Antibacterial compounds and methods of use thereof

Abstract

Compounds useful as antibacterial agents, pharmaceutical compositions comprising the compounds, and methods of using the same. The compounds are believed to target bacterial RNA polymerase and NusG.

Description

Cross-reference to related applications

[0001] This application claims priority to U.S. Provisional Patent Application No. 63 / 654,205, filed May 31, 2024, which is incorporated herein by reference in its entirety. Technical Field

[0002] This disclosure generally relates to compounds that can be used as antibacterial agents, pharmaceutical compositions comprising said compounds, and methods of using them. Background Technology

[0003] Bacterial transcription is a key biological process in prokaryotes, comprising initiation, elongation, and termination phases. It involves the synthesis of RNA molecules complementary to the DNA strand of the bacterial genome, which are programmed by the major enzyme RNA polymerase (RNAP). The RNA products serve as templates for subsequent protein synthesis, influencing bacterial cellular physiology and metabolic activities.

[0004] Bacterial transcription is a fundamental biological process in prokaryotes, comprising initiation, elongation, and termination phases. It involves the synthesis of RNA molecules complementary to the DNA template strand of the bacterial genome, which are programmed by the major enzyme RNA polymerase (RNAP). The RNA product is further used as a template for subsequent protein synthesis, influencing bacterial cellular physiology and metabolic activities.

[0005] A group of small proteins called transcription factors are essential for regulating transcription steps, such as σ, NusB, NusE, and NusG. NusG is one of the essential bacterial transcription factors, a key and ubiquitous component of the bacterial transcription machinery, and plays a defined role in both transcriptional elongation and termination. Several studies have revealed that NusG enhances the overall efficiency and regulation of transcription by building communication bridges between elongated RNAPs and downstream elements or factors. Regarding transcriptional termination, NusG has been shown to promote the function of Rho (a termination factor that disrupts the transcription complex). NusG interacts with Rho to promote its function, enhancing Rho-dependent termination. Furthermore, NusG has been found to participate in the arrangement of transcription-translation coupling, thereby protecting the integrity of the mRNA molecule.

[0006] The structure of NusG highlights a dual-domain architecture, featuring an N-terminal domain (NTD) and a C-terminal Kyrpides-Ouzounis-Woese (KOW) domain. Fine structural analysis reveals that the NTD of NusG employs a β-barrel fold, while the C-terminal KOW domain exhibits a triple-stranded β-sheet, connected to two α-helices on one side. Indeed, NusG primarily interacts directly with RNAP through its N-terminal domain. Structural studies have revealed that association between NusG and RNAP mainly occurs at the β' subunit of RNAP, particularly in the region known as the clamp-helix (CH). Figure 1 (A on the left). This conformation promotes NusG-RNAP protein-protein interactions (PPIs) and leads to movement within the "clamp" region to establish a stable elongation complex, thereby helping to protect the transcriptional vesicle and effectively enhance the transcriptional connection between RNAP and the DNA template.

[0007] Furthermore, the homolog of NusG, known as Spt5 in archaea and eukaryotic cells, is the only conserved transcription factor present in all three domains of life. Unlike bacteria where NusG functions as a monomeric transcription factor, in archaea and eukaryotic cells Spt5 forms a heterodimeric complex with Spt4 via NTD. This complex couples RNA processing and chromatin modification to transcriptional elongation. In humans, the Spt4-Spt5 complex is known as DSIF (DRB-sensitive inducible factor), and it regulates the sustained synthesis of RNA polymerase II (Pol II) and the activation of transcription. Although NusG homologs share functions, sequences, and structures across all domains of life, NusG theoretically represents a viable target for inhibiting bacterial growth without affecting human cells. This is because the binding site and amino acid sequence we have selected for drug design are conserved only in bacteria, making it a truly selective target for antibacterial drug discovery.

[0008] In summary, bacterial transcription is a crucial biological process for gene expression in prokaryotes, and NusG serves as an indispensable regulator, providing tools to improve efficiency during transcription while also promoting sequence-specific termination of the process. Considering the important role of NusG in bacterial transcription, we designed and synthesized a series of inhibitors targeting RNAP-NusG PPIs in this study and confirmed their target specificity and antimicrobial activity. Summary of the Invention

[0009] In the first aspect, this article provides a compound of Formula 1: 1 Or pharmaceutically acceptable salts, wherein: m is an integer selected from 1 to 4; n is an integer selected from 1 to 4; X 1 It is -O-, -S-, -OCH2-, -SCH2-, -(NR 5 - or -SO2(NR) 5 )-; X 2 It is -O-, -S-, -OCH2-, -SCH2-, -(NR 5 - or -SO2(NR) 5 )-; Ar 1 Selected from: , , , , , and ; Ar 2 Selected from: , , , , , and ; Each time R appears, it is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, and aralkyl; or R appearing twice together with their covalently bonded atoms form 3-6 membered cycloalkyl or heterocycloalkyl groups. R 1 Each occurrence is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, perhaloalkoxy, halogen, nitrile, azido, -OR 5 -SR 5 -N(R) 5 )2、-C(O)R 5 -C(O)OR 5 -OC(O)R 5 -N(R) 5 )C(O)R 5 -C(O)N(R) 5 )2、-N(R 5 )C(O)OR 5 -OC(O)N(R) 5 )-、-OC(O)OR 5 -N(R)5 )C(O)N(R 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 and -N(R 5 )S(O)2R 5 ; R 2 Each occurrence is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, perhaloalkoxy, halogen, nitrile, azido, -OR 5 -SR 5 -N(R) 5 )2、-C(O)R 5 -C(O)OR 5 -OC(O)R 5 -N(R) 5 )C(O)R 5 -C(O)N(R) 5 )2、-N(R 5 )C(O)OR 5 -OC(O)N(R) 5 )-、-OC(O)OR 5 -N(R) 5 )C(O)N(R 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 and -N(R 5 )S(O)2R 5 ; R 3 It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, -C(O)R 5 -C(O)OR 5 -C(O)N(R) 5 )2、-S(O)2R 5 or -S(O)2N(R) 5 )2; R 4 It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, -C(O)R 5 -C(O)OR 5 -C(O)N(R) 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 or -P(O)(OR 5 )2; and R 5 Each occurrence of R is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, and aralkyl; or two occurrences of R. 5 Together with the covalently bonded atoms, they form 3-6 membered cycloalkyl or heterocycloalkyl groups.

[0011] In some implementations, m is an integer selected from 1 to 2; and n is an integer selected from 1 to 2.

[0012] In some implementations, X 1 It is -O-, -S-, -(NR) 5 - or -SO2(NR) 5 )-.

[0013] In some implementations, Ar 1 Selected from: , and ;and Ar 2 yes: .

[0014] In some implementations, at least one R 1 It is CF3; and at least one R 2 It's CF3.

[0015] In some embodiments, the compound has Formula 2: 2 Or its pharmaceutically acceptable salt, wherein: m is an integer selected from 1 to 4; n is an integer selected from 1 to 4; X 1 It is -O-, -S-, -OCH2-, -SCH2-, -(NR 5 - or -SO2(NR) 5 )-; X 2 It is -O-, -S-, -OCH2-, -SCH2-, -(NR 5 - or -SO2(NR) 5 )-; Ar 1 Selected from: , , , , , and ; Ar 2 Selected from: , , , , , and ; Each time R appears, it is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, and aralkyl; or R appearing twice together with their covalently bonded atoms form 3-6 membered cycloalkyl or heterocycloalkyl groups. R 1 Each occurrence is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, perhaloalkoxy, halogen, nitrile, azido, -OR 5 -SR 5 -N(R) 5 )2、-C(O)R 5 -C(O)OR 5 -OC(O)R 5 -N(R) 5 )C(O)R 5 -C(O)N(R) 5 )2、-N(R 5 )C(O)OR 5 -OC(O)N(R) 5 )-、-OC(O)OR 5 -N(R) 5 )C(O)N(R 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 and -N(R 5 )S(O)2R 5 ; R 2 Each occurrence is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, perhaloalkoxy, halogen, nitrile, azido, -OR 5 -SR 5 -N(R) 5 )2、-C(O)R 5 -C(O)OR 5 -OC(O)R 5 -N(R)5 )C(O)R 5 -C(O)N(R) 5 )2、-N(R 5 )C(O)OR 5 -OC(O)N(R) 5 )-、-OC(O)OR 5 -N(R) 5 )C(O)N(R 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 and -N(R 5 )S(O)2R 5 ; R 3 It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, -C(O)R 5 -C(O)OR 5 -C(O)N(R) 5 )2、-S(O)2R 5 or -S(O)2N(R) 5 )2; R 4 It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, -C(O)R 5 -C(O)OR 5 -C(O)N(R) 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 or -P(O)(OR 5 )2; and R 5 Each occurrence of R is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, and aralkyl; or two occurrences of R. 5 Together with the covalently bonded atoms, they form 3-6 membered cycloalkyl or heterocycloalkyl groups.

[0017] In some implementations, Ar 1 Selected from: , and ;and Ar 2 yes: .

[0018] In some implementations, at least one R 1It is CF3; and at least one R 2 It's CF3.

[0019] In some implementations, R 3 It is hydrogen and R 4 It is hydrogen.

[0020] In some embodiments, the compound has Formula 3: 3 Or its pharmaceutically acceptable salt, wherein: m is an integer selected from 1 to 2; n is an integer selected from 1 to 2; A is C, CH, or N; X 1 It is -O-, -S-, -(NR) 5 - or -SO2(NR) 5 )-; X 2 It is -O-, -S-, -(NR) 5 -, -OCH2- or -SCH2-; Each time R appears, it is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, and aralkyl; or R appearing twice together with their covalently bonded atoms form 3-6 membered cycloalkyl or heterocycloalkyl groups. R 1 Each occurrence is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, perhaloalkoxy, halogen, nitrile, azido, -OR 5 -SR 5 -N(R) 5 )2、-C(O)R 5 -C(O)OR 5 -OC(O)R 5 -N(R) 5 )C(O)R 5 -C(O)N(R) 5 )2、-N(R 5 )C(O)OR 5 -OC(O)N(R) 5 )-、-OC(O)OR 5 -N(R) 5 )C(O)N(R 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 and -N(R 5)S(O)2R 5 ; R 2 Each occurrence is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, perhaloalkoxy, halogen, nitrile, azido, -OR 5 -SR 5 -N(R) 5 )2、-C(O)R 5 -C(O)OR 5 -OC(O)R 5 -N(R) 5 )C(O)R 5 -C(O)N(R) 5 )2、-N(R 5 )C(O)OR 5 -OC(O)N(R) 5 )-、-OC(O)OR 5 -N(R) 5 )C(O)N(R 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 and -N(R 5 )S(O)2R 5 ; R 3 It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, -C(O)R 5 -C(O)OR 5 -C(O)N(R) 5 )2、-S(O)2R 5 or -S(O)2N(R) 5 )2; R 4 It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, -C(O)R 5 -C(O)OR 5 -C(O)N(R) 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 or -P(O)(OR 5 )2; and R 5 Each occurrence of R is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, and aralkyl; or two occurrences of R. 5Together with the covalently bonded atoms, they form 3-6 membered cycloalkyl or heterocycloalkyl groups.

[0022] In some implementations, R 3 It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, or aralkyl.

[0023] In some implementations, R 3 It is hydrogen.

[0024] In some implementations, R 4 It is hydrogen.

[0025] In some embodiments, the compound is selected from: , , , , , , , , , , , , and its pharmaceutically acceptable salt, where m is an integer selected from 1 to 2; and n is an integer selected from 1 to 2.

[0026] In some implementations, m is 1 and n is 1.

[0027] In some embodiments, the compound is selected from: , , , , , , , , , , , , , And its pharmaceutically acceptable salts.

[0028] In some embodiments, the compound is selected from: , , , , And its pharmaceutically acceptable salts.

[0029] In a second aspect, this document provides a pharmaceutical composition comprising the compound described herein and at least one pharmaceutically acceptable carrier or pharmaceutically acceptable excipient.

[0030] In a third aspect, this article provides a method for treating a bacterial infection in a subject in need, the method comprising administering to the subject a therapeutically effective amount of the compound described herein.

[0031] In some implementations, the bacterial infection is caused by Gram-positive bacteria.

[0032] In some implementations, the bacterial infection is caused by Gram-negative bacteria.

[0033] In some embodiments, the bacterial infection is caused by bacteria selected from Enterococcus faecalis, Staphylococcus aureus, Streptococcus pneumoniae, Enterobacter cloacae, Escherichia coli, Acinetobacter baumannii, Staphylococcus epidermidis, Staphylococcus saprophyticus, Streptococcus pyogenes, and Streptococcus agalactiae.

[0034] In some embodiments, the bacterial infection is caused by bacteria selected from methicillin-resistant Staphylococcus aureus, vancomycin-resistant Staphylococcus aureus, aminoglycoside-resistant Staphylococcus aureus, macrolide-resistant Staphylococcus aureus, and fluoroquinolone-resistant Staphylococcus aureus. Attached Figure Description

[0035] The foregoing aspects and many of the accompanying advantages of the invention will become more readily apparent and understood when taken in conjunction with the accompanying drawings and the following detailed description.

[0036] Figure 1 The structure and sequence analysis of the core enzyme and NusG of Escherichia coli RNA polymerase were depicted. (A) Core enzyme (subunit) of Escherichia coli RNA polymerase bound to NusG (PDB5TBZ). ββ’ Crystal structure of the bacterial transcription complex. Left: Overall structure of the bacterial transcription complex, highlighting key elements of the core enzyme (blue: RNAP β' subunit; orange-red: NusG). Right: Key hydrogen bond interactions between NusG and the β'CH region.

[0037] Figure 2 Left side: AW00783 structure; Middle: AW00783 docking with the pharmacophore model (green sphere: hydrogen bond donor; gray sphere: exclusive region); Right side: AW00783 docking with β'CH (surface view).

[0038] Figure 3 Scheme 1. Synthetic routes for compounds 1-39 are described. a . aReagents and conditions: (a) N,N-diisopropylethylamine (DIPEA), EtOH, reflux, 8 h.

[0039] Figure 4 Scheme 2 is described. The synthetic route for substrates A1-A18 is depicted. a . a Reagents and conditions: (a) For A1-A9: ethylenediamine monohydrate, K2CO3, THF, reflux, overnight; For A10-A15: ethylenediamine monohydrate, CuCl, Cs2CO3, DMSO, 120℃, 8h; For A16: ethylenediamine monohydrate, triethylamine (Et3N), DCM, rt, overnight; For A17-A18: 2-(Boc-amino)ethyl bromide, K2CO3, DMF, 65℃, 8h; (b) CF3COOH, DCM, reflux, 8h.

[0040] Figure 5 Scheme 3 is described. The synthetic route for substrates B1-B17 is depicted. a . a Reagents and conditions: (a) For B1-B10: epichlorohydrin, KI, Cs2CO3, DMF, 80°C, overnight; For B11: (i) epichlorohydrin, Zn(SO3CF3)2, CHCl3, 60°C, 12 h; (ii) KI, MeCN, 80°C, 6 h; For B12: epichlorohydrin, K2CO3, MeCN, 85°C, 8 h; For B13-B16: epichlorohydrin, TBAB, 1M NaOH aqueous solution, rt, 18 h; For B17: epichlorohydrin, KOH, water:dioxane = 1:1, rt, 8 h.

[0041] Figure 6 Table 1 shows the antimicrobial activity (MIC µg / mL) of the lead compound and compounds 1-24.

[0042] Figure 7 The antimicrobial activities (MIC µg / mL) of compounds 25-29 are described in Table 2.

[0043] Figure 8 The antimicrobial activities (MIC µg / mL) of compounds 30-34 are described in Table 3.

[0044] Figure 9 The antimicrobial activities (MIC µg / mL) of compounds 35-39 are described in Table 4.

[0045] Figure 10 Table 5 describes the evaluation of the antimicrobial activity of the derivatives against representative pathogenic Gram-positive bacteria. SEPI: Staphylococcus epidermidis ATCC ®12228, SSAP: Staphylococcus saprophyticus ATCC ® 15305, SPYO: Streptococcus pyogenes (Group A Streptococcus) ATCC ® 19615, SAGA: Agalactiae Streptococcus (Group B Streptococcus) ATCC ® 12386. Van: Vancomycin, Cip: Ciprofloxacin, Oxa: Oxacillin, Gen: Gentamicin.

[0046] Figure 11 Table 6 describes the evaluation of the antimicrobial activity of the derivatives against a range of Streptococcus pneumoniae strains, including clinical isolates. SPNE a Streptococcus pneumoniae ATCC ® 49619, SPNE b Streptococcus pneumoniae strain TCH8431 (HM-145), SPNE c Streptococcus pneumoniae strain NP112 (NR-19213). Van: vancomycin, Cip: ciprofloxacin, Oxa: oxacillin, Gen: gentamicin.

[0047] Figure 12 A heatmap of the MIC (µg / mL) of the compounds and commercially available antibiotics against a group of antibiotic-resistant Staphylococcus aureus was constructed. Our compounds showed strong antimicrobial activity against a range of MRSA and VRSA strains, with MIC values ​​comparable to the type strains. Van: vancomycin; Oxa: oxacillin; Cip: ciprofloxacin; Gen: gentamicin.

[0048] Figure 13 A table depicting the MIC and MBC values ​​of compound 38 and the control drug against Staphylococcus aureus strains is presented.

[0049] Figure 14 The effects of compound 38 on (A) Staphylococcus aureus ATCC were described in CA-MHB medium at ¼×, 1×, 4× and 16× MIC. ® Effects of time-killing kinetics between strains 25923 and (B) CA-MRSA strain USA300. Triple copies were used in the experiment. 25923: Staphylococcus aureus ATCC ® 25923; USA300: CA-MRSA strain USA300.

[0050] Figure 15 The inhibition curve of compound 38, measured by protein complementation assay, was plotted. 50 The calculated value is 166.9 ± 47.24 µM.

[0051] Figure 16Fluorescence microscopy images depicting Bacillus subtilis cells expressing transcription-related proteins (A) NusG-GFP and (B) RpoC-GFP in the presence of rifampin, chloramphenicol, and compound 38 are shown. GFP fluorescence (green), nucleoids stained with DAPI (red), and overlays of GFP and DAPI signals are displayed. Scale bar: 4 µm.

[0052] Figure 17 Fluorescence microscopy images of Bacillus subtilis cells expressing (A) AtpA-GFP and (B) RpsB-GFP proteins in the presence of rifampin, chloramphenicol, colistin, and compound 38 are depicted. GFP fluorescence (green), nucleoids stained with DAPI (red), and overlays of GFP and DAPI signals are shown. Scale bar: 4 µm. Detailed Implementation

[0053] definition The following terms will be used to describe the present invention. In the absence of specific definitions set forth herein, the terms used to describe the present invention should be interpreted in accordance with their common meanings as understood by one of ordinary skill in the art.

[0054] Throughout this disclosure, unless the context otherwise requires, the word "comprise" or its variations such as "comprises" or "comprising" will be understood to imply inclusion of the specified integers or groups of integers, but not to exclude any other integers or groups of integers. It should also be noted that in this disclosure, and particularly in the claims and / or paragraphs, terms such as "comprises," "comprised," "comprising," etc., may have the meanings accorded to them under U.S. patent law; for example, they may mean "includes," "included," "including," etc.; and terms such as "consisting essentially of" and "consists essentially of" have the meanings accorded to them under U.S. patent law, for example, they allow elements not expressly stated, but exclude elements found in the prior art or elements affecting the essential or novel features of the invention.

[0055] Furthermore, throughout this disclosure and the claims, unless the context otherwise requires, the word “include” or variations such as “includes” or “including” will be understood to imply inclusion of the specified integer or group of integers, but not to exclude any other integer or group of integers.

[0056] The use of the singular in this document includes the plural (and vice versa), unless otherwise expressly stated. Furthermore, where the term "about" is used before a quantity value, the present teaching also includes the specific quantity value itself, unless otherwise expressly stated. As used herein, the term "about" means a variation of ±10%, ±7%, ±5%, ±3%, ±1%, or ±0% of the nominal value, unless otherwise specified or inferred.

[0057] As used herein, the terms “treat,” “treating,” “treatment,” etc., refer to the reduction or improvement of a condition / disease and / or its associated symptoms. It will be understood that, although not excluded, treating a condition or disease does not require the complete elimination of the condition, disease, or its associated symptoms. In some implementations, treatment includes the prevention of a condition or disease and / or its associated symptoms. As used herein, the terms “prevention” or “prevent” refer to any action that inhibits or at least delays the occurrence of a condition, disease, or its associated symptoms. Prevention may include primary, secondary, and tertiary levels of prevention, wherein: a) primary prevention avoids the occurrence of disease; b) secondary prevention activities aim to treat disease early, thereby increasing opportunities for intervention to prevent disease progression and the onset of symptoms; and c) tertiary prevention reduces the negative effects of existing disease by restoring function and reducing disease-related complications.

[0058] As used herein, the term "subject" refers to an animal, typically a mammal or human, who will be or has been the subject of treatment, observation, and / or experimentation. When the term is used in conjunction with the administration of the compounds described herein, the subject is already the subject of treatment, observation, and / or administration of the compounds described herein.

[0059] As used herein, the term "therapeutic effective amount" means the amount of a compound or agent that elicits a biological and / or medical response in a cell culture, tissue system, subject, animal, or human, a response sought by an investigator, veterinarian, clinician, or physician, including the reduction of symptoms of the disease, condition, or ailment being treated.

[0060] The term "composition" is intended to include products containing specified amounts of specified ingredients, as well as any products produced directly or indirectly from combinations of specified amounts of specified ingredients.

[0061] The term "pharmaceutically acceptable carrier" refers to a medium used to prepare the desired dosage form of a compound. Pharmaceutically acceptable carriers may include one or more solvents, diluents or other liquid media; dispersants or suspending agents; surfactants; isotonic agents; thickeners or emulsifiers; preservatives; solid binders; lubricants, etc. Remington's Pharmaceutical Sciences, 15th edition, EW Martin (Mack Publishing Co., Easton, Pa., 1975) and Handbook of Pharmaceutical Excipients, 3rd edition, AH Kibbe ed. (American Pharmaceutical Association, 2000) disclose various carriers for formulating pharmaceutical compositions and known techniques for their preparation.

[0062] As used herein, unless otherwise specified, the terms “halogenated” or “halogenated” include fluorinated, chlorinated, brominated, or iodinated compounds.

[0063] As used herein, "alkyl" refers to a straight-chain or branched saturated hydrocarbon group. Examples of alkyl groups include methyl-, ethyl-, propyl (e.g., n-propyl and isopropyl), butyl (e.g., n-butyl, isobutyl, sec-butyl, tert-butyl), pentyl (e.g., 1-methylbutyl, 2-methylbutyl, isopentyl, tert-pentyl, 1,2-dimethylpropyl, neopentyl, and 1-ethylpropyl), hexyl, etc. In various embodiments, the alkyl group may have 1-40 carbon atoms (i.e., C1-40 alkyl), for example, 1-30 carbon atoms (i.e., C1-30 alkyl). In some embodiments, the alkyl group may have 1-6 carbon atoms and may be referred to as "lower alkyl". Examples of lower alkyl groups include methyl, ethyl, propyl (e.g., n-propyl and isopropyl), and butyl (e.g., n-butyl, isobutyl, sec-butyl, tert-butyl). In some embodiments, the alkyl group may optionally be substituted, as described herein. Alkyl groups are not typically substituted by another alkyl, alkenyl, or ynyl group.

[0064] As used herein, "alkenyl" refers to a straight-chain or branched alkyl group having one or more carbon-carbon double bonds. Examples of alkenyl groups include vinyl, propenyl, butenyl, pentenyl, hexenyl, butadienyl, pentadienyl, hexadienyl, etc. The one or more carbon-carbon double bonds can be internal (e.g., in 2-butene) or terminal (e.g., in 1-butene). In various embodiments, the alkenyl group can have 2-40 carbon atoms (i.e., C2-40 alkenyl), for example 2-20 carbon atoms (i.e., C2-20 alkenyl). In some embodiments, the alkenyl group can be substituted, as described herein. The alkenyl group is generally not substituted by another alkenyl, alkyl, or ynyl group.

[0065] As used herein, "cycloalkyl" itself, or as part of another substituent, unless otherwise specified, means a monocyclic hydrocarbon having 3-12 carbon atoms in a ring system and includes hydrogen, straight-chain, branched, and / or cyclic substituents. Exemplary cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, etc.

[0066] As used herein, "heteroatoms" refers to atoms of any element other than carbon or hydrogen, and includes, for example, nitrogen, oxygen, silicon, sulfur, phosphorus, and selenium.

[0067] As used herein, the term "heterocyclic alkyl" includes references to saturated heterocyclic moieties having 3, 4, 5, 6, or 7 ring carbon atoms and 1, 2, 3, 4, or 5 ring heteroatoms selected from nitrogen, oxygen, phosphorus, and sulfur. The group can be a polycyclic ring system, but is more commonly monocyclic. The term includes references to groups such as azirrobutyl, pyrrolidinyl, tetrahydrofuranyl, piperidinyl, oxadiazolidinyl, pyrazolyl, imidazolyl, indolizidyl, piperazinyl, thiazolyl, morpholinyl, thiomorpholinyl, quinolizidyl, etc.

[0068] As used herein, "aryl" refers to an aromatic monocyclic or polycyclic ring system in which two or more aromatic rings are fused together (i.e., have a common bond) or at least one aromatic monocyclic ring is fused to one or more cycloalkyl and / or heterocyclic alkyl rings. An aryl group may have 6-24 carbon atoms in its ring system (e.g., C6-24 aryl), and may include multiple fused rings. In some embodiments, a polycyclic aryl group may have 8-24 carbon atoms. Any suitable ring position of the aryl group may be covalently linked to a defined chemical structure. Examples of aryl groups having only aromatic carbon rings include phenyl, 1-naphthyl (bicyclic), 2-naphthyl (bicyclic), anthracene (tricyclic), phenanthrene (tricyclic), and pentaphenyl (pentacyclic) groups. Examples of polycyclic systems in which at least one aromatic carbide ring is fused to one or more cycloalkyl and / or heteroalkyl rings include, but are not limited to, benzo[i](x) derivatives of cyclopentane (i.e., indenyl, which is a 5,6-bicyclocycloalkyl / aromatic ring system), benzo[i](x) derivatives of cyclohexane (i.e., tetrahydronaphthalene, which is a 6,6-bicyclocycloalkyl / aromatic ring system), benzo[i](x) derivatives of imidazoline (i.e., benzimidazolinyl, which is a 5,6-bicyclocycloalkyl / aromatic ring system), and benzo[i](x) derivatives of pyran (i.e., benzopyranyl, which is a 6,6-bicyclocycloalkyl / aromatic ring system). Other examples of aryl groups include benzodioxane-hexyl, benzodioxane-pentyl, benzodihydropyranyl, indololinyl, etc. In some embodiments, the aryl group may optionally be substituted.

[0069] The term "aralkyl" refers to an alkyl group that has been substituted with an aryl group.

[0070] As used herein, "heteroaryl" refers to an aromatic monocyclic or polycyclic ring system containing at least one cyclic heteroatom selected from oxygen (O), nitrogen (N), sulfur (S), silicon (Si), and selenium (Se), wherein at least one ring in the ring system is aromatic and contains at least one cyclic heteroatom. Polycyclic heteroaryls include those having two or more heteroaryl rings fused together, and those having at least one monocyclic heteroaryl ring fused to one or more aromatic carbocyclic, non-aromatic, and / or non-aromatic cyclic heteroalkyl rings. Heteroaryls as a whole may have, for example, 5-24 ring atoms and contain 1-5 cyclic heteroatoms (i.e., 5-20 membered heteroaryls). Heteroaryls may be attached to a defined chemical structure at any heteroatom or carbon atom that results in a stable structure. Typically, heteroaryl rings do not contain OO, SS, or SO bonds. However, one or more N or S atoms in a heteroaryl group may be oxidized (e.g., pyridine N-oxide, thiophene S-oxide, thiophene S,S-dioxide). Examples of heteroaryl groups include, for example, 5- or 6-membered monocyclic and 5- to 6-membered bicyclic ring systems shown below:

[0071] Where T is O, S, NH, N-alkyl, N-aryl, N-(arylalkyl) (e.g., N-benzyl), SiH2, SiH(alkyl), Si(alkyl)2, SiH(arylalkyl), Si(arylalkyl)2, or Si(alkyl)(arylalkyl). Examples of such heteroaryl rings include pyrroloyl, furanyl, thiopheneyl, pyridinyl, pyrimidinyl, pyridazinyl, triazolyl, tetrazolyl, pyrazolyl, imidazolyl, isothiazolyl, thiazolyl, thiadiazolyl, isoxazolyl, oxazolyl, oxadiazolyl, indolyl, isoyndolyl, benzofuranyl, benzothiopheneyl, quinolinyl, 2-methylquinolinyl, isoquinolinyl, quinoxalinyl, quinazolinyl, benzotriazolyl, benzimidazolyl, benzothiazolyl, Benzisothiazolyl, benzisoxazolyl, benzoxadiazolyl, benzoxazolyl, cenolinyl, 1H-inzolyl, 2H-inzolyl, indoleazinyl, isobenzofuranyl, naphridinyl, phthalazinyl, pteridinyl, purineyl, oxazolopyridinyl, thiazopyridinyl, imidazopyridinyl, furanopyridinyl, thienopyridinyl, pyridinopyrimidinyl, pyridinopyrazinyl, pyridinopyridazinyl, thienothiazoyl, thienooxazolyl, thienoimidazoyl, etc. Further examples of heteroaryl groups include 4,5,6,7-tetrahydroindolyl, tetrahydroquinolinyl, benzothienopyridinyl, benzofuranopyridinyl, etc. In some embodiments, the heteroaryl group may be substituted, as described herein. In some embodiments, the heteroaryl group may optionally be substituted.

[0072] The term "optional substitution" refers to a chemical group, such as alkyl, cycloalkyl, aryl, etc., in which one or more hydrogen atoms may be replaced by substituents as described herein, such as halogens, azides, alkyl, aralkyl, alkenyl, alkynyl, cycloalkyl, hydroxyl, alkoxy, amino, nitro, mercapto, imino, amide, phosphonate, hypophosphite, carbonyl, carboxyl, silyl, ether, alkylthio, sulfonyl, sulfonamide, ketone, aldehyde, ester, heterocyclic, aromatic or heteroaromatic moiety, -CF3, -CN, etc.

[0073] The term "nitro" is generally accepted in the art and refers to -NO2; the term "halogen" is generally accepted in the art and refers to -F, -Cl, -Br, or -I; the term "mercapto" is generally accepted in the art and refers to -SH; the term "hydroxyl" means -OH; and the terms "sulfonyl" and "sulfone" are generally accepted in the art and refer to -SO2-. "Halide" refers to the corresponding anion of the halogen.

[0074] The symbol "in chemical structure" "or" "or" "or" "" indicates the position where a specified chemical structure is bonded to another chemical structure.

[0075] As used herein, the term "pharmaceutically acceptable salt" means that, within reasonable medical judgment, it is suitable for use in contact with the tissues of a subject without excessive toxicity, irritation, allergic reactions, etc., and is commensurate with a reasonable benefit / risk ratio. Pharmaceutically acceptable salts are well known in the art. For example, Berge et al. describe pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences (1977) 66:1-19. Pharmaceutically acceptable salts of the compounds provided herein include those derived from suitable inorganic and organic acids and bases. Examples of pharmaceutically acceptable non-toxic acid addition salts are salts of amino groups formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid, and perchloric acid, or with organic acids such as acetic acid, oxalic acid, maleic acid, tartaric acid, citric acid, succinic acid, or malonic acid, or by using other methods used in the art, such as ion exchange. Other pharmaceutically acceptable salts include adipic acid salts, alginate salts, ascorbate salts, aspartate salts, benzenesulfonate salts, besylate salts, benzoate salts, hydrogen sulfate salts, borate salts, butyrate salts, camphorate salts, camphor sulfonate salts, citrate salts, cyclopentanepropionate salts, diglucuronate salts, dodecyl sulfate salts, ethanesulfonate salts, formate salts, fumarate salts, glucohepanoate salts, glycerophosphate salts, glucuronate salts, hemisulfate salts, heptanate salts, and hexanoate salts. The organic acids that can be derived from salts include, for example, acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, dihydroxynaphthalate, pectate, persulfate, 3-phenylpropionate, phosphate, picrate, neopentanoate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valerate, etc. In some embodiments, the organic acids that can be derived from salts include, for example, acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, etc.

[0076] Pharmaceutically acceptable salts derived from suitable alkalis include alkali metals, alkaline earth metals, ammonium, and nitrogen. + (C 1-4Alkyl)4 salts. Representative alkali metal or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, iron, zinc, copper, manganese, aluminum, etc. Further pharmaceutically acceptable salts, where appropriate, include non-toxic ammonium, quaternary ammonium, and amine cations formed using anti-charge ions such as halide, hydroxide, carboxyl, sulfate, phosphate, nitrate, lower alkyl sulfonates, and aryl sulfonates. Organic bases from which salts can be derived include, for example, primary, secondary, and tertiary amines, substituted amines (including naturally occurring substituted amines), cyclic amines, basic ion exchange resins, etc., such as isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, and ethanolamine. In some embodiments, pharmaceutically acceptable base addition salts are selected from ammonium, potassium, sodium, calcium, and magnesium salts.

[0077] This disclosure provides a compound of Formula 1: 1 Or its pharmaceutically acceptable salt, wherein: m is an integer selected from 1 to 4; n is an integer selected from 1 to 4; X 1 It is -O-, -S-, -OCH2-, -SCH2-, -(NR 5 - or -SO2(NR) 5 )-; X 2 It is -O-, -S-, -OCH2-, -SCH2-, -(NR 5 - or -SO2(NR) 5 )-; Ar 1 Selected from: , , , , , and ; Ar 2 Selected from: , , , , , and ; Each time R appears, it is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, and aralkyl; or R appearing twice together with their covalently bonded atoms form 3-6 membered cycloalkyl or heterocycloalkyl groups. R 1 Each occurrence is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, perhaloalkoxy, halogen, nitrile, azido, -OR 5 -SR 5 -N(R) 5 )2、-C(O)R 5 -C(O)OR 5 -OC(O)R 5 -N(R) 5 )C(O)R 5 -C(O)N(R) 5 )2、-N(R 5 )C(O)OR 5 -OC(O)N(R) 5 )-、-OC(O)OR 5 -N(R) 5 )C(O)N(R 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 and -N(R 5 )S(O)2R 5 ; R 2 Each occurrence is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, perhaloalkoxy, halogen, nitrile, nitro, azide, -OR 5 -SR 5 -N(R) 5 )2、-C(O)R 5 -C(O)OR 5 -OC(O)R 5 -N(R) 5 )C(O)R 5 -C(O)N(R) 5 )2、-N(R 5 )C(O)OR 5 -OC(O)N(R) 5 )-、-OC(O)OR 5 -N(R) 5 )C(O)N(R 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 and -N(R 5 )S(O)2R 5 ; R 3It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, -C(O)R 5 -C(O)OR 5 -C(O)N(R) 5 )2、-S(O)2R 5 or -S(O)2N(R) 5 )2; R 4 It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, -C(O)R 5 -C(O)OR 5 -C(O)N(R) 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 or -P(O)(OR 5 )2; and R 5 Each occurrence of R is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, and aralkyl; or two occurrences of R. 5 Together with the covalently bonded atoms, they form 3-6 membered cycloalkyl or heterocycloalkyl groups.

[0079] In some embodiments, the compound of formula 1 does not include: .

[0080] The compounds described herein may include isomers at different positions indicated by bonds that are not connected to the vertices of the chemical structure, for example, using the model structure illustration below: .

[0081] In this model structure, where the valence allows, the group R can be attached to any atom on the ring structure, i.e., carbon 2, 3, 4, 5 or 6 in the above structure.

[0082] In some implementations, m is an integer selected from 1-4, 1-3, 2-3, 3-4, and 1-2. In some implementations, m is 1, 2, 3, or 4.

[0083] In some implementations, n is an integer selected from 1-4, 1-3, 2-3, 3-4, and 1-2. In some implementations, n is 1, 2, 3, or 4.

[0084] In some implementations, X 1 It is -O-, -S-, -(NR) 5 - or -SO2(NR)5 In some implementations, X 1 Yes - (NR) 5 )-, where R 5 It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, or aralkyl. In some embodiments, X 1 It is -(NH)-.

[0085] In some implementations, X 2 It is -O-, -S-, -(NR) 5 -, -OCH2-, or -SCH2-. In some implementations, X 2 It is -O-, -S-, -(NR) 5 -, -OCH2- or -SCH2-, where R 5 It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, or aralkyl. In some embodiments, X 2 It is -O-, -S-, -(NH)-, -OCH2- or -SCH2-.

[0086] In some implementations, Ar 1 yes: , , , , , or .

[0087] In some implementations, Ar 1 yes: , or .

[0088] In some implementations, Ar 1 yes: or .

[0089] In some implementations, Ar 1 yes: , , , , , or .

[0090] In some implementations, Ar 1 yes: or .

[0091] In some implementations, Ar 2 yes: , , , , , or .

[0092] In some implementations, Ar 2 yes: .

[0093] In some implementations, Ar 2 yes: , or .

[0094] Each occurrence of R can be independently selected from hydrogen, alkyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl; or two occurrences of R together with their covalently bonded atoms can form a 3-6 membered cycloalkyl or heterocycloalkyl group. In some embodiments, each occurrence of R is independently hydrogen, alkyl, cycloalkyl, aryl, or heteroaryl. In some embodiments, each occurrence of R is independently hydrogen or alkyl. In some embodiments, R is hydrogen.

[0095] R 1 Each occurrence can be independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, perhaloalkoxy, halogen, nitrile, nitro, azide, -OR 5 -N(R) 5 )2、-C(O)R 5 -C(O)OR 5 -OC(O)R 5 -N(R) 5 )C(O)R 5 and -C(O)N(R) 5 2. In some implementations, R 1 Each time it appears independently, it is hydrogen, alkyl, haloalkyl, perhaloalkyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, halogen, nitrile, nitro, azide, -OR 5 -N(R) 5 )2、-C(O)R 5-C(O)OR 5 -OC(O)R 5 -N(R) 5 )C(O)R 5 and -C(O)N(R) 5 2. In some implementations, R 1 At least one instance is -CF3.

[0096] R 2 Each occurrence can be independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, perhaloalkoxy, halogen, nitrile, nitro, azide, -OR 5 -N(R) 5 )2、-C(O)R 5 -C(O)OR 5 -OC(O)R 5 -N(R) 5 )C(O)R 5 and -C(O)N(R) 5 2. In some implementations, R 2 Each time it appears independently, it is hydrogen, alkyl, haloalkyl, perhaloalkyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, halogen, nitrile, nitro, azide, -OR 5 -N(R) 5 )2、-C(O)R 5 -C(O)OR 5 -OC(O)R 5 -N(R) 5 )C(O)R 5 and -C(O)N(R) 5 2. In some implementations, R 2 At least one instance is -CF3.

[0097] R 3 It can be hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, -C(O)R 5 -C(O)OR 5 -C(O)N(R) 5 )2、-S(O)2R 5 or -S(O)2N(R) 5 2. In some implementations, R 3 It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, or aralkyl. In some embodiments, R 3It is hydrogen, C1-C6 alkyl, C1-C5 alkyl, C1-C4 alkyl, C1-C3 alkyl, or C1-C2 alkyl. In some embodiments, R 3 It is hydrogen.

[0098] R 4 It can be hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, -C(O)R 5 -C(O)OR 5 -C(O)N(R) 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 or -P(O)(OR 5 2. In some implementations, R 4 It is hydrogen, -C(O)R 5 -C(O)OR 5 or -P(O)(OR 5 2. In some implementations, R 4 It is hydrogen or -P(O)(OH)2. In some embodiments, R 4 It is hydrogen.

[0099] R 5 Each occurrence of R can be independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, and aralkyl; or two occurrences of R. 5 Together with the covalently bonded atoms, they form 3-6, 4-6, or 5-6 membered cycloalkyl or heterocycloalkyl groups. In some embodiments, R 5 Each time it appears, it is independently hydrogen, alkyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, or aralkyl.

[0100] In some embodiments, the compound has Formula 2: 2 Or its pharmaceutically acceptable salt, wherein Ar 1 Ar 2 X 1 X 2 R 3 and R 4 Each is independent as defined in any of the implementations described herein.

[0102] In some embodiments, the compound has Formula 3: 3 Or a pharmaceutically acceptable salt thereof, wherein: A is C, CH or N and m, n, X1 X 2 R 1 R 2 R 3 and R 4 Each is independently defined as in any of the embodiments described herein. In the case where A in the compound of Formula 3 is C, when C is with R... 1 or CF 3 During bonding, it will exist as C-R1 or C-CF3, while when C is not bonded to R... 1 or CF 3 During bonding, it will exist as CH.

[0104] In some embodiments, the compound is selected from: , , , , , , , , , , , , , and its pharmaceutically acceptable salt, where m is an integer selected from 1 to 2; n is an integer selected from 1 to 2, and R 1 R 2 R 3 and R 4 Each is independent as defined in any of the embodiments described herein. In some embodiments, R 3 and R 4 Each is hydrogen.

[0105] In some embodiments, the compound is selected from: , , , , and its pharmaceutically acceptable salt, where m is an integer selected from 1 to 2; n is an integer selected from 1 to 2, and R 1 R 2 R 3 and R 4 Each is independent as defined in any of the embodiments described herein. In some embodiments, R 3 and R 4 Each is hydrogen.

[0106] In some embodiments, the compound is selected from: , , , , , , , , , , , , And its pharmaceutically acceptable salts.

[0107] In some embodiments, the compound is selected from: , , , , And its pharmaceutically acceptable salts.

[0108] This disclosure also provides a pharmaceutical composition comprising the compound described herein and at least one pharmaceutically acceptable excipient and / or pharmaceutically acceptable carrier.

[0109] The compounds described herein and their pharmaceutically acceptable salts may be administered to subjects, alone or in combination with a pharmaceutically acceptable carrier or diluent, in a pharmaceutical composition, in accordance with standard pharmaceutical practice. The compounds may be administered parenterally. Parenterally administration includes intravenous, intramuscular, intraperitoneal, subcutaneous, and topical administration.

[0110] Therefore, this disclosure provides pharmaceutically acceptable compositions comprising a therapeutically effective amount of the compound described herein, formulated with one or more pharmaceutically acceptable carriers (additives) and / or diluents. The pharmaceutical compositions of this disclosure may be specifically formulated for administration in liquid form, including those suitable for: (1) parenteral administration, such as as a sterile solution or suspension, for example, via intravenous administration.

[0111] As illustrated herein, certain embodiments of the compounds described herein may contain a basic functional group, such as an amino group, and thus be able to form pharmaceutically acceptable salts with pharmaceutically acceptable acids. In this regard, the term "pharmaceutically acceptable salt" refers to a relatively non-toxic inorganic and organic acid addition salt of the compounds of this disclosure. These salts may be prepared in situ during the manufacture of an application medium or dosage form, or by reacting a purified compound of the invention in its free basic form with a suitable organic or inorganic acid, and separating the resulting salt during subsequent purification. Representative salts include bromides, chlorides, sulfates, bisulfates, carbonates, bicarbonates, nitrates, acetates, valerates, oleates, palmitates, stearates, laurates, benzoates, lactates, phosphates, toluenesulfonates, citrates, maleates, fumarates, succinates, tartrates, naphthalates, methanesulfonates, glucono-p-ethylates, lacturonates, and laurylsulfonates, etc.

[0112] Pharmaceutically acceptable salts of the compounds disclosed herein include conventional non-toxic salts or quaternary ammonium salts of the compounds, such as those derived from non-toxic organic or inorganic acids. For example, such conventional non-toxic salts include those derived from inorganic acids, such as hydrochloric acid, hydrobromic acid, sulfuric acid, aminosulfonic acid, phosphoric acid, nitric acid, etc.; and salts prepared from organic acids, such as acetic acid, propionic acid, succinic acid, glycolic acid, stearic acid, lactic acid, malic acid, tartaric acid, citric acid, ascorbic acid, palmitic acid, maleic acid, hydroxymaleic acid, phenylacetic acid, glutamic acid, benzoic acid, salicylic acid, p-aminobenzenesulfonic acid, 2-acetoxybenzoic acid, fumaric acid, toluenesulfonic acid, methanesulfonic acid, ethanedisulfonic acid, oxalic acid, hydroxyethylsulfonic acid, etc.

[0113] In other cases, the compounds described herein may contain one or more acidic functional groups, thus enabling them to form pharmaceutically acceptable salts with pharmaceutically acceptable bases. In these cases, the term "pharmaceutically acceptable salt" refers to a relatively non-toxic inorganic or organic base addition salt of the compounds of the present invention. These salts can be prepared in situ during the manufacture of the administration medium or dosage form, or by reacting a purified compound in its free acid form with a suitable base, such as a hydroxide, carbonate, or bicarbonate of a pharmaceutically acceptable metal cation, with ammonia, or with a pharmaceutically acceptable primary, secondary, or tertiary organic amine. Representative alkali metal or alkaline earth metal salts include lithium, sodium, potassium, calcium, magnesium, and aluminum salts. Representative organic amines that can be used to form base addition salts include ethylamine, diethylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine, etc.

[0114] Wetting agents, emulsifiers and lubricants (such as sodium lauryl sulfate and magnesium stearate), as well as colorants, releasing agents, coating agents, sweeteners, flavoring agents and aromas, preservatives, solubilizers, buffers and antioxidants may also be present in the composition.

[0115] Methods for preparing a pharmaceutical product comprising the compound described herein include the steps of combining the compound described herein with a carrier and optionally one or more excipients. Generally, a formulation is prepared by uniformly and tightly combining the compound described herein with a liquid carrier (liquid formulation), the liquid carrier followed by lyophilization (powder formulation, for reconstitution with sterile water, etc.), or a finely powdered solid carrier, or both, and then shaping or packaging the product if desired.

[0116] The pharmaceutical compositions suitable for parenteral administration according to this disclosure comprise one or more of the compounds described herein, combined with one or more pharmaceutically acceptable sterile isotonic aqueous or non-aqueous solutions, dispersions, suspensions or emulsions, or sterile powders (which may be reconstituted into sterile injectable solutions or dispersions immediately prior to use), which may contain sugars (e.g., sucrose), alcohols, nonionic surfactants (e.g., Tween® 20), antioxidants, buffers, antibacterial agents, chelating agents, solutes or suspending agents or thickeners that make the formulation isotonic with the blood of the intended recipient.

[0117] Examples of suitable aqueous and non-aqueous carriers that can be used in the pharmaceutical compositions of this disclosure include water, ethanol, polyols (e.g., glycerol, propylene glycol, polyethylene glycol, etc.) and suitable mixtures thereof, vegetable oils such as olive oil, and injectable organic esters such as ethyl oleate. For example, in the case of dispersions, appropriate flowability can be maintained by using a coating material such as lecithin, by maintaining the desired particle size, and by using a surfactant.

[0118] These compositions may also contain adjuvants such as preservatives, wetting agents, emulsifiers, and dispersants. Prevention of microbial action on the compounds of this disclosure can be ensured by including various antibacterial and antifungal agents, such as parabens, chlorobutanol, phenol, sorbic acid, etc. Isotonic agents, such as sugars, sodium chloride, etc., may also be included in the composition. Furthermore, prolonged absorption of injectable drug forms can be achieved by including agents that delay absorption, such as aluminum monostearate and gelatin.

[0119] On the other hand, this article provides a method for treating a bacterial infection in a subject in need, the method comprising the step of administering a therapeutically effective amount of the compound described herein to the subject.

[0120] Exemplary bacterial infections include, but are not limited to, pneumonia, otitis media, sinusitis, bronchitis, tonsillitis, and mastoiditis associated with infections of *Streptococcus pneumoniae*, *Haemophilus influenzae*, *Moraxella catarrhalis*, *Staphylococcus aureus*, *Enterococcus faecalis*, *Enterococcus faecium*, *Enterococcus pyogenes*, *Staphylococcus epidermidis*, *Staphylococcus hemolyticus*, or *Peptostreptococcus* species; pharyngitis, rheumatic fever, and glomerulonephritis associated with infections of *Streptococcus pyogenes*, group C and group G streptococci, *Corynebacterium diphtheriae*, or *Actinomyces hemolyticus*; respiratory infections associated with infections of *Mycoplasma pneumoniae*, *Legionella pneumophila*, *Streptococcus pneumoniae*, *Haemophilus influenzae*, or *Chlamydia pneumoniae*; and infections caused by *Staphylococcus aureus*, *Staphylococcus hemolyticus*, *Enterococcus faecalis*, *Streptococcus pneumoniae*, *Enterococcus faecalis*, *Streptococcus pneumoniae*, *Haemophilus influenzae*, or *Chlamydia pneumoniae*. Enterococcus faecalis and Enterococcus tenuisae, including strains resistant to known antimicrobial agents (such as, but not limited to, β-lactams, vancomycin, aminoglycosides, quinolones, chloramphenicol, tetracyclines, and macrolides), causing blood and tissue infections, including endocarditis and osteomyelitis; uncomplicated skin and soft tissue infections and abscesses associated with Staphylococcus aureus, coagulase-negative staphylococci (i.e., Staphylococcus epidermidis, Staphylococcus hemolyticus, etc.), Streptococcus pyogenes, Streptococcus agalactiae, Group CF Streptococci (small colony streptococci), viridans streptococci, Corynebacterium microphyllum, Clostridium species, or Bartonella henselae, as well as puerperal fever; infections associated with Staphylococcus aureus and coagulase-negative staphylococci. Uncomplicated acute urinary tract infections associated with infection with *Enterococcus* species or *Enterococcus* spp.; urethritis and cervicitis; sexually transmitted infections associated with *Chlamydia trachomatis*, *Haemophilus ducreyi*, *Treponema pallidum*, *Ureaplasma urealyticum*, or *Neisseria gonorrhoeae*; toxin-related illnesses associated with *Staphylococcus aureus* (food poisoning and toxic shock syndrome) or *Streptococcus* spp. A, B, and C; ulcers associated with *Helicobacter pylori* infection; systemic febrile syndrome associated with *Relapsing fever* infection; Lyme disease associated with *Borrelia burgdorferi* infection; infections associated with *Chlamydia trachomatis*, *Neisseria gonorrhoeae*, *Staphylococcus aureus*, *Streptococcus pneumoniae*, *Streptococcus pyogenes*, *Haemophilus influenzae*, or *Listeria* species. Infection-related conjunctivitis, keratitis, and dacryocystitis; disseminated mycobacterial complex (MAC) disease associated with infection with Mycobacterium avium or intracellular mycobacteria; infections caused by Mycobacterium tuberculosis, Mycobacterium leprae, Mycobacterium paratuberculosis, Mycobacterium kansas, or Mycobacterium cypriniforme; gastroenteritis associated with infection with Campylobacter jejuni; intestinal protozoan infections associated with infection with Cryptosporidium species; odontogenic infections associated with infection with Streptococcus viridans; persistent cough associated with infection with Bordetella pertussis; gas gangrene associated with infection with Clostridium perfringens or Bacteroides species; and atherosclerosis or cardiovascular disease associated with infection with Helicobacter pylori or Chlamydia pneumoniae.Bacterial infections and related conditions that can be treated or prevented in animals include the following: bovine respiratory diseases associated with infections of Pasteurella multocida, Pasteurella multocida, Mycoplasma bovis, or Bordetella species; bovine intestinal diseases associated with infections of Escherichia coli or protozoa (i.e., coccidia, Cryptosporidium, etc.); dairy cow mastitis associated with infections of Staphylococcus aureus, Streptococcus lactis, Streptococcus agalactiae, Streptococcus dysgalactiae, Klebsiella species, Corynebacterium species, or Enterococcus species; swine respiratory diseases associated with infections of Actinobacillus pleuropneumoniae, Pasteurella multocida, or Mycoplasma species; swine intestinal diseases associated with infections of Escherichia coli, Lawsonia intracellularis, Salmonella species, or Spirochetes swine dysenteriae; and diseases associated with infections of *Fusobacterium*. Bovine foot rot associated with infection of fungal species; bovine metritis associated with infection of *Escherichia coli*; bovine hairy warts associated with infection of *Fusobacterium necrophorum* or *Bacteroides nodosum*; bovine conjunctivitis associated with infection of *Moraxella bovis*; bovine premature birth and abortion associated with infection of protozoa (i.e., *Neospora*); urinary tract infections associated with infection of *Escherichia coli* in dogs and cats; skin and soft tissue infections associated with infection of *Staphylococcus epidermidis*, *Staphylococcus intermedius*, coagulase-negative staphylococci*, or *Pasteurella multocida* in dogs and cats; and dental or oral infections associated with infection of species of *Alcaligenes*, *Bacteroides*, *Clostridium*, *Enterobacter*, *Eubacterium*, *Peptostreptococcus*, *Porphyromonas*, or *Prevotella* in dogs and cats. Other bacterial infections and conditions associated with said infections that can be treated or prevented according to the method of the present invention are mentioned in JP Sanford et al., “The Sanford Guide To Antimicrobial Therapy,” 26th edition, (Antimicrobial Therapy, Inc., 1996).

[0121] In some embodiments, the bacteria are Gram-positive bacteria. In some embodiments, the bacteria are Gram-negative bacteria.

[0122] In some embodiments, the bacteria are antibiotic-resistant bacteria. In some embodiments, the antibiotic-resistant bacteria are selected from methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant Staphylococcus aureus (RVSA), aminoglycoside-resistant Staphylococcus aureus (MRSA), macrolide-resistant Staphylococcus aureus (MRSA), and fluoroquinolone-resistant Staphylococcus aureus (FRA).

[0123] In some embodiments, the bacteria are selected from Enterococcus faecalis, Staphylococcus aureus, Streptococcus pneumoniae, Enterobacter cloacae, Escherichia coli, Acinetobacter baumannii, Staphylococcus epidermidis, Staphylococcus saprophyticus, Streptococcus pyogenes, and Streptococcus agalactiae. In some embodiments, the bacteria are Enterococcus faecalis ATCC. ® 19433, Staphylococcus aureus ATCC ®25923, Staphylococcus aureus ATCC ® 29213, Streptococcus pneumoniae ATCC ® 49619, Enterobacter cloacae ATCC ® 13047, Acinetobacter baumannii ATCC ® 19606, Enterobacter cloacae ATCC ® 13047, Escherichia coli ATCC ® 25922, Staphylococcus epidermidis ATCC ® 12228, Streptococcus pyogenes ATCC ® 19615, Agalactia cholecystis (ATCC) ® 12386, W-231 ST45, ATCC ® BAA-43, ATCC ® BAA-44, ST239, W-45 ST59, W-47 ST30, ST22, USA300, SA-RN4220-pUL5054 or SA-1199B.

[0124] The subject can be a canine, feline, bovine, equine, non-human primate, or human. In some embodiments, the subject is a human.

[0125] In some embodiments, the compounds described herein are bactericidal or bacteriostatic.

[0126] On the other hand, this article provides the use of the compounds described herein in the preparation of medicaments for treating bacterial infections.

[0127] On the other hand, this document provides a method for treating a bacterial infection in a subject in need, the method comprising the step of co-administering a therapeutically effective amount of the compound described herein and an antibacterial agent to the subject. The antibacterial agent may be any antibacterial agent known in the art.

[0128] Design and computer screening X-ray crystal structure of *E. coli* NusG complexed with RNAP (PDB: 5TBZ) was used to generate a pharmacophore model. The study showed that the binding of NusG to β'CH is a crucial interaction site in the RNAP-NusG PPI, where NusG interacts with β'CH at sites closely adjacent to or even overlapping with σ. NusG uses the carbonyl groups G17, T24, and T46 in the main chain to bind the guanine residues R281, R278, and R270 of β'CH. Figure 1(A right side). Our previous research has shown that the three arginine residues are important for RNAP binding σ (an essential transcription initiation factor). We believe that these hydrogen bond interactions are also important for NusG function during transcription, therefore we chose G17, T24, and T46 of NusG to create a pharmacophore model, supplemented with dedicated regions to avoid spatial conflicts.

[0129] Structure-activity relationship study Based on the docking model of AW00783 and the pharmacophore model ( Figure 2 (On the right side), we decided to first modify the two terminal aryl rings by changing a variety of common and available substituents in the drug, followed by isosteric changes in the middle straight chain, especially heteroatoms that can affect the construction of the molecular structure.

[0130] A group of bacteria was used to evaluate the antimicrobial activity of compounds 1-39, comprising eight pathogens from the WHO Priority Pathogens List for Guiding the Development of New Antibiotics. This group consisted of four Gram-positive strains: Enterococcus faecalis ATCC. ® 19433 (EFAE), Staphylococcus aureus ATCC ® 25923 (SAUR a ) and 29213 (SAUR b Streptococcus pneumoniae ATCC ® 49619 (SPNE); and five Gram-negative strains: Klebsiella pneumoniae ATCC ® 700603 (KPNE), Acinetobacter baumannii ATCC ® 19606 (ABAU), Pseudomonas aeruginosa ATCC ® 27853 (PAER), Enterobacter cloacae ATCC ® 13047 (ECLO) and Escherichia coli ATCC ® Composition 25922 (ECOL). To determine the minimum inhibitory concentration (MIC) of the tested compounds, a broth microdilution assay was performed according to guidelines established by the Clinical and Laboratory Standards Institute (CLSI). The lead compound AW00783 showed mild antimicrobial activity against Staphylococcus aureus and Streptococcus pneumoniae, with MICs ranging from 128 to 256 μg / mL.

[0131] As shown in Table 1, initial modifications were made to the two terminal aromatic rings of the lead compound. Compound 1, lacking a substituent on Ar1, did not show antimicrobial activity. This result suggests that the presence of a trifluoromethyl group (rather than the nitrogen atom of pyrimidine) on Ar1 of AW00783 may promote interaction with R281. This hypothesis was confirmed by the moderate activity of compound 8, which contains a trifluoromethyl group on the benzene ring of Ar1 lacking pyrimidine nitrogen. Furthermore, compounds 2–6, with trifluoromethyl substitutions at different positions on the pyridine ring, indicate that the relative position of the trifluoromethyl group on the aromatic ring also affects antimicrobial activity. Among these derivatives, 4 and 8 showed promising activity and were selected for further modification. Meanwhile, evaluation of derivatives with different substituents on Ar2 confirmed that trifluoromethyl remains the preferred choice with other hydrogen bond acceptors, as shown in Table 1. Figure 6 The activity of compounds 10-13, with alternative hydrogen bond acceptors, was not improved, suggesting that other substituents may be incompatible with binding to R270. Although the nitro group promotes both hydrogen bonding and electrostatic interactions, it shows a weaker affinity for β'CH compared to trifluoromethyl, as observed in the binding model of AW00783. Additionally, compounds 4, 8, and 14 demonstrated some antimicrobial activity against Gram-negative bacteria.

[0132] When Ar2 is substituted with a trifluoromethyl group, the substituents in Ar1 are further modified. Compounds 17 and 19, with trifluoromethyl groups at the para and ortho positions of the benzene ring in Ar1, demonstrated the greatest activity among this group of analogs, especially 19. This compound showed the most potent antimicrobial activity against Gram-positive bacteria, particularly Streptococcus pneumoniae, with a MIC of 4 μg / mL. It also exhibited enhanced activity against Gram-negative bacteria such as Acinetobacter baumannii, Enterobacter cloacae, and Escherichia coli, with a MIC of approximately 16 μg / mL (Table 1). Figure 6 In contrast, when the cyano group, known as a hydrogen bond acceptor and commonly used in rational drug design, was substituted at the same position, the antimicrobial activity of compound 21 decreased. Meanwhile, modifications at the CF3 position of Ar2 from compounds 19 through 22 and 23 did not yield more promising derivatives. This finding suggests that trifluoromethyl groups at the para or ortho position of Ar1 and the para position of Ar2 are optimal substitutions on the terminal aromatic ring, acting as hydrogen bond acceptors.

[0133] In addition to modifications on Ar1 and Ar2, we also investigated the length of the X2 linker. Compound 25, with X2 being NH, showed a slightly improved activity compared to 15. (See Table 2). Figure 7As shown in Figure 1, a methylene group is inserted between oxygen and Ar2. Compared to phenyl ether 17, benzyl ether 27 exhibits twice the activity against Acinetobacter baumannii and comparable activity against other Gram-positive and Gram-negative bacteria. Furthermore, modification of Ar2 by altering the CF3 substitution position resulted in changes in activity (27-29), indicating that para substitution is advantageous, similar to the trend observed in previously synthesized analogs (19, 22, 23). The absence of CF3, however, reduced activity (26).

[0134] When we changed X2 from a methylene ether to a thioether, compound 30 showed better antimicrobial activity than 15, demonstrating the best antimicrobial activity among pyridine derivatives. This improvement in activity against both Gram-positive and Gram-negative bacteria was comparable to that of compound 19, as shown in Table 3. Figure 8 As shown in the diagram. Other modifications (32-34) at the CF3 position on Ar1, while retaining X2 as a methylene ether, show similar active effects to 27-29.

[0135] After obtaining SAR data, we hypothesized that isosteric variations in the intermediate chain could further enhance antimicrobial activity. X1 and X2 were modified isosterically by maintaining Ar1 and Ar2 as ortho and para CF3, respectively. See Table 4 ( Figure 9 As shown, when X1 in compound 19 changes from N to O or S, the antimicrobial activity of 36 or 37 against both Gram-positive and Gram-negative bacteria is retained, except for the activity of 37 against Enterobacter cloacae. Similar to compound 30, compound 38, with X2 changing from O to S, demonstrated maximum activity against Gram-positive bacteria (particularly Streptococcus pneumoniae), with a MIC of 1 μg / mL. When both X1 and X2 are changed to S, 39 maintains its antimicrobial activity against Gram-positive bacteria compared to compound 37.

[0136] Antimicrobial activity against representative bacterial pathogens Following the initial evaluation of the antimicrobial activity of our compounds, we further screened the selected compounds against a panel of clinically relevant pathogens to confirm their clinical potential. Among the pathogens tested, *Streptococcus pneumoniae* showed particular sensitivity to our compound family, prompting us to expand our antimicrobial activity assay to include Group A and Group B *Streptococcus* spp.: *Streptococcus pyogenes* (Group A Streptococcus, GAS), known to cause septic pharyngitis, localized skin infections, and necrotizing fasciitis; and *Streptococcus agalactiae* (Group B Streptococcus: GBS), causing neonatal infections. Furthermore, we evaluated the effects of our compounds against clinically significant Gram-positive pathogens *Staphylococcus epidermidis* and *Staphylococcus saprophyticus*. To further confirm the antimicrobial activity of our compounds against *Streptococcus pneumoniae*, we included several clinical isolates of this bacterium, designated CUHK-X01-04. The identity of these isolates was determined using the Bruker MALDI Biotyper. ® Confirmed, and listed in the support information.

[0137] Table 5 ( Figure 10 The results in Table 1-4 confirm that our compounds exhibit good antimicrobial activity against clinically challenging pathogens, with MICs ranging from 2 μg / mL to 16 μg / mL. Notably, compound 38 demonstrated robust antimicrobial activity against Staphylococcus saprophyticus, with an MIC comparable to vancomycin (2 μg / mL). Furthermore, all other compounds showed antimicrobial activity against the listed pathogens, with MICs comparable to those in Table 1-4. Figure 6-9 The other Gram-positive bacteria shown in the figure are comparable.

[0138] We expanded our research to include a group of drug-resistant Staphylococcus aureus strains, including both methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Staphylococcus aureus (VRSA). In particular, MRSA strains pose a significant medical challenge due to their resistance to multiple antibiotics, including vancomycin, a widely used “last-line” antimicrobial agent in the clinical setting. To evaluate the efficacy of our compounds against MRSA, we tested their activity against a diverse set of clinically relevant strains representing both hospital-acquired (HA-MRSA) and community-acquired (CA-MRSA) strains, as well as VRSA isolates.

[0139] HA-MRSA strains include W-231 ST45, ATCC BAA-43, ATCC BAA-44, and ST239, while CA-MRSA strains consist of W-45 ST59, W-47 ST30, ST22, and USA300. Additionally, we evaluated our compounds against laboratory strains characterized by specific resistance mechanisms, including SA-APH2”-AAC6’, SA-APH3’, and SA-ANT4’, which exhibit aminoglycoside resistance due to aminoglycoside-modifying enzymes (AMEs). Similarly, we included a macrolide-resistant strain, SA-RN4220-pUL5054, and a fluoroquinolone-resistant strain, SA-1199B.

[0140] Our findings confirm that our compounds exhibit potent antimicrobial activity against a broad spectrum of MRSA and VRSA strains, achieving MIC values ​​comparable to or better than those of reference strains. Significantly, these compounds outperform commonly used antibiotics, including ciprofloxacin, gentamicin, and oxacillin, particularly against strains resistant to these agents. For example, most of the MRSA strains tested were resistant to oxacillin (a first-line treatment for MRSA infections in the United States) and gentamicin (a protein synthesis inhibitor). These results suggest that our compounds may act through a different mechanism of action, minimizing cross-resistance with existing antibiotics.

[0141] For the evaluation of VRSA, we used strains provided by the Staphylococcus aureus Antimicrobial Resistance Network (NARSA), allocated by BEI Resources, NIAID, NIH, and managed by ATCC (Manassas, VA, USA). The VRSA strains tested included NR-46410 (HIP11714, VRS1), NR-46413 (HIP13419, VRS3b), NR-46419 (AIS 080003, VRS9), and NR-46423 (1002434, VRS12). Figure 12 As shown, our compounds exhibit strong antimicrobial activity against these VRSA strains, with MIC values ​​as low as 4 µg / mL. In contrast, these strains show resistance or reduced sensitivity to common cell wall synthesis inhibitors, such as vancomycin and oxacillin.

[0142] Compounds effective against MRSA strains consistently exhibited antimicrobial activity against VRSA strains, with MIC values ​​ranging from 4 to 16 µg / mL. This broad efficacy against VRSA strains with multiple resistance mechanisms demonstrates the potential of our compounds as promising candidates for further development. These findings highlight their potential utility as novel therapeutic agents against serious drug-resistant Staphylococcus aureus infections, particularly those caused by VRSA.

[0143] bactericidal properties To determine whether compound 38 exhibits antibacterial or bactericidal activity, a minimum bactericidal concentration (MBC) assay was performed. Figure 13 An overview of compound 38 against Staphylococcus aureus ATCC ® The MIC and MBC values ​​of 25923 and CA-MRSA USA300 were determined. The results confirmed that compound 38 exerted a bactericidal effect against both strains, consistent with the bactericidal activity observed in the time-kinetic kinetic assay.

[0144] Time-sterilization kinetics Time-kinetic kinetic assays, performed according to guidelines established by CLSI, are used to evaluate the in vitro activity of antimicrobial agents against specific bacterial strains within defined time ranges. In this study, the activity against Staphylococcus aureus subtype ATCC was evaluated in liquid cultures. ® 25923 ( Figure 14 A) and community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) strain USA300 ( Figure 14 B) The time-kickdown kinetics of compound 38 were measured to simplify the experimental requirements for testing Streptococcus pneumoniae. Significantly, treatment with compound 38 at a concentration of 1× MIC confirmed growth inhibition after 2 hours, followed by progressively decreasing bacterial counts after extended treatments at 4 and 6 hours. Furthermore, when higher concentrations of compound 38 (4× and 16× MIC) were used in the experiments, both subtypes of Staphylococcus aureus were completely and effectively eradicated to below-theoretical detection levels (200 CFU / mL) after 6 hours. Specifically, compound 38 showed efficacy against Staphylococcus aureus ATCC. ® Bacterial counts of strain 25923 decreased to below the detectable threshold within 2 hours, while strain USA300 required 4 hours to achieve the same level of bacterial elimination. In summary, our findings indicate that compound 38 exhibits bacterial growth inhibition at a concentration of 1 × MIC, while higher concentrations effectively eliminate bacteria to below the detectable threshold.

[0145] NusG-β'CH PPI inhibition To confirm the disruptive effect of compound 38 on the interaction between β'CH and NusG from Bacillus subtilis strain 168, we used an internally developed competitive protein complementation assay to determine the 50% inhibitory concentration (IC50) of the compound. 50 In this assay, NusG and β'CH are fused with two complementary fragments of Nano-Luc luciferase. When a specific inhibitor targeting the NusG-β'CH PPI is introduced into the reaction, efficient reformation of the native luciferase complex is inhibited. Therefore, the catalytic reaction of the reconstituted luciferase is suppressed, resulting in changes in luminescence levels. These changes are used as a measure of the inhibitory activity exhibited by the compound tested. Figure 15 As shown, compound 38 demonstrated inhibitory activity against NusG-β'CH PPI, IC50. 50 The value was 167 ± 47.24 μM. This protein complementation assay confirmed that compound 38 disrupts the interaction between NusG and β'CH. Note that inhibitors targeting PPIs typically show high IC50 values ​​due to weak interactions. 50 The value indicates that the weak interaction is sufficient to block biological processes associated with a specific PPI.

[0146] Epifluorescence microscopy To confirm the role of compound 38 as a transcriptional inhibitor targeting NusG-β'CH interaction, a group of Bacillus subtilis reporter strains were used for bacterial cytological analysis. These strains express GFP-tagged proteins involved in key cellular processes, including transcription (transcription factors NusG and RNA polymerase β' subunit RpoC) and translation (ribosomal subunit RpsB). 7 Cells were treated with compound 38 at 1× MIC or a control antibiotic and then incubated for 30 minutes. To visualize nucleoids, 4',6-diamidinyl-2-phenylindole (DAPI) was added to a final concentration of 1 µg / mL, and epifluorescence microscopy was used to examine the cellular localization of the tagged proteins.

[0147] Analysis of the report strain revealed that compound 38 disrupts bacterial transcription, such as that of GFP-tagged NusG ( Figure 16 A) and RpoC ( Figure 16 The delocalization pattern in B) is similar to that observed with rifampin (a known RNA polymerase inhibitor that targets bacterial transcription).

[0148] To investigate the effect of compound 38 on membrane morphology, ATP synthase (AtpA-GFP) located in the membrane was used as a reporter. Neither rifampin nor compound 38 caused membrane invagination or delocalization of AtpA. Figure 17A). In contrast, colistin (a membrane-targeting antibiotic) causes AtpA-GFP to delocalize from the membrane, resulting in a loss of its fluorescence signal in the membrane region.

[0149] When examining the ribosomal protein RpsB-GFP, both compound 38 and rifampin resulted in similar protein delocalization effects. Figure 17 B). This observation is consistent with previous research showing that rifampicin treatment of Bacillus subtilis leads to the expansion of nucleoid bodies, producing diffuse structures that extend throughout the cell. In contrast, protein synthesis inhibitors, such as tetracycline and chloramphenicol, alter the localization of RpsB by shifting it from the cell poles to the mid-cell. Figure 17 B).

[0150] Interestingly, no significant nucleoid body expansion was observed in cells treated with compound 38. This is likely due to the short treatment duration, as transcriptional inhibitors are known to initially cause nucleoid body contraction followed by expansion later. However, the exact mechanism by which transcriptional inhibitors induce nucleoid body expansion remains unclear.

[0151] In summary, bacterial cytological analysis indicates that compound 38 primarily targets transcription by disrupting the interaction between NusG and RNA polymerase. The subcellular delocalization of NusG, RpoC, and RpsB without affecting ATP synthase localization or membrane morphology suggests that compound 38 exerts its role in transcription rather than causing membrane disruption.

[0152] Chemical Option 1 ( Figure 3 The general synthetic methods for synthesizing target compounds 1-39 are described, which are obtained through nucleophilic substitution reactions between specific amines A and epoxides B. Four series of amine substrates A were prepared by substitution reactions of different aromatic rings Ar1 and amine side chains (Scheme 2, ...). Figure 4 Commercially available chloroaromatic hydrocarbons react with ethylenediamine to form diamines A1-A15. For the synthesis of sulfonamide A16, nucleophilic acyl substitution is performed to form a sulfonamide structure. Ether A17 and thioether A18 are synthesized by nucleophilic substitution of phenol and thiophenol, respectively, followed by deprotection of the BOC group.

[0153] Epoxide substrate B is prepared via nucleophilic substitution reactions of substituted phenols, anilines, thiophenols, benzyl alcohols, and benzyl thiols with epichlorohydrins (Scheme 3). Figure 5 To synthesize amine B11, the weakly nucleophilic p-trifluoromethylaniline is preferably reacted with the epoxide group of epichlorohydrin. In the presence of a substoichiometric zinc salt, the intermediate 1-chloro-3-((4-(trifluoromethyl)phenyl)amino)prop-2-ol is first obtained. Subsequently, an intramolecular nucleophilic substitution reaction occurs to provide the desired compound.

[0154] in conclusion In previous studies, we employed structure-based drug design and successfully discovered two series of PPI inhibitors targeting RNAP-σ and NusB-NusE PPIs, optimizing their antimicrobial activity to levels comparable to commercially available antibiotics. In this study, we report the discovery and evaluation of novel analogs of inhibitors targeting bacterial RNAP and the transcription factor NusG. Using a rational design approach based on pharmacophore models, followed by analog synthesis, we successfully identified a series of compounds with inhibitory activity against the interaction between RNAP β'CH and NusG. These compounds, particularly compound 38, exhibited potent antimicrobial activity against both Gram-positive and Gram-negative bacteria. Notably, they demonstrated superior efficacy in inhibiting the growth of Streptococcus pneumoniae, with MICs as low as 1 μg / mL. Time-kinetic kinetics also revealed that compound 38 gradually eradicated bacteria at concentrations of 1 × MIC or higher, exhibiting bactericidal characteristics. Finally, protein-protein inhibition assays and fluorescence microscopy images confirmed the interaction of compound 38 with NusG and β'CH, confirming its mechanism of action. Based on these findings, we conclude that our transcriptional inhibitor effectively stops bacterial transcription by disrupting the interaction between β'CH and NusG, leading to inhibition of bacterial growth.

[0155] Experimental Section General Principles Unless otherwise specified, all chemicals and reagents used in the synthesis were commercially available and required no purification. All reactions were visualized by thin-layer chromatography (TLC) on glass slides (silica gel F) under UV light. 254 Monitoring was performed using a silica gel (200-300 mesh) column for rapid chromatographic purification. 1 H-NMR (400 MHz or 600 MHz), 13 C-NMR (100 MHz or 150 MHz) and 19 F-NMR (565 MHz) spectra were measured on a Bruker Advance-III spectrometer, with TMS as an internal standard. Chemical shifts were expressed as... δ (ppm) represents the coupling constant ( J(Indicated in Hz). High-resolution mass spectrometry (HR-MS) spectra were measured using an Agilent 6540 liquid chromatography-electrospray ionization (LC-EI) QTOF mass spectrometer. The purity of all product compounds for bioactivity testing was >95%, determined by analytical HPLC performed on a Waters HPLC system comprising a 2535 quaternary gradient module, a 2707 autosampler, a 2998 photodiode array (PDA) detector, and an XBridge C18 (4.6 × 100 mm, 5 mM particle size).

[0156] Option 2 ( Figure 4 Synthesis method of ) Used in the synthesis of compound A1-A 17 A general approach. The title compound was obtained using four different methods: Method A: N 1 -(pyrimidin-2-yl)ethane-1,2-diamine (A1) At room temperature, ethylenediamine monohydrate (177.6 mL, 2.18 mmol) was added to a THF solution of 2-chloropyrimidine (50 mg, 0.44 mmol) and K₂CO₃ (72.4 mg, 0.52 mmol). The mixture was refluxed and stirred overnight, and the reaction was monitored for completion. After cooling to room temperature and filtration, the filtrate was concentrated and purified by silica gel column chromatography with eluent DCM / MeOH (50:1–20:1, with a few additional drops of ammonia) to provide A1, a pale yellow oil, 50.1 mg, in 83.1% yield. 1 H NMR (600 MHz, DMSO- d 6) δ 8.24 (d, J = 4.8 Hz, 2H), 7.24 (t, J = 5.7 Hz, 1H), 6.53 (t, J = 4.8 Hz, 1H), 3.30 (q, J = 6.2 Hz, 2H), 2.72 (t, J = 6.5 Hz, 2H).

[0157] N 1 -(5-(trifluoromethyl)pyrimidin-2-yl)ethane-1,2-diamine (A2) The title compound was prepared from 2-chloro-5-(trifluoromethyl)pyrimidine (50 mg, 0.27 mmol) and ethylenediamine monohydrate (111.5 mL, 1.37 mmol) in a similar manner as described for compound A1. Eluent DCM / MeOH (50:1–20:1, with a few additional drops of ammonia) was used for chromatography. A pale yellow oil was obtained, 49.7 mg, in 87.9% yield. 1 H NMR (400 MHz, DMSO- d 6) δ 8.62–8.52 (m, 2H), 8.12 (t, J = 5.7 Hz, 1H), 3.37 (s, 2H), 2.74 (d, J = 12.9 Hz, 2H).

[0158] N 1 -(4-(trifluoromethyl)pyrimidin-2-yl)ethane-1,2-diamine (A3) The title compound was prepared from 2-chloro-4-(trifluoromethyl)pyrimidine (50 mg, 0.27 mmol) and ethylenediamine monohydrate (111.5 mL, 1.37 mmol) in a similar manner as described for compound A1. Eluent DCM / MeOH (50:1–20:1, with a few additional drops of ammonia) was used for chromatography. A pale yellow oil was obtained, 48.2 mg, in 85.4% yield. 1 H NMR (600 MHz, DMSO- d 6) δ 8.59 (d, J = 26.5 Hz, 1H), 7.87 (d, J = 29.7 Hz, 1H), 6.95 (d, J = 4.9 Hz, 1H), 3.58 (s, 4H).

[0159] N 1 -(6-(trifluoromethyl)pyridin-2-yl)ethane-1,2-diamine (A4) The title compound was prepared from 2-chloro-6-(trifluoromethyl)pyridine (50 mg, 0.28 mmol) and ethylenediamine monohydrate (112.1 mL, 1.38 mmol) in a similar manner as described for compound A1. Eluent DCM / MeOH (100:1–30:1, with a few additional drops of ammonia) was used for chromatography. A pale yellow oil was obtained, 48.1 mg, in 85.1% yield. 1 H NMR (600 MHz, chloroform-) d ) δ 7.52 (t, J = 7.9 Hz, 1H), 6.92 (d, J = 7.2 Hz, 1H), 6.57 (d, J = 8.5 Hz, 1H), 5.18 (s, 1H), 3.42 (q, J = 5.8 Hz, 2H), 2.97 (t, J = 5.9 Hz, 2H).

[0160] N 1 -(5-(trifluoromethyl)pyridin-2-yl)ethane-1,2-diamine (A5)The title compound was prepared from 2-chloro-5-(trifluoromethyl)pyridine (50 mg, 0.28 mmol) and ethylenediamine monohydrate (112.1 mL, 1.38 mmol) in a similar manner as described for compound A1. Eluent DCM / MeOH (100:1–30:1, with a few additional drops of ammonia) was used for chromatography. A pale yellow oil was obtained, 47.6 mg, in 84.2% yield. 1 H NMR (400 MHz, chloroform-) d ) δ 8.34 (d, J = 2.2 Hz, 1H), 7.58 (dd, J = 8.8, 2.4 Hz, 1H), 6.46 (d, J = 8.8 Hz, 1H), 5.38 (s, 1H), 3.45(q, J = 5.8 Hz, 2H), 2.99 (s, 2H).

[0161] N 1 -(4-(trifluoromethyl)pyridin-2-yl)ethane-1,2-diamine (A6) The title compound was prepared from 2-chloro-4-(trifluoromethyl)pyridine (50 mg, 0.28 mmol) and ethylenediamine monohydrate (112.1 mL, 1.38 mmol) in a similar manner as described for compound A1. Eluent DCM / MeOH (100:1–30:1, with a few additional drops of ammonia) was used for chromatography. A pale yellow oil was obtained, 48.4 mg, in 85.6% yield. 1 H NMR (600 MHz, chloroform-) d ) δ 8.22 (d, J = 5.3 Hz, 1H), 6.75 (dd, J = 5.3, 1.4 Hz, 1H), 6.61 (s, 1H), 5.23 (s, 1H), 3.43 (q, J = 5.8Hz, 2H), 2.99 (t, J = 5.9 Hz, 2H).

[0162] N 1 -(3-(trifluoromethyl)pyridin-2-yl)ethane-1,2-diamine (A7) The title compound was prepared from 2-chloro-3-(trifluoromethyl)pyridine (50 mg, 0.28 mmol) and ethylenediamine monohydrate (112.1 mL, 1.38 mmol) in a similar manner as described for compound A1. Eluent DCM / MeOH (100:1–30:1, with a few additional drops of ammonia) was used for chromatography. A pale yellow oil was obtained, 46.4 mg, in 82.2% yield.1 H NMR (600 MHz, chloroform-) d ) δ 8.28-8.24 (m, 1H), 7.66 (dd, J = 7.6, 1.7 Hz, 1H), 6.63 (dd, J = 7.5, 5.0 Hz, 1H), 5.38 (s, 1H), 3.58(q, J = 5.7 Hz, 2H), 2.98 (t, J = 6.0 Hz, 2H).

[0163] N 1 -(5-Fluoropyridin-2-yl)ethane-1,2-diamine (A8) The title compound was prepared from 2-chloro-5-fluoropyridine (50 mg, 0.38 mmol) and ethylenediamine monohydrate (154.7 mL, 1.9 mmol) in a similar manner as described for compound A1. Eluent DCM / MeOH (100:1–30:1, with a few additional drops of ammonia) was used for chromatography. A pale yellow oil was obtained, 46.1 mg, in 78.2% yield.

[0164] N 1 -(5-nitropyridin-2-yl)ethane-1,2-diamine (A9) The title compound was prepared from 2-chloro-5-nitropyridine (50 mg, 0.32 mmol) and ethylenediamine monohydrate (128.3 mL, 1.58 mmol) in a manner similar to that described for compound A1. Eluent DCM / MeOH (100:1–30:1, with a few additional drops of ammonia) was used for chromatography. Yellow oil, 49.2 mg, 85.6% yield. 1 H NMR (600 MHz, DMSO- d 6) δ 8.91 (d, J = 2.8 Hz, 1H), 8.15-8.05(m, 1H), 6.62 (d, J = 9.3 Hz, 1H), 3.50 (s, 2H), 2.85 (t, J = 6.2 Hz, 2H).

[0165] Method B: N 1 -(2-(trifluoromethyl)phenyl)ethane-1,2-diamine (A 10 )Ethylenediamine monohydrate (74.6 mL, 0.92 mmol) was added to a DMSO solution of 1-iodo-2-(trifluoromethyl)benzene (50 mg, 0.18 mmol), CuCl (3.6 mg, 0.038 mmol), and Cs₂CO₃ (119.8 mg, 0.37 mmol) at room temperature. The mixture was stirred at 120 °C for 8 hours, and the reaction was monitored for completion. After cooling to room temperature and filtration, water was added, and the filtrate was extracted three times with ethyl acetate and washed with saturated brine. The combined organic layers were dried over anhydrous Na₂SO₄, concentrated, and purified by silica gel column chromatography with eluent DCM / MeOH (200:1–100:1, with a few drops of additional ammonia) to provide A. 10 Light yellow oil, 26.4 mg, 70.3% yield. 1 H NMR (400 MHz, chloroform-) d ) δ 7.46 (d, J = 7.7 Hz, 1H), 7.38 (t, J = 7.9 Hz, 1H), 6.79-6.71 (m, 2H), 4.79 (s, 1H), 3.27 (q, J = 5.7 Hz, 2H), 3.02 (t, J = 5.9 Hz, 2H).

[0166] N 1 -([1,1'-biphenyl]-4-yl)ethane-1,2-diamine (A 11 ) For example, for compound A 10 The title compound was prepared from 4-iodo-1,1'-biphenyl (50 mg, 0.18 mmol) and ethylenediamine monohydrate (72.6 mL, 0.89 mmol) in a similar manner as described. Eluent DCM / MeOH (200:1–100:1, with a few additional drops of ammonia) was used for chromatography. A pale yellow oil was obtained, 28.5 mg, in 75.2% yield. 1 H NMR (400 MHz, chloroform-) d ) δ 8.26 (d, J = 1.6 Hz, 1H), 7.57-7.54 (m, 2H), 7.49-7.46 (m, 2H), 7.42 (t, J = 7.7 Hz, 2H), 7.31 (d, J = 1.3Hz, 2H), 6.75-6.70 (m, 2H), 3.62 (q, J = 6.0 Hz, 2H), 3.40 (t,J = 5.8 Hz, 2H).

[0167] N 1 -(4-(trifluoromethyl)phenyl)ethane-1,2-diamine (A 12 ) For example, for compound A 10 The title compound was prepared from 1-iodo-4-(trifluoromethyl)benzene (50 mg, 0.18 mmol) and ethylenediamine monohydrate (74.8 mL, 0.92 mmol) in a similar manner as described. Eluent DCM / MeOH (200:1–100:1, with a few additional drops of ammonia) was used for chromatography. A pale yellow oil was obtained, 27.1 mg, in 72.2% yield. 1 H NMR (400 MHz, chloroform-) d ) δ 8.27 (s, 1H), 7.43 (d, J = 8.4 Hz, 2H), 6.64 (d, J = 8.4 Hz, 2H), 5.89 (s, 1H), 4.50 (s, 1H), 3.61 (q, J = 6.0 Hz, 2H), 3.37 (q, J = 5.6 Hz, 2H).

[0168] N 1 -(3-(trifluoromethyl)phenyl)ethane-1,2-diamine (A 13 ) For example, for compound A 10 The title compound was prepared from 1-iodo-3-(trifluoromethyl)benzene (50 mg, 0.18 mmol) and ethylenediamine monohydrate (74.8 mL, 0.92 mmol) in a similar manner as described. Eluent DCM / MeOH (200:1–100:1, with a few additional drops of ammonia) was used for chromatography. A pale yellow oil was obtained, 28.1 mg, in 74.8% yield. 1 H NMR (400 MHz, chloroform-) d ) δ 7.26 (d, J = 7.9 Hz, 1H), 6.95 (d, J = 7.6 Hz, 1H), 6.84 (d, J = 2.4 Hz, 1H), 6.79 (dd, J = 8.2, 2.3Hz, 1H), 4.34 (s, 1H), 3.22 (t, J = 5.8 Hz, 2H), 3.00 (t, J = 5.8 Hz, 2H).

[0169] N 1 -(2-nitrophenyl)ethane-1,2-diamine (A 14 )For example, for compound A 10 The title compound was prepared from 1-iodo-2-nitrobenzene (50 mg, 0.2 mmol) and ethylenediamine monohydrate (79.6 mL, 1 mmol) in a similar manner as described. Eluent DCM / MeOH (200:1–100:1 with a few additional drops of ammonia) was used for chromatography. Yellow oil, 27.8 mg, 76.4% yield. 1 H NMR (400 MHz, chloroform-) d ) δ 8.27 (s, 1H), 8.18 (dt, J = 8.6, 2.8 Hz, 1H), 7.44 (m, J = 8.9, 5.3, 2.1 Hz, 1H), 6.88 (dd, J = 8.7, 2.6 Hz, 1H), 6.65 (m, J = 8.8, 7.3, 3.5 Hz, 1H), 3.44-3.36 (m, 2H), 3.07 (td, J = 6.1, 3.1 Hz, 2H).

[0170] 2-((2-aminoethyl)amino)benzonitrile (A) 15 ) For example, for compound A 10 The title compound was prepared from 2-iodobenzonitrile (50 mg, 0.22 mmol) and ethylenediamine monohydrate (86.6 mL, 1.1 mmol) in a similar manner as described. Eluent DCM / MeOH (200:1–100:1 with a few additional drops of ammonia) was used for chromatography. A pale yellow oil was obtained, 27.6 mg, in 78.5% yield. 1 H NMR (600 MHz, chloroform-) d ) δ 7.40 (td, J = 7.6, 1.5 Hz, 2H), 6.72-6.68 (m,2H), 3.40 (t, J = 5.8 Hz, 2H), 3.07 (t, J = 5.8 Hz, 2H).

[0171] Method C: N -(2-aminoethyl)-2-(trifluoromethyl)benzenesulfonamide (A) 16 )2-(trifluoromethyl)benzenesulfonyl chloride (50 mg, 0.2 mmol) was added dropwise to a DCM solution of ethylenediamine monohydrate (16.7 mL, 0.2 mmol) and triethylamine (61.4 mL, 0.61 mmol) in an ice bath. The mixture was heated to room temperature and stirred overnight. After the reaction was completed, the mixture was concentrated and purified by silica gel column chromatography with eluent DCM / MeOH (200:1–50:1, with a few additional drops of ammonia) to provide A. 16 Yellow oil, 50.5 mg, 92.1% yield. 1 1H NMR (400 MHz, M ethanol-) d 4) δ 8.23-8.17 (m, 1H), 7.98 (dd, J = 6.9, 2.2 Hz, 1H), 7.83 (m, J = 6.1, 5.3, 2.7 Hz, 2H), 3.05 (t, J = 6.2 Hz, 2H), 2.78 (t, J = 6.2 Hz, 2H).

[0172] Method D: 2-(2-(trifluoromethyl)phenoxy)ethyl-1-amine (A) 17 ) At room temperature, tert-butyl (2-bromoethyl)carbamate (82.9 mg, 0.37 mmol) was added to a DMF solution of 2-(trifluoromethyl)phenol (50 mg, 0.31 mmol) and K₂CO₃ (426.3 mg, 3.08 mmol). The mixture was stirred at 65 °C for 8 hours, and the reaction was monitored for completion. After cooling to room temperature and filtration, water was added, and the filtrate was extracted three times with ethyl acetate and washed with saturated brine. The combined organic layers were dried over anhydrous Na₂SO₄ and purified by silica gel column chromatography with hexane / EA (10:1–5:1) eluent to obtain tert-butyl (2-(2-(trifluoromethyl)phenoxy)ethyl)carbamate, a pale yellow solid, 83.6 mg, 88.8% yield. 1 1H NMR (400 MHz, M ethanol-) d 4) δ 7.55 (m, J = 14.0, 7.7, 1.7 Hz, 2H), 7.16 (d, J = 8.3 Hz, 1H), 7.06 (t, J =7.6 Hz, 1H), 4.13 (t, J = 5.9 Hz, 2H), 3.48 (m, J= 5.9, 4.6 Hz, 2H), 1.45(s, 9H).

[0173] TFA (110.1 mL, 1.64 mmol) was added to a DCM solution of (50 mg, 0.16 mmol) tert-butyl carbamate (2-(2-(trifluoromethyl)phenoxy)ethyl)carbamate at room temperature. The mixture was refluxed and stirred overnight, and the reaction was monitored for completion. After cooling to room temperature, an aqueous (1 M) solution of NaOH was added in an ice bath to adjust the pH to 8–10. The mixture was extracted three times with DCM and washed with saturated brine. The combined organic layers were dried over anhydrous Na₂SO₄ and concentrated to give title compound A. 17 Pale yellow oil, 32.1 mg, 95.6% yield. 1 H NMR (600 MHz, chloroform-) d ) δ 7.62 (dd, J = 7.8,1.5 Hz, 1H), 7.56-7.52 (m, 1H), 7.11 (t, J = 7.6 Hz, 1H), 7.01 (d, J = 8.3Hz, 1H), 4.22 (t, J = 5.0 Hz, 2H), 3.86 (q, J = 5.4 Hz, 2H).

[0174] 2-((2-(trifluoromethyl)phenyl)thio)ethyl-1-amine (A) 18 ) For example, for compound A 17 The title compound was prepared in a similar manner to that described, using (trifluoromethyl)thiophenol (50 mg, 0.28 mmol) and (2-bromoethyl)carbamate tert-butyl ester (75.5 mg, 0.34 mmol) to provide (2-((2-(trifluoromethyl)phenyl)thio)ethyl)carbamate tert-butyl ester. Hexane / EA (10:1–5:1) was used as the eluent for chromatography. A pale yellow solid, 58.8 mg, 65.2% yield. 1 1H NMR (400 MHz, M ethanol-) d 4) δ 7.67 (t, J = 7.8 Hz, 2H), 7.55 (t, J = 7.8 Hz, 1H), 7.33 (t, J =7.7 Hz, 1H), 4.85 (d, J= 1.9 Hz, 1H), 3.32-3.24 (m, 2H), 3.11 (dd, J = 8.3, 6.1 Hz, 2H), 1.44 (s, 9H). Compound A was prepared from tert-butyl (2-((2-(trifluoromethyl)phenyl)thio)ethyl)carbamate (50 mg, 0.15 mmol) by deprotection with trifluoroacetic acid (104.6 mL, 1.56 mmol). 18 Pale yellow oil, 33.1 mg, 96.3% yield. 1 H NMR (600 MHz, chloroform-) d ) δ 7.67-7.63 (m, 1H), 7.53 (d, J = 7.9 Hz, 1H), 7.47 (t, J = 7.7 Hz, 1H), 7.29 (d, J = 15.3 Hz, 1H), 3.07 (td, J = 6.3, 1.6 Hz, 2H), 2.92 (td, J = 6.3, 1.7 Hz, 2H).

[0175] Option 3 ( Figure 5 Synthesis method of ) Used in the synthesis of compound B1-B 17 A general approach. Four different methods are used to obtain the title compound: Method A: 2-((4-nitrophenoxy)methyl)ethylene oxide (B1) A DMF solution of 4-nitrophenol (50 mg, 0.36 mmol), Cs₂CO₃ (175.7 mg, 0.54 mmol), and KI (11.9 mg, 0.072 mmol) was stirred at room temperature for 30 minutes, followed by the addition of 2-(chloromethyl)ethylene oxide (84.5 mL, 1.08 mmol). The solution was stirred overnight at 80 °C, and the reaction was monitored for completion. After cooling to room temperature and filtration, water was added, and the filtrate was extracted three times with ethyl acetate and washed with saturated brine. The combined organic layers were dried over anhydrous Na₂SO₄ and purified by silica gel column chromatography with hexane / EA (200:1–50:1) eluent to give compound B1, a pale yellow oil, 57.9 mg, 82.6% yield. 1 H NMR (400 MHz, DMSO- d 6) δ 8.25-8.19 (m, 2H), 7.22-7.16 (m, 2H), 4.53 (dd, J= 11.5, 2.5 Hz, 1H), 4.00(dd, J = 11.5, 6.6 Hz, 1H), 3.39 (m, J = 6.8, 5.0, 2.6 Hz, 1H), 2.88 (t, J =4.6 Hz, 1H), 2.75 (dd, J = 5.1, 2.6 Hz, 1H).

[0176] 2-(phenoxymethyl)ethylene oxide (B2) The title compound was prepared from phenol (50 mg, 0.53 mmol) and 2-(chloromethyl)ethylene oxide (124.9 mL, 1.59 mmol) in a similar manner as described for compound B1. Hexane / EA (200:1–50:1) was used as the eluent for chromatography. A pale yellow oil was obtained, 67.2 mg, in 84.2% yield. 1 H NMR (400 MHz, chloroform-) d ) δ 7.36-7.30 (m, 2H), 7.04-6.94 (m, 3H), 4.25 (dd, J = 11.0, 3.1 Hz, 1H), 3.97 (dd, J = 11.0, 5.7 Hz, 1H), 3.38 (m, J = 5.7, 4.1, 2.8 Hz, 1H),2.92 (t, J = 4.5 Hz, 1H), 2.78 (dd, J = 5.0, 2.7 Hz, 1H).

[0177] Methyl 4-(ethylene oxide-2-ylmethoxy)benzoate (B3) The title compound was prepared from methyl 4-hydroxybenzoate (50 mg, 0.33 mmol) and 2-(chloromethyl)ethylene oxide (77.2 mL, 0.99 mmol) in a manner similar to that described for compound B1. Hexane / EA (200:1–50:1) was used as the eluent for chromatography. A pale yellow oil was obtained, 54 mg, in 78.9% yield. 1 HNMR (600 MHz, chloroform- d ) δ 8.00-7.98 (m, 2H), 6.95-6.92 (m, 2H), 4.30 (dd, J =11.0, 3.0 Hz, 1H), 3.98 (dd, J= 11.0, 5.9 Hz, 1H), 3.88 (s, 3H), 3.38 (m, J = 5.7, 4.0, 2.7 Hz, 1H), 2.93 (t, J = 4.5 Hz, 1H), 2.77 (dd, J = 4.9, 2.6 Hz, 1H).

[0178] 2-((4-fluorophenoxy)methyl)ethylene oxide (B4) The title compound was prepared from 4-fluorophenol (50 mg, 0.45 mmol) and 2-(chloromethyl)ethylene oxide (104.8 mL, 1.34 mmol) in a manner similar to that described for compound B1. Hexane / EA (200:1–50:1) was used as the eluent for chromatography. A pale yellow oil was obtained, 62.6 mg, in 83.5% yield. 1 H NMR (400MHz, chloroform-) d ) δ 7.56 (d, J = 8.3 Hz, 2H), 7.00 (d, J = 8.3 Hz, 2H), 4.32 (m, J = 11.1, 2.0 Hz, 1H), 3.98 (m, J = 11.1, 6.0, 1.9 Hz, 1H), 3.38 (m, J = 2.5Hz, 1H), 2.98-2.91 (m, 1H), 2.78 (m, J = 4.6, 2.1 Hz, 1H).

[0179] 2-((3,4-difluorophenoxy)methyl)ethylene oxide (B5) The title compound was prepared from 3,4-difluorophenol (50 mg, 0.38 mmol) and 2-(chloromethyl)ethylene oxide (90.3 mL, 1.15 mmol) in a manner similar to that described for compound B1. Hexane / EA (200:1–50:1) was used as the eluent for chromatography. A pale yellow oil was obtained, 58.6 mg, in 81.9% yield. 1 H NMR (600 MHz, chloroform-) d ) δ 7.07 (q, J = 9.3 Hz, 1H), 6.76 (m, J = 11.9, 6.5, 3.0 Hz,1H), 6.63 (m, J= 8.5, 3.2, 1.7 Hz, 1H), 4.23 (dd, J = 11.0, 2.8 Hz, 1H), 3.87 (dd, J = 11.0, 5.9 Hz, 1H), 3.35 (m, J = 6.0, 4.0, 2.8 Hz, 1H), 2.93 (t, J = 4.5 Hz, 1H), 2.76 (dd, J = 4.8, 2.6 Hz, 1H).

[0180] 2-((4-methoxyphenoxy)methyl)ethylene oxide (B6) The title compound was prepared from 4-methoxyphenol (50 mg, 0.4 mmol) and 2-(chloromethyl)ethylene oxide (94.6 mL, 1.21 mmol) in a manner similar to that described for compound B1. Hexane / EA (200:1–50:1) was used as the eluent for chromatography. Yellow oil, 55.3 mg, 76.2% yield. 1 H NMR (400 MHz, chloroform-) d ) δ 7.28 (t, J = 3.4 Hz, 2H), 6.92-6.87 (m, 2H), 4.23 (dd, J = 11.1, 3.1 Hz, 1H), 3.96 (dd, J = 11.1, 5.7 Hz, 1H), 3.39-3.33 (m, 1H), 2.93(t, J = 4.5 Hz, 1H), 2.77 (dd, J = 4.9, 2.7 Hz, 1H), 2.47 (s, 3H).

[0181] 2-((4-(trifluoromethyl)phenoxy)methyl)ethylene oxide (B7) The title compound was prepared from 4-(trifluoromethyl)phenol (50 mg, 0.31 mmol) and 2-(chloromethyl)ethylene oxide (72.5 mL, 0.93 mmol) in a manner similar to that described for compound B1. Hexane / EA (200:1–50:1) was used as the eluent for chromatography. A pale yellow oil was obtained, 57.4 mg, in 85.3% yield. 1 H NMR (400 MHz, chloroform-) d ) δ 7.60-7.49 (m, 2H), 7.04-6.93 (m, 2H), 4.30(m,J = 11.1, 2.2 Hz, 1H), 3.94 (m, J = 10.9, 5.9, 3.1 Hz, 1H), 3.35 (m, J =5.6, 4.1, 2.6 Hz, 1H), 2.90 (dd, J = 5.7, 3.3 Hz, 1H), 2.75 (m, J = 4.8, 2.5Hz, 1H).

[0182] 2-((3-(trifluoromethyl)phenoxy)methyl)ethylene oxide (B8) The title compound was prepared from 3-(trifluoromethyl)phenol (50 mg, 0.31 mmol) and 2-(chloromethyl)ethylene oxide (72.5 mL, 0.93 mmol) in a manner similar to that described for compound B1. Hexane / EA (200:1–50:1) was used as the eluent for chromatography. A pale yellow oil was obtained, 55 mg, in 81.8% yield. 1 H NMR (400 MHz, chloroform-) d ) δ 7.41 (t, J = 8.0 Hz, 1H), 7.25 (s, 1H), 7.17(d, J = 2.1 Hz, 1H), 7.12 (dd, J = 8.3, 2.5 Hz, 1H), 4.31 (dt, J = 10.9, 2.2Hz, 1H), 3.97 (m, J = 11.1, 5.9, 1.5 Hz, 1H), 3.39 (m, J = 8.7, 4.4, 2.1, 1.7Hz, 1H), 2.94 (td, J = 4.5, 1.4 Hz, 1H), 2.79 (dt, J = 4.4, 2.0 Hz, 1H).

[0183] 2-((2-(trifluoromethyl)phenoxy)methyl)ethylene oxide (B9) The title compound was prepared from 2-(trifluoromethyl)phenol (50 mg, 0.31 mmol) and 2-(chloromethyl)ethylene oxide (72.5 mL, 0.93 mmol) in a manner similar to that described for compound B1. Hexane / EA (200:1–50:1) was used as the eluent for chromatography. A pale yellow oil was obtained, 56.2 mg, in 83.5% yield. 1 H NMR (400 MHz, chloroform-)d ) δ 7.59 (dd, J = 7.6, 1.6 Hz, 1H), 7.50 (td, J =8.0, 1.6 Hz, 1H), 7.05 (t, J = 8.4 Hz, 2H), 4.35 (dd, J = 11.2, 2.9 Hz, 1H), 4.12 (dd, J = 11.2, 5.0 Hz, 1H), 3.39 (m, J = 5.0, 2.9 Hz, 1H), 2.92 (t, J =4.6 Hz, 1H), 2.85 (dd, J = 5.0, 2.6 Hz, 1H).

[0184] 4-(ethylene oxide-2-ylmethoxy)benzonitrile (B 10 ) The title compound was prepared from 4-hydroxybenzonitrile (50 mg, 0.42 mmol) and 2-(chloromethyl)ethylene oxide (98.6 mL, 1.26 mmol) in a similar manner as described for compound B1. Hexane / EA (200:1–50:1) was used as the eluent for chromatography. A pale yellow oil was obtained, 60.9 mg, in 82.8% yield. 1 H NMR (400 MHz, chloroform-) d ) δ 7.63-7.54 (m, 2H), 7.02-6.95 (m, 2H), 4.34 (dd, J = 11.1, 2.8 Hz, 1H), 3.96 (dd, J = 11.2, 5.9 Hz, 1H), 3.37 (m, J = 6.8, 5.5, 2.8 Hz,1H), 2.93 (t, J = 4.5 Hz, 1H), 2.77 (dd, J = 4.8, 2.6 Hz, 1H).

[0185] Method B: N -(ethylene oxide-2-ylmethyl)-4-(trifluoromethyl)aniline (B 11 )A solution of 4-(trifluoromethyl)aniline (50 mg, 0.31 mmol) and Zn(SO3CF3)2 (13.3 mg, 0.062 mmol) in CHCl3 was added dropwise to 2-(chloromethyl)ethylene oxide (24.3 mL, 0.31 mmol) in an ice bath. After heating to room temperature, the solution was stirred at 60 °C for 12 hours, and the reaction was monitored for completion. After cooling to room temperature and filtration, water was added, and the filtrate was extracted three times with ethyl acetate and washed with saturated brine. The combined organic layers were dried over anhydrous Na2SO4 and concentrated to obtain the crude product. A solution of 1-chloro-3-((4-(trifluoromethyl)phenyl)amino)prop-2-ol (67.3 mg, 0.27 mmol) and KI (8 mg, 0.05 mmol) in MeCN was stirred at 80 °C for 6 hours, and the reaction was monitored for completion. After cooling to room temperature and filtration, water was added and the filtrate was extracted three times with ethyl acetate and washed with saturated brine. The combined organic layers were dried over anhydrous Na₂SO₄ and purified by silica gel column chromatography with DCM / MeOH (200:1–100:1) as eluent to give compound B. 11 Yellow oil, 42.5 mg, 73.8% yield. 1 H NMR (400 MHz, chloroform-) d ) δ 7.44 (d, J = 8.5 Hz, 2H), 6.67 (d, J = 8.4 Hz, 2H), 4.23(s, 1H), 3.63 (m, J = 13.8, 5.8, 2.7 Hz, 1H), 3.35-3.18 (m, 2H), 2.86 (t, J =4.3 Hz, 1H), 2.71 (dd, J = 4.8, 2.6 Hz, 1H).

[0186] Method C: 2-(((4-(trifluoromethyl)phenyl)thio)methyl)ethylene oxide (B 12 ) 2-(chloromethyl)ethylene oxide (43.9 mL, 0.56 mmol) was added to a solution of 4-(trifluoromethyl)thiophenol (50 mg, 0.28 mmol) and K₂CO₃ (58.2 mg, 0.42 mmol) in acetonitrile at room temperature. The solution was stirred at 85 °C for 8 hours, and the reaction was monitored for completion. After cooling to room temperature and filtration, water was added, and the filtrate was extracted three times with ethyl acetate and washed with saturated brine. The combined organic layers were dried over anhydrous Na₂SO₄ and purified by silica gel column chromatography with hexane / EA (200:1–50:1) eluent to give compound B. 12Yellow oil, 53.8 mg, 81.8% yield. 1 H NMR (400 MHz, chloroform-) d ) δ 7.55 (d, J = 8.3 Hz, 2H), 7.48 (d, J = 8.3 Hz, 2H), 3.25-3.18 (m, 2H), 3.12 (dd, J = 15.5, 6.5 Hz,1H), 2.86-2.82 (m, 1H), 2.64 (dd, J = 4.9, 2.3 Hz, 1H).

[0187] Method D: 2-((benzyloxy)methyl)ethylene oxide (B 13 ) Benzyl alcohol (50 mg, 0.46 mmol) was added dropwise to a solution of 2-(chloromethyl)ethylene oxide (43.4 mL, 0.56 mmol) and TBAB (16.3 mg, 0.023 mmol) in 50% NaOH aqueous solution in an ice bath. The solution was heated to room temperature and stirred for 18 hours, and the reaction was monitored for completion. The reaction was quenched with ice water and extracted three times with ethyl acetate and washed with saturated brine. The combined organic layers were dried over anhydrous Na2SO4 and purified by silica gel column chromatography with hexane / EA eluent (200:1–100:1) to provide compound B. 13 Pale yellow oil, 68.1 mg, 89.7% yield.

[0188] 2-(((4-(trifluoromethyl)benzyl)oxy)methyl)ethylene oxide (B 14 ) For example, for compound B 13 The title compound was prepared in a similar manner to that described above from 2-(chloromethyl)ethylene oxide (26.7 mL, 0.34 mmol) and (4-(trifluoromethyl)phenyl)methanol (50 mg, 0.28 mmol). Hexane / EA (200:1–50:1) was used as the eluent for chromatography. A pale yellow oil was obtained, 59.5 mg, in 90.2% yield. 1 H NMR (400 MHz, chloroform-) d ) δ 7.56 (d, J = 8.5 Hz, 2H), 7.00 (d, J = 8.5 Hz, 2H), 4.31 (dd, J = 11.1, 2.8 Hz, 1H), 3.99-3.91 (m, 1H), 3.38 (m, J = 5.5, 3.9, 2.7 Hz, 1H), 2.93 (t,J = 4.5 Hz, 1H), 2.78 (dd, J = 4.9, 2.6 Hz, 1H).

[0189] 2-(((3-(trifluoromethyl)benzyl)oxy)methyl)ethylene oxide (B 15 ) For example, for compound B 13 The title compound was prepared in a similar manner to that described above from 2-(chloromethyl)ethylene oxide (26.7 mL, 0.34 mmol) and (3-(trifluoromethyl)phenyl)methanol (50 mg, 0.28 mmol). Hexane / EA (200:1–50:1) was used as the eluent for chromatography. A pale yellow oil was obtained, 57.7 mg, in 87.6% yield. 1 H NMR (400 MHz, chloroform-) d ) δ 7.56 (tt, J = 24.8, 5.9 Hz, 4H), 4.66(m, J = 12.3, 5.2 Hz, 2H), 3.86 (m, J = 11.5, 2.9 Hz, 1H), 3.47 (dt, J =11.4, 5.7 Hz, 1H), 3.23 (m, J = 6.1, 3.0 Hz, 1H), 2.84 (q, J = 5.1 Hz, 1H), 2.65 (dt, J = 5.1, 2.7 Hz, 1H).

[0190] 2-(((2-(trifluoromethyl)benzyl)oxy)methyl)ethylene oxide (B 16 ) For example, for compound B 13 The title compound was prepared from 2-(chloromethyl)ethylene oxide (26.7 mL, 0.34 mmol) and (2-(trifluoromethyl)phenyl)methanol (50 mg, 0.28 mmol) in a similar manner as described. Hexane / EA (200:1–50:1) was used as the eluent for chromatography. A pale yellow oil was obtained, 60.1 mg, in 91.2% yield. 1 H NMR (400 MHz, chloroform-) d ) δ 7.78–7.55 (m, 3H), 7.40 (t, J = 7.7Hz, 1H), 4.85-4.73 (m, 2H), 3.86 (dd, J = 11.3, 3.0 Hz, 1H), 3.52 (dd, J=11.4, 5.9 Hz, 1H), 3.25 (m, J = 5.9, 4.1, 2.8 Hz, 1H), 2.85 (t, J = 4.6 Hz, 1H), 2.68 (dd, J = 5.0, 2.7 Hz, 1H).

[0191] Method D: 2-(((4-(trifluoromethyl)benzyl)thio)methyl)ethylene oxide (B) 17 ) To a solution of 2-(chloromethyl)ethylene oxide (61.1 mL, 0.78 mmol) and KOH (43.8 mg, 0.78 mmol) in dioxane and water (1:1), (4-(trifluoromethyl)phenyl)methanethiol (50 mg, 0.26 mmol) was added dropwise in an ice bath. The solution was heated to room temperature and stirred for 8 hours, and the reaction was monitored for completion. The mixture was extracted three times with ethyl acetate and washed with saturated brine. The combined organic layers were dried over anhydrous Na₂SO₄ and purified by silica gel column chromatography with hexane / EA eluent (200:1–100:1) to provide compound B. 17 Yellow oil, 52.8 mg, 81.8% yield. 1 H NMR (400 MHz, chloroform-) d ) δ 7.60 (d, J = 8.0 Hz, 2H), 7.49 (d, J = 8.0 Hz, 2H), 3.93-3.82 (m, 2H), 3.14 (m, J = 6.3, 4.2, 2.7 Hz,1H), 2.79 (t, J = 4.4 Hz, 1H), 2.66-2.51 (m, 3H).

[0192] Synthesis method A general method for synthesizing compounds 1-39. To the selected compounds A1-A 17 (1 equivalent) and compound B1-B 17 A solution of (1 equivalent) EtOH in DIPEA (2 equivalents) was added. The solution was refluxed and stirred for 8 hours while monitoring for completion. After cooling to room temperature, the mixture was concentrated and purified by column chromatography to provide the title compound.

[0193] 1-(4-Nitrophenoxy)-3-((2-(pyrimidin-2-ylamino)ethyl)amino)prop-2-ol(1)The title compound was prepared from A1 (30 mg, 0.22 mmol) and B1 (42.4 mg, 0.22 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). Pale yellow solid, 52.9 mg, 73.2% yield, mp 144–146 °C. 1 H NMR (400 MHz, chloroform-) d ) d 8.27 (d, J = 4.8 Hz, 2H), 8.23-8.18 (m,2H), 7.00-6.95 (m, 2H), 6.55 (dd, J = 5.4, 4.3 Hz, 1H), 5.46 (s, 1H), 4.11-4.08 (m, 3H), 3.57 (q, J = 5.8 Hz, 2H), 2.97-2.91 (m, 3H), 2.86-2.80 (m, 1H); 13 C NMR (101 MHz, chloroform-) d ) d 163.6, 162.5, 158.1, 141.8, 125.9, 114.6, 110.8, 71.1, 68.2, 51.3, 49.1, 41.2. HRMS(ESI): For C 15 H 19 N5O4, [M + H] + Calculated value: 334.1515; Measured value: 334.1514. HPLC purity: 97.06%.

[0194] 1-(4-Nitrophenoxy)-3-((2-((5-(trifluoromethyl)pyrimidin-2-yl)amino)ethyl)amino)prop-2-ol (2) The title compound was prepared from A2 (30 mg, 0.15 mmol) and B1 (28.4 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). Yellow solid, 41.5 mg, 70.8% yield, mp 133–134 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 8.47 (s, 2H), 8.25-8.17 (m, 2H),7.01-6.95 (m, 2H), 6.01 (s, 1H), 4.14-4.07 (m, 3H), 3.62 (q,J = 5.8 Hz, 2H), 2.95 (m, J = 9.5, 3.8, 3.0 Hz, 3H), 2.84 (dd, J = 12.3, 7.3 Hz, 1H), 2.39 (s, 2H). 13 C NMR (101 MHz, chloroform-) d ) δ 163.5, 163.4, 155.8, 155.8, 155.8, 155.7, 141.8, 125.9, 125.3, 114.5, 114.2, 71.0, 68.3, 51.3, 48.6, 41.2. 19 F NMR (565MHz, DMSO- d 6) δ -59.26. HRMS(ESI): For C 16 H 18 F3N5O4, [M + H] + Calculated value: 402.1389; Measured value: 402.1382. HPLC purity: 97.06%.

[0195] 1-(4-Nitrophenoxy)-3-((2-((6-(trifluoromethyl)pyridin-2-yl)amino)ethyl)amino)prop-2-ol (3) The title compound was prepared from A4 (30 mg, 0.15 mmol) and B1 (28.5 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). Yellow solid, 38.9 mg, 66.2% yield, mp 110–111 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 8.18 (d, J = 8.9 Hz, 2H), 7.50(t, J = 7.9 Hz, 1H), 6.94 (dd, J = 15.7, 8.1 Hz, 3H), 6.54 (d, J = 8.5 Hz, 1H), 5.16 (t, J = 5.6 Hz, 1H), 4.09 (d, J = 7.0 Hz, 3H), 3.48 (q, J = 5.8 Hz, 2H), 2.93 (dd, J= 9.6, 4.4 Hz, 3H), 2.82 (dd, J = 12.3, 7.2 Hz, 1H). 13 C NMR (101 MHz, chloroform-) d ) δ 163.6, 158.6, 146.6, 146.3, 141.7, 138.1, 125.9, 123.0, 120.3, 114.5, 110.3, 109.1, 109.1, 71.1, 68.2, 51.3, 48.8, 41.5. 19 F NMR (565MHz, DMSO- d 6) δ -67.29. HRMS(ESI): For C 17 H 19 F3N4O4, [M + H] + Calculated value: 401.1437; Measured value: 401.1431. HPLC purity: 98.85%.

[0196] 1-(4-Nitrophenoxy)-3-((2-((5-(trifluoromethyl)pyridin-2-yl)amino)ethyl)amino)prop-2-ol (4) The title compound was prepared from A5 (30 mg, 0.15 mmol) and B1 (28.5 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). Yellow solid, 37.7 mg, 64.7% yield, mp 131–132 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 8.29 (s, 1H), 8.15 (d, J = 8.8Hz, 2H), 7.55 (d, J = 8.8 Hz, 1H), 6.93 (d, J = 8.8 Hz, 2H), 6.42 (d, J = 8.8Hz, 1H), 5.59 (t, J = 5.7 Hz, 1H), 4.19-4.04 (m, 3H), 3.49 (q, J = 5.3, 4.8Hz, 2H), 2.97-2.89 (m, 3H), 2.83 (dd, J = 12.3, 7.9 Hz, 1H). 13 C NMR (101 MHz, chloroform-)d ) δ 163.6, 160.3, 145.9, 145.8, 141.7, 134.4, 125.9, 123.2, 115.6, 115.3, 114.5, 106.6, 71.2, 68.3, 51.5, 48.6, 41.4. 19 F NMR (565 MHz, DMSO- d 6) δ-59.18. HRMS(ESI): For C 17 H 19 F3N4O4, [M + H] + Calculated value: 401.1437; Measured value: 401.1433. HPLC purity: 96.36%.

[0197] 1-(4-Nitrophenoxy)-3-((2-((4-(trifluoromethyl)pyridin-2-yl)amino)ethyl)amino)prop-2-ol (5) The title compound was prepared from A6 (30 mg, 0.15 mmol) and B1 (28.5 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). Yellow solid, 42.1 mg, 71.9% yield, mp 120–122 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 8.25–8.16 (m, 3H), 6.97 (dd, J =9.0, 1.5 Hz, 2H), 6.75 (d, J = 5.3 Hz, 1H), 6.58 (s, 1H), 5.15 (s, 1H), 4.14-4.07 (m, 3H), 3.49 (q, J = 5.7 Hz, 2H), 3.00-2.91 (m, 3H), 2.84 (dd, J =12.4, 7.1 Hz, 1H). 13 C NMR (101 MHz, chloroform-) d ) δ 163.5, 158.9, 149.4, 141.8, 139.8, 139.5, 125.9, 124.4, 121.6, 114.5, 108.2, 108.2, 71.0, 68.3, 51.4, 48.7, 41.7. 19 F NMR (565 MHz, DMSO-d 6) δ -64.04. HRMS(ESI): For C 17 H 19 F3N4O4, [M +H] + Calculated value: 401.1437; Measured value: 401.1431. HPLC purity: 98.44%.

[0198] 1-(4-Nitrophenoxy)-3-((2-((3-(trifluoromethyl)pyridin-2-yl)amino)ethyl)amino)prop-2-ol (6) The title compound was prepared from A7 (30 mg, 0.15 mmol) and B1 (28.5 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). Yellow solid, 40.1 mg, 68.5% yield, mp 115–117 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 8.28–8.15 (m, 3H), 7.65 (d, J =7.6 Hz, 1H), 7.00-6.93 (m, 2H), 6.62 (dd, J = 7.6, 5.0 Hz, 1H), 5.41 (d, J =5.4 Hz, 1H), 4.11 (q, J = 3.9 Hz, 3H), 3.63 (q, J = 5.6 Hz, 2H), 2.97 (m, J =12.4, 10.4, 4.8 Hz, 3H), 2.84 (dd, J = 12.3, 6.5 Hz, 1H), 2.57(s, 2H). 13 C NMR (101 MHz, chloroform-) d ) δ 163.6, 154.7, 151.6, 141.7, 135.2, 135.1, 135.1, 135.0, 125.9, 125.8, 123.1, 120.4, 114.5, 111.5, 108.8, 108.5, 108.2, 71.0, 68.3, 51.1, 48.6, 40.9. 19 F NMR (565 MHz, DMSO- d 6) δ -62.80. HRMS(ESI): For C 17 H19 F3N4O4,[M + H] + Calculated value: 401.1437; Measured value: 401.143. HPLC purity: 98.7%.

[0199] 1-((2-((5-fluoropyridin-2-yl)amino)ethyl)amino)-3-(4-nitrophenoxy)prop-2-ol(7) The title compound was prepared from A8 (30 mg, 0.19 mmol) and B1 (37.7 mg, 0.19 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). Yellow solid, 47.1 mg, 69.5% yield, mp 139–141 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 8.25-8.13 (m, 4H), 7.01-6.94 (m, 2H), 5.50 (t, J = 5.7 Hz, 1H), 4.09 (s, 3H), 3.53 (q, J = 5.8 Hz, 2H), 2.96-2.90(m, 3H), 2.82 (dd, J = 12.3, 6.8 Hz, 1H), 2.30 (s, 2H). 13 C NMR (101 MHz, chloroform-) d ) δ 163.6, 159.5, 153.4, 150.9, 145.7, 145.4, 141.8, 125.9, 114.5, 71.1, 68.2, 51.3, 48.9, 41.8. 19 F NMR (565 MHz, DMSO- d 6) δ -157.53. HRMS(ESI): For C 15 H 18 FN5O4, [M + H] + Calculated value: 352.1469; Measured value: 352.1478. HPLC purity: 95.72%.

[0200] 1-(4-Nitrophenoxy)-3-((2-((2-(trifluoromethyl)phenyl)amino)ethyl)amino)prop-2-ol(8) From A 10The title compound was prepared by silica gel column chromatography using B1 (30 mg, 0.15 mmol) and B1 (28.7 mg, 0.15 mmol) with eluent DCM / MeOH (200:1–100:1, with a few additional drops of ammonia). Yellow solid, 42.8 mg, 73.1% yield, mp 102–103 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 8.25–8.17 (m, 2H), 7.46 (d, J =7.6 Hz, 1H), 7.39 (t, J = 7.9 Hz, 1H), 7.01-6.95 (m, 2H), 6.76 (dd, J = 8.2,5.2 Hz, 2H), 4.91 (s, 1H), 4.13 (s, 3H), 3.30 (m, J = 6.8, 3.4 Hz, 2H), 3.08-2.94 (m, 3H), 2.89-2.82 (m, 1H). 13 C NMR (101 MHz, chloroform-) d ) δ 163.6, 145.7, 145.7, 141.8, 133.2, 129.3, 126.8, 126.7, 126.7, 126.6, 125.9, 123.9, 121.2, 116.1, 114.5, 114.0, 113.7, 113.4, 113.1, 111.9, 71.0, 68.5, 51.0, 48.0, 42.8. 19 F NMR (565 MHz, DMSO- d 6) δ -61.42. HRMS(ESI): For C 18 H 20 F3N3O4, [M + H] + Calculated value: 400.1484; Measured value: 400.1491. HPLC purity: 95.37%.

[0201] 1-Phenoxy-3-((2-((4-(trifluoromethyl)pyrimidin-2-yl)amino)ethyl)amino)prop-2-ol(9)The title compound was prepared from A3 (30 mg, 0.15 mmol) and B2 (21.9 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). Pale yellow solid, 38.6 mg, 74.5% yield, mp 113–115 °C. 1 H NMR (400 MHz, chloroform-) d ) d 8.44 (s, 1H), 7.27 (d, J = 7.6 Hz,2H), 6.99-6.84 (m, 3H), 6.77 (s, 1H), 6.53 (s, 1H), 4.15 (s, 1H), 3.98 (s,2H), 3.59 (s, 2H), 2.96-2.80 (m, 4H). 13 C NMR (101 MHz, chloroform-) d ) d 162.5, 160.4, 158.6, 156.9, 156.5, 156.2, 155.8, 129.5, 121.9, 121.1, 119.2, 116.4, 114.5, 105.3, 70.6, 68.7, 51.8, 48.6, 41.1. 19 F NMR (565 MHz, chloroform- d δ -70.83. HRMS(ESI): For C 16 H 19 F3N4O2, [M + H] + Calculated value: 357.1538; Measured value: 357.153. HPLC purity: 97.91%.

[0202] 4-(2-hydroxy-3-((2-((4-(trifluoromethyl)pyrimidin-2-yl)amino)ethyl)amino)propoxy)benzoic acid Methyl ester (10) The title compound was prepared from A3 (30 mg, 0.15 mmol) and B3 (30.3 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). Pale yellow solid, 41.4 mg, 68.3% yield, mp 124–125 °C. 1 H NMR (400 MHz, chloroform-) d ) d 8.40 (s, 1H), 7.91 (d, J = 8.5 Hz, 2H), 6.84 (d,J = 8.5 Hz, 2H), 6.74 (d, J = 4.9 Hz, 1H), 6.50 (t, J = 5.9 Hz, 1H), 4.12 (m, J = 9.3, 4.9 Hz, 1H), 4.02-3.95 (m, 2H), 3.84 (s,3H), 3.56 (q, J = 5.8 Hz, 4H), 2.92-2.83 (m, 3H), 2.79 (dd, J = 12.1, 8.1 Hz, 1H). 13 C NMR (101 MHz, chloroform-) d ) d 166.7, 162.5, 162.3, 160.3, 156.8, 156.5, 156.1, 131.5, 122.8, 121.8, 119.1, 114.0, 105.4, 105.3, 105.3, 70.7, 68.4, 51.8, 51.6, 48.6, 41.0. 19 F NMR (565 MHz, chloroform- d δ -70.83. HRMS(ESI): For C 18 H 21 F3N4O4, [M + H] + Calculated value: 415.1593; Measured value: 415.1586. HPLC purity: 98.5%.

[0203] 1-(4-fluorophenoxy)-3-((2-((4-(trifluoromethyl)pyrimidin-2-yl)amino)ethyl)amino)prop-2-ol (11) The title compound was prepared from A3 (30 mg, 0.15 mmol) and B4 (24.5 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). Pale yellow solid, 39.4 mg, 72.1% yield, mp 116–117 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 8.46 (s, 1H), 6.96 (t, J =8.4 Hz, 2H), 6.90-6.72 (m, 3H), 6.17 (s, 1H), 4.15-4.04 (m, 1H), 3.95 (d, J=5.2 Hz, 2H), 3.60 (q, J = 6.0 Hz, 2H), 3.10-2.52 (m, 6H). 13 C NMR (101 MHz, chloroform-) d ) δ 162.5, 160.4, 158.6, 157.0, 156.6, 156.2, 155.9, 154.7, 154.7, 121.9, 119.1, 116.4, 115.9, 115.7, 115.6, 115.5, 105.5, 105.5, 71.2, 68.6, 51.6, 48.6, 41.1. 19 F NMR (565 MHz, chloroform- d δ -70.82, -123.52. HRMS(ESI): For C 16 H 18 F4N4O2, [M + H] + Calculated value: 375.1444; Measured value: 375.1438. HPLC purity: 98.74%.

[0204] 1-(3,4-Difluorophenoxy)-3-((2-((4-(trifluoromethyl)pyrimidin-2-yl)amino)ethyl)amino)prop-2- Alcohol (12) The title compound was prepared from A3 (30 mg, 0.15 mmol) and B5 (27.1 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). Pale yellow solid, 42.2 mg, 73.3% yield, mp 109–111 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 8.46 (d, J = 4.9 Hz, 1H), 7.04 (q, J = 9.4 Hz, 1H), 6.81 (d, J = 4.9 Hz, 1H), 6.72 (m, J = 12.1, 6.6,3.0 Hz, 1H), 6.60 (dd, J = 8.9, 4.2 Hz, 1H), 6.18 (t, J = 5.8 Hz, 1H), 4.09(m, J = 9.1, 4.7 Hz, 1H), 3.93 (d, J= 5.1 Hz, 2H), 3.60 (q, J = 5.8 Hz, 2H), 2.95-2.70 (m, 6H). 13 C NMR (101 MHz, chloroform-) d ) δ 162.5, 160.4, 157.0, 156.6, 156.2, 154.9, 154.9, 154.9, 154.8, 151.7, 151.6, 149.2, 149.1, 146.4, 146.3, 144.0, 143.9, 124.6, 121.8, 119.1, 117.3, 117.3, 117.1, 117.1, 116.4, 109.8, 109.8, 109.8, 109.7, 105.6, 105.5, 105.5, 104.3, 104.1, 71.3, 68.4, 51.5, 48.6, 41.1. 19 F NMR (565 MHz, chloroform- d δ -70.84, -135.30 (d, J = 21.4 Hz), -147.81 (d, J = 21.5 Hz). HRMS(ESI): for C 16 H 17 F5N4O2, [M + H] + Calculated value: 393.135; Measured value: 393.1343. HPLC purity: 98.98%.

[0205] 1-(4-Methoxyphenoxy)-3-((2-((4-(trifluoromethyl)pyrimidin-2-yl)amino)ethyl)amino)prop-2- Alcohols (13) 45 The title compound was prepared from A3 (30 mg, 0.15 mmol) and B6 (26.2 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). Pale yellow solid, 40.1 mg, 71.3% yield, mp 116–117 °C. 1 H NMR (400 MHz, chloroform-) d ) d 8.52-8.41 (m, 1H), 6.82 (m, J = 11.8, 3.7 Hz, 5H), 6.14 (s, 1H), 4.09 (m, J = 9.0, 5.1 Hz, 1H), 3.94(d, J= 5.2 Hz, 2H), 3.77 (s, 3H), 3.59 (q, J = 5.8 Hz, 2H), 2.95-2.87 (m,3H), 2.81 (m, J = 12.1, 7.8 Hz, 1H). 13 C NMR (101 MHz, chloroform-) d ) d 162.5, 160.4, 156.6, 156.2, 154.1, 152.7, 121.9, 119.1, 115.5, 114.7, 105.5, 105.5, 71.3, 68.8, 55.7, 51.7, 48.6, 41.2. 19 F NMR (565 MHz, chloroform- d δ -70.83. HRMS(ESI): For C 17 H 21 F3N4O3, [M + H] + Calculated value: 387.1644; Measured value: 387.1644. HPLC purity: 99.02%.

[0206] 1-(4-(trifluoromethyl)phenoxy)-3-((2-((4-(trifluoromethyl)pyrimidin-2-yl)amino)ethyl)amino) Propyl-2-ol (14) The title compound was prepared from A3 (30 mg, 0.15 mmol) and B7 (31.8 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). Pale yellow solid, 41.5 mg, 67.2% yield, mp 105–107 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 8.46 (s, 1H), 7.53 (d, J = 8.6 Hz, 2H), 6.96 (d, J = 8.6 Hz, 2H), 6.81 (d, J = 4.9 Hz, 1H), 6.20(t, J = 5.7 Hz, 1H), 4.14 (m, J = 9.2, 5.0 Hz, 1H), 4.03 (d, J = 5.1 Hz, 2H), 3.61 (q, J= 5.7 Hz, 2H), 3.08-2.74 (m, 6H). 13 C NMR (101 MHz, chloroform-) d ) δ 162.5,161.0, 160.4, 156.6, 156.2, 127.0, 126.9, 126.9, 126.9, 125.7, 123.4, 123.1,123.0, 121.8, 119.1, 114.5, 105.6, 105.5, 70.6, 68.4, 51.5, 48.7, 41.1. 19 F NMR (565 MHz, chloroform-) d δ -61.54, -70.84. HRMS(ESI): For C 17 H 18 F6N4O2, [M + H] + Calculated value: 425.1412; Measured value: 425.1406. HPLC purity: 96.47%.

[0207] 1-(4-(trifluoromethyl)phenoxy)-3-((2-((5-(trifluoromethyl)pyridin-2-yl)amino)ethyl)amino) Propyl-2-ol (15) The title compound was prepared from A5 (30 mg, 0.15 mmol) and B7 (31.9 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). Pale yellow solid, 42.2 mg, 68.7% yield, mp 118–120 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 8.36–8.27 (m, 1H), 7.55 (t, J = 8.7 Hz, 3H), 6.96 (d, J = 8.5 Hz, 2H), 6.41 (d, J = 8.8 Hz, 1H), 5.28 (s, 1H), 4.10 (m, J = 10.0, 5.8, 2.8 Hz, 1H), 4.06-4.00 (m, 2H), 3.49(q, J = 5.7 Hz, 2H), 2.97-2.89 (m, 3H), 2.82 (dd, J = 12.2, 7.7 Hz, 1H). 13 CNMR (101 MHz, chloroform-d ) δ 161.0, 160.3, 146.1, 146.1, 146.0, 146.0, 134.4, 134.4, 134.3, 130.9, 127.0, 127.0, 126.9, 126.9, 125.9, 125.7, 123.5, 123.2, 123.0, 115.8, 115.4, 114.5, 106.6, 70.5, 68.5, 51.4, 48.6, 41.5. 19 F NMR (565MHz, DMSO- d 6) δ -59.20, -59.77. HRMS(ESI): For C 18 H 19 F6N3O2, [M + H] + Calculated value: 424.146; Measured value: 424.1454. HPLC purity: 99.4%.

[0208] 1-((2-([1,1'-biphenyl]-4-ylamino)ethyl)amino)-3-(4-(trifluoromethyl)phenoxy)prop-2-ol (16) From A 11 The title compound was prepared by silica gel column chromatography using B7 (30 mg, 0.14 mmol) and B7 (30.8 mg, 0.14 mmol) with eluent DCM / MeOH (200:1–100:1, with a few additional drops of ammonia). The product was a white solid, 42.2 mg, 70.6% yield, mp 149–151 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 7.56 (d, J = 8.1 Hz, 4H), 7.47 (d, J = 8.2 Hz, 2H), 7.43 (d, J = 7.6 Hz, 2H), 7.29 (d, J = 8.2 Hz, 1H), 6.99 (d, J = 8.4 Hz, 2H), 6.73 (d, J = 8.3 Hz, 2H), 4.16-4.03 (m, 4H), 3.33(q, J = 5.1 Hz, 2H), 3.01-2.91 (m, 3H), 2.85 (dd, J = 12.2, 7.5 Hz, 1H). 13CNMR (101 MHz, chloroform- d ) δ 161.0, 147.6, 141.1, 130.6, 128.7, 128.0, 127.0, 127.0, 127.0, 126.9, 126.3, 126.1, 125.7, 123.5, 123.1, 123.0, 114.5, 113.2, 70.6, 68.5, 51.5, 48.8, 43.7. 19 F NMR (565 MHz, DMSO- d 6) δ -59.72. HRMS(ESI): For C 24 H 25 F3N2O2, [M + H] + Calculated value: 431.1946; Measured value: 431.1939. HPLC purity: 99.02%.

[0209] 1-(4-(trifluoromethyl)phenoxy)-3-((2-((4-(trifluoromethyl)phenyl)amino)ethyl)amino)prop-2- Alcohols (17) From A 12 The title compound was prepared by silica gel column chromatography using B7 (32.1 mg, 0.15 mmol) and B8 (30 mg, 0.15 mmol) and purified by silica gel column chromatography with DCM / MeOH (200:1–100:1 eluent, with a few drops of ammonia) as the eluent. White solid, 42.2 mg, 68.8% yield, mp 108–109 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 7.54 (d, J = 8.5 Hz, 2H), 7.39 (d, J = 8.3 Hz, 2H), 6.95 (d, J = 8.4 Hz, 2H), 6.61 (d, J = 8.3 Hz, 2H), 4.47 (t, J = 5.5 Hz, 1H), 4.12 (m, J = 9.9, 7.8, 4.3 Hz, 1H), 4.06-3.98 (m,2H), 3.28 (q, J = 5.5 Hz, 2H), 2.97-2.87 (m, 3H), 2.82 (dd, J = 12.2, 7.8 Hz, 1H). 13 C NMR (101 MHz, chloroform-)d ) δ 160.9, 150.7, 128.4, 127.0, 127.0, 127.0, 126.9, 126.7, 126.6, 126.6, 126.6, 126.3, 125.7, 123.6, 123.5, 123.2, 123.0, 119.1, 118.7, 114.5, 111.9, 70.5, 68.5, 51.5, 48.4, 43.0. 19 F NMR (565 MHz, DMSO- d 6) δ -58.81, -59.75. HRMS(ESI): For C 19 H 20 F6N2O2, [M + H] + Calculated value: 423.1507; Measured value: 423.1503. HPLC purity: 97.3%.

[0210] 1-(4-(trifluoromethyl)phenoxy)-3-((2-((3-(trifluoromethyl)phenyl)amino)ethyl)amino)prop-2- Alcohols (18) From A 13 The title compound was prepared by silica gel column chromatography using B7 (32.1 mg, 0.15 mmol) and B7 (30 mg, 0.15 mmol) and purified by eluent DCM / MeOH (200:1–100:1 with a few drops of ammonia). White solid, 44.3 mg, 71.1% yield, mp 89–91 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 7.57 (d, J = 8.4 Hz, 2H), 7.29 (d, J = 4.4 Hz, 1H), 6.98 (t, J = 8.8 Hz, 3H), 6.88-6.74 (m, 2H), 4.29 (s, 1H), 4.17-4.02 (m, 3H), 3.30 (s, 2H), 3.06-2.81 (m, 4H). 13 C NMR (101 MHz, chloroform-) d ) δ161.0, 148.4, 131.8, 131.4, 131.1, 129.6, 128.4, 127.0, 127.0, 126.9, 126.9, 125.7, 123.5, 123.2, 123.0, 116.0, 114.5, 114.0, 113.9, 113.9, 113.9, 108.9, 108.9, 108.9, 70.5, 68.5, 51.4, 48.5, 43.3. 19 F NMR (565 MHz, DMSO- d 6) δ -59.74, -61.33. HRMS(ESI): For C 19 H 20 F6N2O2, [M + H] + Calculated value: 423.1507; Measured value: 423.1520. HPLC purity: 98.83%.

[0211] 1-(4-(trifluoromethyl)phenoxy)-3-((2-((2-(trifluoromethyl)phenyl)amino)ethyl)amino)prop-2- Alcohols (19) From A 10 The title compound was prepared by silica gel column chromatography using B7 (32.1 mg, 0.15 mmol) and B7 (30 mg, 0.15 mmol) and purified by eluent DCM / MeOH (200:1–100:1 with a few drops of ammonia). The product was a white solid, 41.3 mg, 66.1% yield, mp 65–66 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 7.56 (d, J = 8.5 Hz, 2H), 7.46 (d, J = 7.8 Hz, 1H), 7.39 (t, J = 7.8 Hz, 1H), 6.99 (d, J = 8.5 Hz, 2H), 6.76 (d, J = 7.8 Hz, 2H), 4.93 (d, J = 5.2 Hz, 1H), 4.15-4.05 (m, 3H), 3.30(q, J = 6.0 Hz, 2H), 3.07-2.93 (m, 3H), 2.85 (dd, J = 12.3, 6.6 Hz, 1H). 13 CNMR (101 MHz, chloroform- d) δ 161.0, 145.7, 133.1, 127.0, 126.9, 116.1, 114.5, 111.9, 70.5, 68.7, 51.2, 48.1, 42.9. 19 F NMR (565 MHz, DMSO- d 6) δ -59.75, -61.46. HRMS(ESI): For C 19 H 20 F6N2O2, [M + H] + Calculated value: 423.1507; Measured value: 423.1503. HPLC purity: 98.13%.

[0212] 1-((2-((2-nitrophenyl)amino)ethyl)amino)-3-(4-(trifluoromethyl)phenoxy)prop-2-ol (20) From A 14 The title compound was prepared by silica gel column chromatography using B7 (36.1 mg, 0.17 mmol) and B8 (30 mg, 0.17 mmol) and purified by eluent DCM / MeOH (200:1–100:1 with a few drops of ammonia). Yellow solid, 41.9 mg, 63.9% yield, mp 97–99 °C. 1 H NMR (600 MHz, chloroform-) d ) δ 8.40 (d, J = 5.1 Hz, 1H), 8.19 (dd, J = 8.6, 1.5 Hz, 1H), 7.55 (d, J = 8.4 Hz, 2H), 7.45 (m, J = 8.5, 7.0, 1.5Hz, 1H), 7.01 (d, J = 8.5 Hz, 2H), 6.86 (d, J = 8.6 Hz, 1H), 6.67 (m, J =8.4, 6.9, 1.1 Hz, 1H), 4.12 (m, J = 16.0, 8.2, 4.4 Hz, 3H), 3.44 (q, J = 5.9Hz, 2H), 3.13-2.97 (m, 3H), 2.88 (dd, J = 12.2, 6.2 Hz, 1H). 13 C NMR (101 MHz, chloroform-) d ) δ161.0, 145.4, 136.2, 132.1, 127.0, 127.0, 126.9, 126.9, 125.7, 123.4, 123.1, 123.0, 115.4, 114.5, 113.8, 70.4, 68.7, 51.1, 47.9, 42.4. 19 F NMR (565 MHz, DMSO- d 6) δ -59.74. HRMS(ESI): For C 18 H 20 F3N3O4, [M + H] + Calculated value: 400.1484; Measured value: 400.1490. HPLC purity: 99.68%.

[0213] 2-((2-((2-hydroxy-3-(4-(trifluoromethyl)phenoxy)propyl)amino)ethyl)amino)benzonitrile(21) From A 15 The title compound was prepared by silica gel column chromatography using B7 (30 mg, 0.19 mmol) and B7 (40.6 mg, 0.19 mmol) with eluent DCM / MeOH (200:1–100:1, with a few additional drops of ammonia). The product was a white solid, 47.4 mg, 66.4% yield, mp 157–158 °C. 1 H NMR (600 MHz, chloroform-) d ) δ 7.56 (d, J = 8.5 Hz, 2H), 7.40 (m, J = 7.5, 5.9, 1.8 Hz, 2H), 7.02 (d, J = 8.5 Hz, 2H), 6.73-6.67 (m, 2H), 5.13(d, J = 5.9 Hz, 1H), 4.12 (m, J = 13.1, 4.1 Hz, 3H), 3.33 (m, J = 5.6, 2.0Hz, 2H), 3.06-2.95 (m, 3H), 2.86 (dd, J = 12.2, 6.5 Hz, 1H). 13 C NMR (151 MHz, chloroform-) d ) δ161.0, 150.4, 134.3, 132.8, 127.0, 127.0, 126.9, 126.9, 125.3, 123.5, 123.4, 123.2, 118.0, 116.7, 114.6, 110.8, 96.0, 70.5, 68.8, 51.2, 48.1, 42.7. HRMS(ESI): For C 19 H 20 F3N3O2, [M + H] + Calculated value: 380.1586; Measured value: 380.1593. HPLC purity: 98.38%.

[0214] 1-(3-(trifluoromethyl)phenoxy)-3-((2-((2-(trifluoromethyl)phenyl)amino)ethyl)amino)prop-2- Alcohols (22) From A 10 The title compound was prepared by silica gel column chromatography using B8 (32.1 mg, 0.15 mmol) and B8 (30 mg, 0.15 mmol) and purified by eluent DCM / MeOH (200:1–100:1 with a few drops of ammonia). White solid, 42.4 mg, 68.6% yield, mp 119–121 °C. 1 H NMR (600 MHz, chloroform-) d ) δ 7.49–7.44 (m, 1H), 7.40 (m, J = 10.6, 7.9 Hz, 2H), 7.27 (d, J = 20.1 Hz, 1H), 7.17 (t, J = 2.0 Hz, 1H), 7.10 (dd, J = 8.3, 2.5 Hz, 1H), 6.78-6.73 (m, 2H), 4.93 (s, 1H), 4.15-4.05(m, 3H), 3.31 (m, J = 5.8, 5.1 Hz, 2H), 3.06-2.94 (m, 3H), 2.85 (dd, J =12.2, 6.9 Hz, 1H). 13 C NMR (101 MHz, chloroform-) d ) δ158.7, 145.7, 133.1, 132.1, 131.7, 130.0, 127.9, 126.8, 126.7, 126.6, 126.6, 125.2, 123.9, 122.5, 121.2, 117.9, 117.8, 117.8, 117.7, 116.0, 114.0, 113.7, 113.5, 111.9, 111.4, 111.4, 111.4, 111.3, 70.6, 68.7, 51.2, 48.1, 42.9. 19 F NMR (565 MHz, DMSO- d 6) δ -61.18, -61.52. HRMS(ESI): For C 19 H 20 F6N3O2, [M + H] + Calculated value: 423.1507; Measured value: 423.1518. HPLC purity: 96.63%.

[0215] 1-(2-(trifluoromethyl)phenoxy)-3-((2-((2-(trifluoromethyl)phenyl)amino)ethyl)amino)prop-2- Alcohols (23) From A 10 The title compound was prepared by silica gel column chromatography using B9 (32.1 mg, 0.15 mmol) and B9 (30 mg, 0.15 mmol) and purified by eluent DCM / MeOH (200:1–100:1 with a few additional drops of ammonia). The product was a white solid, 40.6 mg, 65.2% yield, mp 78–80 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 7.59 (d, J = 7.7 Hz, 1H), 7.51 (t, J = 8.0 Hz, 1H), 7.45 (d, J = 7.8 Hz, 1H), 7.38 (t, J = 7.9 Hz, 1H),7.08-7.00 (m, 2H), 6.74 (t, J = 8.3 Hz, 2H), 4.96 (s, 1H), 4.14 (d, J = 2.5Hz, 3H), 3.29 (q, J = 5.3 Hz, 2H), 3.04-2.94 (m, 3H), 2.90 (dd, J = 10.9, 4.1Hz, 1H).13 C NMR (101 MHz, chloroform-) d ) δ 156.4, 145.8, 133.4, 133.1, 129.3, 127.1, 127.1, 127.0, 127.0, 126.7, 126.6, 126.6, 126.5, 125.1, 123.9, 122.4, 120.5, 118.9, 118.6, 115.9, 113.7, 113.4, 113.1, 112.9, 111.9, 71.0, 68.6, 51.0, 48.1, 42.8. 19 F NMR (565 MHz, DMSO- d 6) δ -60.89, -61.55. HRMS(ESI): For C 19 H 20 F6N3O2, [M + H] + Calculated value: 423.1507; Measured value: 423.1509. HPLC purity: 96.79%.

[0216] 4-(2-hydroxy-3-((2-((2-(trifluoromethyl)phenyl)amino)ethyl)amino)propoxy)benzonitrile (24) From A 10 (30 mg, 0.15 mmol) and B 10 The title compound was prepared (25.7 mg, 0.15 mmol) by silica gel column chromatography with DCM / MeOH (200:1–100:1 eluent, with a few additional drops of ammonia) as the eluent. White solid, 42.1 mg, 75.4% yield, mp 128–130 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 7.63–7.57 (m, 2H), 7.46 (d, J =7.8 Hz, 1H), 7.39 (t, J = 7.9 Hz, 1H), 7.00-6.95 (m, 2H), 6.79-6.72 (m, 2H), 4.91 (s, 1H), 4.15-4.06 (m, 3H), 3.34-3.25 (m, 2H), 3.07-2.93 (m, 3H), 2.84(dd, J = 12.3, 6.3 Hz, 1H). 13 C NMR (101 MHz, chloroform-) d ) δ161.8, 145.7, 145.7, 134.0, 133.2, 126.8, 126.7, 126.7, 126.6, 123.9, 119.1, 116.1, 115.2, 113.7, 113.4, 111.9, 104.4, 70.6, 68.5, 51.1, 48.1, 42.9. 19 F NMR (565 MHz, DMSO- d 6) δ-61.43. HRMS(ESI): For C 19 H 20 F3N3O2, [M + H] + Calculated value: 380.1586; Measured value: 380.1590. HPLC purity: 96.25%.

[0217] 1-((4-(trifluoromethyl)phenyl)amino)-3-((2-((5-(trifluoromethyl)pyridin-2-yl)amino)ethyl)amino (25)propane-2-ol From A5 (30 mg, 0.15 mmol) and B 11 The title compound was prepared (31.8 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). A pale yellow solid, 36.2 mg, 58.8% yield, mp 130–132 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 8.32 (s, 1H), 7.59 (d, J = 8.8 Hz, 1H), 7.40 (d, J = 8.3 Hz, 2H), 6.62 (d, J = 8.3 Hz, 2H), 6.42 (d, J = 8.9 Hz, 1H), 5.40 (t, J = 5.7 Hz, 1H), 4.58 (s, 1H), 3.94 (m, J = 8.0, 3.8 Hz, 1H), 3.48 (q, J = 5.8 Hz, 2H), 3.30 (dd, J = 12.9, 3.7 Hz, 1H), 3.13 (dd, J = 12.8, 7.1 Hz, 1H), 2.99-2.39 (m, 6H). 13 C NMR (101 MHz, chloroform-)d ) δ 160.3, 150.7, 146.0, 146.0, 145.9, 134.5, 134.4, 128.9, 126.7, 126.6, 126.6, 126.6, 126.2, 125.9, 123.6, 123.2, 119.3, 119.0, 118.6, 115.9, 115.5, 115.2, 112.1, 106.5, 68.3, 52.8, 52.8, 48.7, 47.3, 47.3, 41.6. 19 F NMR (565MHz, chloroform-) d ) δ -61.05, -61.18. HRMS(ESI): For C 18 H 21 F6N4O, [M + H] + Calculated value: 423.1620; Measured value: 423.1614. HPLC purity: 98.89%.

[0218] 1-(benzyloxy)-3-((2-((5-(trifluoromethyl)pyridin-2-yl)amino)ethyl)amino)prop-2-ol (26) From A5 (30 mg, 0.15 mmol) and B 13 The title compound was prepared (24 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). A pale yellow solid, 39.9 mg, 73.3% yield, mp 64–69 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 8.33 (d, J = 2.1 Hz, 1H), 7.57 (dd, J = 8.9, 2.4 Hz, 1H), 7.41-7.29 (m, 5H), 6.42 (d, J = 8.8 Hz, 1H), 5.42 (s,1H), 4.57 (s, 2H), 3.94 (m, J = 7.6, 4.1 Hz, 1H), 3.57-3.44 (m, 4H), 2.91 (t, J = 5.8 Hz, 2H), 2.75 (m, J = 12.2, 5.8 Hz, 2H). 13 C NMR (101 MHz, chloroform-) d ) δ160.3, 146.0, 146.0, 145.9, 137.8, 134.3, 134.2, 134.2, 134.2, 128.5, 127.9, 127.8, 125.9, 123.3, 115.6, 115.3, 106.8, 73.5, 72.7, 69.0, 51.7, 48.5, 41.3. HRMS(ESI): For C 18 H 22 F3N3O2, [M + H] + Calculated value: 370.1742; Measured value: 370.1734. HPLC purity: 95.09%.

[0219] 1-((4-(trifluoromethyl)benzyl)oxy)-3-((2-((5-(trifluoromethyl)pyridin-2-yl)amino)ethyl)amino (27) Propyl-2-ol From A5 (30 mg, 0.15 mmol) and B 14 The title compound was prepared (33.9 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). A pale yellow solid, 37.1 mg, 58.7% yield, mp 90–91 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 8.32 (d, J = 2.3 Hz,1H), 7.65-7.53 (m, 3H), 7.44 (d, J = 8.0 Hz, 2H), 6.41 (d, J = 8.8 Hz, 1H), 5.50 (t, J = 5.5 Hz, 1H), 4.61 (s, 2H), 3.95 (m, J = 7.8, 6.1, 3.9 Hz, 1H),3.58-3.49 (m, 2H), 3.45 (q, J = 5.7 Hz, 2H), 2.90 (t, J = 5.8 Hz, 2H), 2.80-2.69 (m, 2H). 13 C NMR (101 MHz, chloroform-) d ) δ160.3, 146.0, 145.9, 142.0, 134.3, 134.3, 130.1, 129.8, 127.6, 125.9, 125.4, 125.4, 125.4, 125.3, 123.2, 122.7, 115.6, 115.2, 106.6, 73.2, 72.6, 69.1, 51.7, 48.5, 41.4. 19 F NMR (565 MHz, chloroform- d ) δ -61.16, -62.53. HRMS(ESI): For C 19 H 21 F6N3O2, [M + H] + Calculated value: 438.1616; Measured value: 438.1612. HPLC purity: 97.4%.

[0220] 1-((3-(trifluoromethyl)benzyl)oxy)-3-((2-((5-(trifluoromethyl)pyridin-2-yl)amino)ethyl)amino (28)propane-2-ol From A5 (30 mg, 0.15 mmol) and B 15 The title compound was prepared (33.9 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). Pale yellow solid, 33.8 mg, 53.5% yield, mp 66–67 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 8.33 (d, J = 2.3 Hz,1H), 7.61-7.46 (m, 5H), 6.42 (d, J = 8.8 Hz, 1H), 5.36 (s, 1H), 4.62 (s, 2H), 3.95 (m, J = 7.9, 6.3, 3.9 Hz, 1H), 3.59-3.52 (m, 2H), 3.47 (q, J = 5.7 Hz, 2H), 2.92 (t, J = 5.8 Hz, 2H), 2.83-2.69 (m, 2H). 13 C NMR (101 MHz, chloroform-) d ) δ160.3, 139.0, 130.8, 128.9, 124.6, 124.2, 115.3, 73.1, 72.7, 69.1, 51.7, 48.5, 41.4. HRMS(ESI): For C 19 H 21 F6N3O2, [M + H] + Calculated value: 438.1616; Measured value: 438.1629. HPLC purity: 95.18%.

[0221] 1-((2-(trifluoromethyl)benzyl)oxy)-3-((2-((5-(trifluoromethyl)pyridin-2-yl)amino)ethyl)amino (29) Propyl-2-ol From A5 (30 mg, 0.15 mmol) and B 16 The title compound was prepared (33.9 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). Pale yellow solid, 34.2 mg, 54.1% yield, mp 71–73 °C. 1 H NMR (600 MHz, chloroform-) d ) δ 8.33-8.30 (m, 1H), 7.66 (t, J = 7.6 Hz, 2H), 7.58-7.53 (m, 2H), 7.40 (t, J = 7.7 Hz, 1H), 6.42(d, J = 8.8 Hz, 1H), 5.49 (s, 1H), 4.74 (s, 2H), 3.97 (m, J = 10.1, 7.5, 3.2Hz, 1H), 3.59 (dd, J = 9.6, 3.9 Hz, 1H), 3.55 (dd, J = 9.6, 6.3 Hz, 1H),3.48-3.44 (m, 2H), 2.91 (t, J = 5.8 Hz, 2H), 2.79 (dd, J = 12.2, 3.7 Hz, 1H), 2.74 (dd, J = 12.2, 8.0 Hz, 1H). 13 C NMR (151 MHz, chloroform-) d ) δ160.3, 146.0, 146.0, 146.0, 145.9, 136.5, 134.3, 134.2, 134.2, 134.2, 132.0, 129.1, 128.0, 127.8, 127.6, 127.4, 127.3, 127.0, 125.9, 125.9, 125.9, 125.8, 125.5, 125.2, 123.7, 123.4, 121.9, 121.6, 115.7, 115.5, 115.2, 115.0, 73.4, 69.5, 69.1, 51.7, 48.5, 41.4. 19 F NMR (565 MHz, chloroform-d) δ -60.07, -62.64. HRMS (ESI): for C 19 H 21 F6N3O2, [M + H] + Calculated value: 438.1616; Measured value: 438.1628. HPLC purity: 95.9%.

[0222] 1-((4-(trifluoromethyl)benzyl)thio)-3-((2-((5-(trifluoromethyl)pyridin-2-yl)amino)ethyl)amino propyl-2-ol (30) From A5 (30 mg, 0.15 mmol) and B 17 The title compound was prepared (36.3 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). Yellow solid, 43.7 mg, 65.9% yield, mp 64–66 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 8.38–8.28 (m, 1H), 7.58 (dd, J = 8.6, 3.2 Hz, 3H), 7.45 (d, J = 8.0 Hz, 2H), 6.43 (d, J = 8.8Hz, 1H), 5.38 (s, 1H), 3.81 (s, 3H), 3.46 (q, J = 5.7 Hz, 2H), 2.89 (m, J =5.7, 1.4 Hz, 2H), 2.78 (dd, J = 12.1, 3.3 Hz, 1H), 2.65-2.51 (m, 3H). 13 C NMR (101 MHz, chloroform-)d ) δ 160.3, 146.1, 146.0, 146.0, 146.0, 142.3, 134.4, 134.3, 134.3, 134.3, 129.6, 129.3, 129.2, 125.9, 125.6, 125.5, 125.5, 125.5, 125.4, 123.2, 122.7, 115.7, 115.3, 106.6, 68.7, 53.9, 48.5, 41.5, 36.3, 36.2. 19 F NMR (565 MHz, chloroform-d) δ -61.16, -62.48. HRMS (ESI): for C 19 H 21 F6N3OS, [M + H] + Calculated value: 454.1388; Measured value: 454.1385. HPLC purity: 97.15%.

[0223] 1-((2-((5-nitropyridin-2-yl)amino)ethyl)amino)-3-((4-(trifluoromethyl)benzyl)oxy)prop- 2-Alcohol(31) From A9 (30 mg, 0.16 mmol) and B 14 The title compound was prepared (38.2 mg, 0.16 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (100:1–50:1, with a few additional drops of ammonia). Yellow solid, 43.1 mg, 69.7% yield, mp 80–81 °C. 1 H NMR (600 MHz, chloroform-) d ) δ 9.02 (d, J = 2.7 Hz, 1H), 8.18 (d, J = 9.1 Hz, 1H), 7.63 (d, J = 8.0 Hz, 2H), 7.46 (d, J = 7.9 Hz, 2H), 6.38 (d, J = 9.2 Hz, 1H), 5.95 (s, 1H), 4.63 (s, 2H), 3.96 (m, J = 10.4, 3.8Hz, 1H), 3.59-3.50 (m, 4H), 2.95 (t, J = 5.8 Hz, 2H), 2.81 (dd, J = 12.2, 3.7Hz, 1H), 2.74 (dd,J = 12.2, 7.9 Hz, 1H). 13 C NMR (151 MHz, chloroform-) d ) δ 161.1,147.0, 141.9, 135.8, 130.2, 129.9, 127.6, 125.5, 125.4, 125.4, 125.0,123.2, 121.4, 73.1, 72.7, 69.2, 51.6, 48.2, 41.5. 19 F NMR (565 MHz, chloroform- d ) δ -62.52. HRMS(ESI): For C 18 H 21 F3N4O4, [M + H] + Calculated value: 415.1593; Measured value: 415.1601. HPLC purity: 98.35%.

[0224] 1-((4-(trifluoromethyl)benzyl)oxy)-3-((2-((2-(trifluoromethyl)phenyl)amino)ethyl)amino) Propyl-2-ol (32) From A 10 (30 mg, 0.15 mmol) and B 14 The title compound was prepared by silica gel column chromatography (34.1 mg, 0.15 mmol) with DCM / MeOH (200:1–100:1 eluent, with a few drops of ammonia) as the eluent. White solid, 39.9 mg, 62.2% yield, mp 67–69 °C. 1 H NMR (600 MHz, chloroform-) d ) δ 7.61 (d, J = 8.0 Hz, 2H), 7.46 (d, J = 7.9 Hz, 3H), 7.41-7.36 (m, 1H), 6.75 (d, J = 7.9 Hz, 2H), 4.94(d, J = 5.4 Hz, 1H), 4.62 (s, 2H), 3.94 (m, J = 6.8, 4.0 Hz, 1H), 3.56 (m, J = 9.7, 5.1 Hz, 2H), 3.27 (q, J = 5.6 Hz, 2H), 3.00-2.92 (m, 2H), 2.81 (m, J=11.9, 4.1, 1.5 Hz, 1H), 2.73 (m, J = 12.1, 7.3, 1.1 Hz, 1H). 13 C NMR (151 MHz, chloroform-) d ) δ 145.8, 142.1, 133.1, 120.0, 129.8, 127.6, 126.7, 126.7, 126.6, 126.6, 126.1, 125.4, 125.4, 125.4, 125.3, 125.0, 124.3, 123.2, 115.9, 113.6, 113.4, 111.9, 73.1, 72.6, 69.3, 51.4, 48.0, 42.9. 19 F NMR (565 MHz, chloroform- d ) δ -62.51, -62.58. HRMS(ESI): For C 20 H 22 F6N2O2, [M + H] + Calculated value: 437.1664; Measured value: 437.1669. HPLC purity: 95.05%.

[0225] 1-((3-(trifluoromethyl)benzyl)oxy)-3-((2-((2-(trifluoromethyl)phenyl)amino)ethyl)amino) Propyl-2-ol (33) From A 10 (30 mg, 0.15 mmol) and B 15 The title compound was prepared by silica gel column chromatography (34.1 mg, 0.15 mmol) with DCM / MeOH (200:1–100:1 eluent, with a few drops of ammonia) as the eluent. White solid, 38.1 mg, 59.4% yield, mp 77–79 °C. 1 H NMR (600 MHz, chloroform-) d ) δ 7.62 (s, 1H), 7.57 (d, J =7.8 Hz, 1H), 7.53 (d, J = 7.7 Hz, 1H), 7.49-7.44 (m, 2H), 7.40-7.36 (m, 1H), 6.74 (t, J = 8.1 Hz, 2H), 4.94 (t, J = 5.1 Hz, 1H), 4.61 (s, 2H), 3.94 (m, J= 7.9, 6.5, 4.0 Hz, 1H), 3.57 (m, J = 9.7, 5.1 Hz, 2H), 3.26 (q, J = 5.7 Hz, 2H), 2.96 (m, J = 5.9 Hz, 2H), 2.81 (dd, J = 12.1, 4.1 Hz, 1H), 2.72 (dd, J =12.1, 7.3 Hz, 1H). 13 C NMR (101 MHz, chloroform-) d ) δ 145.8, 145.8, 145.8, 139.1,133.1, 130.8, 130.6, 128.9, 128.8, 126.7, 126.6, 126.6, 126.5, 125.5, 124.6,124.5, 124.5, 124.5, 124.4, 124.3, 124.3, 124.2, 124.2, 123.9, 122.8, 115.9,113.9, 113.7, 113.6, 113.4, 113.3, 113.1, 111.9, 111.8, 73.1, 72.7, 69.2, 51.4, 48.0, 42.8. 19 F NMR (565 MHz, chloroform-d) δ -62.61, -62.62. HRMS (ESI): for C 20 H 22 F6N2O2, [M + H] + Calculated value: 437.1664; Measured value: 437.1664. HPLC purity: 98.51%.

[0226] 1-((2-(trifluoromethyl)benzyl)oxy)-3-((2-((2-(trifluoromethyl)phenyl)amino)ethyl)amino) Propyl-2-ol (34) From A 10 (30 mg, 0.15 mmol) and B 16 The title compound was prepared by silica gel column chromatography (34.1 mg, 0.15 mmol) with DCM / MeOH (200:1–100:1 eluent, with a few drops of ammonia) as the eluent. White solid, 42.9 mg, 67.6% yield, mp 64–65 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 7.67 (d, J= 7.9 Hz, 2H), 7.56 (t, J = 7.6 Hz, 1H), 7.42 (m, J = 24.0, 7.9 Hz, 3H), 6.74 (t, J = 8.1Hz, 2H), 4.94 (d, J = 5.3 Hz, 1H), 4.76 (s, 2H), 3.96 (m, J = 6.2, 4.1 Hz, 1H), 3.60 (m, J = 5.8 Hz, 2H), 3.27 (q, J = 5.6 Hz, 2H), 2.97 (m, J = 5.7,2.0 Hz, 2H), 2.86-2.71 (m, 2H). 13 C NMR (101 MHz, chloroform-) d ) δ 145.8, 145.8,145.8, 136.7, 136.7, 133.1, 132.0, 129.3, 129.1, 128.4, 128.2, 127.9, 127.6,127.3, 126.7, 126.7, 126.6, 126.6, 126.5, 125.9, 125.9, 125.8, 125.8, 125.7,123.9, 123.0, 120.2, 115.9, 114.0, 113.7, 113.4, 113.1, 111.9, 73.3, 69.5,69.2, 51.5, 48.0, 42.9. 19 F NMR (565 MHz, chloroform-d) δ -60.07, -62.64. HRMS (ESI): for C 20 H 22 F6N2O2, [M + H] + Calculated value: 437.1664; Measured value: 437.1663. HPLC purity: 99.4%.

[0227] N -(2-((2-hydroxy-3-(4-(trifluoromethyl)phenoxy)propyl)amino)ethyl)-2-(trifluoromethyl)benzenesulfonate Amide (35) From A 16The title compound was prepared by silica gel column chromatography using B7 (30 mg, 0.11 mmol) and B7 (24.4 mg, 0.11 mmol) with DCM / MeOH (100:1–50:1, with a few additional drops of ammonia) as eluent. The product was a white solid, 37.7 mg, 69.3% yield, mp 95–97 °C. 1 H NMR (600 MHz, DMSO- d 6) δ 8.11 (dd, J = 8.1, 3.0 Hz, 1H), 7.98 (dd, J = 8.1, 2.6 Hz, 1H), 7.89 (t, J = 7.7 Hz, 1H), 7.83 (t, J =7.7 Hz, 1H), 7.63 (dd, J = 9.0, 2.8 Hz, 2H), 7.11 (d, J = 8.5 Hz, 2H), 5.06(s, 1H), 4.02 (m, J = 9.9, 4.4, 1.6 Hz, 1H), 3.93 (m, J = 9.3, 6.2, 2.5 Hz,1H), 3.84 (m, J = 5.6 Hz, 1H), 2.99-2.93 (m, 2H), 2.61 (m, J = 12.9, 5.1, 4.2Hz, 3H), 2.54 (m, J = 10.7, 6.5, 3.7 Hz, 1H). 13 C NMR (151 MHz, DMSO- d 6) δ 162.0, 140.0, 133.7, 133.7, 133.2, 133.2, 130.4, 128.9, 128.9, 128.8, 128.8, 127.4, 127.3, 127.3, 127.3, 126.7, 126.5, 126.5, 126.1, 125.9, 124.3, 124.1, 122.5, 121.8, 121.5, 121.3, 121.3, 115.4, 71.4, 68.5, 52.3, 49.2, 43.3. 19F NMR (565 MHz, chloroform-d) δ -57.95, -61.54. HRMS (ESI): for C 19 H 20 F6N2O4S, [M + H] + Calculated value: 487.1126; Measured value: 487.1132. HPLC purity: 99.23%.

[0228] 1-(4-(trifluoromethyl)phenoxy)-3-((2-(2-(trifluoromethyl)phenoxy)ethyl)amino)prop-2-ol (36) From A 17 The title compound was prepared from B7 (30 mg, 0.15 mmol) and B7 (31.9 mg, 0.15 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (200:1–100:1, with a few drops of additional ammonia). White solid, 37.4 mg, 60.3% yield, mp 83–85 °C. 1 H NMR (600 MHz, DMSO- d 6) δ 7.62 (dd, J = 16.9, 8.2 Hz, 4H), 7.27 (d, J = 8.4 Hz, 1H), 7.10 (t, J = 8.2 Hz, 3H), 5.09 (d, J = 5.0 Hz, 1H), 4.16 (t, J = 5.6 Hz, 2H), 4.04 (dd, J = 9.8, 4.5 Hz, 1H), 3.96 (dd, J =9.8, 6.1 Hz, 1H), 3.90 (t, J = 5.9 Hz, 1H), 2.94 (t, J = 5.6 Hz, 2H), 2.76(dd, J = 12.0, 4.8 Hz, 1H), 2.68 (dd, J = 11.9, 6.7 Hz, 1H). 113 C NMR (151 MHz, DMSO- d 6) δ162.0, 156.9, 134.7, 130.1, 127.4, 127.4, 127.3, 127.3, 127.1, 127.1, 127.1, 126.0, 125.2, 124.2, 123.4, 121.5, 121.3, 120.7, 117.8, 117.6, 117.4, 115.4, 114.1, 71.4, 69.0, 68.5, 52.5, 48.6. 19 F NMR (565 MHz, chloroform- d ) δ-61.51, -62.26. HRMS(ESI): For C 19 H 19 F6NO3, [M + H] + Calculated value: 424.1347; Measured value: 424.1355. HPLC purity: 95.69%.

[0229] 1-(4-(trifluoromethyl)phenoxy)-3-((2-((2-(trifluoromethyl)phenyl)thio)ethyl)amino)prop-2- Alcohols (37) From A 18 The title compound was prepared from B7 (30 mg, 0.14 mmol) and B7 (29.6 mg, 0.14 mmol) and purified by silica gel column chromatography with eluent DCM / MeOH (200:1–100:1, with a few additional drops of ammonia). White solid, 34.5 mg, 58.1% yield, mp 77–78 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 7.68 (d, J = 7.9 Hz, 1H),7.59-7.46 (m, 4H), 7.33 (t, J = 7.6 Hz, 1H), 6.99 (d, J = 8.4 Hz, 2H), 4.08-4.01 (m, 3H), 3.17 (m, J = 6.4, 2.2 Hz, 2H), 2.99-2.87 (m, 3H), 2.83-2.76 (m,1H). 13 C NMR (101 MHz, chloroform-) d ) δ161.1, 135.4, 132.0, 131.6, 130.6, 130.3, 127.0, 127.0, 127.0, 126.9, 126.9, 126.9, 126.3, 125.7, 125.1, 123.4, 123.1, 123.0, 122.4, 120.3, 114.5, 70.5, 68.2, 51.1, 47.9, 35.2. HRMS(ESI): For C 19 H 19 F6NO2S, [M + H] + Calculated value: 440.1119; Measured value: 440.1120. HPLC purity: 98.46%.

[0230] 1-((2-((2-(trifluoromethyl)phenyl)amino)ethyl)amino)-3-((4-(trifluoromethyl)phenyl)thio) Propyl-2-ol (38) From A 10 (30 mg, 0.15 mmol) and B 12 The title compound was prepared by silica gel column chromatography (34.4 mg, 0.15 mmol) with DCM / MeOH (200:1–100:1 eluent, with a few drops of ammonia) as the eluent. White solid, 34.3 mg, 70.7% yield, mp 65–66 °C. 1 H NMR (400 MHz, chloroform-) d ) δ 7.53 (d, J = 8.2 Hz, 2H), 7.42 (m, J = 15.6, 7.9 Hz, 4H), 6.75 (dd, J = 8.1, 5.4 Hz, 2H), 4.87 (s, 1H), 3.86 (m, J = 7.3, 3.9 Hz, 1H), 3.34-3.22 (m, 2H), 3.20-3.09 (m, 2H), 3.03-2.88 (m, 3H), 2.72 (dd, J = 12.2, 7.5 Hz, 1H). 13 C NMR (101 MHz, chloroform-) d ) δ145.7, 145.7, 141.3, 133.1, 128.4, 128.1, 128.0, 127.7, 126.8, 126.7, 126.7, 126.6, 126.6, 125.8, 125.8, 125.7, 125.7, 125.4, 123.9, 122.7, 116.1, 113.7, 113.4, 111.9, 68.5, 53.4, 48.0, 42.9, 37.7. 19 F NMR (565 MHz, chloroform-d) δ -62.48, -62.55. HRMS (ESI): for C 19 H 20 F6N2OS, [M + H] + Calculated value: 439.1279; Measured value: 439.1279. HPLC purity: 96.13%.

[0231] 1-((4-(trifluoromethyl)phenyl)thio)-3-((2-((2-(trifluoromethyl)phenyl)thio)ethyl)amino) Propyl-2-ol (39) From A 18 (30 mg, 0.14 mmol) and B 12 The title compound was prepared by silica gel column chromatography (31.8 mg, 0.14 mmol) with DCM / MeOH (200:1–100:1 eluent, with a few drops of ammonia) as the eluent. White solid, 46.8 mg, 66.4% yield, mp 73–75 °C. 1 H NMR (600 MHz, chloroform-) d ) δ 7.70 -7.67 (m, 1H), 7.54 (t, J = 8.6 Hz, 3H), 7.49 (t, J = 7.7 Hz, 1H), 7.43 (d, J = 8.2 Hz, 2H), 7.33(t, J = 7.6 Hz, 1H), 3.82 (m, J = 12.0, 6.1, 3.6 Hz, 1H), 3.18-3.10 (m, 4H), 2.93-2.84 (m, 3H), 2.65 (dd, J = 12.2, 8.1 Hz, 1H). 13 C NMR (151 MHz, chloroform-) d ) δ141.5, 135.4, 132.0, 131.6, 130.6, 130.4, 127.9, 127.9, 127.7, 127.5, 127.0, 127.0, 126.9, 126.9, 126.3, 125.8, 125.8, 125.7, 125.7, 125.0, 124.6, 123.2, 122.8, 68.2, 53.3, 47.8, 37.5, 35.2. 19 F NMR (565 MHz, chloroform-d) δ -60.78, -62.46. HRMS (ESI): for C 19 H 19 F6NOS2, [M + H] + Calculated value: 456.0890; Measured value: 456.0893. HPLC purity: 95.63%.

[0232] MIC determination Protein overproduction, purification, and inhibitor testing were performed using the previously established protocol. Briefly, C-SmBiT-CH (40 μL, 100 nM, in PBS) was added to each well of a 96-well plate and then mixed with 20 μL of the desired concentration of the compound. The mixture was incubated at 37°C for 10 minutes. Then, N-LgBiT-NusG (40 μL, 100 nM, in PBS) was added to each well, followed by incubation at 37°C for 10 minutes. After this final incubation step, an equal volume of Promega Nano-Glo was added. ® The luciferase assay substrate (Promega, Madison, Wisconsin, United States) was added to the reaction mixture. The emitted light was measured using a Victor X3 Multilabel plate reader (Waltham, Massachusetts, United States). For consistent results, the experiment was repeated in triplicate.

[0233] MBC Measurement MBC values ​​were determined according to the Clinical and Laboratory Standards Institute (CLSI) guidelines. After establishing the MIC, solutions ranging from 1×MIC to 16×MIC were prepared, and 10 µL aliquots were spotted onto Columbia blood agar plates. The plates were incubated at 37°C for 24 hours, and viable bacterial colonies were counted to calculate colony forming units / mL (CFU / mL). MBC was established by comparing CFU / mL values ​​with untreated controls. All experiments were performed in duplicate to ensure reproducibility.

[0234] Combined with inhibition assay Protein overproduction, purification, and inhibitor testing were performed using the previously established protocol. Briefly, C-SmBiT-CH (40 μL, 100 nM, in PBS) was added to each well of a 96-well plate and then mixed with 20 μL of the desired concentration of the compound. The mixture was incubated at 37°C for 10 minutes. Then, N-LgBiT-NusG (40 μL, 100 nM, in PBS) was added to each well, followed by incubation at 37°C for 10 minutes. After this final incubation step, an equal volume of Promega Nano-Glo was added. ® The luciferase assay substrate (Promega, Madison, Wisconsin, United States) was added to the reaction mixture. The emitted light was measured using a Victor X3 Multilabel plate reader (Waltham, Massachusetts, United States). For consistent results, the experiment was repeated in triplicate.

[0235] Time-sterilization kinetics The dose- and time-dependent antimicrobial effects of the compound against Staphylococcus aureus strains under aerobic conditions were evaluated by adapting to relevant CLSI guidelines. Staphylococcus aureus cells were suspended to approximately 1.5 × 10⁻⁶ MIC in CA-MHB broth supplemented with different concentrations of the compound (i.e., ¼ ×, 1 ×, 4 ×, and 16 × MIC) during the logarithmic growth phase. 6 CFU / mL (colony-forming units / mL). As an untreated control, bacteria were incubated in CA-MHB broth without the compound. Cultures were grown at 37°C with shaking at 180 rpm. 20 µL samples were taken from each treatment group at defined time intervals (i.e., 0, 2, 4, 6 h) and then serially diluted 10-fold in sterile phosphate-buffered saline (PBS). For each dilution, 5 µL of sample was spotted onto Columbia blood agar plates. The plates were then incubated overnight at 37°C, after which the number of viable bacteria in each sample was counted and expressed as CFU / mL. The entire experiment was performed in triplicate.

[0236] Epifluorescence microscopy At 37°C Bacillus subtilis Multiple strains were cultured overnight on selective LB agar plates (Lennox formulation). Single colonies from each plate were inoculated into LB medium containing a suitable selective antibiotic and incubated overnight at 37°C with shaking at 180 rpm. The overnight cultures were diluted to 0.05 OD. 600The mixture was stirred and grown at 37°C until an OD of 0.50 was reached. 600 At this point, the antibiotic and compound 38 (1× MIC), along with xylose, were added to the culture, followed by incubation at 37°C with shaking for 30 minutes. To visualize the nuclei, 4',6-diamidinyl-2-phenylindole (DAPI) was added to a final concentration of 1 µg / mL.

[0237] For microscopic analysis, 2.5 µL of treated cell culture was placed on a freshly prepared 1.2% agarose pad, covered with a coverslip, and then imaged. Fluorescence images were captured using an ECLIPSE Ti2-E live-cell fluorescence imaging system (Nikon) equipped with a 100× / 1.45 oil immersion objective. GFP signals were visualized using a FITC filter (525 / 50 emission), and DAPI signals were visualized using a DAPI filter (460 / 50 emission). Digital images were processed and analyzed using ImageJ software.

Claims

1. A compound of formula 1: 1 Or pharmaceutically acceptable salts, wherein: m is an integer selected from 1 to 4; n is an integer selected from 1 to 4; X 1 It is -O-, -S-, -OCH2-, -SCH2-, -(NR 5 - or -SO2(NR) 5 )-; X 2 It is -O-, -S-, -OCH2-, -SCH2-, -(NR 5 - or -SO2(NR) 5 )-; Ar 1 Selected from: 、 、 、 , , and ; Ar 2 Selected from: 、 、 、 , , and ; Each time R appears, it is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, and aralkyl; or R appearing twice together with their covalently bonded atoms form 3-6 membered cycloalkyl or heterocycloalkyl groups. R 1 Each occurrence is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, perhaloalkoxy, halogen, nitrile, azido, -OR 5 -SR 5 -N(R) 5 )2、-C(O)R 5 -C(O)OR 5 -OC(O)R 5 -N(R) 5 )C(O)R 5 -C(O)N(R) 5 )2、-N(R 5 )C(O)OR 5 -OC(O)N(R) 5 )-、-OC(O)OR 5 -N(R) 5 )C(O)N(R 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 and -N(R 5 )S(O)2R 5 ; R 2 Each occurrence is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, perhaloalkoxy, halogen, nitrile, azido, -OR 5 -SR 5 -N(R) 5 )2、-C(O)R 5 -C(O)OR 5 -OC(O)R 5 -N(R) 5 )C(O)R 5 -C(O)N(R) 5 )2、-N(R 5 )C(O)OR 5 -OC(O)N(R) 5 )-、-OC(O)OR 5 -N(R) 5 )C(O)N(R 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 and -N(R 5 )S(O)2R 5 ; R 3 It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, -C(O)R 5 -C(O)OR 5 -C(O)N(R) 5 )2、-S(O)2R 5 or -S(O)2N(R 5 )2; R 4 It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, -C(O)R 5 -C(O)OR 5 -C(O)N(R) 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 or -P(O)(OR 5 )2; and R 5 Each occurrence of R is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, and aralkyl; or two occurrences of R. 5 Together with the covalently bonded atoms, they form 3-6 membered cycloalkyl or heterocycloalkyl groups.

2. The compound according to claim 1, wherein m is an integer selected from 1-2; and n is an integer selected from 1-2.

3. The compound according to claim 1 or 2, wherein X 1 It is -O-, -S-, -(NR) 5 - or -SO2(NR) 5 )-.

4. The compound according to any one of claims 1-3, wherein Ar 1 Selected from: , and ;and Ar 2 yes: 。 5. The compound according to any one of claims 1-4, wherein at least one R 1 It is CF3; and at least one R 2 It's CF3.

6. The compound according to claim 1, wherein the compound has formula 2: 2 Or its pharmaceutically acceptable salt, wherein: m is an integer selected from 1 to 4; n is an integer selected from 1 to 4; X 1 It is -O-, -S-, -OCH2-, -SCH2-, -(NR 5 - or -SO2(NR) 5 )-; X 2 It is -O-, -S-, -OCH2-, -SCH2-, -(NR 5 - or -SO2(NR) 5 )-; Ar 1 Selected from: 、 、 、 , , and ; Ar 2 Selected from: 、 、 、 , , and ; Each time R appears, it is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, and aralkyl; or R appearing twice together with their covalently bonded atoms form 3-6 membered cycloalkyl or heterocycloalkyl groups. R 1 Each occurrence is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, perhaloalkoxy, halogen, nitrile, azido, -OR 5 -SR 5 -N(R) 5 )2、-C(O)R 5 -C(O)OR 5 -OC(O)R 5 -N(R) 5 )C(O)R 5 -C(O)N(R) 5 )2、-N(R 5 )C(O)OR 5 -OC(O)N(R) 5 )-、-OC(O)OR 5 -N(R) 5 )C(O)N(R 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 and -N(R 5 )S(O)2R 5 ; R 2 Each occurrence is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, perhaloalkoxy, halogen, nitrile, azido, -OR 5 -SR 5 -N(R) 5 )2、-C(O)R 5 -C(O)OR 5 -OC(O)R 5 -N(R) 5 )C(O)R 5 -C(O)N(R) 5 )2、-N(R 5 )C(O)OR 5 -OC(O)N(R) 5 )-、-OC(O)OR 5 -N(R) 5 )C(O)N(R 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 and -N(R 5 )S(O)2R 5 ; R 3 It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, -C(O)R 5 -C(O)OR 5 -C(O)N(R) 5 )2、-S(O)2R 5 or -S(O)2N(R 5 )2; R 4 It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, -C(O)R 5 -C(O)OR 5 -C(O)N(R) 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 or -P(O)(OR 5 )2; and R 5 Each occurrence of R is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, and aralkyl; or two occurrences of R. 5 Together with the covalently bonded atoms, they form 3-6 membered cycloalkyl or heterocycloalkyl groups.

7. The compound according to claim 6, wherein Ar 1 Selected from: , and ;and Ar 2 yes: 。 8. The compound according to claim 6 or 7, wherein at least one R 1 It is CF3; and at least one R 2 It's CF3.

9. The compound according to any one of claims 6-8, wherein R 3 It is hydrogen and R 4 It is hydrogen.

10. The compound according to claim 1, wherein the compound has formula 3: 3 Or its pharmaceutically acceptable salt, wherein: m is an integer selected from 1 to 2; n is an integer selected from 1 to 2; A is C, CH, or N; X 1 It is -O-, -S-, -(NR) 5 - or -SO2(NR) 5 )-; X 2 It is -O-, -S-, -(NR) 5 -, -OCH2- or -SCH2-; Each time R appears, it is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, and aralkyl; or R appearing twice together with their covalently bonded atoms form 3-6 membered cycloalkyl or heterocycloalkyl groups. R 1 Each occurrence is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, perhaloalkoxy, halogen, nitrile, azido, -OR 5 -SR 5 -N(R) 5 )2、-C(O)R 5 -C(O)OR 5 -OC(O)R 5 -N(R) 5 )C(O)R 5 -C(O)N(R) 5 )2、-N(R 5 )C(O)OR 5 -OC(O)N(R) 5 )-、-OC(O)OR 5 -N(R) 5 )C(O)N(R 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 and -N(R 5 )S(O)2R 5 ; R 2 Each occurrence is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, perhaloalkoxy, halogen, nitrile, azido, -OR 5 -SR 5 -N(R) 5 )2、-C(O)R 5 -C(O)OR 5 -OC(O)R 5 -N(R) 5 )C(O)R 5 -C(O)N(R) 5 )2、-N(R 5 )C(O)OR 5 -OC(O)N(R) 5 )-、-OC(O)OR 5 -N(R) 5 )C(O)N(R 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 and -N(R 5 )S(O)2R 5 ; R 3 It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, -C(O)R 5 -C(O)OR 5 -C(O)N(R) 5 )2、-S(O)2R 5 or -S(O)2N(R 5 )2; R 4 It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, aralkyl, -C(O)R 5 -C(O)OR 5 -C(O)N(R) 5 )2、-S(O)2R 5 -S(O)2N(R) 5 )2 or -P(O)(OR 5 )2; and R 5 Each occurrence of R is independently selected from hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, and aralkyl; or two occurrences of R. 5 Together with the covalently bonded atoms, they form 3-6 membered cycloalkyl or heterocycloalkyl groups.

11. The compound according to claim 10, wherein R 3 It is hydrogen, alkyl, haloalkyl, perhaloalkyl, alkenyl, ynyl, cycloalkyl, heterocycloalkyl, aryl, heteroaryl, or aralkyl.

12. The compound according to claim 10, wherein R 3 It is hydrogen.

13. The compound according to claim 12, wherein R 4 It is hydrogen.

14. The compound according to claim 1, wherein the compound is selected from: 、 、 、 、 、 、 、 、 , , , , and its pharmaceutically acceptable salt, where m is an integer selected from 1 to 2; and n is an integer selected from 1 to 2.

15. The compound according to claim 14, wherein m is 1 and n is 1.

16. The compound according to claim 1, wherein the compound is selected from: , , , , , , , , , , , , , And its pharmaceutically acceptable salts.

17. The compound according to claim 1, wherein the compound is selected from: , , , , And its pharmaceutically acceptable salts.

18. A pharmaceutical composition comprising a compound according to any one of claims 1-17 and at least one pharmaceutically acceptable carrier or pharmaceutically acceptable excipient.

19. A method for treating a bacterial infection in a subject in need, the method comprising administering to the subject a therapeutically effective amount of the compound according to any one of claims 1-17.

20. The method of claim 19, wherein the bacterial infection is caused by Gram-positive bacteria.

21. The method of claim 19, wherein the bacterial infection is caused by Gram-negative bacteria.

22. The method of claim 19, wherein the bacterial infection is caused by a bacterium selected from Enterococcus faecalis (… Enterococcus faecalis Staphylococcus aureus ( Staphylococcus aureus Streptococcus pneumoniae () Streptococcus pneumonia Enterobacter cloacae () Enterobacter cloacae ), Escherichia coli ( Escherichia coli Acinetobacter baumannii ( Acinetobacter baumannii Staphylococcus epidermidis ( S. epidermidis ), saprophytic Staphylococcus ( S. saprophyticus ), Streptococcus pyogenes ( S. pyogenes ) and agalactococcus ( S. agalactiae Caused by bacteria.

23. The method of claim 19, wherein the bacterial infection is caused by bacteria selected from methicillin-resistant Staphylococcus aureus, vancomycin-resistant Staphylococcus aureus, aminoglycoside-resistant Staphylococcus aureus, macrolide-resistant Staphylococcus aureus, and fluoroquinolone-resistant Staphylococcus aureus.

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