67.83%) was obtained as a white solid. LC / MS: C 23 H 27 Calculated mass for N5O2S: 437.19, Found: 438.10 [M+H] + .

[0891] Step 3: Synthesis of tert-butyl (S)-2-(4-(4-(16-bromohexadecanamido)phenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate

[0892] 16-Bromohexadecanoic acid (191.59 mg, 0.57 mmol, 1.00 equiv.) was dissolved in DMF (3.00 mL). NMI (93.82 mg, 1.14 mmol, 2.00 equiv.), TCFH (320.62 mg, 1.14 mmol, 2.00 equiv.), and tert-butyl (S)-2-(4-(4-aminophenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate (250.00 mg, 0.57 mmol, 1.00 equiv.) were added sequentially to the solution at 0 °C. The mixture was allowed to warm to room temperature and stirred at room temperature for 1.0 h. Upon completion of the reaction, the mixture was added dropwise to ice water (7 mL). A solid formed, which was filtered, washed with water (2x5 mL) and dried under vacuum to give tert-butyl (S)-2-(4-(4-(16-bromohexadecanamido)phenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate (450.00 mg, 90.77%) as a pale yellow solid. LC / MS: C 39 H 56 Calculated mass for BrNOS: 753.32, Found: 754.55, 756.55 [M+H, M+2+H] + .

[0893] Step 4: Synthesis of tert-butyl (S)-2-(4-(4-(16-((2-(1H-indol-3-yl)ethyl)amino)hexadecanamido)phenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate

[0894] To a solution of tert-butyl (S)-2-(4-(4-(16-bromohexadecanamido)phenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate (480.00 mg, 0.64 mmol, 1.00 equiv) and tryptamine (101.88 mg, 0.64 mmol, 1.00 equiv) in ACN (3.00 mL) was added KCO (527.29 mg, 3.82 mmol, 6.00 equiv), and the resulting mixture was stirred at 70 °C for 17.0 h. The resulting mixture was filtered, and the filter cake was washed with CHCN (3 × 5 mL). The filtrate was concentrated under reduced pressure and purified by reverse flash chromatography using the following conditions: column, C18 silica gel; mobile phase, 0.05% NH4HCO3 and CH3CN in water, 50% to 60% gradient over 15 minutes; detector, UV 254-220 nm. Fractions were combined and concentrated to give tert-butyl (S)-2-(4-(4-(16-((2-(1H-indol-3-yl)ethyl)amino)hexadecanamido)phenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate (100.00 mg, 15.08% yield) as a white solid. LC / MS: C 49 H 67 Calculated mass for N7O3S: 833.50, Found: 834.75 [M+H] + .

[0895] Step 5: Synthesis of tert-butyl (S)-2-(4-(4-(16-((2-(1H-indol-3-yl)ethyl)(((9H-fluoren-9-yl)methoxy)carbonyl)amino)hexadecanamido)phenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate

[0896] To a stirred solution of tert-butyl (S)-2-(4-(4-(16-((2-(1H-indol-3-yl)ethyl)amino)hexadecanamido)phenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate (90.00 mg, 0.11 mmol, 1.00 equiv) in THF (2.00 mL) was added DIEA (41.83 mg, 0.32 mmol, 3.00 equiv), and 2,5-dioxopyrrolidin-1-yl 9H-fluoren-9-ylmethyl carbonate (36.39 mg, 0.11 mmol, 1.00 equiv), and the mixture was stirred at room temperature for 4.0 h. The resulting mixture was concentrated under vacuum, and the residue was purified by preparative TLC (DCM:MeOH=10:1) to give tert-butyl (S)-2-(4-(4-(16-((2-(1H-indol-3-yl)ethyl)(((9H-fluoren-9-yl)methoxy)carbonyl)amino)hexadecanamido)phenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate (80.00 mg, 56.15% yield) as a white solid. LC / MS: C 64 H 77 Calculated mass for N7O5S: 1055.57, Found: 1056.50 [M+H] + .

[0897] Step 6: Synthesis of (S)-2-(4-(4-(16-((2-(1H-indol-3-yl)ethyl)(((9H-fluoren-9-yl)methoxy)carbonyl)amino)hexadecanamido)phenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetic acid

[0898] The procedure was the same as for methyl 4-[4-(4-amino-1-methylpyrrol-2-amido)-1-methylimidazol-2-amido]-1-methylpyrrole-2-carboxylate (Example 2, Step 4). 70.00 mg of tert-butyl (S)-2-(4-(4-(16-((2-(1H-indol-3-yl)ethyl)(((9H-fluoren-9-yl)methoxy)carbonyl)amino)hexadecanamido)phenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate was used to obtain 70.00 mg of the crude desired product as a pale yellow oil. LC / MS: C 60 H 69 Calculated mass for N7O5S: 999.50, Measured mass: 1000.80 [M+H] + .

[0899] Example 17. Synthesis of (S)-2-(2,3,9-trimethyl-4-(4-octanamidophenyl)-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetic acid

[0900] [ka]

[0901] Step 1: Synthesis of tert-butyl (S)-2-(2,3,9-trimethyl-4-(4-octanamidophenyl)-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate

[0902] The procedure was the same as for tert-butyl (S)-2-(4-(4-(16-bromohexadecanamido)phenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate. 100.00 mg of tert-butyl (S)-2-(4-(4-aminophenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate was used to obtain 117.00 mg of crude desired product as a yellow solid. LC / MS: C 31 H 41 Calculated mass for N5O3S: 563.29, Found: 564.45 [M+H] + .

[0903] Step 2: Synthesis of (S)-2-(2,3,9-trimethyl-4-(4-octanamidophenyl)-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetic acid

[0904] The procedure was the same as for methyl 4-[4-(4-amino-1-methylpyrrol-2-amido)-1-methylimidazol-2-amido]-1-methylpyrrole-2-carboxylate (Example 2, Step 4). 110.00 mg of tert-butyl (S)-2-(2,3,9-trimethyl-4-(4-octanamidophenyl)-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate was used to obtain 110.00 mg of crude desired product as a yellow oil. LC / MS: C 27 H 33 Calculated mass for N5O3S: 507.23, Found: 508.35 [M+H] + .

[0905] Example 18. Synthesis of (S)-2-(2,3,9-trimethyl-4-(4-palmitamidophenyl)-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetic acid

[0906] [ka]

[0907] Step 1: Synthesis of tert-butyl (S)-2-(2,3,9-trimethyl-4-(4-palmitamidophenyl)-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate

[0908] The procedure was the same as for ethyl 4-(1-methylimidazol-2-amido)-1-[26-(2,5,8,11,14,17,20,23-octaoxapentacosan-25-yl)-2,5,8,11,14,17,20,23-octaoxa-26-azahentriacontan-31-yl]pyrrole-2-carboxylate, but the reaction time was 2.0 h. Using 60.00 mg of palmitic acid, 120.00 mg of the desired product was obtained as a yellow oil (75.87% yield). LC / MS: C 39 H 57 Calculated mass for N5O3S: 675.42, Found: 676.65 [M+H] + .

[0909] Step 2: Synthesis of (S)-2-(2,3,9-trimethyl-4-(4-palmitamidophenyl)-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetic acid

[0910] The procedure was the same as for methyl 4-[4-(4-amino-1-methylpyrrol-2-amido)-1-methylimidazol-2-amido]-1-methylpyrrole-2-carboxylate (Example 2, Step 4). 110.00 mg of tert-butyl (S)-2-(2,3,9-trimethyl-4-(4-palmitamidophenyl)-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetate was used to obtain 110.00 mg of crude desired product as a yellow oil. LC / MS: C 35 H 49Calculated mass for N5O3S: 619.36, Measured: 620.50 [M+H] + .

[0911] Example 19. Synthesis of 4-[2-(2,4-difluorophenoxy)-5-methanesulfonylphenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylic acid

[0912] [ka]

[0913] Step 1: Synthesis of 1-(2,4-difluorophenoxy)-4-methanesulfonyl-2-nitrobenzene

[0914] To a stirred solution of 2,4-difluorophenol (1.78 g, 13.68 mmol, 1.00 equiv) in DMSO (50.00 mL) was added 1-fluoro-4-methanesulfonyl-2-nitrobenzene (3.00 g, 13.682 mmol, 1.00 equiv) and K2CO3 (1.89 g, 13.68 mmol, 1.00 equiv). The resulting mixture was stirred at 120 °C for 1.0 h. The reaction mixture was poured into ice water (120 mL) and extracted with EA (3 × 150 mL). The combined organic phase was washed with HO (100 mL) and NaCl (100 mL) and dried over anhydrous Na2SO4. The solid was filtered and the filtrate was concentrated to give 1-(2,4-difluorophenoxy)-4-methanesulfonyl-2-nitrobenzene (4.20 g, 88.56% yield) as a yellow solid. 1 H NMR(400MHz,DMSO-d6)δ:8.61(s,1H), 8.15(d,J=8.8Hz,1H), 7.55-7.66(m,2H), 7.24-7.30(m,2H), 3.34(s,3H).

[0915] Step 2: Synthesis of 2-(2,4-difluorophenoxy)-5-methanesulfonylaniline

[0916] The procedure was the same as for 2-(2,4-difluorophenoxy)-5-methanesulfonylaniline (INT-444-2), except the reaction time was 1.0 h. 500.00 mg of 1-(2,4-difluorophenoxy)-4-methanesulfonyl-2-nitrobenzene was used to give 420.00 mg of 2-(2,4-difluorophenoxy)-5-methanesulfonylaniline as a colorless oil. LC / MS: C 13 H 11 Calculated mass for F2NO3S: 299.04, Measured mass: 300.05 [M+H] + .

[0917] Step 3: Synthesis of 1-(2,4-difluorophenoxy)-2-iodo-4-methanesulfonylbenzene

[0918] The procedure was the same as for 5-fluoro-2-(2-iodo-4-methanesulfonylphenoxy)-1,3-dimethylbenzene (Example 13, Step 5), except that the reaction time was 1.0 h after adding KI. Using 420.00 mg of 5-fluoro-2-(2-iodo-4-methanesulfonylphenoxy)-1,3-dimethylbenzene, 440.00 mg of 1-(2,4-difluorophenoxy)-2-iodo-4-methanesulfonylbenzene was obtained as a yellow solid (78.57% yield). 1 H NMR(400MHz,DMSO-d6)δ:8.38(s,1H), 7.86 d,J=8.4Hz,1H), 7.50-7.61(m,1H), 7.41-7.49(m,1H), 7.15-7.25(m,1H), 6.90(d,J=8.8Hz,1H), 3.26(s,3H).

[0919] Step 4: Synthesis of ethyl 4-[2-(2,4-difluorophenoxy)-5-methanesulfonylphenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylate

[0920] The procedure was the same as for ethyl 4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-methanesulfonylphenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylate (INT-444-4), except the reaction temperature was 75° C. and the reaction time was 1.0 h. Using 420.00 mg of 1-(2,4-difluorophenoxy)-2-iodo-4-methanesulfonylbenzene, 340.00 mg of ethyl 4-[2-(2,4-difluorophenoxy)-5-methanesulfonylphenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylate was obtained as a white solid (62.11% yield). LC / MS: C 24 H 20 Calculated mass for F2N2O6S: 502.10, Measured mass: 503.25 [M+H] + .

[0921] Step 5: Synthesis of 4-[2-(2,4-difluorophenoxy)-5-methanesulfonylphenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylic acid

[0922] The procedure was the same as for 4-[3-[(tert-butoxycarbonyl)amino]propanamido]-1-methylimidazole-2-carboxylic acid (Example 1, Step 3), except that the reaction time was 1.0 h. Using 320.00 mg of ethyl 4-[2-(2,4-difluorophenoxy)-5-methanesulfonylphenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylate, 290.00 mg of 4-[2-(2,4-difluorophenoxy)-5-methanesulfonylphenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxylic acid was obtained as a white solid (92.15% yield). LC / MS: C 22 H 16 Calculated mass for F2N2O6S: 474.07, Measured mass: 475.20 [M+H] + .

[0923] Synthesis of Representative Compounds of the Present Disclosure

[0924] Example 20. N-(5-{[2-({2-[(2-{[26-(4-{1-ethyl-4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxopyrrolo[2,3-c]pyridin-2-amido}phenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosane-1 Synthesis of [1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido]imidazole-2-carboxamide (Compound 196)

[0925] [ka]

[0926] Step 1: Synthesis of benzyl N-(4-hydroxyphenyl)carbamate

[0927] A solution of aminophenol (10.00 g, 91.64 mmol, 1.00 equiv.), benzyl 2,5-dioxopyrrolidin-1-yl carbonate (27.00 g, 108.34 mmol, 1.18 equiv.), and DIEA (29.02 g, 224.51 mmol, 2.45 equiv.) in THF (60.00 mL) was stirred at room temperature for 2.0 hours. The resulting mixture was extracted with EtOAc (3×150 mL). The combined organic layers were washed with brine (3×50 mL) and dried over anhydrous NaSO. After filtration, the filtrate was concentrated under reduced pressure. The resulting pink solid was washed with 3×30 mL of ACN. This afforded benzyl N-(4-hydroxyphenyl)carbamate (15.00 g, 63.93% yield) as a white solid. LC / MS: C 14 H 13 Calculated mass for NO3: 243.09, Measured mass: 244.10 [M+H] + .

[0928] Step 2: Synthesis of benzyl N-[4-({26-[(tert-butoxycarbonyl)amino]-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl}oxy)phenyl]carbamate

[0929] The procedure was the same as for ethyl 1-(5-bromopentyl)-4-[(tert-butoxycarbonyl)amino]pyrrole-2-carboxylate, but the reaction time was 17.0 hours. Using 2.80 g of benzyl N-(4-hydroxyphenyl)carbamate, 7.00 g of benzyl N-[4-({26-[(tert-butoxycarbonyl)amino]-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl}oxy)phenyl]carbamate was obtained as a colorless oil (82.31% yield). LC / MS: C 37 H 58 N2O 13 Calculated mass: 738.39, Measured mass: 756.55 [M+H2O] + .

[0930] Step 3: Synthesis of benzyl N-{4-[(26-amino-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl)oxy]phenyl}carbamate

[0931] The procedure was the same as for methyl 4-[4-(3-aminopropanamido)-1-methylimidazol-2-amido]-1-methylpyrrole-2-carboxylic acid hydrochloride, except that the reaction solvent was 4 M HCl / DCM (1:1) in dioxane. Using 6.50 g of benzyl N-[4-({26-[(tert-butoxycarbonyl)amino]-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl}oxy)phenyl]carbamate, 6.50 g of crude benzyl N-{4-[(26-amino-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl)oxy]phenyl}carbamate was obtained as a colorless oil. LC / MS: C 32 H 50 N2O 11Calculated mass: 638.34, Measured mass: 639.40 [M+H] + .

[0932] Step 4: Synthesis of benzyl N-[4-({26-[3-({1-methyl-4-[3-({1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazol-2-amido]pyrrol-2-yl}formamido)propanamido]imidazol-2-yl}formamido)propanamido]-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl}oxy)phenyl]carbamate

[0933] The procedure was the same as for methyl 4-(4-{4-[(tert-butoxycarbonyl)amino]-1-methylpyrrol-2-amido}-1-methylimidazol-2-amido)-1-methylpyrrole-2-carboxylate (Example 2, Step 3). 9.00 g of benzyl N-{4-[(26-amino-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl)oxy]phenyl}carbamate was used to prepare 15.50 g of benzyl N-[4-({26-[3-({1-methyl-4-[3-({1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido) Imidazole-2-amido)pyrrol-2-yl}formamido)propanamido)imidazol-2-yl}formamido)propanamido)imidazol-2-yl}formamido)propanamido)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl}oxy)phenyl]carbamate was obtained as a yellow solid (75.84% yield). LC / MS: C 68 H 91 N 17 O 19 Calculated mass: 1449.67, Measured mass: 1450.90 [M+H] + .

[0934] Step 5: Synthesis of N-(5-{[2-({2-[(2-{[26-(4-aminophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamoyl}ethyl)carbamoyl]-1-methylimidazol-4-yl}carbamoyl)ethyl]carbamoyl}-1-methylpyrrol-3-yl)-1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-carboxamide

[0935] To a solution of benzyl N-[4-({26-[3-({1-methyl-4-[3-({1-methyl-4-[1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamide)imidazol-2-amido]pyrrol-2-yl}formamide)propanamide]imidazol-2-yl}formamide)propanamide]-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl}oxy)phenyl]carbamate (5.50 g, 3.792 mmol, 1.00 equiv) in DMF (40.00 mL) was added Pd / C (1.10 g, 20% w / w). The H atmosphere was then exchanged three times. The mixture was stirred under an H atmosphere at room temperature for 2.0 h. The Pd / C was filtered and washed with MeOH, and the filtrate was concentrated and lyophilized. The resulting mixture was poured into water / ice (60 mL). The precipitated solid was collected by filtration, washed with HO (3 x 20 mL), and lyophilized. N-(5-{[2-({2-[(2-{[26-(4-aminophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamoyl}ethyl)carbamoyl]-1-methylimidazol-4-yl}carbamoyl)ethyl]carbamoyl}-1-methylpyrrol-3-yl)-1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-carboxamide (4.08 g, 81.74%) was obtained as a white solid. LC / MS: C 60 H 85 N 17 O17 Calculated mass: 1315.63, Measured mass: 1317.10 [M+H] + .

[0936] Step 6: Synthesis of Compound 196

[0937] A 25 mL flask was charged with 1-ethyl-4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxopyrrolo[2,3-c]pyridine-2-carboxylic acid (280.00 mg, 0.57 mmol, 1.00 equiv.) and DMF (3.00 mL). The mixture was cooled to 0 °C, and then PyBOP (443.74 mg, 0.85 mmol, 1.50 equiv.) and N-(5-{[2-({2-[(2-{[26-(4-aminophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamoyl}ethyl)carbamoyl]-1-methylimidazol-4-yl}carbamoyl)ethyl]carbamoyl {moyl}-1-methylpyrrol-3-yl)-1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-carboxamide (748.36 mg, 0.57 mmol, 1.00 equiv) was added, followed by the portionwise addition of DIEA (220.41 mg, 1.70 mmol, 3.00 equiv). The reaction was stirred at room temperature for 1.0 h. The reaction mixture was filtered, and the filtrate in DMF (3.00 mL) was purified by preparative HPLC under the following conditions: Column: XSelect CSH Prep C18 OBD column, 19*250 mm, 5 μm; Mobile phase A: water (10 mmol / L NH4HCO3 + 0.1% NH3.H2O), Mobile phase B: ACN; Flow rate: 25 mL / min; Gradient: 48% B to 56% B, 56% B in 10 min; Wavelength: 254 nm; RT1 (min): 9.02; Run number: 0. The fractions were combined and directly lyophilized.This resulted in the production of N-(5-{[2-({2-[(2-{[26-(4-{1-ethyl-4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxopyrrolo[2,3-c]pyridin-2-amido}phenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamoyl}ethyl)carbamoyl]-1-methylimidazol-4-yl}carbamoyl)ethyl]carbamoyl}-1-methylpyrrol-3-yl)-1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-carboxamide (171.10 mg, 16.53% yield) was obtained as a white solid. HRMS: C. 88 H 112 FN 19 O 21 Calculated mass: 1789.8264, Measured mass: 1790.8420 [M+H] + .

[0938] Examples 21 to 27 were prepared according to the procedure of Example 20.

[0939] Example 21. N-(5-{[2-({2-[(2-{[26-(4-{4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(prop-1-en-2-yl)phenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridin-2-amido}phenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl Synthesis of ]carbamoyl}ethyl)carbamoyl]-1-methylimidazol-4-yl}carbamoyl)ethyl]carbamoyl}-1-methylpyrrol-3-yl)-1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-carboxamide (Compound 197)

[0940] Using 300.00 mg of N-(5-{[2-({2-[(2-{[26-(4-aminophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamoyl}ethyl)carbamoyl]-1-methylimidazol-4-yl}carbamoyl)ethyl]carbamoyl}-1-methylpyrrol-3-yl)-1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-carboxamide, 232.40 mg of the desired product was obtained as a white solid (56.28% yield). HRMS: C 86 H 106 FN 19 O 20 Calculated mass: 1743.7845, Measured mass: 1744.7899 [M+H] + .

[0941] Example 22. Synthesis of 1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamide}propanamido)-N-(1-methyl-5-{[2-({1-methyl-2-[(2-{[26-(4-{6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridin-2-amido}phenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamoyl}ethyl)carbamoyl]imidazol-4-yl}carbamoyl)ethyl]carbamoyl}pyrrol-3-yl)imidazole-2-carboxamide (Compound 198)

[0942] Using 400.00 mg of N-(5-{[2-({2-[(2-{[26-(4-aminophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamoyl}ethyl)carbamoyl]-1-methylimidazol-4-yl}carbamoyl)ethyl]carbamoyl}-1-methylpyrrol-3-yl)-1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-carboxamide, 242.00 mg of the desired product was obtained as a white solid (51.22% yield). HRMS: C 69 H 91 N 19 O 19 Calculated mass: 1489.6738, Measured mass: 1490.6778 [M+H] + .

[0943] Example 23. (S)-1-methyl-4-(3-(1-methyl-4-(1-methyl-1H-imidazole-2-carboxamido)-1H-pyrrole-2-carboxamido)propanamide)-N-(1-methyl-5-((3-((1-methyl-2-((28-oxo-1-(4-(2-(2,3,9-trimethyl-4-(4-octanamidophenyl)-6H-thieno[3,2-f][1,2,4] Synthesis of triazolo[4,3-a][1,4]diazepin-6-yl)acetamido)phenoxy)-3,6,9,12,15,18,21,24-octaoxa-27-azatriacontan-30-yl)carbamoyl)-1H-imidazol-4-yl)amino)-3-oxopropyl)carbamoyl)-1H-pyrrol-3-yl)-1H-imidazole-2-carboxamide (Compound 203)

[0944] Using 50.00 mg of (S)-2-(2,3,9-trimethyl-4-(4-octanamidophenyl)-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetic acid, 39.70 mg of the desired product was obtained as a white solid (21.94% yield). HRMS: C 87 H 116 N22 O 19 Calculated mass for S: Accurate mass: 1804.8508, Measured mass: 1805.8650 [M+H] + .

[0945] Example 24. (S)-1-methyl-4-(3-(1-methyl-4-(1-methyl-1H-imidazole-2-carboxamido)-1H-pyrrole-2-carboxamido)propanamide)-N-(1-methyl-5-((3-((1-methyl-2-((28-oxo-1-(4-(2-(2,3,9-trimethyl-4-(4-palmitamidophenyl)-6H-thieno[3,2-f][1,2,4]trimethyl)-2 ... Synthesis of riazolo[4,3-a][1,4]diazepin-6-yl)acetamido)phenoxy)-3,6,9,12,15,18,21,24-octaoxa-27-azatriacontan-30-yl)carbamoyl)-1H-imidazol-4-yl)amino)-3-oxopropyl)carbamoyl)-1H-pyrrol-3-yl)-1H-imidazole-2-carboxamide (Compound 204)

[0946] Using 110.00 mg of (S)-2-(2,3,9-trimethyl-4-(4-palmitamidophenyl)-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetic acid, 61.10 mg of the desired product was obtained as a white solid (17.25% yield). HRMS: C 95 H 132 N 22 O 19 Calculated mass for S: 1916.9760, Measured: 1917.9832 [M+H] + .

[0947] Example 25. Synthesis of N-(5-{[2-({2-[(2-{[26-(4-{4-[2-(2,4-difluorophenoxy)-5-methanesulfonylphenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridin-2-amido}phenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamoyl}ethyl)carbamoyl]-1-methylimidazol-4-yl}carbamoyl)ethyl]carbamoyl}-1-methylpyrrol-3-yl)-1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-carboxamide (Compound 207)

[0948] Using 150.00 mg of N-(5-{[2-({2-[(2-{[26-(4-aminophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamoyl}ethyl)carbamoyl]-1-methylimidazol-4-yl}carbamoyl)ethyl]carbamoyl}-1-methylpyrrol-3-yl)-1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-carboxamide, 42.30 mg of the desired product was obtained as a white solid (20.40% yield). HRMS: C 82 H 99 F2N 19 O 22 Calculated mass for S: Accurate mass: 1771.6900, Measured mass: 1772.6914 [M+H] + .

[0949] Example 26. (R)—N-(5-((3-((2-((1-(4-(2-(4-(4-chlorophenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetamido)phenoxy)-28-oxo-3,6,9,12,15,18,21,24-octaoxa-27-azatriacontan-30-yl) Synthesis of (carbamoyl)-1-methyl-1H-imidazol-4-yl)amino)-3-oxopropyl)carbamoyl)-1-methyl-1H-pyrrol-3-yl)-1-methyl-4-(3-(1-methyl-4-(1-methyl-1H-imidazole-2-carboxamide)-1H-pyrrole-2-carboxamide)propanamide)-1H-imidazole-2-carboxamide (Compound 231)

[0950] Using 87.40 mg of N-(5-{[2-({2-[(2-{[26-(4-aminophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamoyl}ethyl)carbamoyl]-1-methylimidazol-4-yl}carbamoyl)ethyl]carbamoyl}-1-methylpyrrol-3-yl)-1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-carboxamide, 43.70 mg of the desired product was obtained as a white solid (38.51% yield). HRMS: C 79 H 100 ClN 21 O 18 Calculated mass for S: 1697.6964, Measured: 1698.7020 [M+H] + .

[0951] Example 27 (S)—N-(5-((3-((2-((1-(4-(2-(4-(4-(16-((2-(1H-indol-3-yl)ethyl)amino)hexadecanamido)phenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetamido)phenoxy)-28-oxo-3,6,9,12,15,18,21,24-octaoxa- Synthesis of 27-azatriacontan-30-yl)carbamoyl)-1-methyl-1H-imidazol-4-yl)amino)-3-oxopropyl)carbamoyl)-1-methyl-1H-pyrrol-3-yl)-1-methyl-4-(3-(1-methyl-4-(1-methyl-1H-imidazole-2-carboxamide)-1H-pyrrole-2-carboxamide)propanamide)-1H-imidazole-2-carboxamide (Compound 200)

[0952] To a stirred solution of (S)-2-(4-(4-(16-((2-(1H-indol-3-yl)ethyl)(((9H-fluoren-9-yl)methoxy)carbonyl)amino)hexadecanamido)phenyl)-2,3,9-trimethyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a][1,4]diazepin-6-yl)acetic acid 60.00 mg, 0.06 mmol, 1.00 equiv) in DMF (2.00 mL) was added PyBOP (31.21 mg, 0.06 mmol, 1.00 equiv), N-(5-{[2-({2-[(2-{[26-(4-aminophenoxy)-3, To the solution at 0° C. were added sequentially 6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamoyl}ethyl)carbamoyl]-1-methylimidazol-4-yl}carbamoyl)ethyl]carbamoyl}-1-methylpyrrol-3-yl)-1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-carboxamide (78.96 mg, 0.06 mmol, 1.00 equiv.) and DIEA (23.26 mg, 0.180 mmol, 3.00 equiv.). The mixture was warmed to room temperature and stirred at room temperature for 1.0 h. Then, piperidine (0.20 mL) (volume ratio of piperidine to DMF = 1:10) was added, and the mixture was stirred at room temperature for 1.0 h. Upon completion of the reaction, the mixture was added dropwise to ice water (5 mL). The resulting solid was filtered, washed with water (2 × 3 mL), and dried under vacuum. The crude product (80.00 mg) was dissolved in DMF (2.00 mL), and the resulting mixture was filtered. The filtrate was purified by preparative HPLC using the following conditions: Column: XBridge Shield RP18 OBD column, 19 × 250 mm, 10 μm; Mobile phase A: water (0.05% TFA), Mobile phase B: ACN; Flow rate: 25 mL / min; Gradient: 40% B to 53% B, 53% B in 10 min; Wavelength: 254 nm; RT1 (min): 9.83; Run number: 0. The fractions were combined and directly lyophilized to give the desired product (15.00 mg, 20.13%) as a white solid. HRMS: C 105 H 142 N 24 O 19Calculated mass for S: 2075.0603, Measured: 2076.0676 [M+H] + .

[0953] Example 28. N-(5-{[2-({2-[(2-{[26-(4-{4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridin-2-amido}phenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamoyl}ethyl)carbamoyl]-1-methylimidazo Synthesis of {1-methyl-4-(3-{[1-methyl-4-(1-methylimidazol-2-amido)pyrrol-2-yl]formamido}propanamido)imidazole-2-carboxamide (Compound 241)

[0954] [ka]

[0955] Step 1: Synthesis of tert-butyl N-[26-(4-nitrophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamate

[0956] To a stirred solution of 4-fluoronitrobenzene (329.66 mg, 2.34 mmol, 1.20 equiv) in THF (15.00 mL) was added tert-butyl N-(26-hydroxy-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl)carbamate (1.00 g, 1.95 mmol, 1.00 equiv) and CsCO (1.90 g, 5.84 mmol, 3.00 equiv). The resulting mixture was stirred at 55 °C for 16.0 h. The resulting mixture was concentrated in vacuo. The residue was purified by silica gel column chromatography (0-10% MeOH / DCM) to give tert-butyl N-[26-(4-nitrophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamate (1.10 g, 89.01% yield) as a yellow oil. LC / MS: C 29 H 50 N2O 13 Calculated mass: 634.33, Measured mass: 635.55 [M+H] + .

[0957] Step 2: Synthesis of 26-(4-nitrophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-amine

[0958] The procedure was the same as for methyl 4-[4-(4-amino-1-methylpyrrol-2-amido)-1-methylimidazol-2-amido]-1-methylpyrrole-2-carboxylate (Example 2, Step 4). 1.10 g of tert-butyl N-[26-(4-nitrophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamate was used to obtain 920.0 mg of crude 26-(4-nitrophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-amine as a yellow oil. LC / MS: C 24 H 42 N2O 11 Calculated mass: 534.27, Measured mass: 535.45 [M+H] + .

[0959] Step 3: Synthesis of benzyl N-[26-(4-nitrophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamate

[0960] To a stirred solution of 26-(4-nitrophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-amine (920.00 mg, 1.72 mmol, 1.00 equiv) in DCM (6.00 mL) was added benzyl 2,5-dioxopyrrolidin-1-yl carbonate (514.66 mg, 2.07 mmol, 1.20 equiv) and DIEA (667.25 mg, 5.16 mmol, 3.00 equiv). The resulting mixture was stirred at room temperature for 1.0 h. The resulting mixture was concentrated in vacuo. The residue was purified by silica gel column chromatography (0-10% MeOH / DCM) to give benzyl N-[26-(4-nitrophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamate (1.10 g, 95.58% yield) as a yellow oil. LC / MS: C 32 H 48 N2O 13 Calculated mass: 668.31, Measured mass: 669.25 [M+H] + .

[0961] Step 4: Synthesis of benzyl N-[26-(4-aminophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamate

[0962] To a stirred solution of benzyl N-[26-(4-nitrophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamate (1.10 g, 1.65 mmol, 1.00 equiv) in EtOH (6.00 mL) and HO (2.00 mL) was added Fe (1.38 g, 24.67 mmol, 15.00 equiv) and NH4Cl (1.32 g, 24.68 mmol, 15.00 equiv). The resulting mixture was stirred at 70 °C for 1.0 h. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (0-10% MeOH / DCM) to give benzyl N-[26-(4-aminophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamate (960.00 mg, 91.37% yield) as a yellow oil. LC / MS: C 32 H 50 N2O 11 Calculated mass: 638.34, Measured mass: 639.35 [M+H] + .

[0963] Step 5: Synthesis of benzyl N-[26-(4-{4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridin-2-amido}phenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamate

[0964] The procedure was the same as for methyl 4-(4-{4-[(tert-butoxycarbonyl)amino]-1-methylpyrrol-2-amido}-1-methylimidazol-2-amido)-1-methylpyrrole-2-carboxylate (Example 2, step 3). Using 930.00 mg of benzyl N-[26-(4-aminophenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamate, 1.50 g of benzyl N-[26-(4-{4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridin-2-amido}phenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamate was obtained as a yellow oil (94.93% yield). LC / MS: C 58 H 73 FN4O 15 Calculated mass: 1084.50, Measured mass: 1086.05 [M+H] + .

[0965] Step 6: Synthesis of N-{4-[(26-amino-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl)oxy]phenyl}-4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridine-2-carboxamide

[0966] To a stirred solution of benzyl N-[26-(4-{4-[2-(4-fluoro-2,6-dimethylphenoxy)-5-(2-hydroxypropan-2-yl)phenyl]-6-methyl-7-oxo-1H-pyrrolo[2,3-c]pyridin-2-amido}phenoxy)-3,6,9,12,15,18,21,24-octaoxahexacosan-1-yl]carbamate (1.50 g, 1.38 mmol, 1.00 equiv) in DMF (8.00 mL) was added Pd / C (150.00 mg, 10% w / w). The resulting mixture was stirred at room temperature under an H atmosphere for 3.0 h. The resulting mixture was filtered and directly lyophilized to give N-{4-[(26-am...

Claims

1. 1. A transcription modulator molecule, or a pharma- ceutically acceptable salt thereof, having a first end, a second end, and an oligomeric backbone, a) the first end comprises a DNA binding moiety capable of non-covalently binding to the nucleotide repeat sequence GAA, the first end having the structure of formula (A-2): 【Chemistry 1】 Formula (A-2) During the ceremony, m1 is 1 to 4; n1 is 0 to 2; each Y, Y, Y, and Y is independently CH or N; each Z1, Z2, Z3, and Z4 is independently O, S, or NR1D; each L3 is an optionally substituted C1-C6 alkylene, an optionally substituted C3-C7 cycloalkylene, an optionally substituted 3- to 7-membered heterocyclene, or an optionally substituted 5- to 6-membered heteroarylene; Each R30 is hydrogen or C1-C6 alkyl; or each R30 and L3, together with the atoms to which they are attached, combine to form a 4- to 7-membered heterocycle; W1 is hydrogen, optionally substituted C1-C6 alkyl, -NR1E-C(O)-NR1ER1F, -C(O)-NR1ER1F, or (AA)1-10; W2 is hydrogen, optionally substituted C1-C6 alkyl, —C(O)—NR1ER1F, or (AA)1-10; each R1D and R1E is independently hydrogen, optionally substituted C1-C50 alkyl, optionally substituted C1-C50 heteroalkyl, or optionally substituted PEG1-50; R1F is hydrogen, optionally substituted C1-C20 alkyl, optionally substituted C1-C20 heteroalkyl, optionally substituted PEG1-20, or one or more AA; each AA is independently a naturally occurring amino acid; b) the second end comprises a protein-binding moiety capable of binding to a regulatory molecule that regulates expression of a gene comprising the nucleotide repeat sequence GAA; c) a transcription modulator molecule, or a pharma- ceutically acceptable salt thereof, wherein said oligomeric backbone comprises a linker between said first end and said second end.

2. The first end comprises a structure of formula (A-3), or a pharma- ceutically acceptable salt thereof: 【Chemistry 2】 Formula (A-3) During the ceremony, m 1 is 1 to 4; n 1 is 0 to 2; each Y 1 , Y 2 , Y 3 , and Y 4 is independently CH or N; each Z 1 , Z 2 , Z 3 , and Z 4 is independently O, S, or NR 1D ; W 1 is hydrogen, optionally substituted C 1 -C 6 alkyl, —NR 1E —C(O)—NR 1E R 1F , —C(O)—NR 1E R 1F , or (AA) 1-10 ; W 2 is hydrogen, optionally substituted C 1 -C 6 alkyl, —C(O)—NR 1E R 1F , or (AA) 1-10 ; each R 1D and R 1E is independently hydrogen, optionally substituted C 1 -C 50 alkyl, optionally substituted C 1 -C 50 heteroalkyl, or optionally substituted PEG 1-50 ; R 1F is hydrogen, optionally substituted C 1 -C 20 alkyl, optionally substituted C 1 -C 20 heteroalkyl, optionally substituted PEG 1-20 , or one or more AA; 2. The transcription modulator molecule of claim 1, or a pharma- ceutically acceptable salt thereof, wherein each AA is independently an amino acid residue selected from β-alanine, lysine, and arginine.

3. The first end is represented by formula (A-4): 【Chemistry 3】 Formula (A-4) 3. The transcription modulator molecule of claim 2, comprising the structure: 。 4. The transcription modulator molecule of any one of claims 1 to 3, or a pharma- ceutically acceptable salt thereof, wherein each Z1, Z2, Z3, and Z4 is independently NR1D and each R1D is independently optionally substituted C1-C6 alkyl.

5. The transcription modulator molecule of claim 1, wherein each Y 1 and Y 3 is N, and each Y 2 and Y 4 is each independently CH or N, or a pharma- ceutically acceptable salt thereof.

6. The transcription modulator molecule of claim 1, or a pharma- ceutically acceptable salt thereof, wherein the linker has a length of less than about 50 angstroms or comprises 5 to 50 chain atoms.

7. The linker comprises a multimer having 2 to 50 spacing moieties, the spacing moieties being independently selected from -((CR3aR3b)x-O)y-, -((CR3aR3b)x-NR4a)y-, -((CR3aR3b)x-CH=CH-(CR3aR3b)x-O)y-, optionally substituted C1-12 alkyl, optionally substituted C2-10 alkenyl, optionally substituted C2-10 alkynyl, optionally substituted C6-10 arylene, optionally substituted C3-7 alkyl, optionally substituted C2-7 alkenyl, optionally substituted C2-7 alkynyl, optionally substituted C6-10 arylene, optionally substituted C3-7 alkyl, optionally substituted C4-7 alkyl, optionally substituted C5-7 alkyl, optionally substituted C6-7 alkyl, optionally substituted C7-7 alkyl, optionally substituted C8-7 alkyl, optionally substituted C9-8 alkyl, optionally substituted C1-12 alkyl, optionally substituted C2-10 alkenyl, optionally substituted C2-10 alkynyl, optionally substituted C6-10 arylene, optionally substituted C3-7 alkyl, optionally substituted C4-7 alkyl, optionally substituted C5-7 alkyl, optionally substituted C6-7 alkyl, optionally substituted C7-7 alkyl, optionally substituted C8-7 alkyl, optionally substituted C9-8 alkyl, optionally substituted C1-12 alkyl, optionally substituted C2-10 ...1-12 alkyl, optionally substituted C2-10 alkyl, optionally substituted C1-12 alkyl, optionally substituted C2-10 alkyl selected from the group consisting of cycloalkylene, optionally substituted 5- to 10-membered heteroarylene, optionally substituted 4- to 10-membered heterocycloalkylene, an amino acid residue, -O-, -C(O)NR 4a -, -NR 4a C(O)-, -C(O)-, -NR 4a -, and any combination thereof; each x is independently 2 to 4; each y is independently 1 to 10; each R 3a and R 3b is independently selected from hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted alkoxy, optionally substituted amino, carboxyl, carboxyl ester, acyl, acyloxy, acylamino, aminoacyl, optionally substituted alkylamido, sulfonyl, optionally substituted thioalkoxy, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted cycloalkyl, and optionally substituted heterocyclyl; 7. The transcription modulator molecule of any one of claims 1 to 6, or a pharma- ceutically acceptable salt thereof, wherein each R 4a is independently hydrogen or optionally substituted C 1-6 alkyl.

8. The linker is -(CH2-C(O)N(R'')-(CH2)q-N(R')-(CH2)q-N(R'')C(O)-(CH2)x-C(O)N(R'')-A-, -(CH2)x-C(O)N(R'')-(CH2CH2O)y(CH2)x-C(O)N(R'')-A-, -C(O)N(R'')-(CH2)q-N(R')-(CH2)q-N(R'')C(O)-(CH2)x-A-, -(CH2)x-O-(CH2CH2O)y-(CH2)x and each q is independently an integer from 2 to 10; and each A is independently a bond, optionally substituted C 1-12 alkyl, optionally substituted C 6-10 arylene, optionally substituted C 3-7 aryl, optionally substituted C 6-7 arylene, optionally substituted C 7-8 alkyl, or optionally substituted C 7-8 aryl.

7. The transcription modulator molecule of any one of claims 1 to 6, wherein the heteroaryl group is selected from cycloalkylene, optionally substituted 5- to 10-membered heteroarylene, and optionally substituted 4- to 10-membered heterocycloalkylene, or a pharma- ceutically acceptable salt thereof.

9. The transcription modulator molecule of any one of claims 1 to 8, or a pharma- ceutically acceptable salt thereof, wherein the linker is attached to the second end by a group selected from -CO-, -NR 1a -, C 1-12 alkyl, -CONR 1a -, and -NR 1a CO-, and wherein each R 1a is independently hydrogen, or optionally substituted C 1-6 alkyl, or optionally substituted C 1-12 alkylene, optionally substituted C 2-10 alkenylene, optionally substituted C 2-10 alkynylene, optionally substituted C 6-10 arylene, optionally substituted C 3-7 cycloalkylene, optionally substituted 5- to 10-membered heteroarylene, and optionally substituted 4- to 10-membered heterocycloalkylene.

10. The linker is attached to the second end at a moiety having a structure of formula (C-1), or a pharma- ceutically acceptable salt thereof, 【Chemistry 4】 Formula (C-1) During the ceremony, Ring D is absent, arylene, or heterocycloacrylene; L 1 is absent, —CH 2 —, —CH 2 CH 2 —, —C≡C—, or —C≡C-C≡C—; each X 3 and X 4 is independently CH or N; p 1 is 0 to 3; 9. The transcription modulator molecule of any one of claims 1 to 8, or a pharma- ceutically acceptable salt thereof, wherein ** indicates a bond to the second end.

11. The linker is bonded to the second end at a moiety having a structure of formula (C-2) or a pharma- ceutically acceptable salt thereof, 【Chemistry 5】 Formula (C-2) During the ceremony, L 1 is absent, —CH 2 —, —CH 2 CH 2 —, —C≡C—, or —C≡C-C≡C—; X 4 is independently CH or N; each X 5 and X 6 is independently N or CH; 11. The transcription modulator molecule of claim 10, or a pharma- ceutically acceptable salt thereof, wherein ** indicates binding to the second end.

12. The linker is bonded to the second end at a moiety having a structure of formula (C-3), or a pharma- ceutically acceptable salt thereof, 【Chemistry 6】 Formula (C-3) During the ceremony, p 1 is 0 to 3; r 1 is 1 to 3; R 27 is optionally substituted C 1-50 alkyl, optionally substituted C 1-50 heteroalkyl, —C(O)(C 1-50 alkyl), or —C(O)(C 1-50 heteroalkyl), each alkyl and heteroalkyl being optionally substituted; R 1G is hydrogen or C 1 -C 3 alkyl; 9. The transcription modulator molecule of any one of claims 1 to 8, or a pharma- ceutically acceptable salt thereof, wherein ** indicates a bond to the second end.

13. The second end comprises a compound having a structure of formula (12-A) or a pharma- ceutically acceptable salt thereof, 【Chemistry 7】 Formula (12-A) During the ceremony, each R 20 , R 21 , and R 22 is independently hydrogen, halogen, optionally substituted C 1 -C 6 alkyl, optionally substituted C 1 -C 6 haloalkyl, or optionally substituted C 1 -C 6 hydroxyalkyl; 13. The transcription modulator molecule of any one of claims 1 to 12, wherein each z 1 , z 2 , and z 3 is independently 1 to 4, or a pharma- ceutically acceptable salt thereof.

14. The second end is represented by formula (12-B) or formula (12-C): 【Chemistry 8】 14. The transcription modulator molecule of claim 13, comprising a compound having the structure: or a pharma- ceutically acceptable salt thereof.

15. The transcriptional modulator molecule of claim 1, 【Chemistry 9】 15. The transcription modulator molecule of any one of claims 1 to 14, which is:

16. A pharmaceutical composition comprising a transcription modulator molecule according to any one of claims 1 to 15, or a pharma- ceutically acceptable salt thereof, and a pharma- ceutically acceptable excipient.

17. Use of a transcription modulator molecule, or a pharma- ceutically acceptable salt thereof, according to any one of claims 1 to 15 in the manufacture of a medicament for regulating the expression of fxn.

18. Use of a transcription modulator molecule according to any one of claims 1 to 15, or a pharma- ceutically acceptable salt thereof, in the manufacture of a medicament for treating a disease or condition caused by expression of a defective fxn.

19. The use described in claim 18, wherein the disease is Friedreich's ataxia (FA).

20. Use of a transcription modulator molecule, or a pharma- ceutically acceptable salt thereof, according to any one of claims 1 to 15 in the manufacture of a medicament for treating Friedreich's ataxia (FA).

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