Compositions and methods for treating microbial infections

Abstract

Described are analogs of aztreonam and disulfiram of Formula I, II, and III. These compounds can be used to treat bacterial infections in a subject. Also disclosed is the use of aztreonam and disulfiram for treating H. pylori infections. Combinations of the disclosed analogs and / or aztreonam and disulfiram with additional antibiotics and proton pump inhibitors are also disclosed.

Description

[0001] Attorney Docket No. 10063-110WO1

[0002] COMPOSITIONS AND METHODS FOR TREATING MICROBIAL INFECTIONS

[0003] CROSS REFERENCE TO RELATED APPLICATIONS

[0004] This application claims the benefit of priority to U. S. Provisional Application 63 / 719,849, filed November 13, 2024, which is incorporated by reference herein in its entirety.

[0005] ACKNOWLEDGEMENT OF GOVERNMENT SUPPORT

[0006] This invention was made with government support under grant no. P20GM121344 awarded by the National Institutes of Health. The government has certain rights in the invention.

[0007] BACKGROUND

[0008] H. pylori is a Gram-negative bacterium, which survives in the acidic gastric environment by neutralizing acid through urease secretion and penetrating the gastric mucus layer. It also elicits host immune responses, causing cell death and utilizing nutrients for continuous colonization. About half of the global population harbors H. pylori (Nguyen, T. N., et al., Host determinants of Helicobacter pylori infection and its clinical outcome. Helicobacter, 1999. 4(3): p. 185-197) which spreads through fecal-oral or oral-oral routes due to poor hygiene and among family members (Peek, R. M., et al., Helicobacter pylori and gastrointestinal tract adenocarcinomas. Nature Reviews Cancer, 2002. 2(1): p. 28-37; Ansari, S., et al., Helicobacter pylori Virulence Factors Exploiting Gastric Colonization and its Pathogenicity. Toxins, 2019. 11(11): p. 677; Ansari, S., et al., Survival of Helicobacter pylori in gastric acidic territory. Helicobacter, 2017. 22(4): p.

[0009] 10.1111 / hel.12386). The virulent strains of H. pylori produce two major toxins - the oncogenic cytotoxic-associated antigen A (CagA) and the vacuolating toxin A (VacA) (Knorr, J., et al., Classification of Helicobacter pylori Virulence Factors: Is CagA a. Toxin or Not? Trends Microbiol, 2019. 27(9): p. 731-738; Kim, S.-H., et al., Inhibitory effects of anthocyanins on secretion of Helicobacter pylori CagA and VacA toxins. International journal of medical sciences, 2012. 9(10): p. 838; Ansari, S. et al., Role of vacuolating cytotoxin A in Helicobacter pylori infection and its impact on gastric pathogenesis. Expert Rev Anti Infect Ther, 2020. 18(10): p. 987-996). CagA is directly injected into gastric epithelial cells and is associated with 89.0%-95.5% of gastric mucosa-assisted lymphoid tissue lymphoma cases (Eek, M., et al., MALT-type lymphoma of the stomach is associated with Helicobacter pylori strains expressing the CagA protein. Gastroenterology, 1997. Attorney Docket No. 10063-110WO1

[0010] 112.(5): p. 1482-6). CagA triggers inflammation through diverse mechanisms, including the infiltration of bacterial peptidoglycans into gastric epithelial cells, and activates the cytoplasmic pathogen recognition receptor Nodi, subsequently inducing NF-KB and triggering the production of inflammatory cytokines (Viala, J., et al., Nodi responds to peptidoglycan delivered by the Helicobacter pylori cag pathogenicity’ island. Nat Immunol, 2004. 5(11): p. 1166-74). VacA, classified as a pore-forming toxin due to its capacity to create anion-selective channels in planar lipid bilayers (Czajkowsky, D. M., et al., The vacuolating toxin from Helicobacter pylori forms hexameric pores in lipid bilayers at low pH. Proc Natl Acad Sci U S A, 1999. 96(5): p. 2001-6), can be translocated across the outer membrane, released into the extracellular space, or retained on the bacterial surface (Foegeding, N. J., et al., An Overview of Helicobacter pylori VacA Toxin Biology. Toxins, 2016. 8(6): p. 173). Upon internalization into gastric epithelial cells, VacA accumulates in various cellular compartments, leading to cell vacuolation, depolarization of membrane potential, and dissipation of mitochondrial transmembrane potential (Zhang, K., et al., Cryo-EM structures of Helicobacter pylori vacuolating cytotoxin A oligomeric assemblies at near-atomic resolution. Proceedings of the National Academy of Sciences, 2019.

[0011] 116(14): p. 6800-6805). These events result in mitochondrial dysfunction, cell apoptosis, and the activation of mitogen-activated protein kinases (Czajkowsky, D. M., et al., 1999). Additionally, VacA induces autophagy, inhibits T cell activities, and activates the proapoptotic factor Bax, promoting apoptosis and cell death (Ansari, S., et al., 1999). H. pylori infection often originates in childhood and can cause gastritis, which can develop into gastric cancer (Weyermann, M., et al., Acquisition of Helicobacter pylori infection in early childhood: independent contributions of infected mothers, fathers, and siblings. Official journal of the American College of Gastroenterology | ACG, 2009. 104(1): p. 182-189) and is responsible for approximately 75% of all cases of this malignancy (Hatakeyama, M., Structure and function of Helicobacter pylori CagA, the first-identified bacterial protein involved in human cancer. Proc Jpn Acad Ser B Phys Biol Sci, 2017. 93(4): p. 196-219).

[0012] Effective therapy for H pylori infection requires the sequential administration of at least two clinical antibiotics (amoxicillin, clarithromycin, metronidazole, or nitrofurantoin) and a proton-pump inhibitor (PPI) (omeprazole, rabeprazole, esomeprazole, or lansoprazole) (Romano, M., et al., Eradication of Helicobacter pylori: a clinical update. Medscape General Medicine, 2004. 6(1): p. 19). However, standard triple therapy regimens fail to clear H. pylori infection due to increasing antibiotic resistance rates (Shiota, S., et al., Antibiotic resistance of Helicobacter pylori among male United States veterans. Clinical

[0013] 9 Attorney Docket No. 10063-110WO1

[0014] Gastroenterology and Hepatology, 2015. 13(9): p. 1616-1624). For example, H. pylori resistance to clarithromycin among male U. S. veterans increased from 16% to 24% from 2009 to 2013, and resistance to metronidazole is also high (id.). Hence, novel therapeutic approaches, involving compounds either from biotic sources or synthesized, are urgently needed.

[0015] Furthermore, pathogen-specific antibiotics can mitigate antimicrobial resistance (AMR). Developing an antibiotic is often expensive, time-consuming, laborious, and high- risk, which has slowed the pace of research on new antimicrobials, with most pharmaceutical industries focusing on therapies for chronic diseases and disorders (Dutescu, LA., et al., Encouraging the Development of New Antibiotics: Are Financial Incentives the Right Way Forward? A Systematic Review and Case Study. Infect Drug Resist, 2021. 14: p.

[0016] 415-434). Also, 90% of novel drug candidates fail due to a variety of reasons, including a lack of clinical efficacy (Sun, D., et al., Why 90% of clinical drug development fails and how to improve it? Acta Pharm Sin B, 2022. 12(7): p. 3049-3062). The rapid rise of AMR and emerging new pathogens poses a global health threat. Fatalities caused by drug-resistant bacteria are estimated to surpass cancer-related casualties by 2050 (de Kraker, M. E. A., et al., Will 10 Million People Die a Year due to Antimicrobial Resistance by 2050? PLOS Medicine, 2016. 13(11): p. el002184). The World Health Organization has declared H. pylori a class 1 gastric carcinogen (Peek, R. M., et al., Helicobacter pylori and gastrointestinal tract adenocarcinomas. Nature Reviews Cancer, 2002. 2(1): p. 28-37) and a type-2, high-priority pathogen to promote research and development of new antibiotics against drug-resistant H. pylori, including clarithromycin-resistant H. pylori.

[0017] While antibiotic treatments recover the host from infection, they disturb the gut microbiome. Antibiotics disturb the balance and communication within the bacterial community and between bacteria and the host. Antibiotic-induced alterations in microbial composition negatively affect host health by reducing diversity, modifying the functional aspects of the microbiota, and promoting the formation and selection of antibiotic -resistant strains (Patangia, D. V., et al., Impact of antibiotics on the human microbiome and consequences for host health. Microbiologyopen, 2022. 11(1): p. e1260). These changes make hosts more vulnerable to infections by other gastric pathogens (id.), which further complicates clinical treatment. Given the intricate connection between the host and microbiota, judicious antibiotic use is crucial, emphasizing the need to explore antibiotic alternatives to minimize unintended side effects. Efforts should focus on designing, Attorney Docket No. 10063-110WO1

[0018] developing, and implementing new alternatives, along with efficient methodologies to preserve and restore the microbial community following antibiotic-associated disruptions.

[0019] As noted above, there is an urgent need for new compositions and treatment methods for H. pylori and drug resistant variants thereof. This need similarly applies to other microbial infections as well. The compositions and methods disclosed herein address these and other needs.

[0020] SUMMARY

[0021] Disclosed herein, in one aspect, are compounds, compositions, and methods for making and using the disclosed compounds and compositions. In a more specific aspect, disclosed herein are compounds having Formula I or II:

[0022]

[0023] wherein,

[0024] n is 0 to 4;

[0025] — is a bond that is present or absent,

[0026] X1is O or S;

[0027] X2is S, CH2, or CHR4;

[0028] X3and X4are, independently, CH2, O, S, NH, NR4, CHR4, CR4, or C(R4)2;

[0029] R1and R2are, independently, H, branched or straight chain Ci-C4alkyl, aryl, heteroaryl, (Ci- C4alkyl)aryl, (Ci-C4aikyl)heteroaryl, C3-C6cycloalkyl, C3-C6cycloheteroalkyl, (C1-C4alkyl)C3-C6cycloalkyl, or (Ci-C4alkyl)C3-C6cycloheteroalkyl, wherein R1and R2are not both H;

[0030] R3is branched or straight chain C1-C12alkyl, C1-C12heteroalkyl, aryl, heteroaryl, (Ct- C4alkyl)aryl, (Ci-C4alkyl)heteroaryl, Cs-Cecycloalkyl, Cs-Cecycloheteroalkyl, (Ci- C4alkyl)C3-C6cycloalkyl, or (C1-C4alkyl)C3-C6cycloheteroalkyl;

[0031] R4is branched or straight chain Ci-C4alkyl, C2-C4alkenyl, Ci-C^heteroalkyl, C2- C4heteroalkenyl, or two R4's bond together to form a fused Cs-Cecycloalkyl, fused C3-C6cycloheteroalkyl, fused aryl, fused heteroaryl, spiro Cs-Cecycloalkyl, or spiro C2-C6cycloheteroalkyl, any of which are optionally substituted with oxo, sulfonate, sulfonyl, halo, hydroxyl, thiol, carboxylate, carbonyl, Ci-Cealkyl, or C1-C6haloalkyl; Attorney Docket No. 10063-110WO1

[0032] or a pharmaceutically acceptable salt thereof;

[0033] or a pharmaceutically acceptable salt thereof. In a specific example, the compound is not Formula II where X1and X2are both S, R3is C8alkyl, n is 1, and X ' and X4are both CH2.

[0034] In other aspect, disclosed herein are compounds having Formula III:

[0035]

[0036] SO3H

[0037] III

[0038] wherein,

[0039] Lis null, -NHC(O)C(CH3)2-, -NR5C(O)C(CH3)2, -or -OC(O)C(CH3)2-;

[0040] R5is branched or straight chain Ci-Ci2alkyl, Ci-Ci2heteroalkyl, aryl, heteroaryl, (Ci- CralkyDaryl, (Ci-C4alkyl)heteroaryl, C3-C6cycloalkyl, C3-C6cycloheteroalkyl, (C1-C4alkyl)C3-C6cycloalkyl, or (Ci -C4alkyl)C3-C6cycloheteroalkyl;

[0041] or a pharmaceutically acceptable salt thereof.

[0042] Compositions comprising combinations of these compounds are also disclosed, as are combinations of these compounds with aztreonam and / or disulfiram, optionally other antibiotics and proton pump inhibitors. Methods of treating bacterial infections by administering the disclosed compositions are also disclosed.

[0043] Additional advantages will be set forth in part in the following description and in part will be obvious from the description or may be learned by practicing the aspects described below. The advantages described below will be realized and attained by the elements and combinations pointed out in the appended claims. It is to be understood that the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive.

[0044] BRIEF DESCRIPTION OF THE FIGURES

[0045] Figures 1 -1G show antibacterial activity of aztreonam and disulfiram. Fig. 1A. Killing kinetics- / / , pylori 60190 was treated with test drugs at lx or 4x MIC and total CFU Attorney Docket No. 10063-110WO1

[0046] demonstrated that the drug inhibited bacterial multiplication at high-dose. Fig. IB.

[0047] Adhesion inhibition- / / . pylori adhesin to gastric epithelial cells was monitored. The test drugs administration cleared adhered H. pylori. Figs. 1C-1D. G. mellonella larvae were infected with H. pylori. 60190 and treated with different doses of test drugs injected at the last proleg of the larvae. Aztreonam at 25 mg / kg and disulfiram at 20 mg / kg promoted 50% of H. pylori- infected larvae survival. Figs. 1E-1F. Motility assay-Bacterial motility was monitored in soft agar impregnated with test drugs to demonstrate that test drugs hindered H. pylori motility. Fig. 1G. Vacuolation assay- / / . pylori 60190 infected gastric epithelial cells in the presence and absence of test drugs. The light microscopy was used to visualize VacA-mediated vacuolation and aztreonam or disulfiram treatment hindered vacuolation.

[0048] Figures 2A-2G show inflammation scores are based on histological findings.

[0049] C57BL / C mouse model was used to test the in vivo efficacy. H. pylori SS1-infected mice (Fig. 2D-Single therapy) or H. pylori PMSS1 -infected mice (Fig. 2E-Triple therapy) were treated with test drugs, and gastric tissues were sectioned and H& E-stained. The blinded slides were scored by Gl-Histopathologist based on leucocyte infiltration for inflammation score. Aztreonam and disulfiram decreased H. pylori-induced inflammation significantly (pcO. OOl) in single therapy (Fig. 2D). Gastric tissues samples were stained using H& E (Figs. 2F-Fig. 2G-above panel) and H. pylori polyclonal immunohistochemistry (Figs. 2F-Fig. 2G -below panel) suggested the drugs clear bacterial adherence (IHC), resulting in decreased inflammation (H& E ) (Fig. 2D; Fig. 2E).

[0050] Figures 3A-3J show in vivo toxicity, weight change and blood cells count during the shorter triple therapy. Figs. 3A-3B. In vivo kidney and liver toxicity during DIS& AZ alone or in combination therapy was determined using the BUN and ALT analysis. Fig. 3C. Weight change was monitored during DIS& AZ alone or in combination therapy. Figs. 3D-3I. Blood cells count (WBC and RBC) was determined using automated total blood count analyzer in the clinical laboratory. Fig. 3J. Histopathology studies of liver (right) and kidney tissues of treated mice.

[0051] Figures 4A-4D show observed diversity of gut microbiome after treatment. Figs. 4B-4C. Principal Coordinate Analysis (PCoA) based on bray Curtis and Unifrac analysis to study the changes in taxonomy level to study abundance and distance showed that AB, AZPA, and DISPA have similar microbiome profiles compared to H. pylori infected group. Fig. 4D. Gut-microbiome abundance between groups. Based on the stack plots, the abundance of Akkermansia and Bacteroides increased in treatment groups compared to controls. However, when the Wilcoxon test was applied, only the standard antibiotic therapy Attorney Docket No. 10063-110WO1

[0052] group showed a significantly higher abundance of Akkermansia and Bifidobacterium. The abundance of Bacteroides was increased in the standard antibiotic and AZ DIS triple therapy groups.

[0053] Figure 5 shows metabolomics analysis of bacterial cell lysate. H. pylori was treated with sub-MIC of aztreonam and disulfiram for 6 h, and cells were frozen until analysis. Metabolic pathways were significantly associated with the treatment. Bubble plots display altered metabolic pathways. Mummichog pathway enrichment analysis (PEA) was conducted for both ESI - and + modes with the Staphylococcus aureus N315 (MRSA / VSSA) [KEGG] network. The x-axis represents the gamma p- value.

[0054] Figure 6 shows RNA sequencing analysis of aztreonam and disulfiram treatment during a host-pathogen interaction. MKN28 gastric cells were infected with H. pylori cells and treated with 4xMIC of aztreonam or disulfiram for 22 h. Total RNA was isolated and subjected to RNA sequencing. Top left is a PCoA plot. The observation H. pylori infected MKN cells was distinct from the other control and treatment samples suggests the significant differences in gene expression patterns during infection compared to the control and treatment conditions. This indicates that the transcriptome undergoes substantial changes in response to infection, highlighting the importance of understanding the molecular mechanisms underlying the infection process. Top right shows volcano plots provide a visual representation of the relationship between fold change and statistical significance for each gene. Bottom shows a heat map analysis of known logCPM-valued genes are shown. We identified 2848 significantly up-regulated genes and 2758 significantly downregulated genes (P-value < 0.05) in the infection samples (MKN). The genes MUC21, TPSD1, and NKG7 were among the top upregulated genes observed in MKN samples, whereas downregulated genes such as AC083843.1, AC009123.1, and A2M-AS1 were identified in control and treatment samples. Gene Ontology (GO) pathway enrichment analysis revealed that the upregulated genes were associated with inflammatory response, chemokine production, and cell death pathways. Specifically, genes such as ILIA, TIMP1, HM0X1, TGFB1, HIF1A, and F2R were implicated in the inflammatory response pathway. Conversely, the downregulated genes were mainly involved in DNA replication, cell proliferation, and WNT signaling pathways. For instance, genes involved in the Wnt signaling pathway, such as WNT6, TLE5, FZD1, and LGR4, were downregulated.

[0055] Figure 7 shows scanning electron microscopic pictures of H. pylori treated with DIS or AZ at lOx MIC for 4h and visualized in scanning electron microscope. Attorney Docket No. 10063-110WO1

[0056] Figure 8 shows scanning electron microscopic pictures of S. aureus treated with DIS or DIS 5-2 or vancomycin at lOx MIC for 4h and visualized in scanning electron microscope.

[0057] Figure 9 shows killing kinetics: S. aureus MW strain was treated with DIS analogs at 4x MIC and total CFU demonstrated that the drug inhibited bacterial multiplication demonstrated as bacteriostatic.

[0058] Figure 10 shows mutational frequency of H. pylori against DIS and AZ.

[0059] Figure 11 shows killing kinetics: N. gonorrhoea 1145734 (BEI reference strain) was treated with DIS or AZ or its analogs at 4x MIC and total CFU demonstrated that the drug inhibited bacterial multiplication.

[0060] Figure 12 shows checkerboard assays. Combinatorial drugs were treated with A7. gonorrhoea 1145734 (BET reference strain) and the growth inhibition was measured.

[0061] Figure 13 shows scanning electron microscopic pictures of A. gonorrhoea 1145734 (BEI reference strain) treated with DIS or AZ or its analogs at lOx MIC for 4h and visualized in scanning electron microscope.

[0062] Figure 14 shows data from an acid stability study where selected analogs were exposed to pH 2.

[0063] Figure 15 shows metabolomic profiling of H. pylori exposed to aztreonam (AZ) and disulfiram (DIS) revealed distinct yet overlapping stress signatures in wild-type and drugresistant mutant cells. Both drugs induced nucleotide depletion, amino-acid perturbation, and altered acyl-carnitine levels, indicating global metabolic stress and membrane remodeling. In AZ-treated wild-type cells, central-carbon fluxes (acetyl-CoA, PEP, TCA intermediates) were redirected to support peptidoglycan precursor synthesis, whereas AZ-resistant mutants maintained sustained nucleotide turnover and membrane lipid compensation. DIS treatment in wild-type cells caused pleiotropic stress, with membrane perturbation, redox imbalance (NADP), and host metabolite changes (GABA, dopamine), whereas DIS-resistant mutants relied on lipid remodeling and nucleotide / redox buffering. Comparative analysis showed that both drugs share core antibacterial stress effects, but AZ primarily disrupts cell envelope and iron metabolism, while DIS targets thiol-redox homeostasis and protein synthesis. Resistance-associated mutations (e.g., fur, ftsl, bamAfor AZ; wecA, lepA, infA for DIS) align with these metabolic adaptations. Amino-acid shifts, acyl-carnitine accumulation, and nucleotide fluctuations suggest compensatory pathways in resistant clones. Overall, these data provide a mechanistic link between drug-specific Attorney Docket No. 10063-110WO1

[0064] metabolic stress and genetic adaptations in H. pylori. This integrated approach highlights distinct metabolic vulnerabilities for rational combination therapies.

[0065] Figure 16 shows comparative lipidomic heatmaps showing drug-induced membrane remodeling in H. pylori mutants and wild-type strains. Heatmaps illustrate relative abundance changes of phospholipid species in H. pylori strains under aztreonam (AZ) or disulfiram (DIS) treatment compared to untreated (NC) controls. (1) AZ-mutant ± AZ: Long-chain unsaturated phosphatidylcholines (PCs), including PC(18:1..22:6)+AcO and PC(16:0„20:l)+AcO, are highly enriched under AZ treatment, while shorter-chain PCs / PEs show moderate increases and LPG(14:0) is depleted. (2) DIS-mutant ± DIS: Elevated levels of PE(18:1..22:4) and PC(14:0..16: l)+Ac() indicate pronounced remodeling of PE and PC classes under DIS exposure; long-chain PCs remain low in untreated samples. (3) WT ± AZ: Strong upregulation of PCs (e.g., PC(14:0_16:l)+AcO, PC(14:0_18:l)+AcO) with reduced PE and PG abundance highlights drug-induced shifts toward PC-dominant membrane composition. (4) WT ± DIS: Distinct increases in PG and lysophosphatidylethanolamine (LPE) species -particularly LPE(18:2) and LPE(22:4)--suggest activation of membrane turnover and adaptive remodeling during DIS treatment. Color scale represents log-transformed relative abundance (blue = low; red = high). Data reflect the mean of biological replicates, normalized to total lipid content.

[0066] Figure 17 shows global transcriptional response of H. pylori AZ and DIS mutants compared to wild type upon antibiotic treatment. Heatmap illustrating differential gene expression profiles of H. pylori strains following exposure to aztreonam (AZ) and DIS compounds. Each column represents a biological replicate (1-3) for the AZ mutant, DIS mutant, and their corresponding wild-type controls under treated (AZ or DIS) and untreated (NC) conditions. Genes are hierarchically clustered based on log2fold change in expression (red: up-regulated; blue: down-regulated). The AZ mutant show's strong induction of stress- responsive and efflux-associated genes (hslU, hefA, hop A, gyrA), coupled with repression of ribosomal and translational components (rplV, rpmC, rpsH, tig), indicating translational slowdown and activation of survival pathways. In contrast, the w'ild-type strain treated with AZ exhibits moderate suppression of ribosomal genes (rpmB, rpsO, rplK) but elevated expression of oxidative stress and membrane stabilization factors (sodB, vacA, accB). Upon DIS treatment, both mutant and wild-type strains display pronounced down-regulation of translational machinery (rpmB, fabZ, tig, secE) and up-regulation of virulence-associated loci (cagC, cagF, vacA, ureA) and antioxidant defenses (trxA, sodB), suggesting that DIS exposure triggers virulence activation and redox adaptation. Collectively, the transcriptomic Attorney Docket No. 10063-110WO1

[0067] trends reveal that while wild-type strains primarily mount oxidative and metabolic stress responses, the AZ and DIS mutants undergo compensatory activation of efflux pumps, virulence islands, and membrane biogenesis pathways, reflecting mutation -driven resistance and adaptive survival strategies under antibiotic pressure.

[0068] DETAILED DESCRIPTION

[0069] The following description of the disclosure is provided as an enabling teaching of the disclosure in its best, currently known aspects. Many modifications and other aspects disclosed herein will come to mind to one skilled in the art to which the disclosed compositions and methods pertain, having the benefit of the teachings presented in the foregoing descriptions and the associated drawings. Therefore, it is to be understood that the disclosures are not to be limited to the specific aspects disclosed and that modifications and other aspects are intended to be included within the scope of the appended claims. The skilled artisan will recognize many variants and adaptations of the aspects described herein. These variants and adaptations are intended to be included in the teachings of this disclosure and to be encompassed by the claims herein.

[0070] Although specific terms are employed herein, they are used in a generic and descriptive sense only and not for purposes of limitation.

[0071] As can be apparent to those of skill in the art upon reading this disclosure, each of the individual aspects described and illustrated herein has discrete components and features that may be readily separated from or combined with the features of any of the other several aspects without departing from the scope or spirit of the present disclosure.

[0072] Any recited method can be carried out in the order of events recited or in any other order that is logically possible. That is, unless otherwise expressly stated, it is in no way intended that any method or aspect set forth herein be construed as requiring that its steps be performed in a specific order. Accordingly, where a method claim does not specifically state in the claims or descriptions that the steps are to be limited to a specific order, it is in no way intended that an order be inferred in any respect. This holds for any possible non¬ express basis for interpretation, including matters of logic with respect to the arrangement of steps or operational flow, plain meaning derived from grammatical organization or punctuation, or the number or type of aspects described in the specification.

[0073] All publications mentioned herein are incorporated herein by reference to disclose and describe the methods and / or materials in connection with which the publications are cited. The publications discussed herein are provided solely for their disclosure prior to the Attorney Docket No. 10063-110WO1

[0074] filing date of the present application. Further, the dates of publication provided herein can be different from the actual publication dates, which can require independent confirmation.

[0075] It is also to be understood that the terminology used herein is for the purpose of describing particular aspects only and is not intended to be limiting. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the disclosed compositions and methods belong. It can be further understood that terms, such as those defined in commonly used dictionaries, should be interpreted as having a meaning that is consistent with their meaning in the context of the specification and relevant art and should not be interpreted in an idealized or overly formal sense unless expressly defined herein.

[0076] Prior to describing the various aspects of the present disclosure, the following definitions are provided and should be used unless otherwise indicated. Additional terms may be defined elsewhere in the present disclosure.

[0077] Definitions

[0078] As used herein, “comprising” is to be interpreted as specifying the presence of the stated features, integers, steps, or components as referred to but does not preclude the presence or addition of one or more features, integers, steps, or components, or groups thereof. Moreover, each of the terms “by,” “comprising,” “comprises,” “comprised of,” “including,” “includes,” “included,” “involving,” “involves,” “involved,” and “such as” are used in their open, non-limiting sense and may be used interchangeably. Further, the term “comprising” is intended to include examples and aspects encompassed by the terms “consisting essentially of’ and “consisting of.” Similarly, the term “consisting essentially of’ is intended to include examples encompassed by the term “consisting of.

[0079] As used in the specification and the appended claims, the singular forms “a,” “an,” and “the” include plural referents unless the context clearly dictates otherwise.

[0080] It should be noted that ratios, concentrations, amounts, and other numerical data can be expressed herein in a range format. It can be further understood that the endpoints of each of the ranges are significant both in relation to the other endpoint and independently of the other endpoint. It is also understood that there are a number of values disclosed herein and that each value is also herein disclosed as “about” that particular value in addition to the value itself. For example, if the value “10” is disclosed, then “about 10” is also disclosed. Ranges can be expressed herein as from “about” one particular value and / or to “about” another particular value. Similarly, when values are expressed as approximations, by use of Attorney Docket No. 10063-110WO1

[0081] the antecedent “about,” it can be understood that the particular value forms a further aspect. For example, if the value “about 10” is disclosed, then “10” is also disclosed.

[0082] When a range is expressed, a further aspect includes from the one particular value and / or to the other particular value. For example, where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the disclosure, e.g., the phrase “x to y” includes the range from ‘x’ to ‘y’ as well as the range greater than ‘x’ and less than ‘y’. The range can also be expressed as an upper limit, e.g., ‘about x, y, z, or less’ and should be interpreted to include the specific ranges of ‘about x,’ ‘about y’, and ‘about z’ as well as the ranges of ‘less than x,’ less than y’, and ‘less than z’. Likewise, the phrase ‘about x, y, z, or greater’ should be interpreted to include the specific ranges of ‘about x,’ ‘about y,’ and ‘about z’ as well as the ranges of ‘greater than x,’ greater than y’, and ‘greater than z.’ In addition, the phrase “about ‘x’ to ‘y’,” where ‘x’ and ‘y’ are numerical values, includes “about ‘x’ to about ‘y’.”

[0083] It is to be understood that such a range format is used for convenience and brevity and, thus, should be interpreted in a flexible manner to include not only the numerical values explicitly recited as the limits of the range but also all the individual numerical values or sub-ranges encompassed within that range as if each numerical value and sub¬ range is explicitly recited. To illustrate, a numerical range of “about 0.1% to 5%” should be interpreted to include not only the explicitly recited values of about 0.1% to about 5% but also include individual values (e.g., about 1%, about 2%, about 3%, and about 4%) and the sub-ranges (e.g., about 0.5% to about 1.1%; about 5% to about 2.4%; about 0.5% to about 3.2%, and about 0.5% to about 4.4%, and other possible sub-ranges) within the indicated range.

[0084] As used herein, the terms “about,” “approximate,” “at or about,” and “substantially” mean that the amount or value in question can be the exact value or a value that provides equi valent results or effects as recited in the claims or taught herein. 'That is, it is understood that amounts, sizes, formulations, parameters, and other quantities and characteristics are not and need not be exact but may be approximate and / or larger or smaller, as desired, reflecting tolerances, conversion factors, rounding off, measurement error and the like, and other factors known to those of skill in the art such that equivalent results or effects are obtained. In some circumstances, the value that provides equivalent results or effects cannot be reasonably determined. In such cases, it is generally understood, as used herein, that “about” and “at or about” mean the nominal value indicated ±10% variation unless otherwise indicated or inferred. In general, an amount, size, formulation, parameter, or other Attorney Docket No. 10063-110WO1

[0085] quantity or characteristic is “about,” “approximate,” or “at or about” whether or not expressly stated to be such. It is understood that where “about,” “approximate,” or “at or about” is used before a quantitative value, the parameter also includes the specific quantitative value itself unless specifically stated otherwise.

[0086] As used herein, the term “therapeutically effective amount” refers to an amount that is sufficient to achieve the desired therapeutic result or to have an effect on undesired symptoms but is generally insufficient to cause adverse side effects. The specific therapeutically effective dose level for any particular patient will depend upon a variety of factors, including the disorder being treated and the severity of the disorder; the specific composition employed; the age, body weight, general health, sex, and diet of the patient; the time of administration; the route of administration; the rate of excretion of the specific compound employed; the duration of the treatment; drags used in combination or coincidental with the specific compound employed and like factors within the knowledge and expertise of the health practitioner and which may be well known in the medical arts. In the case of treating a particular disease or condition, in some instances, the desired response can be inhibiting the progression of the disease or condition. This may involve only slowing the progression of the disease temporarily. However, in other instances, it may be desirable to halt the progression of the disease permanently. Tills can be monitored by routine diagnostic methods known to one of ordinary skill in the art for any particular disease. The desired response to treatment of the disease or condition can also be delaying the onset or even preventing the onset of the disease or condition.

[0087] For example, it is well within the skill of the art to start doses of a compound at levels lower than those required to achieve the desired therapeutic effect and to gradually increase the dosage until the desired effect is achieved. If desired, the effective daily dose can be divided into multiple doses for purposes of administration. Consequently, single¬ dose compositions can contain such amounts or submultiples thereof to make up the daily dose. The dosage can be adjusted by the individual physician in the event of any contraindications. It is generally preferred that a maximum dose of the pharmacological agents of the disclosure (alone or in combination with other therapeutic agents) be used, that is, the highest safe dose according to sound medical judgment. It will be understood by those of ordinary skill in the art, however, that a patient may insist upon a lower dose or tolerable dose for medical reasons, psychological reasons, or virtually any other reasons.

[0088] A response to a therapeutically effective dose of a disclosed compound or composition can be measured by determining the physiological effects of the treatment or Attorney Docket No. 10063-110WO1

[0089] medication, such as the decrease or lack of disease symptoms following administration of the treatment or pharmacological agent. Other assays will be known to one of ordinary skill in the art and can be employed for measuring the level of the response. The amount of a treatment may be varied, for example, by increasing or decreasing the amount of a disclosed compound and / or pharmaceutical composition, changing the disclosed compound and / or pharmaceutical composition administered, changing the route of administration, changing the dosage timing, and so on. Dosage can vary and can be administered in one or more doses daily for one or several days. Guidance can be found in the literature for appropriate dosages for given classes of pharmaceutical products.

[0090] As used herein, the terms “optional” or “optionally” mean that the subsequently described event or circumstance can or cannot occur and that the description includes instances where said event or circumstance occurs and instances where it does not.

[0091] As used interchangeably herein, “subject,” “individual,” or “patient” can refer to a vertebrate organism, such as a mammal (e.g., human). " Subject" can also refer to a cell, a population of cells, a tissue, an organ, or an organism, preferably to humans and constituents thereof.

[0092] As used herein, the terms "treating" and "treatment" can refer generally to obtaining a desired pharmacological and / or physiological effect. The effect can be, but does not necessarily have to be, prophylactic in terms of preventing or partially preventing a disease, symptom, or condition thereof, such as a bacterial infections. The effect can be therapeutic in terms of a partial or complete cure of a disease, condition, symptom, or adverse effect attributed to the disease, disorder, or condition. The term "treatment" as used herein can include any treatment of a disorder in a subject, particularly a human, and can include any one or more of the following: (a) preventing the disease from occurring in a subject which may be predisposed to the disease but has not yet been diagnosed as having it; (b) inhibiting the disease, i.e., arresting its development; and (c) relieving the disease, i.e., mitigating or ameliorating the disease and / or its symptoms or conditions. The term "treatment," as used herein, can refer to both therapeutic treatment alone, prophylactic treatment alone, or both therapeutic and prophylactic treatment. Those in need of treatment (subjects in need thereof) can include those already with the disorder and / or those in which the disorder is to be prevented. As used herein, the term "treating" can include inhibiting the disease, disorder, or condition, e.g., impeding its progress, and relieving the disease, disorder, or condition, e.g., causing regression of the disease, disorder, or condition. Treating the disease, disorder, or condition can include ameliorating at least one symptom of the particular disease, disorder, Attorney Docket No. 10063-110WO1

[0093] or condition, even if the underlying pathophysiology is not affected, e.g., such as treating the pain of a subject by administration of an analgesic agent even though such agent does not treat the cause of the pain.

[0094] As used herein, “dose,” “unit dose,” or “dosage” can refer to physically discrete units suitable for use in a subject, each unit containing a predetermined quantity of a disclosed compound and / or a pharmaceutical composition thereof calculated to produce the desired response or responses in association with its administration.

[0095] As used herein, “therapeutic” can refer to heating, healing, and / or ameliorating a disease, disorder, condition, or side effect or to decreasing the rate of advancement of a disease, disorder, condition, or side effect.

[0096] Chemical Definitions

[0097] Compounds are described using standard nomenclature. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as is commonly understood by one of skill in the art to which this disclosure belongs.

[0098] The compounds described herein include enantiomers, mixtures of enantiomers, diastereomers, tautomers, racemates, and other isomers, such as retainers, as if each is specifically described unless otherwise indicated or otherwise excluded by context. It is to be understood that the compounds provided herein may contain chiral centers. Such chiral centers may be of either the ( / ?-) or (S'-) configuration. The compounds provided herein may either be enantiomerically pure or diastereomeric or enantiomeric mixtures. It is to be understood that the chiral centers of the compounds provided herein may undergo epimerization in vivo. As such, one of skill in the art will recognize that administration of a compound in its ( / ?-) form is equivalent, for compounds that undergo epimerization in vivo, to administration of the compound in its (S-) form. Unless stated to the contrary, a formula with chemical bonds shown only as solid lines and not as wedges or dashed lines contemplates each possible isomer, e.g., each enantiomer, diastereomer, and meso compound, and a mixture of isomers, such as a racemic or scalemic mixture.

[0099] Compounds described herein may contain one or more double bonds and, thus, potentially give rise to cis / trans (E / Z) isomers, as well as other conformational isomers. Unless stated to the contrary, all such possible isomers are contemplated, as well as mixtures of such isomers.

[0100] Compounds described herein may also present as an equilibrium of tautomers. For example, ketones with an a-hydrogen can exist in an equilibrium of the keto form and the enol form. Likewise, amides with an N-hydrogen can exist in an equilibrium of the amide Attorney Docket No. 10063-110WO1

[0101] form and the iniidic acid form. Unless stated to the contrary, all possible tautomers of the compounds described herein are contemplated.

[0102] A dash

[0103]

[0104] that is not between two letters or symbols is used to indicate a point of attachment for a substituent. For example, -(C=O)NH2 is attached through the carbon of the keto (C=O) group.

[0105] _ i

[0106] As used herein, the symbol “? ” (which hereinafter can be referred to as “a point of attachment bond”) denotes a bond that is a point of attachment between two chemical entities, one of which is depicted as being attached to the point of attachment bond and the other of which is not depicted as being attached to the point of attachment bond. For y y --- example, “ « ” indicates that the chemical entity “XY” is bonded to another chemical entity via the point of attachment bond. Furthermore, the specific point of attachment to the non-depicted chemical entity can be specified by inference. For example, the compound XY _ i

[0107] CH3-R3, wherein R3is H or “?,” infers that when R3is “XY,” the point of attachment bond is the same bond as the bond by which R3is depicted as being bonded to CII3.

[0108] The term “substituted,” as used herein, means that any one or more hydrogens on the designated atom or group are replaced with a moiety selected from the indicated group, provided that the designated atom’s normal valence is not exceeded and the resulting compound is stable. For example, when the substituent is oxo (i.e., =0), two hydrogens on the atom are replaced. For example, a pyridyl group substituted by oxo is a pyridine.

[0109] Combinations of substituents and / or variables are permissible only if such combinations result in stable compounds or useful synthetic intermediates. A stable active compound refers to a compound that can be isolated and can be formulated into a dosage form with a shelf life of at least one month. A stable manufacturing intermediate or precursor to an active compound is stable if it does not degrade within the period needed for reaction or other use. A stable moiety or substituent group is one that does not degrade, react, or fall apart within the period necessary for use. Non-limiting examples of unstable moieties are those that combine heteroatoms in an unstable arrangement, as typically known and identifiable to those of skill in the art.

[0110] Any suitable group may be present on a “substituted” or “optionally substituted” position that forms a stable molecule and meets the desired purpose of the disclosure and includes, but is not limited to: alkyl, haloalkyl, alkoxy, alkenyl, alkynyl, aryl, heteroaryl, Attorney Docket No. 10063-110WO1

[0111] cycloalkyl, heterocycle, aldehyde, amino, carboxylic acid, ester, ether, halo, hydroxy, keto, nitro, cyano, azido, oxo, silyl, sulfo-oxo, sulfonyl, sulfone, sulfoxide, sulfonylamino, or thiol.

[0112] The terms for various functional groups as used herein are not intended to be limited to monovalent radicals and may include polyvalent radical groups as appropriate, such as divalent, tri valent, tetravalent, pentavalent, and hexavalent groups, and the like, based on the position and location of such groups in the compounds described herein as would be readily understood by the skilled person in the context in which said functional groups are recited.

[0113] “Alkyl” is a straight chain or branched saturated aliphatic hydrocarbon group. In certain aspects, the alkyl is C1-C2, C1-C3, or Ci-Ce (i.e., the alkyl chain can be 1, 2, 3, 4, 5, or 6 carbons in length). The specified ranges used herein indicate an alkyl group with length of each member of the range described as an independent species. For example, Ci-Cealkyl as used herein indicates an alkyl group having from 1, 2, 3, 4, 5, or 6 carbon atoms and is intended to mean that each of these is described as an independent species and Ci-C ralkyl as used herein indicates an alkyl group having from 1, 2, 3, or 4 carbon atoms and is intended to mean that each of these is described as an independent species. When Co- Cnalkyl is used herein in conjunction with another group, for example (C3-C?cycloalkyI)Co-C4alkyl, or -Co-CrCCs-Cvcycloalkyl), the indicated group, in this case cycloalkyl, is either directly bound by a single covalent bond (Coalkyl), or attached by an alkyl chain, in this case 1, 2, 3, or 4 carbon atoms. Alkyls can also be attached via other groups such as heteroatoms, as in -O-Co-CralkyKCs-Cvcycloalkyl). Examples of alkyl include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, t-butyl, n-pentyl, isopentyl, tert-pentyl, neopentyl, n-hexyl, 2-methylpentane, 3-methylpentane, 2,2- dimethylbutane, and 2,3-dimethylbutane. In one aspect, the alkyl group is optionally substituted as described herein.

[0114] The term “heteroalkyl” refers to straight chain or branched alkyl group having one or more additional heteroatoms in the backbone, where the heteroatoms may be selected from N, O, and S. An example of a heteroalkyl group is an ethoxyethylether, methoxyethylether, and the like.

[0115] “Cycloalkyl” is a saturated mono- or multi-cyclic hydrocarbon ring system. When composed of two or more rings, the rings may be joined together in a fused or bridged fashion. Non-limiting examples of typical cycloalkyl groups include cyclopropyl, Attorney Docket No. 10063-110WO1

[0116] cyclobutyl, cyclopentyl, cyclohexyl, and cycloheptyl. In one aspect, the cycloalkyl group is optionally substituted as described herein.

[0117] The term “heterocycloalkyl” refers to a cycloalkyl group having one or more additional heteroatoms in the backbone, where the heteroatoms may be selected from N, O, and S. Examples of a heterocycloalkyl group is a pyran, morpholine, piperadine, piperazine, pyrazine, pyrrolidine, and the like.

[0118] “Alkenyl” is a straight or branched chain aliphatic hydrocarbon group having one or more carbon-carbon double bonds, each of which is independently either cis or trans, that may occur at a stable point along the chain. Non-limiting examples include Cz-CTalkenyl and C2-Cealkenyl (i.e., having 2, 3, 4, 5, or 6 carbons). The specified ranges as used herein indicate an alkenyl group having each member of the range described as an independent species, as described above for the alkyl moiety. Examples of alkenyl include, but are not limited to, ethenyl and propenyl. In one aspect, the alkenyl group is optionally substituted as described herein.

[0119] A “heteroalkenyl” refers to an alkenyl group having one or more additional heteroatoms in the backbone, where the heteroatoms may be selected from N, O, and S.

[0120] “Alkynyl” is a straight or branched chain aliphatic hydrocarbon group having one or more carbon-carbon triple bonds that may occur at any stable point along the chain, for example, C2-C4alkynyl or C -Cealkynyl (i.e., having 2, 3, 4, 5, or 6 carbons). The specified ranges as used herein indicate an alkynyl group having each member of the range described as an independent species, as described above for the alkyl moiety. Examples of alkynyl include, but are not limited to, ethynyl, propynyl, 1-butynyl, 2-butynyl, 3-butynyl, 1-pentynyl, 2-pentynyl, 3-pentynyl, 4-pentynyl, 1 -hexynyl, 2 -hexynyl, 3-hexynyl, 4-hexynyl, and 5-hexynyl. In one aspect, the alkynyl group is optionally substituted as described herein.

[0121] “Alkoxy” is an alkyl group as defined above covalently bound through an oxygen bridge (-O-). Examples of alkoxy include, but are not limited to, methoxy, ethoxy, n- propoxy, isopropoxy, n-butoxy, 2-butoxy, tert-butoxy, n-pentoxy, 2 -pentoxy, 3-pentoxy, isopentoxy, neopentoxy, n-hexoxy, 2 -hexoxy, 3-hexoxy, and 3-methylpentoxy. Similarly, an “alkylthio” or “thioalkyl” group is an alkyl group as defined above with the indicated number of carbon atoms covalently bound through a sulfur bridge (-S-). In one aspect, the alkoxy group is optionally substituted as described herein.

[0122] “Alkanoyl” is an alkyl group as defined above covalently bound through a carbonyl (C=O) bridge. The carbonyl carbon is included in the number of carbons, for example Attorney Docket No. 10063-110WO1

[0123] C2alkanoyi is a CH?(C=O)- group. In one aspect, the alkanoyl group is optionally substituted as described herein.

[0124] “Halo” or “halogen” indicates, independently, any of fluoro, chloro, bromo or iodo. “Aryl” indicates an aromatic group containing only carbon in the aromatic ring or rings. In one aspect, the aryl group contains 1 to 3 separate or fused rings and is 6 to 14 or 18 ring atoms, without heteroatoms as ring members. When indicated, such aryl groups may be further substituted with carbon or non-carbon atoms or groups. Such substitution may include fusion to a 4- to 7- or 5- to 7 -membered saturated or partially unsaturated cyclic group that optionally contains 1, 2, or 3 heteroatoms independently selected from N, O, B, P, Si and S. to form, for example, a 3,4-methylenedioxyphenyl group. Aryl groups include, for example, phenyl and naphthyl, including 1 -naphthyl and 2-naphthyl. In one aspect, aryl groups are pendant. An example of a pendant ring is a phenyl group substituted with a phenyl group. In one aspect, the aryl group is optionally substituted as described herein.

[0125] The term “heterocycle” refers to saturated and partially saturated heteroatom¬ containing ring radicals, where the heteroatoms may be selected from N, O, and S. The term heterocycle includes monocyclic 3-12 members rings, as well as bicyclic 5-16 membered ring systems ( which can include fused, bridged, or spiro bicyclic ring systems). It does not include rings containing -O-O-, -O-S-, and -S-S- portions. Examples of saturated heterocycle groups including saturated 4- to 7-membered monocyclic groups containing 1 to 4 nitrogen atoms [e.g., pyrrolidinyl, imidazolidinyl, piperidinyl, pyrrolinyl, azetidinyl, piperazinyl, and pyrazolidinyl]; saturated 4- to 6-membered monocyclic groups containing 1 to 2 oxygen atoms and 1 to 3 nitrogen atoms [e.g,, morpholinyl]; and saturated 3- to 6-membered heteromonocyclic groups containing 1 to 2 sulfur atoms and 1 to 3 nitrogen atoms [e.g., thiazolidinyl], Examples of partially saturated heterocycle radicals include, but are not limited, dihydrothienyl, dihydropyranyl, dihydrofuryl, and dihydrothiazolyl.

[0126] Examples of partially saturated and saturated heterocycle groups include, but are not limited to, pyrrolidinyl, imidazolidinyl, piperidinyl, pyrrolinyl, pyrazolidinyl, piperazinyl, morpholinyl, tetrahydropyranyl, thiazolidinyl, dihydrothienyl, 2,3-dihydro-benzo[l,4]dioxanyl, indolinyl, isoindolinyl, dihydrobenzothienyl, dihydrobenzofuryl, isochromanyl, chromanyl, 1,2-dihydroquinolyl, 1,2,3,4-tetrahydro-isoquinolyl, 1, 2,3,4-tetraliydro-quinolyl, 2,3,4,4a,9,9a-hexaliydro-lH-3-aza-fluorenyl, 5, 6,7 -trihydro-1, 2,4-triazolo[3,4-a]isoquinolyl, 3,4-dihydro-2H-benzo[ 1,4]oxazinyl, benzo[l,4]dioxanyl, 2,3,- dihydro-! II-benzo[d]isothazol-6-yl, dihydropyranyl, dihydrofuryl, and dihydrothiazolyl. Bicyclic heterocycle includes groups wherein the heterocyclic radical is fused with an aryl Attorney Docket No. 10063-110WO1

[0127] radical wherein the point of attachment is the heterocycle ring. Bicyclic heterocycle also includes heterocyclic radicals that are fused with a carbocyclic radical. Representative examples include, but are not limited to, partially unsaturated condensed heterocyclic groups containing 1 to 5 nitrogen atoms, for example indoline and isoindoline, partially unsaturated condensed heterocyclic groups containing 1 to 2 oxygen atoms and 1 to 3 nitrogen atoms, partially unsaturated condensed heterocyclic groups containing 1 to 2 sulfur atoms and 1 to 3 nitrogen atoms, and saturated condensed heterocyclic groups containing 1 to 2 oxygen or sulfur atoms.

[0128] “Heteroaryl” refers to a stable monocyclic, bicyclic, or multicyclic aromatic ring that contains from 1 to 4, or in some aspects 1, 2, or 3 heteroatoms selected from N, 0, S, B, and P (and typically selected from N, O, and S) with remaining ring atoms being carbon, or a stable bicyclic or tricyclic system containing at least one 5, 6, or 7 membered aromatic ring which contains from 1 to 4, or in some aspects from 1 to 3 or from 1 to 2, heteroatoms selected from N, (), S, B, or P, with remaining ring atoms being carbon. In one aspect, the only heteroatom is nitrogen. In one aspect, the only heteroatom is oxygen. In one aspect, the only heteroatom is sulfur. Monocyclic heteroaryl groups typically have from 5 to 6 ring atoms. In some aspects, bicyclic heteroaryl groups are 8- to 10-membered heteroaryl groups, such as groups containing 8 or 10 ring atoms in which one 5-, 6-, or 7-membered aromatic ring is fused to a second aromatic or non-aromatic ring, wherein the point of attachment is the aromatic ring. When the total number of S and O atoms in the heteroaryl group excess 1, these heteroatoms are not adjacent to one another. In one aspect, the total number of S and O atoms in the heteroaryl group is not more than 2. In another aspect, the total number of S and O atoms in the heteroaryl group is not more than 1. Examples of heteroaryl groups include, but are not limited to, pyridinyl, imidazolyl, imidazopyridinyl, pyrimidinyi, pyrazoiyl, triazolyl, pyrazinyl, furyl, thienyl, isoxazolyl, thiazolyl, oxadiazolyl, oxazolyl, isothiazolyl, pyrrolyl, quinolinyl, isoquinolinyl, tetrahydroisoquinolinyl, indolyl, benzimidazolyl, benzofuranyl, cinnolinyl, indazolyl, indolizinyl, phthalazinyl, pyridazinyl, triazinyl, isoindolyl, pteridinyl, purinyl, triazolyl, thiadiazolyl, furazanyl, benzofurazanyl, benzothiophenyl, benzothiazolyl, benzoxazolyl, quinazolinyl, quinoxalinyl, naphthyridinyl, and furopyridinyl.

[0129] A “pharmaceutically acceptable salt” is a derivative of the disclosed compound in which the parent compound is modified by making inorganic and organic, pharmaceutically acceptable, acid or base addition salts thereof. The salts of the present compounds can be synthesized from a parent compound that contains a basic or acidic moiety by conventional Attorney Docket No. 10063-110WO1

[0130] chemical methods. Generally, such salts can be prepared by reacting free acid forms of these compounds with a stoichiometric amount of the appropriate base (such as Na, Ca, Mg, or K hydroxide, carbonate, bicarbonate, or the like) or by reacting free base forms of these compounds with a stoichiometric amount of the appropriate acid. Such reactions are typically carried out in water, an organic solvent, or a mixture of the two. Generally, non¬ aqueous media like ether, ethyl acetate, ethanol, isopropanol, or acetonitrile are typical, where practicable. Salts of the present compounds further include solvates of the compounds and of the compound salts. Examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic residues such as amines; alkali or organic salts of acidic residues such as carboxylic acids; and the like. The pharmaceutically acceptable salts include salts that are acceptable for human consumption and the quaternary ammonium salts of the parent compound formed, for example, from inorganic or organic salts. Example of such salts include, but are not limited to, those derived from inorganic acids such as hydrochloric, hydrobromic, sulfuric, sulfamic, phosphoric, nitric, and the like; and the salts prepared from organic acids such as acetic, propionic, succinic, glycolic, stearic, lactic, malic, tartaric, citric, ascorbic, palmoic, maleic, hydroxymaleic, phenylacetic, glutamic, benzoic, salicyclic, mesylic, esylic, besylic, sulfanilic, 2-acetoxybenzoic, fumaric, toluenesulfonic, methanesulfonic, ethane disulfonic, oxalic, isethionic, HOOC-(CH2)I-4-COOH, and the like, or using a different acid that produced the same counterion. Lists of additional suitable salts may be found, e.g., in Remington’s Pharmaceutical Sciences, 17thed., Mack Publishing Company, Easton, PA., p.

[0131] 1418 (1985).

[0132] As used herein, the term “derivative” refers to a compound having a structure derived from the structure of a parent compound (e.g., a compound disclosed herein) and whose structure is sufficiently similar to those disclosed herein and based upon that similarity would be expected by one skilled in the art to exhibit the same or similar activities and utilities as the claimed compounds, or to induce, as a precursor, the same or similar activities and utilities as the claimed compound. Exemplary derivatives include but are not limited to, salts, esters, amides, salts of esters or amides, and N-oxides of a parent compound.

[0133] As used herein, substantially pure means sufficiently homogeneous to appear free of readily detectable impurities as determined by standard methods of analysis, such as thin layer chromatography (TLC), nuclear magnetic resonance (NMR), gel electrophoresis, high performance liquid chromatography (HPLC) and mass spectrometry (MS), gas- Attorney Docket No. 10063-110WO1

[0134] chromatography mass spectrometry (GC-MS), and similar, used by those of skill in the art to assess such purity, or sufficiently pure such that further purification would not delectably alter the physical and chemical properties, such as enzymatic and biological activities, of the substance. Both traditional and modern methods for purification of the compounds to produce substantially chemically pure compounds are known to those of skill in the art. A substantially chemically pure compound may, however, be a mixture of stereoisomers.

[0135] Certain materials, compounds, compositions, and components disclosed herein can be obtained commercially or readily synthesized using techniques generally known to those of skill in the art. For example, the starting materials and reagents used in preparing the disclosed compounds and compositions are either available from commercial suppliers such as Sigma- Aldrich (formally MilliporeSigma, Burlington, MA) or Thermo Fisher Scientific Inc. (Waltham, MA) or are prepared by methods known to those skilled in the art following procedures set forth in references such as Fieser and Fieser's Reagents for Organic Synthesis (John Wiley and Sons, 2007); Organic Reactions (John Wiley and Sons, 2004); March's Advanced Organic Chemistry, (John Wiley and Sons, 8thEdition); and Larock's Comprehensive Organic Transformations (John Wiley and Sons, 3rdedition, 2017).

[0136] Compounds

[0137] Disclosed herein, in one aspect, are compounds having Formula I or II:

[0138]

[0139] wherein,

[0140] n is 0 to 4;

[0141] — - is a bond that is present or absent,

[0142] X1is O or S;

[0143] X2is S, CH2, or CHR4;

[0144] X3and X4are, independently, CH2, O, S, NH, NR4, CHR4, CR4, or C(R4)2;

[0145] R1and R2are, independently, H, branched or straight chain Ci-C4alkyl, aryl, heteroaryl, (Ci- C4alkyl)aryl, (Ci-C4alkyl)heteroaryl, Cs-Cecycloalkyl, -Cecycloheteroalkyl, (Ci- C4alkyl)C3-C6cycloalkyl, or (Ci-C4alkyl)C3-C6cycloheteroalkyl, wherein R1and R2are not both H; Attorney Docket No. 10063-110WO1

[0146] R3is branched or straight chain C1-C12alkyl, C1-C12heteroalkyl, aryl, heteroaryl, (Ci- C4alkyl)aryl, (Ci-C4alkyl)heteroaryl, Cs-Cecycloalkyl, Cs-Cecycloheteroalkyl, (Ci- C4alkyl)C3-C6cycloalkyl, or (Ci-C4alkyl)C3-C6cycloheteroalkyl;

[0147] R4is branched or straight chain Ci-C4alkyl, C2-C4alkenyl, Ci-C4heteroalkyl, C2-C4heteroalkenyl, or two R4’s bond together to form a fused Cs-Cecycloalkyl, fused Cs-Cecycloheteroalkyl, fused aryl, fused heteroaryl, spiro Cs-Cecycloalkyl, or spiro C2-Cecycloheteroalkyl, any of which are optionally substituted with oxo, sulfonate, sulfonyl, halo, hydroxyl, thiol, carboxylate, carbonyl, Ci-Cealkyl, or Ci-Cghaloalkyl; or a pharmaceutically acceptable salt thereof;

[0148] or a pharmaceutically acceptable salt thereof.

[0149] In a specific example, the disclosed compounds do not include a compound of Formula II where X1and X2are both S, R3is Csalkyl, n is 1, and X3and X4are both CH2? In one example, in either Formula I or II, X1is O. In another example, X1is S. In one example of Formula I or II, X2is S. In another example, X2is CH2.

[0150] In some examples of Formula 1 or II, R is straight chain C3-C10 alkyl, for example propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, or decyl. In other examples, R3is Ci- C4alkyl)aryl, such a benzyl or ethylphenyl.

[0151] In some examples in Formula I, R1and R are both branched or straight chain Ci- C4alkyl, e.g., methyl, ethyl, isopropyl, n-propyl. In another, one of R1or R2is (Ci- C4alkyl)aryl, e.g., benzyl or ethylphenyl. In other examples, R1or R2is H.

[0152] In one example of Formula 11, n is 1. In another example, n is 2. In yet another example, n is 3. In still another example, n is 4. In other examples, n is 0. In one example of Formula II, — is a bond that is present. In another example, — - is a bond that is absent.

[0153] In some examples of Formula II, X3and X4are both CH2. So when n is 1, the compound contains a pyrrolidine, and when n is 2, the compound contains a piperidine. In one example of Formula II, X3is O. So when n is 2 and X4is CH2, the compound contains a morpholine. In other examples, X3and X4are both CHR4and the two R4’s bond together to form a fused Cs-Cgcycloalkyl. In still other examples, X3and X4are both CR4and the two R4’s bond together to form a fused aryl. In still another example, X4is C(R4)2. and the two R4’s bond together to form a spiro Cs-Cecycloheteroalkyl.

[0154] In specific examples, the compound is selected from Attorney Docket No. 10063-110WO1

[0155] DIS8-2 DIS10-2

[0156] S

[0157] DIS15-2

[0158]

[0159] DIS3-3 DIS4-3 DIS5-3 DIS6-3 Attorney Docket No. 10063-110WO1

[0160]

[0161] Attorney Docket No. 10063-110WO1

[0162] LHM"14LHM-15

[0163] LHM-20 Attorney Docket No. 10063-110WO1 LHM-26

[0164] LHM-33

[0165]

[0166] Attorney Docket No. 10063-110WO1

[0167]

[0168] L. HM-53

[0169] Disclosed herein, in another aspect, are compounds having Formula III: Attorney Docket No. 10063-110WO1

[0170]

[0171] SO3H

[0172] III

[0173] wherein,

[0174] L is null, -NHC(O)C(CH3)2-, -NR5C(O)C(CH3)2-, or -OC(O)C(CH3)2-;

[0175] R5is branched or straight chain Ci-Ci2alkyl, Ci-Cnheteroalkyl, aryl, heteroaryl, (Ci- C4alkyl)aryl, (Ci-CAalkyl)heteroaryl, C3-C6cycloalkyl, C3-C6cycloheteroalkyl, (Ci- C4alkyl)C3-C6cycloalkyl, or (Ci-C4alkyl)C3-C6cycloheteroalkyl;

[0176] or a pharmaceutically acceptable salt thereof.

[0177] In one example, L is null; that is R5is bound directly to O. In another example, L is -NHC(O)C(CH3)2-. In another example, L is -NR5C(O)C(CH3)2-. In another example, L is -OC(O)C(CH3)2-.

[0178] In some examples, R is branched or straight chain C2-Cioalkyl or C3-Cioheteroalkyl, e.g., propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, isopropyl, secbutyl, tertbutyl, 1-propylbutyl, ethyloxyethyl, or methoxyethoxylethyl. In other examples, R3is C3- Cscycloalkyl or C3-C6cycloheteroalkyl, e.g., cyclopentyl, cyclohexyl, pyrrolinyl, or piperidinyl. In still other examples, R5is (Ci-C4alkyl)C3-C6Cycloalkyl or (Ci-C4alkyl)C3-Cecycloheteroalkyl. In yet other examples, R5is (Ci-C4alkyl)aryl, e.g., benzyl or ethylphenyl. In one example, when L is -NR5C(O)C(CH3)2-, each R5is a branched or straight chain C2-Csalkyl.

[0179] In specific examples, the compound is selected from Attorney Docket No. 10063-110WO1

[0180] AZ1 AZ2 AZ4

[0181] AZ5 AZ6 AZ7

[0182] OH AZ8 AZ9 AZ10

[0183]

[0184] AZ11 AZ12 Attorney Docket No. 10063-110WO1 AZ1-2 AZ2-2

[0185] AZ5-2 AZ6-2

[0186]

[0187] Pharmaceutically acceptable salts of the above compounds. Attorney Docket No. 10063-110WO1

[0188] Pharmaceutical Compositions

[0189] The compounds as used in the methods described herein can be administered by any suitable method and technique presently or prospectively known to those skilled in the art. For example, the active components described herein can be formulated in a physiologically- or pharmaceutically-acceptable form and administered by any suitable route known in the art, including, for example, oral and parenteral routes of administering. As used herein, the term “parenteral” includes subcutaneous, intradermal, intravenous, intramuscular, intraperitoneal, and intrasternai administration, such as by injection.

[0190] Administration of the active components of their compositions can be a single administration, or at continuous and distinct intervals as can be readily determined by a person skilled in the art.

[0191] Compositions, as described herein, comprising an active compound and a pharmaceutically acceptable carrier or excipient of some sort, may be useful in a variety of medical and non-medical applications. For example, pharmaceutical compositions comprising an active compound and an excipient may be useful for the treatment or prevention of a bacterial infection in a subject in need thereof.

[0192] " Pharmaceutically acceptable carrier" (sometimes referred to as a "carrier") means a carrier or excipient that is useful in preparing a pharmaceutical or therapeutic composition that is generally safe and non-toxic and includes a carrier that is acceptable for veterinary and / or human pharmaceutical or therapeutic use. The terms "carrier" or "pharmaceutically acceptable carrier" can include, but are not limited to, phosphate buffered saline solution, water, emulsions (such as an oil / water or water / oil emulsion), and / or various types of wetting agents. As used herein, the term "carrier" encompasses, but is not limited to, any excipient, diluent, filler, salt, buffer, stabilizer, solubilizer, lipid, stabilizer, or other material well-known in the art for use in pharmaceutical formulations and as described further herein.

[0193] “Excipients” include any and all solvents, diluents or other liquid vehicles, dispersion or suspension aids, surface active agents, isotonic agents, thickening or emulsifying agents, preservatives, solid binders, lubricants, and the like, as suited to the particular dosage form desired. General considerations in formulation and / or manufacture can be found, for example, in Remington's Pharmaceutical Sciences, Sixteenth Edition, E. W. Martin (Mack Publishing Co., Easton, Pa., 1980), and Remington: The Science and Practice of Pharmacy, 21st Edition (Lippincott Williams & Wilkins, 2005). Attorney Docket No. 10063-110WO1

[0194] Exemplary excipients include but are not limited to, any non-toxic, inert solid, semisolid or liquid filler, diluent, encapsulating material or formulation auxiliary of any type. Some examples of materials which can serve as excipients include, but are not limited to, sugars such as lactose, glucose, and sucrose; starches such as com starch and potato starch; cellulose and its derivatives such as sodium carboxymethyl cellulose, ethyl cellulose, and cellulose acetate; powdered tragacanth; malt; gelatin; talc; excipients such as cocoa butter and suppository waxes; oils such as peanut oil, cottonseed oil; safflower oil; sesame oil; olive oil; com oil and soybean oil; glycols such as propylene glycol; esters such as ethyl oleate and ethyl laurate; agar; detergents such as Tween 80; buffering agents such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; Ringer's solution; ethyl alcohol; and phosphate buffer solutions, as well as other non¬ toxic compatible lubricants such as sodium lauryl sulfate and magnesium stearate, as well as coloring agents, releasing agents, coating agents, sweetening, flavoring and perfuming agents, preservatives and antioxidants can also be present in the composition, according to the judgment of the formulator. As would be appreciated by one of skill in this art, the excipients may be chosen based on what the composition is useful for. For example, with a pharmaceutical composition or cosmetic composition, the choice of the excipient will depend on the route of administration, the agent being delivered, time course of delivery of the agent, etc., and can be administered to humans and / or to animals, orally, rectally, paren terally, intracisternally, intravaginally, intranasally, intraperitoneally, topically (as by powders, creams, ointments, or drops), buccally, or as an oral or nasal spray. In some aspects, the active compounds disclosed herein are administered topically.

[0195] Exemplary diluents include calcium carbonate, sodium carbonate, calcium phosphate, dicalcium phosphate, calcium sulfate, calcium hydrogen phosphate, sodium phosphate lactose, sucrose, cellulose, microcrystalline cellulose, kaolin, mannitol, sorbitol, inositol, sodium chloride, dry starch, cornstarch, powdered sugar, etc., and combinations thereof.

[0196] Exemplary granulating and / or dispersing agents include potato starch, corn starch, tapioca starch, sodium starch glycolate, clays, alginic acid, guar gum, citrus pulp, agar, bentonite, cellulose and wood products, natural sponge, cation-exchange resins, calcium carbonate, silicates, sodium carbonate, cross-linked poly(vinyl-pyrrolidone) (crospovidone), sodium carboxymethyl starch (sodium starch glycolate), carboxymethyl cellulose, cross¬ linked sodium carboxymethyl cellulose (croscarmellose), methylcellulose, pregelatinized starch (starch 1500), microcrystalline starch, water insoluble starch, calcium carboxymethyl Attorney Docket No. 10063-110WO1

[0197] cellulose, magnesium aluminum silicate (Veegum), sodium lauryl sulfate, quaternary ammonium compounds, etc., and combinations thereof.

[0198] Exemplary surface active agents and / or emulsifiers include natural emulsifiers (e.g. acacia, agar, alginic acid, sodium alginate, tragacanth, chondrux, cholesterol, xanthan, pectin, gelatin, egg yolk, casein, wool fat, cholesterol, wax, and lecithin), colloidal clays (e.g. bentonite [aluminum silicate] and Veegum [magnesium aluminum silicate]), long chain amino acid derivatives, high molecular weight alcohols (e.g. stearyl alcohol, cetyl alcohol, oleyl alcohol, triacetin monostearate, ethylene glycol distearate, glyceryl monostearate, and propylene glycol monostearate, polyvinyl alcohol), carbomers (e.g. carboxy polymethylene, polyacrylic acid, acrylic acid polymer, and carboxy vinyl polymer), carrageenan, cellulosic derivatives (e.g. carboxymethylcellulose sodium, powdered cellulose, hydroxymethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methylcellulose, methylcellulose), sorbitan fatty acid esters (e.g. polyoxyethylene sorbitan monolaurate [Tween 20], polyoxyethylene sorbitan [Tween 60], polyoxyethylene sorbitan monooleate [Tween 80], sorbitan monopalmitate [Span 40], sorbitan monostearate [Span 60], sorbitan tristearate [Span 65], glyceryl monooleate, sorbitan monooleate [Span 80]), polyoxyethylene esters (e.g. polyoxyethylene monostearate [Myrj 45], polyoxyethylene hydrogenated castor oil, polyethoxylated castor oil, polyoxymethylene stearate, and Solutol), sucrose fatty acid esters, polyethylene glycol fatly acid esters (e.g. Cremophor), polyoxyethylene ethers, (e.g. polyoxyethylene lauryl ether [Brij 30]), poly( vinyl-pyrrolidone), diethylene glycol monolaurate, triethanolamine oleate, sodium oleate, potassium oleate, ethyl oleate, oleic acid, ethyl laurate, sodium lauryl sulfate, Pluronic F 68, Poloxamer 188, cetrimonium bromide, cetylpyridinium chloride, benzalkonium chloride, docusate sodium, etc. and / or combinations thereof. Exemplary binding agents include starch (e.g., cornstarch and starch paste), gelatin, sugars (e.g., sucrose, glucose, dextrose, dextrin, molasses, lactose, lactitol, mannitol, etc.), natural and synthetic gums (e.g., acacia, sodium alginate, extract of Irish moss, panwar gum, ghatti gum, mucilage of isapol husks, carboxymethylcellulose, methylcellulose, ethylcellulose, hydroxyethylcellulose, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, microcrystalline cellulose, cellulose acetate, polyvinylpyrrolidone), magnesium aluminum silicate (Veegum), and larch arabogalactan), alginates, polyethylene oxide, polyethylene glycol, inorganic calcium salts, silicic acid, polymethacrylates, waxes, water, alcohol, etc., and / or combinations thereof. Attorney Docket No. 10063-110WO1

[0199] Exemplary preservatives include antioxidants, chelating agents, antimicrobial preservatives, antifungal preservatives, alcohol preservatives, acidic preservatives, and other preservatives.

[0200] Exemplary antioxidants include alpha tocopherol, ascorbic acid, ascorbyl palmitate, butylated hydroxyanisole, butylated hydroxytoluene, monothioglycerol, potassium metabisulfite, propionic acid, propyl gallate, sodium ascorbate, sodium bisulfite, sodium metabisulfite, and sodium sulfite.

[0201] Exemplary chelating agents include ethylenediaminetetraacetic acid (EDTA) and salts and hydrates thereof (e.g., sodium edetate, disodium edetate, trisodium edetate, calcium disodium edetate, dipotassium edetate, and the like), citric acid and salts and hydrates thereof (e.g., citric acid monohydrate), fumaric acid and salts and hydrates thereof, malic acid and salts and hydrates thereof, phosphoric acid and salts and hydrates thereof, and tartaric acid and salts and hydrates thereof. Exemplary antimicrobial preservatives include benzalkonium chloride, benzethonium chloride, benzyl alcohol, bronopol, cetrimide, cetylpyridinium chloride, chlorhexidine, chlorobutanol, chlorocresol, chloroxylenol, cresol, ethyl alcohol, glycerin, hexetidine, imidurea, phenol, phenoxyethanol, phenylethyl alcohol, phenylmercuric nitrate, propylene glycol, and thimerosal.

[0202] Exemplary antifungal preservatives include butyl paraben, methyl paraben, ethyl paraben, propyl paraben, benzoic acid, hydroxybenzoic acid, potassium benzoate, potassium sorbate, sodium benzoate, sodium propionate, and sorbic acid.

[0203] Exemplary alcohol preservatives include ethanol, polyethylene glycol, phenol, phenolic compounds, bisphenol, chlorobutanol, hydroxybenzoate, and phenylethyl alcohol.

[0204] Exemplary acidic preservatives include vitamin A, vitamin C, vitamin E, betacarotene, citric acid, acetic acid, dehydroacetic acid, ascorbic acid, sorbic acid, and phytic acid. Other preservatives include tocopherol, tocopherol acetate, deteroxime mesylate, cetrimide, butylated hydroxyanisol (BHA), butylated hydroxy toluene (BHT), ethylenediamine, sodium lauryl sulfate (SLS), sodium lauryl ether sulfate (SEES), sodium bisulfite, sodium metabisulfite, potassium sulfite, potassium metabisulfite, Glydant Plus, Phenonip, methylparaben, Germall 115, Germaben II, NeoIone, Kathon, and Euxyl. In certain aspects, the preservative is an anti-oxidant. In other aspects, the preservative is a chelating agent.

[0205] Exemplary buffering agents include citrate buffer solutions, acetate buffer solutions, phosphate buffer solutions, ammonium chloride, calcium carbonate, calcium chloride, calcium citrate, calcium glubionate, calcium gluceptate, calcium gluconate, D-gluconic Attorney Docket No. 10063-110WO1

[0206] acid, calcium glycerophosphate, calcium lactate, propanoic acid, calcium levulinate, pentanoic acid, dibasic calcium phosphate, phosphoric acid, tribasic calcium phosphate, calcium hydroxide phosphate, potassium acetate, potassium chloride, potassium gluconate, potassium mixtures, dibasic potassium phosphate, monobasic potassium phosphate, potassium phosphate mixtures, sodium acetate, sodium bicarbonate, sodium chloride, sodium citrate, sodium lactate, dibasic sodium phosphate, monobasic sodium phosphate, sodium phosphate mixtures, tromethamine, magnesium hydroxide, aluminum hydroxide, alginic acid, pyrogen-free water, isotonic saline, Ringer's solution, ethyl alcohol, etc., and combin ations thereof.

[0207] Exemplary lubricating agents include magnesium stearate, calcium stearate, stearic acid, silica, talc, malt, glyceryl behanate, hydrogenated vegetable oils, polyethylene glycol, sodium benzoate, sodium acetate, sodium chloride, leucine, magnesium lauryl sulfate, sodium lauryl sulfate, etc., and combinations thereof.

[0208] Exemplary natural oils include almond, apricot kernel, avocado, babassu, bergamot, black current seed, borage, cade, chamomile, canola, caraway, carnauba, castor, cinnamon, cocoa butter, coconut, cod liver, coffee, corn, cotton seed, emu, eucalyptus, evening primrose, fish, flaxseed, geraniol, gourd, grape seed, hazel nut, hyssop, isopropyl myristate, jojoba, kukui nut, lavandin, lavender, lemon, litsea cubeba, macademia nut, mallow, mango seed, meadowfoam seed, mink, nutmeg, olive, orange, orange roughy, palm, palm kernel, peach kernel, peanut, poppy seed, pumpkin seed, rapeseed, rice bran, rosemary, safflower, sandalwood, sasquana, savoury, sea buckthorn, sesame, shea butter, silicone, soybean, sunflower, tea tree, thistle, tsubaki, vetiver, walnut, and wheat germ oils. Exemplary synthetic oils include, but are not limited to, butyl stearate, caprylic triglyceride, capric triglyceride, cyclomethicone, diethyl sebacate, dimethicone 360, isopropyl myristate, mineral oil, octyldodecanol, oleyl alcohol, silicone oil, and combinations thereof.

[0209] Additionally, the composition may further comprise a polymer. Exemplary polymers contemplated herein include, but are not limited to, cellulosic polymers and copolymers, for example, cellulose ethers such as methylcellulose (MC), hydroxyethylcellulose (HEC), hydroxypropyl cellulose (HPC), hydroxypropyl methyl cellulose (HPMC), methylhydroxyethylcellulose (MIIEC), methylhydroxypropylcellulose (MHPC), carboxymethyl cellulose (CMC) and its various salts, including, e.g., the sodium salt, hydroxyethylcarboxymethylcellulose (HECMC) and its various salts, carboxymethylhydroxyethylcellulose (CMHEC) and its various salts, other polysaccharides and polysaccharide derivatives such as starch, dextran, dextran derivatives, chitosan, and Attorney Docket No. 10063-110WO1

[0210] alginic acid and its various salts, carageenan, varoius gums, including xanthan gum, guar gum, gum arabic, gum karaya, gum ghatti, konjac and gum tragacanth, glycosaminoglycans and proteoglycans such as hyaluronic acid and its salts, proteins such as gelatin, collagen, albumin, and fibrin, other polymers, for example, polyhydroxyacids such as polylactide, polyglycolide, polyl(lactide-co-glycolide) and poly(.epsilon.-caprolactone-co-glycolide)-, carboxyvinyl polymers and their salts (e.g., carbomer), polyvinylpyrrolidone (PVP), polyacrylic acid and its salts, polyacrylamide, polyacrylic acid / acrylamide copolymer, polyalkylene oxides such as polyethylene oxide, polypropylene oxide, polyethylene oxide¬ propylene oxide), and a Pluronic polymer, polyoxy ethylene (polyethylene glycol), poly anhydrides, polyvinylalcohol, polyethyleneamine and polypyridine, polyethylene glycol (PEG) polymers, such as PEGylated lipids (e.g., PEG-stearate, I,2-Distearoyl-sn-glycero-3- Phosphoethanolamine-N-[Methoxy(Polyethylene glycol)- 1000], 1,2-Distearoyl-sn-glycero-3-Phosphoethanolamine-N-[Methoxy(Polyethylene glycol)-2000], and 1,2-Distearoyl-sn-glycero-3-Phosphoethanolamine-N-[Methoxy(Polyethylene glycol)-5000]), copolymers and salts thereof.

[0211] Additionally, the composition may further comprise an emulsifying agent.

[0212] Exemplary emulsifying agents include, but are not limited to, a polyethylene glycol (PEG), a polypropylene glycol, a polyvinyl alcohol, a poly-N-vinyl pyrrolidone and copolymers thereof, poloxamer nonionic surfactants, neutral water-soluble polysaccharides (e.g., dextran, Ficoll, celluloses), non-cationic poly(meth)acrylates, non-cationic polyacrylates, such as poly (meth) acrylic acid, and esters amide and hydroxy alkyl amides thereof, natural emulsifiers (e.g. acacia, agar, alginic acid, sodium alginate, tragacanth, chondrux, cholesterol, xanthan, pectin, gelatin, egg yolk, casein, wool fat, cholesterol, wax, and lecithin), colloidal clays (e.g. bentonite [aluminum silicate] and Veegum [magnesium aluminum silicate]), long chain amino acid derivatives, high molecular weight alcohols (e.g. stearyl alcohol, cetyl alcohol, oleyl alcohol, triacetin monostearate, ethylene glycol distearate, glyceryl monostearate, and propylene glycol monostearate, polyvinyl alcohol), carbomers (e.g. carboxy polymethylene, polyacrylic acid, acrylic acid polymer, and carboxy vinyl polymer), carrageenan, cellulosic derivatives (e.g. carboxymethylcellulose sodium, powdered cellulose, hydroxymethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methylcellulose, methylcellulose), sorbitan fatty acid esters (e.g. polyoxyethylene sorbitan monolaurate [Tween 20], polyoxyethylene sorbitan [Tween 60], polyoxyethylene sorbitan monooleate [Tween 80], sorbitan monopalmitate [Span 40], sorbitan monostearate [Span 60], sorbitan tristearate [Span 65], glyceryl monooleate, sorbitan monooleate [Span 80]), Attorney Docket No. 10063-110WO1

[0213] polyoxyethylene esters (e.g. polyoxyethylene monostearate [Myrj 45], polyoxyethylene hydrogenated castor oil, polyethoxylated castor oil, polyoxymethylene stearate, and Solutol), sucrose fatty acid esters, polyethylene glycol fatty acid esters (e.g. Cremophor), polyoxyethylene ethers, (e.g. polyoxyethylene lauryl ether [Brij 30]), polyvinyl¬ pyrrolidone), diethylene glycol monolaurate, triethanolamine oleate, sodium oleate, potassium oleate, ethyl oleate, oleic acid, ethyl laurate, sodium lauryl sulfate, Pluronic F 68, Poloxamer 188, cetrimonium bromide, cetylpyridinium chloride, benzalkonium chloride, docusate sodium, etc. and / or combinations thereof. In certain aspects, the emulsifying agent is cholesterol.

[0214] Liquid compositions include emulsions, microemulsions, solutions, suspensions, syrups, and elixirs. In addition to the active compound, the liquid composition may contain inert diluents commonly used in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils (in particular, cottonseed, groundnut, corn, germ, olive, castor, and sesame oils), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, and mixtures thereof. Besides inert diluents, the oral compositions can also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.

[0215] Injectable compositions, such as injectable aqueous or oleaginous suspensions, may be formulated according to the known art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be an injectable solution, suspension, or emulsion in a nontoxic parenterally acceptable diluent or solvent, for example, as a solution in 1,3-butanediol. Among the acceptable vehicles and solvents for pharmaceutical or cosmetic compositions that may be employed are water, Ringer's solution, U. S. P., and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. Any bland fixed oil can be employed, including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid are used in the preparation of injectables. In certain aspects, the particles are suspended in a carrier fluid comprising 1% (w / v) sodium carboxymethyl cellulose and 0.1% (v / v) Tween 80. The injectable composition can be sterilized, for example, by filtration through a bacteria-retaining filter or by incorporating sterilizing agents in the form of sterile solid compositions, which can be dissolved or dispersed in sterile water or other sterile injectable media prior to use. Attorney Docket No. 10063-110WO1

[0216] Compositions for rectal or vaginal administration may be in the form of suppositories which can be prepared by mixing the particles with suitable non-irritating excipients or carriers such as cocoa butter, polyethylene glycol, or a suppository wax which are solid at ambient temperature but liquid at body temperature and therefore melt in the rectum or vaginal cavity and release the particles.

[0217] Solid compositions include capsules, tablets, pills, powders, and granules. In such solid compositions, the particles are mixed with at least one excipient and / or a) fillers or extenders such as starches, lactose, sucrose, glucose, mannitol, and silicic acid, b) binders such as, for example, carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidinone, sucrose, and acacia, c) humectants such as glycerol, d) disintegrating agents such as agar- agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate, e) solution retarding agents such as paraffin, f) absorption accelerators such as quaternary ammonium compounds, g) wetting agents such as for example, cetyl alcohol and glycerol monostearate, h) absorbents such as kaolin and bentonite clay, and i) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and mixtures thereof. In the case of capsules, tablets, and pills, the dosage form may also comprise buffering agents. Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polyethylene glycols and the like.

[0218] Tablets, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings and other coatings well known in the pharmaceutical formulating art. They may optionally contain opacifying agents and can also be of a composition that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes.

[0219] Compositions for topical or transdermal administration include ointments, pastes, creams, lotions, gels, powders, solutions, sprays, inhalants, or patches. The active compound is admixed with an excipient and any needed preservatives or buffers as may be required.

[0220] The ointments, pastes, creams, and gels may contain, in addition to the active compound, excipients such as animal and vegetable fats, oils, waxes, paraffins, starch, tragacanth, cellulose derivatives, polyethylene glycols, silicones, bentonites, silicic acid, talc, and zinc oxide, or mixtures thereof. Attorney Docket No. 10063-110WO1

[0221] Powders and sprays can contain, in addition to the active compound, excipients such as lactose, talc, silicic acid, aluminum hydroxide, calcium silicates, and polyamide powder, or mixtures of these substances. Sprays can additionally contain customary propellants such as chlorofluorohydrocarbons.

[0222] Transdermal patches have the added advantage of providing controlled delivery of a compound to the body. Such dosage forms can be made by dissolving or dispensing the nanoparticles in a proper medium. Absorption enhancers can also be used to increase the flux of the compound across the skin. The rate can be controlled by either providing a rate¬ controlling membrane or by dispersing the particles in a polymer matrix or gel.

[0223] The active ingredient may be administered in such amounts, time, and route deemed necessary in order to achieve the desired result. The exact amount of the active ingredient will vary from subject to subject, depending on the species, age, and general condition of the subject, the severity of the medical disorder, the particular active ingredient, its mode of administration, its mode of activity, and the like. The active ingredient, whether the active compound itself or the active compound in combination with an agent, is preferably formulated in dosage unit form for ease of administration and uniformity of dosage. It will be understood, however, that the total daily usage of the active ingredient will be decided by the attending physician within the scope of sound medical judgment. The specific therapeutically effective dose level for any particular subject will depend upon a variety of factors, including the disorder being treated and the severity of the disorder; the activity of the active ingredient employ ed; the specific composition employed; the age, body weight, general health, sex and diet of the patient; the time of administration, route of administration, and rate of excretion of the specific active ingredient employed; the duration of the treatment; drugs used in combination or coincidental with the specific active ingredient employed; and like factors well known in the medical arts.

[0224] The active ingredient may be administered by any route. In some aspects, the active ingredient is administered via a variety of routes, including oral, intravenous, intramuscular, intra-arterial, intramedullary, intrathecal, subcutaneous, intraventricular, transdermal, interdermal, rectal, intravaginal, intraperitoneal, topical (as by powders, ointments, creams, and / or drops), mucosal, nasal, bucal, enteral, sublingual; by intratracheal instillation, bronchial instillation, and / or inhalation; and / or as an oral spray, nasal spray, and / or aerosol. In general, the most appropriate route of administration will depend upon a variety of factors, including the nature of the active ingredient (e.g., its stability in the environment of Attorney Docket No. 10063-110WO1

[0225] the gastrointestinal tract), the condition of the subject (e.g., whether the subject is able to tolerate oral administration), etc.

[0226] The exact amount of an active ingredient required to achieve a therapeutically or prophylactically effective amount will vary from subject to subject, depending on species, age, and general condition of a subject, severity of the side effects or disorder, identity of the particular compound(s), mode of administration, and the like. The amount to be administered to, for example, a child or an adolescent can be determined by a medical practitioner or person skilled in the art and can be lower or the same as that administered to an adult.

[0227] Useful dosages of the active agents and pharmaceutical compositions disclosed herein can be determined by comparing their in vitro activity and in vivo activity in animal models. Methods for the extrapolation of effective dosages in mice and other animals to humans are known to the ait.

[0228] The dosage ranges for the administration of the compositions are those large enough to produce the desired effect in which the symptoms or disorder are affected. The dosage should not be so large as to cause adverse side effects, such as unwanted cross-reactions, anaphylactic reactions, and the like. Generally, the dosage will vary with the age, condition, sex, and extent of the disease in the patient and can be determined by one of skill in the art. 'The dosage can be adjusted by the individual physician in the event of any counterindications. Dosage can vary and can be administered in one or more doses daily for one or several days.

[0229] Combinations

[0230] The disclosed compounds can also be combined with aztreonam and / or disulfiram, either in a single composition or in separate compositions.

[0231] The disclosed compounds can also be combined with additional antimicrobial agents, either in a single composition or in separate compositions. For example, the disclosed compounds can be combined with one or more of Acedapsone; Acetosulfone Sodium; Alamecin; Alexidine; Amdinocillin; Amdinocillin Pivoxil; Amicycline;

[0232] Amifloxacin; Amifloxacin Mesylate; Amikacin; Amikacin Sulfate; Aminosalicylic acid; Aminosalicylate sodium; Amoxicillin; Amphomycin; Ampicillin; Ampicillin Sodium; Apalcillin Sodium; Apramycin; Aspartocin; Astromicin Sulfate; Avilamycin; Avoparcin; Azithromycin; Azlocillin; Azlocillin Sodium; Bacampicillin Hydrochloride; Bacitracin; Bacitracin Methylene Disalicylate; Bacitracin Zinc; Bambermycins; Benzoylpas Calcium; Berythromycin; Betamicin Sulfate; Biapenem; Biniramycin; Biphenamine Hydrochloride; Attorney Docket No. 10063-110WO1

[0233] Bispyrithione Magsulfex; Butikacin; Butirosin Sulfate; Capreomycin Sulfate; Carbadox; Carbenicillin Disodium; Carbenicillin Indanyl Sodium; Carbenicillin Phenyl Sodium; Carbenicillin Potassium; Carumonam Sodium; Cefaclor; Cefadroxil; Cefamandole;

[0234] Cefamandole Nafate; Cefamandole Sodium; Cefaparole; Cefatrizine; Cefazaflur Sodium; Cefazolin; Cefazolin Sodium; Cefbuperazone; Cefdinir; Cefepime; Cefepime Hydrochloride; Cefetecol; Cefixime; Cefinenoxime Hydrochloride; Cefmetazole;

[0235] Cefmetazole Sodium; Cefonicid Monosodium; Cefonicid Sodium; Cefoperazone Sodium; Ceforanide; Cefotaxime Sodium; Cefotetan; Cefotetan Disodium; Cefotiam Hydrochloride; Cefoxitin; Cefoxitin Sodium; Cefpimizole; Cefpimizole Sodium; Cefpiramide; Cefpiramide Sodium; Cefpirome Sulfate; Cefpodoxime Proxetil; Cefprozil; Cefroxadine; Cefsulodin Sodium; Ceftazidime; Ceftibuten; Ceftizoxime Sodium; Ceftriaxone Sodium; Cefuroxime; Cefuroxime Axetil; Cefuroxime Pivoxetil; Cefuroxime Sodium; Cephacetrile Sodium; Cephalexin; Cephalexin Hydrochloride; Cephaloglycin; Cephaloridine; Cephalothin Sodium; Cephapirin Sodium; Cephradine; Cetocycline Hydrochloride; Cetophenicol;

[0236] Chloramphenicol; Chloramphenicol Palmitate; Chloramphenicol Pantothenate Complex; Chloramphenicol Sodium Succinate; Chlorhexidine Phosphanilate; Chloroxylenol;

[0237] Chlortetracycline Bisulfate; Chlortetracycline Hydrochloride; Cinoxacin; Ciprofloxacin; Ciprofloxacin Hydrochloride; Cirolemycin; Clarithromycin; Clinafloxacin Hydrochloride; Clindamycin; Clindamycin Hydrochloride; Clindamycin Palmitate Hydrochloride;

[0238] Clindamycin Phosphate; Clofazimine; Cioxacillin Benzathine; Cioxacillin Sodium;

[0239] Cloxyquin; Colistimethate Sodium; Colistin Sulfate; Coumennycin; Coumermycin Sodium; Cyclacillin; Cycloserine; Dalfopristin; Dapsone; Daptomycin; Demeclocycline;

[0240] Demeclocycline Hydrochloride; Demecycline; Denofungin; Diaveridine; Dicloxacillin; Dicloxacillin Sodium; Dihydrostreptomycin Sulfate; Dipyrithione; Dirithromycin;

[0241] Doxycycline; Doxycycline Calcium; Doxycycline Fosfatex; Doxycycline Hyclate; Droxacin Sodium; Enoxacin; Epicillin; Epitetracycline Hydrochloride; Erythromycin; Erythromycin Acistrate; Erythromycin Estolate; Erythromycin Ethylsuccinate; Erythromycin Gluceptate; Erythromycin Lactobionate; Erythromycin Propionate; Erythromycin Stearate; Ethambutol Hydrochloride; Ethionamide; Fleroxacin; Floxacillin; Fludalanine; Flumequine;

[0242] Fosfomycin; Fosfomycin Tromethamine; Fumoxicillin; Furazolium Chloride; Furazolium Tartrate; Fusidate Sodium; Fusidic Acid; Gentamicin Sulfate; Gloximonam; Gramicidin; Haloprogin; Hetacillin; Hetacillin Potassium; Hexedine; Ibafloxacin; Imipenem;

[0243] Isoconazole; Isepamicin; Isoniazid; Josamycin; Kanamycin Sulfate; Kitasamycin;

[0244] Levofuraltadone; Levopropylcillin Potassium; Lexithromycin; Lincomycin; Lincomycin Attorney Docket No. 10063-110WO1

[0245] Hydrochloride; Lomefloxacin; Lomefloxacin Hydrochloride; Lomefloxacin Mesylate; Loracarbef; Mafenide; Meclocycline; Meclocycline Sulfosalicylate; Megalomicin Potassium Phosphate; Mequidox; Meropenem; Methacycline; Methacycline Hydrochloride; Methenamine; Methenamine Hippurate; Methenamine Mandelate; Methicillin Sodium; Metioprim; Metronidazole Hydrochloride; Metronidazole Phosphate; Mezlocillin;

[0246] Mezlocillin Sodium; Minocycline; Minocycline Hydrochloride; Mirincamycin Hydrochloride; Monensin; Monensin Sodiumr; Nafcillin Sodium; Nalidixate Sodium; Nalidixic Acid; Natamycin; Nebramycin; Neomycin Palmitate; Neomycin Sulfate;

[0247] Neomycin Undecylenate; Netilmicin Sulfate; Neutramycin; Nifuiradene; Nifuraldezone; Nifuratel; Nifuratrone; Nifurdazil; Nifurimide; Nifiupirinol; Nifurquinazol; Nifurthiazole; Nitrocycline; Nitrofurantoin; Nitromide; Norfloxacin: Novobiocin Sodium; Ofloxacin; Onnetoprim; Oxacillin Sodium; Oximonam; Oximonam Sodium; Oxolinic Acid;

[0248] Oxytetracy cline; Oxytetracycline Calcium; Oxytetracycline Hydrochloride; Paldimycin; Parachlorophenol; Paulomycin; Pefloxacin; Pefloxacin Mesylate; Penamecillin; Penicillin G Benzathine; Penicillin G Potassium; Penicillin G Procaine; Penicillin G Sodium; Penicillin V; Penicillin V Benzathine; Penicillin V Hydrabamine; Penicillin V Potassium; Pentizidone Sodium; Phenyl Aminosalicylate; Piperacillin Sodium; Pirbenicillin Sodium; Piridicillin Sodium; Pirlimycin Hydrochloride; Pivampicillin Hydrochloride; Pivampicillin Pamoate; Pivampicillin Probenate; Polymyxin B Sulfate; Porfiromycin; Propikacin; Pyrazinamide; Pyrithione Zinc; Quindecamine Acetate; Quinupristin; Racephenicol; Ramoplanin;

[0249] Ranimycin; Relomycin; Repromicin; Rifabutin; Rifametane; Rifamexil; Rifamide;

[0250] Rifampin; Rifapentine; Rifaximin; Rolitetracycline; Rolitetracycline Nitrate; Rosaramicin; Rosaramicin Butyrate; Rosaramicin Propionate; Rosaramicin Sodium Phosphate;

[0251] Rosaramicin Stearate; Rosoxacin; Roxarsone; Roxithromycin; Sancycline; Sanfetrinem Sodium; Sarmoxicillin; Sarpicillin; Scopafiingin; Sisomicin; Sisomicin Sulfate;

[0252] Sparfloxacin; Spectinomycin Hydrochloride; Spiramycin; Stallimycin Hydrochloride; Steffimycin; Streptomycin Sulfate; Streptonicozid; Sulfabenz; Sulfabenzamide;

[0253] Sulfacetamide; Sulfacetamide Sodium; Sulfacyline; Sulfadiazine; Sulfadiazine Sodium; Sulfadoxine; Sulfalene; Sulfamerazine; Sulfameter; Sulfamethazine; Sulfamethizole;

[0254] Sulfamethoxazole; Sulfamonomethoxine; Sulfamoxole; Sulfanilate Zinc; Sulfanitran;

[0255] Sulfasalazine; Sulfasomizole; Sulfathiazole; Sulfazamet; Sulfisoxazole; Sulfisoxazole Acetyl; Sulfisboxazole Diolamine; Sulfomyxin; Sulopenem; Sultamricillin; Suncillin Sodium; Talampicillin Hydrochloride; Teicoplanin; Temafloxacin Hydrochloride;

[0256] Temocillin; Tetracycline; Tetracycline Hydrochloride; Tetracycline Phosphate Complex; Attorney Docket No. 10063-110WO1

[0257] Tetroxoprim; Thiamphenicol; Thiphencillin Potassium; Ticarcillin Cresyl Sodium;

[0258] Ticarcillin Disodium; Ticarcillin Monosodium; Ticlatone; Tiodonium Chloride;

[0259] Tobramycin; Tobramycin Sulfate; Tosufloxacin; Trimethoprim; Trimethoprim Sulfate; Trisulfapyrimidines; Troleandomycin; Trospectomycin Sulfate; Tyrothricin; Vancomycin; Vancomycin Hydrochloride; Virginiamycin; or Zorbamycin.

[0260] In some examples, the disclosed compounds can be combined with anti¬ inflammatory agents, either in the same or in separate compositions. Examples of such agents include acetaminophen, aspirin, celecoxib, diclofenac, diflunisal, flurbiprofen, ibuprofen, indomethacin, ketoprofen, ketorolac, meclofenamate, meloxicam, methyl salicylate, nabumetone, naproxen, oxaprozin, piroxicam, sulindac, tolmetin, trolamine.

[0261] In some examples, the disclosed compounds can be combined with proton pump inhibitors, either in the same composition on in separate compositions. Examples of such agents include omeprazole, lansoprazole, pantoprazole, and esomeprazole.

[0262] In a particular aspect, the disclosed compounds of Formulas I, II, and or III, can be combined with, either in the same or separate compositions, one or more of aztreonam, disulfiram, another antibiotic, and a proton pump inhibi tor.

[0263] Administration

[0264] The disclosed compounds can be administered either sequentially or simultaneously in separate or combined pharmaceutical formulations. When one or more of the disclosed compounds is used in combination with a second therapeutic agent the dose of each compound can be either the same as or differ from that when the compound is used alone. Appropriate doses will be readily appreciated by those skilled in the art.

[0265] The term “administration” and variants thereof (e.g., “administering” a compound) in reference to a compound of the invention means introducing the compound or a prodrug of the compound into the system of the animal in need of treatment. When a compound of the invention or prodrug thereof is provided in combination with one or more other active agents (e.g., a cytotoxic agent, etc.), “administration” and its variants are each understood to include concurrent and sequential introduction of the compound or prodrug thereof and other agents.

[0266] In vivo application of the disclosed compounds, and compositions containing them, can be accomplished by any suitable method and technique presently or prospectively known to those skilled in the art. For example, the disclosed compounds can be formulated in a physiologically- or pharmaceutically-acceptable form and administered by any suitable route known in the art including, for example, oral, nasal, rectal, topical, and parenteral Attorney Docket No. 10063-110WO1

[0267] routes of administration. As used herein, the term parenteral includes subcutaneous, intradermal, intravenous, intramuscular, intraperitoneal, and intrasterna] administration, such as by injection. Administration of the disclosed compounds or compositions can be a single administration, or at continuous or distinct intervals as can be readily determined by a person skilled in the art.

[0268] The compounds disclosed herein, and compositions comprising them, can also be administered utilizing liposome technology, slow-release capsules, implantable pumps, and biodegradable containers. These delivery methods can, advantageously, provide a uniform dosage over an extended period. The compounds can also be administered in their salt derivative forms or crystalline forms.

[0269] The compounds disclosed herein can be formulated according to known methods for preparing pharmaceutically acceptable compositions. Formulations are described in detail in a number of sources which are well known and readily available to those skilled in the ait. For example, Remington’s Pharmaceutical Science by E. W. Martin (1995 ) describes formulations that can be used in connection with the disclosed methods. In general, the compounds disclosed herein can be formulated such that an effective amount of the compound is combined with a suitable carrier to facilitate effective administration of the compound. The compositions used can also be in a variety of forms. These include, for example, solid, semi-solid, and liquid dosage forms, such as tablets, pills, powders, liquid solutions or suspension, suppositories, injectable and infusible solutions, and sprays. The preferred form depends on the intended mode of administration and therapeutic application. The compositions also preferably include conventional pharmaceutically-acceptable carriers and diluents which are known to those skilled in the art. Examples of carriers or diluents for use with the compounds include ethanol, dimethyl sulfoxide, glycerol, alumina, starch, saline, and equivalent carriers and diluents. To provide for the administration of such dosages for the desired therapeutic treatment, compositions disclosed herein can advantageously comprise between about 0.1% and 99%, and especially, 1 and 15% by weight of the total of one or more of the subject compounds based on the weight of the total composition including earner or diluent.

[0270] Formulations suitable for administration include, for example, aqueous sterile injection solutions, which can contain antioxidants, buffers, bacteriostats, and solutes that render the formulation isotonic with the blood of the intended recipient; and aqueous and nonaqueous sterile suspensions, which can include suspending agents and thickening agents. The formulations can be presented in unit-dose or multi-dose containers, for Attorney Docket No. 10063-110WO1

[0271] example sealed ampoules and vials, and can be stored in a freeze dried (lyophilized) condition requiring only the condition of the sterile liquid carrier, for example, water for injections, prior to use. Extemporaneous injection solutions and suspensions can be prepared from sterile powder, granules, tablets, etc. It should be understood that in addition to the ingredients particularly mentioned above, the compositions disclosed herein can include other agents conventional in the art having regard to the type of formulation in question.

[0272] Method of Use

[0273] The compounds and compositions disclosed herein can be used to treat infections caused by bacteria and to inhibit the growth of bacteria. In certain examples, disclosed are methods of treating a bacterial infection in a subject, comprising administering to the subject a thereapeutically effective amount of any of the compounds or compositions disclosed herein. Specific examples of infections that can be treated include, but are not limited to, Actinobacter, Actinomycetes, Bacilli, Bortedellen, Clostridia, Corynebacteria, Enterobacter, Enterococci, Helicobacter, Haemophilus, Klebsiella, Listeria, Mycobacteria, Neisseria, Shigella, Salmonella, Streptococci, Staphylococci, tuberculosis bacteria, and Yersinia.

[0274] In some examples, the disclosed compounds and compositions can be used to treat infections caused by H. pylori, including drug resistant H. pylori. Is it disclosed herein that aztreonam and the anti-alcoholic agent disulfiram are active against H. pylori, with low resistance emergence across 50 days. In a mice gastric infection model orally administered aztreonam or disulfiram decreased inflammation and eliminated H. pylori adherence to gastric mucosa and epithelial cells in vivo. Triple therapy with aztreonam, disulfiram, their analogs disclosed herein, or any combination thereof, with a proton pump inhibitor (e.g., omeprazole) and a clinical antibiotic (e.g., amoxicillin) reduced inflammation after seven days. The gut microbiome of the triple therapy-treated mouse group was similar to that of the control antibiotic -treated group. The disclosed aztreonam and disulfiram analogs inhibited various bacterial pathogens including gastric pathogens.

[0275] In some examples, the disclosed compounds and compositions can be used to treat infections caused by resistant G-pos. bacteria such as Methicillin Resistant Staphylococcus aureus (MRSA).

[0276] In some examples, the disclosed compounds and compositions can be used to treat infections caused by resistant G-neg. pathogens such as P. aeruginosa. Infections caused by G-neg. bacteria in general, and MDR P. aeruginosa in particular ( Wagner, S., et al. J. Med. Attorney Docket No. 10063-110WO1

[0277] Chem. 2016, 59, 5929), represent a key need in antibacterial drug discovery that is currently underrepresented by approaches in clinical development.

[0278] In some examples, the disclosed compounds and compositions can be used to treat infections caused by Enterococcus faecium, Klebsiella pneumoniae, Acinetobacter baumannii, various Enterobacter, and Neisseria gonorrhoeae. Further examples include the following diseases include: tuberculosis; Pneumonia; Typhoid; Paratyphoid; Syphilis, Gastritis; Gastroenteritis; Ruhr; Pestilence; Enteritis; extraintestinal infections, peritonitis and appendicitis with E. coli and intestinal infections with EHEC, EPEC, ETEC and EIEC; Cholera, Legionnaires' disease, whooping cough, brucellosis, Lyme disease, leptospirosis, typhus, trachoma, gonorrhea, meningitis, septicemia, leprosy etc. These methods can involve administering a compound disclosed herein to the infected human or animal or the human or animal at risk of being infected.

[0279] In other examples, disclosed herein are methods of treating an infection in a patient, comprising administering to the patient a thereapeutically effective amount of any of the compounds disclosed herein.

[0280] In specific examples, disclosed herein is a method of treating a bacterial infection, comprising administering to a subject in need thereof a composition comprising a compound of any one of Formulas I, II, and / or III. Therapeutically acceptable amounts of the compounds can be used. In some examples, the bacterial infection is caused by a drug¬ resistant bacteria. In other examples, the bacterial infection is caused by H. pylori, A. baumannii, K. pneumoniae, L. monocytogenes. E. faecium, or 5. aureus.

[0281] In some examples, aztreonam and / or disulfiram is also administered to the subject in need thereof along with the compounds of Formula I, II, or III, either as a separate composition or in the same composition. In another aspect, aztreonam and / or disulfiram is administered without the compounds of Formula I, II, or III.

[0282] In some examples, an antibiotic is also administered to the subject in need thereof. Examples of suitable antibiotics are noted above but can include is amoxicillin, clarithromycin, metronidazole, nitrofurantoin, or any combination thereof.

[0283] In some examples, a proton pump inhibitor is also administered, examples of which can include omeprazole, lansoprazole, pantoprazole, or esomeprazole.

[0284] Also disclosed herein is a method of treating a bacterial infection (e.g., H. pylori), comprising: administering to a subject in need thereof aztreonam and / or disulfiram, an antibiotic, and a proton pump inhibitor. The antibiotic is amoxicillin, clarithromycin, Attorney Docket No. 10063-110WO1

[0285] metronidazole, nitrofurantoin, or any combination thereof for example. The proton pump inhibitor is omeprazole, lansoprazole, pantoprazole, or esomeprazole for example.

[0286] Also disclosed is a method of treating abacterial infection, comprising: administering to a subject in need thereof

[0287]

[0288] DIS1-2

[0289] or a pharmaceutically acceptable salt thereof.

[0290] EXAMPLES

[0291] The following examples are set forth below to illustrate the compounds, compositions, and methods claimed herein, along with associated methods and results according to the disclosed subject mater. These examples are not intended to be inclusive of all aspects of the subject matter disclosed herein, but rather to illustrate representative methods and results. These examples are not intended to exclude equivalents and variations of the present disclosure, which are apparent to one skilled in the ait.

[0292] Efforts have been made to ensure accuracy with respect to numbers (e.g., amounts, temperature, etc.), but some errors and deviations should be accounted for. Unless indicated otherwise, parts are parts by weight, temperature is in °C or is at ambient temperature, and pressure is at or near atmospheric. There are numerous variations and combinations of reaction conditions, e.g., component concentrations, temperatures, pressures, and other reaction ranges and conditions that can be used to optimize the product purity and yield obtained from the described process. Only reasonable and routine experimentation will be required to optimize such process conditions.

[0293] The present invention further provides a method of synthesizing compounds according to the present invention, or a pharmaceutically acceptable salt, solvate or prodrug thereof.

[0294] Example 1: Anti-H. pylori activity

[0295] A broth-microdilution assay confirmed the activity of AZ and DIS against multiple H. pylori strains, such as reference strains and clinical strains, with low minimal inhibitory concentrations (MICs). A critical finding was that both drugs are active against clarithromycin -resistant H. pylori, which is a high-priority-2 pathogen. AZ inhibited II. pylori with an MIC of 0.25 nig / L, and DIS had a MIC of 0.125 nig / L (Table 1). When Attorney Docket No. 10063-110WO1

[0296] testing was extended in an acidic environment, AZ was found to retain its growth-inhibitory' activity at an acidic pH of 6.0. Interestingly, DIS is active in acidic pH and its anti-. pylori activity is enhanced in acidic pH tested up to 5.5 (Table 1).

[0297] Table 1: Antibacterial activities of test compounds

[0298] H. pylon-reference H. pylori- pH against 60190

[0299] strains Clinical

[0300] Strains

[0301] Drug 60190 NTCC Tx30a 3713 3719B 7.0 6.5 6.0 5.5

[0302] 11437 A4

[0303] Disulfiram 0.125 0.125 1.0 0.06 0.06 0.125 0.125 0.06 0.06 Aztreonam 0.25 0.5 2.0 0.5 1 0.25 0.25 0.5 1.0 Clarithromycin 0.001 0.001 0.001 0.001 >1 0.006 0.006 0.01 0.03

[0304]

[0305] An antibacterial molecule is either bactericidal or bacteriostatic. AZ and DIS were found to be bacteriostatic against a high dose of H. pylori and bactericidal against a low dose of H. pylori (Figs. 1A-1G). A killing kinetics assay of AZ or DIS against a high dose of cells (109cells / mL) demonstrated that both are bacteriostatic at lx or 4xMIC (Fig. 1A). At IxMIC, both agents inhibited bacterial multiplication, and at 4xMIC, AZ killed the 1-Logio-CFU from an initial inoculum. H. pylori adhesion is an initial step in gastric infection. At 8xMIC, AZ cleared 4-Logio-CFU of H. pylori adhered to gastric epithelial cells and DIS cleared 2-Logio-CFU of adhered bacterial load (Fig. IB). Bacterial motility is a key virulence factor in initial colonization. H. pylori motility testing at sub-MIC of AZ and DIS found that both drugs hindered H. pylori motility at 0.5xMIC (Figs. 1C-1D). VacA is a critical virulence factor and a secreted toxin that translocates into gastric epithelial cells, inducing intracellular vacuolation, including membrane depolarization, autophagy, and inhibition of T cell proliferation. Hence, inhibition of VacA toxin and its activity is critical in treating H. pylori infection. Both drugs, at 4x or 8xMIC inhibited vacuolation, using MKN-28 cells co-cultured with H. pylori infection (Fig. 1G). Thus, the antivirulence profile of the drugs shows that AZ and DIS eliminate H. pylori adherence, hinder H. pylori motility, and inhibit H. pylori-induced vacuolation in mammalian gastric epithelial ceils. Galleria mellonella (G. mellonella) is a simple and cost-effective invertebrate infection model to study H. pylori pathogenesis and drug efficacy. AZ at 25 mg / kg or DIS at 2.0 Attorney Docket No. 10063-110WO1

[0306] mg / kg promotes 50% survival of the H. yfon-infected larvae (Figs. 1E-1F; p<0.0001), similar to the positive control amoxicillin. Together, these findings indicate that AZ and DIS are excellent agents to explore as anti-H. pylori medication. Scanning Electron Microscopy shows morphological changes after DIS and AZ treatment (Fig. 7).

[0307] Example 2: AZ and DIS combination therapy

[0308] Combination therapy is a hallmark of H. pylori therapy. The synergistic approach of AZ and DIS with various clinical agents resulted in partial synergy between them (FICI between 0.5 to 1.0; Table 2).

[0309] Table 2: FICI of test compounds against H. pylori. Fractional inhibitory concentration index (FICI) of AZ and DIS against various PPIs and amoxicillin are shown.

[0310] Lan Rab Eso Onie Panto Amox Clar Bismuth Salicylate

[0311] AZ 0.5 0.5 0.75 1.0 1.0 0.75 1.0 0.5

[0312] DIS 0.75 0.5 0.75 1.0 1.0 1 1.0 1.0

[0313]

[0314] Lan- Lansoprazole; Rab- Rabeprazole; Eso- Esomeprazole; Amox- Amoxicillin

[0315] Examples 3-4: Resistance studies

[0316] Whether H. pylori develops resistance to AZ or DIS was next tested. A mutation frequency assay showed no emergence of resistance to 30 days. Treatment up to 50 days showed AZ and DIS tolerance up to lx to 2x MIC at end of 50th day. We extended exposing the test agents for up to 75 days to generate DIS or AZ resistant mutants to understand its mechanism of action. Exposing up to 75-days show MICs were increased up to 64x against AZ and up to 8x against DIS (Fig. 10). Whole genome sequencing (WGS) showed that changes in H. pylori genome of AZ -resistant clones (Table 3).

[0317] Table 3: Attorney Docket No. 10063-110W01

[0318] AZ resistant mutants / Predicted Sequence- Clone 1

[0319] Evidence Position Mutation Annotation Gene

[0320] d42azl.. O.25

[0321] 606,899 C— *T S351S (AGC^AGT) ctpB Carboxy-terminal processing protease CtpB 627,512 A2 bp intergenic (-8 / +132) cusA / <- FAMPOCIG_00601 Cation efflux system protein CusA / hypothetical protein 706,730 A — -> G M249V (ATG— > GTG) hmuU Hemin transport system permease protein HmuU I3T (ATC-+ACC) rny — > Ribonuclease Y

[0322] 1,069,445 f >( ' VI 57 A (GTA^GCA) virB4_l Type IV secretion system protein virB4 1,435,170 A— > G T312A (ACC— > GCC) ppsA Phosphoenolpyruvate synthase

[0323] d66az__5

[0324] 221,035 intergenic (+189 / -58) btuB_2 -> / -+ FAMPOCIG_00208 Vitamin B12 transporter BtuB / hypothetical protein 259,069 C-»T R30H (CGC— > CAC) rppH <-- RNA pyrophosphohydrolase

[0325] 471,535 (T)8->9 coding (55 / 453 nt) fur Ferric uptake regulation protein

[0326] 606,899 C—> T S351S (AGC— *AGT) ctpB Carboxy-terminal processing protease CtpB 627,512 A2 bp intergenic (-8 / + 132) cusA «-• / <-- FAMPOCIG.0060+1 Cation efflux system protein CusA / hypothetical protein 706,730 A— > G M249V (ATG— > GTG) hmuU —► Hemin transport system permease protein HmuU I3T (ATC— > ACC) rny Ribonuclease Y

[0327] 935,212 A — > G VI 35 A (GTT--> GCT) aroC Chorismate synthase

[0328] 1,001,805 G~> T N847K (AAC— ’■AAA) mdtC Multidrug resistance protein MdtC

[0329] 1,061,351 G— > A G83D (GGC^GAC) rho —> Transcription termination factor Rho

[0330] 1,069,445 T > C VI 57 A (GTA— > GCA) virB4-_I ----> Type IV secretion system protein virB4 1,435,170 A— > G T312A (ACC-+GCC) ppsA — > Phosphoenolpyruvate synthase

[0331] Trifunctional nucleotide phosphoesterase protein 1,454,849 (G) 12^13 intergenic (+144 / -65) yfkN mcp4

[0332] YfkN / Methyl-accepting chemotaxis protein 4 1,596,499 G—> A A498T (GCT— > ACT) ftsl -* Peptidoglycan D, D-transpeptidase FtsI 1,596,628 G— -> A A541T (GCG--> ACG) ft si -> Peptidoglycan D, D-transpeptidase FtsI

[0333] d70az_20

[0334]

[0335] 221,035 intergenic (+ 189 / -58) btuB_2 / -» FAMPOCIGJ)0208 Vitamin B12 transporter BtuB / hypothetical protein Attorney Docket No. 10063-110W01

[0336] 259,069 C^T R30H (CGC^CAC) rppH <— RNA pyrophosphohydrolase

[0337] 471,535 (T / g-.g coding (55 / 453 nt) fur *— Ferric uptake regulation protein

[0338] 606,899 C—> T S351S (AGC^AGT) ctpB Carboxy-terminal processing protease CtpB 706,730 A— > G M249V (ATG— > GTG) hmuU — * Hemin transport system permease protein HmuU 13T (ATC -> ACC) rny --> Ribonuclease Y

[0339] 935,212 A — > G VI 35 A (GTT-> GCT) aroC Chorismate synthase

[0340] 1,001,805 G— > T N847K (AA. C— > AAA) mdtC Multidrug resistance protein MdtC 1,061,351 G -- > A G83D (GGC— > GAC) rho Transcription termination factor Rho 1,069,445 T— > C V157A (GTA— > GCA) virB4_l Type IV secretion system protein virB4 1,435,170 A—> G T312A (ACC— > GCC) ppsA — > Phosphoenolpyruvate synthase

[0341] 1,596,499 G— -> A A498T (GCT— > ACT) ftsl ---> Peptidoglycan D, D-transpeptidase Ftsl 1,596,628 G-> A A541T (GCG-yACG) Peptidoglycan D, D-transpeptidase Ftsl d75az_20

[0342] 221,035 (TW.is intergenic (+189 / -58) btuB.2 / — FAMPOCIG. 00208 Vitamin B12 transporter BtuB / hypothetical protein 259,069 U--> T R30H (CGC— > CAC) rppH <— RNA pyrophosphohydrolase

[0343] 471,535 (T)S^9 coding (557453 nt) Ferric uptake regulation protein

[0344] 606,899 C-»T S351S (AGC--> AGT) ctpB Carboxy-terminal processing protease CtpB 706,730 A-> G M249V ( ATG—*GTG) hmuU —* Hemin transport system permease protein HmuU I3T (ATC— > ACC) rny — > Ribonuclease Y

[0345] 935,212 A— > G VI 35 A (GTT--> GCT) aroC <-- Chorismate synthase

[0346] 943,572 G-> T S621R (AGC-MGA) bamA Outer membrane protein assembly factor BamA 1,001,805 G- 'T N847K (AAC— AAA) mdlC «— Multidrug resistance protein MdtC 1,061,351 G — > A G83D (GGC— > GAC) rho Transcription termination factor Rho 1,069,445 T---> C VI 57 A (GTA--*GCA) virB4_l Type IV secretion system protein virB4 1,435,170 A— > G T312A (ACC— > GCC) ppsA — > Phosphoenolpyruvate synthase

[0347] 1,596,499 G — > A A498T (GCT— ’ACT) Peptidoglycan D, D-transpepddase Ftsl 1,596,628 G--> A A541T (GCG -> ACG) Peptidoglycan D, D-transpeptidase Ftsl

[0348] AZ resistant mutants / Predicted Sequence- Clone 2

[0349]

[0350] Evidence Position Mutation Annotation | Gene Attorney Docket No. 10063-110W01

[0351] d42azj)„25

[0352] 606,899 C--> T S351S (AGC— *AGT) ctpB Carboxy-temiinal processing protease CtpB 706,730 A~> G M249V ( ATG—> GTG) hmuU—* Hemin transport system permease protein HmuU I3T (ATC— > ACC) my Ribonuclease Y

[0353] 935,212 A — -> G VI 35 A (GTT--> GCT) aroC «— Chorismate synthase

[0354] 1,069,445 T- > C VI 57 A (GTA— > GCA) virB4_l Type IV secretion system protein virB4 1,435,170 A— > G T312A (ACC— > GCC) ppsA — > Phosphoenolpyruvate synthase

[0355] d66az...l

[0356] 556,032 (T)!6^;5intergenic (-179 / +172) btuB_3 rocF Vitamin B 12 transporter BtuB / Arginase 706,730 A—> G M249V (ATG— > GTG) hmuU — > Hemin transport system permease protein HmuU I3T (ATC-+ACC) my --> Ribonuclease Y

[0357] 1,062,055 C— *T R318C (CGC-*TGC) rho Transcription termination factor Rho 1,435,170 A — > G T312A (ACC— > GCC) ppsA — > Phosphoenolpyruvate synthase

[0358] 1,596,628 G -- > A A541T (GCG— > ACG) ftsl --> Peptidoglycan D, D-transpeptidase Ftsl d70az_6„25

[0359] 221,035 (T)U^I5intergenic (+189 / -58) btuB_2 -> / FAMPOCIG_00208 Vitamin B 12 transporter BtuB / hypothetical protein 259,069 C-»T R30H (CGC— > CAC) rppH RNA pyrophosphohydrolase

[0360] 471,535 (T)S^9 coding (55 / 453 nt) fur Ferric uptake regulation protein

[0361] 606,899 C^T S351S (AGC^AGT) ctpB Carboxy-terminal processing protease CtpB I3T (ATC--> ACC) rny —> Ribonuclease Y

[0362] 935,212 A—> G VI 35 A (GTT— *GCT) aroC Chorismate synthase

[0363] 1,004,246 G— > C L34V (CTT— > GTT) mdlC Multidrug resistance protein MdtC 1,061,351 G — > A G83D (GGC— > GAC) rho Transcription termination factor Rho 1,069,445 T--> C VI 57 A (GTA— > GCA) virB4_l Type IV secretion system protein virB4 1,435,170 A— > G T312A (ACC— > GCC) ppsA — > Phosphoenolpyruvate synthase Trifunctional nucleotide phosphoesterase protein 1,454,849 (G)12^13 intergenic (+144 / -65) yfkN mcp4

[0364] Y fkN / Methyl-accepting chemotaxis protein 4 1,596,499 G -- > A A498T (GCT--> ACT) ftsl --> Peptidoglycan D, D-transpeptidase Ftsl

[0365]

[0366] 1,596,628 G—> A A541T (GCG— *ACG) fisJ Peptidoglycan D, D-transpeptidase Ftsl Attorney Docket No. 10063-110W01

[0367] d75az__6„25

[0368] 556,032 (T)16— intergenic (-179 / +172) btuB_3 rocF Vitamin B 12 transporter BtuB / Arginase 706,730 A~> G M249V ( ATG—> GTG) hmuU — Hemin transport system permease protein HmuU I3T (ATC— > ACC) my Ribonuclease Y

[0369] 943,572 G— -> T S621R (AGO-* AG A) bamA Outer membrane protein assembly factor BamA 981,779 A— > C intergenic (-35 / +21) ppa — / FAMPOCIG_00958 Inorganic pyrophosphatase / hypothetical protein 1,062,055 C^T R318C (CGC— > TGC) rho — * Transcription termination factor Rho

[0370] 1,435,170 A— > G T312A (ACC— > GCC) ppsA Phosphoenolpyruvate synthase

[0371] 1,596,628 G— > A A541T (GCG— *ACG) Peptidoglycan D, D-transpeptidase FtsI

[0372] AZ resistant mutants / Predicted Sequence- Clone 3

[0373] Evidence Position Mutation Annotation Gene

[0374] d66az_3

[0375] 221,035 intergenic (+189 / -58) btuB_2 -> / -» FAMP()C1G_OO2O8 Vitamin B12 transporter BtuB / hypothetical protein 259,069 C--> T R30H (CGC - -> CAC) rppH <- RNA pyrophosphohydrolase

[0376] 471,535 (T)S^9 coding (55 / 453 nt) fur — Ferric uptake regulation protein

[0377] 606,899 C— *T S351S (AGC-+AGT) ctpB — > Carboxy-terminal processing protease CtpB 706,730 A — -> G M249V (ATG-> GTG) hmuU — * Hemin transport system permease protein HmuU I3T (ATC-*ACC) my Ribonuclease Y

[0378] 935,212 A— > G V135A (GTT^GCT) a roC ■<— Chorismate synthase

[0379] 1,004,246 G— > C L34V (C1T— > GTT) mdtC Multidrug resistance protein MdtC 1,061,351 G— > A G83D (GGC— > GAC) rho —* Transcription termination factor Rho 1,069,445 T > C V157A (GTA-> GCA)virB4_l Type IV secretion system protein virB4 1,435,170 A — *G T312A (ACC-^GCC) ppsA Phosphoenolpyruvate synthase Trifunctional nucleotide phosphoesterase protein 1,454,849 intergenic (+144 / -65) yfkN —> / — > mcp4

[0380] YfkN / Methyl-accepting chemotaxis protein 4 1,596,499 G > A A498T (GCT-*ACT) ftsl^ Peptidoglycan D, D-transpeptidase FtsI 1,596,628 G — > A A541T (GCG-yACG) ft si- Peptidoglycan D, D-transpeptidase FtsI d70az..,25

[0381]

[0382] 95,417 C--> T noncoding (1155 / 1501 nt) FAMPOCIGJ)0089 — 16S ribosomal RNA Attorney Docket No. 10063-110W01

[0383] 95,429 A— > C noncoding (1167 / 1501 nt) FAMPOC1G_00()89 16S ribosomal RNA

[0384] 95,444 +G noncoding (1182 / 1501 nt) FAMPOCIGJ00089 -> 16S ribosomal RNA

[0385] 95,447 Al bp noncoding (1185 / 1501 nt) FAMPOCIG_00089 16S ribosomal RNA

[0386] 221,035 (T)U^I5intergenic ( + 189 / -58) btuB_2 -» / ->• FAMPOCIG_00208 Vitamin B12 transporter BtuB / hypothetical protein 259,069 C-»T R30H (CGC--> CAC) rppH RNA pyrophosphohydrolase

[0387] 471,535 (T)S^9 coding (55 / 453 nt) Jwr «- Ferric uptake regulation protein

[0388] 606,899 C^T S351S (AGC-»AGT) ctpB —> Carboxy-terminal processing protease CtpB 706,730 A— > G M249V (ATG--> GTG) hmuU --> Hemin transport system permease protein HmuU I3T (ATC—> ACC) rny Ribonuclease Y

[0389] 935,212 A—> G V135A (GTT—> GCT) aroC Chorismate synthase

[0390] 1,001,805 G— -> T N847K (AAC--> AAA) mdtC Multi drug resistance protein MdtC

[0391] 1,061,351 G-> A G83D (GGC-+GAC) rho —► Transcription termination factor Rho 1,069,445 V157A (GTA— > GCA) virB4_l > Type I V secretion system protein virB4 1,435,170 A— > G T312A (ACC -> GCC) ppsA — > Phosphoenolpyruvate synthase

[0392] 1,596,499 G—> A A498T (GCT—> ACT) ftsl Peptidoglycan D, D-transpeptidase Ftsl 1,596,628 G— > A A541T (GCG^ACG) ftsl — » Peptidoglycan D, D-transpeptidase Ftsl d75az._25

[0393] 221,035 (T);^15 intergenic (+189 / -58) btuB_2 -» / ->• FAMPOCIG_00208 Vitamin B12 transporter BtuB / hypothetical protein 259,069 C^T R30H (CGC— > CAC) rppH <— RNA pyrophosphohydrolase

[0394] 471,535 (T)«-»9 coding (55 / 453 nt) fur <- Ferric uptake regulation protein

[0395] 606,899 U--> T S351S (AGC-> AGT) ctpB —> Carboxy-terminal processing protease CtpB 706,730 A— > G M249V (ATG^GTG) hmuU —>■ Hemin transport system permease protein HmuU I3T (ATC— *ACC) my — > Ribonuclease Y

[0396] 935,212 A--> G V135A (GTT--> GCT) aroC Chorismate synthase

[0397] 1,004,246 G— C L34V (CTT^GTT) mdtC Multidrug resistance protein MdtC 1,061,351 G — > A G83D (GGC — > GAC) rho Transcription termination factor Rho 1,069,445 T-*C V157A (GTA--> GCA) virB4_l Type IV secretion system protein virB4

[0398]

[0399] 1,435,170 A— > G T312A (ACC— ’•GCC) ppsA —>■ Phosphoenolpyruvate synthase Attorney Docket No. 10063-110W01

[0400] Trifunctional nucleotide phosphoesterase protein 1,454.849 (G)12^13 intergenic (+144 / -65) yfkN — > / — * mcp4

[0401] YfkN / Methyl-accepting chemotaxis protein 4 1,596,499 G -- > A A498T (GCT-> ACT) ftsl --> Peptidoglycan D. D-transpeptidase FtsI

[0402]

[0403] 1,596,628 G—> A A541T (GCG-yACG) ftsl Peptidoglycan D, D-transpeptidase FtsI Attorney Docket No. 10063-110WO1

[0404] Likewise, whole genome sequencing (WGS) showed that changes in H. pylori genome of DIS-resistant clones (Table 4). RNA-seq (LS Sciences, Texas, USA) revealed that both drugs upregulated groES, a heat shock protein, and downregulated kdsA, involved in membrane lipopolysaccharide synthesis, as well as ribH, a protein involved in riboflavin synthesis.

[0405] Table 4: Attorney Docket No. 10063-110W01

[0406] DIS-resistant mutants / Predicted Sequence- Clone 1

[0407] Evidence Position Mutation Annotation Gene

[0408] d52dis_0_25

[0409] 471,535 (T)s ^o coding (55 / 453 nt) fur « - Ferric uptake regulation protein

[0410] 606,899 C—G? S351S (AGC-»AGT) ctpB — > Carboxy-tenninal processing protease CtpB 706,730 A— -> G M249V (ATG--> GTG) hmuU — > Hemin transport system permease protein HmuU I3T (ATC— > ACC) rny Ribonuclease Y

[0411] 935,212 A— *G VI 35 A (GTT-> GCT) aroC Cborismate synthase

[0412] 1,069,445 T > C V157A (GTA—> GCA) virB4_l — * Type IV secretion system protein virB4 1,435,170 A — > G T312A (ACC— > GCC) ppsA Phosphoenolpyruvate synthase

[0413] Trifunctional nucleotide phosphoesterase protein 1,454,849 (G)12-.14 intergenic (+144 / -65) yfkN — > / — > mcp4

[0414] YfkN / Methyl-accepting chemotaxis protein 4 Undecaprenyl-phosphate

[0415] 1,574,975 G— > T G176W (GGG— > TGG) wecA —>■ alpha-N-acetylglucosaminyl 1 -phosphate transferase

[0416] d66disl„l„0

[0417] 471,535 (T)S^9coding (55 / 453 nt) fur <— Ferric uptake regulation protein

[0418] 606,899 C— *T S351S (AGC—AGT) ctpB — > Carboxy-tenninal processing protease CtpB 645,563 C—G? R41C (CGC—> TGC) inf A -» Translation initiation factor IF-1

[0419] 706,730 A— -> G M249V (ATG--> GTG) hmuU Hemin transport system permease protein HmuU I3T (ATC— > ACC) rny Ribonuclease Y

[0420]

[0421] Attorney Docket No. 10063-110W01

[0422] 935,212 A— > G VI 35 A (GTT^GCT) aroC Chorisrnate synthase

[0423] 1,069,445 T— > C V157A (GTA-H-GCA) virB4_l Type IV secretion system protein virB4 1,435,170 A— > G T312A (ACC~*GCC) ppsA — > Phosphoenolpyruvate synthase

[0424] Trifunctional nucleotide phosphoesterase protein 1,454,849 (G)12—.1,1 intergenic (+144 / -65) yfkN — > / mcp4

[0425] YfkNZMethyl-accepting chemotaxis protein 4 Undecaprenyl-phosphate

[0426] 1,574,975 G--> T G176W (GGG- -> TGG) wecA --> alpha-N-acetylglucosaminyl 1 -phosphate transferase

[0427] DIS-resistant mutants / Predicted Sequence- Clone 2

[0428] Evidence Position Mutation Annotation Gene

[0429] d62dislj)_25

[0430] 556,032 (T)16_15 intergenic (-179 / 4-172) btuB_3 i roc. F Vitamin B12 transporter BtuB / Arginase 706,730 A— > G M249V (ATG— > GTG) hmuU — * Hemin transport system permease protein HmuU 832,832 T— > C Y186Y (TAT— > TAC) FAMPOCIG_00821 -> hypothetical protein

[0431] I3T (ATC~»ACC) my Ribonuclease Y

[0432] d66dis..l.. O

[0433] 556,032 (T) 16^15 intergenic (-179 / +172) btuB_3 rocF Vitamin B12 transporter BtuB / Arginase 706,730 A— > G M249V (ATG— > GTG) hmuU -+ Hemin transport system permease protein HmuU 832,832 T—> C Y186Y (TAT-»TAC) FAMPOCIGJ)0821 hypothetical protein

[0434] I3T (ATC—ACC) my Ribonuclease Y

[0435] d70dis_l

[0436] 471,535 (T)S^9 coding (55 / 453 nt) / hr Ferric uptake regulation protein

[0437]

[0438] Attorney Docket No. 10063-110W01

[0439] 606,899 C— T S351S (AGC— AGT) ctpB — Carboxy-terminal processing protease CtpB 706,730 A—G M249V (ATG— GTG) hmuU — Hemin transport system permease protein HmuU I3T (ATC— ACC) my ----> Ribonuclease Y

[0440] 935,212 A—G VI 35 A (GTT— GCT) aroC — Chorismate synthase

[0441] 1,069,445 T— C V157A (GTA— GCA) virB4_l — Type IV secretion system protein virB4 1,201,469 C— A A594S (GCG— TCG) lepA — Elongation factor 4

[0442] 1,435,170 A— > G T312A (ACC— GCC) ppsA ■ Phosphoenolpyruvate synthase

[0443] d75_dis_0_3

[0444] 606,899 C— T S351S (AGC— AGT) ctpB — Carboxy-terminal processing protease CtpB 706,730 A—G M249V (ATG— GTG) hmuU — Hemin transport system permease protein HmuU I3T (ATC— ACC) my Ribonuclease Y

[0445] 935,212 A—G VI 35 A (GTT— GCT) aroC — Chorismate synthase

[0446] 1,069,445 T— C V157A (GTA— GCA) virB4__I — Type IV secretion system protein virB4 1,435,170 A—G T312A (ACC— GCC) ppsA — Phosphoenolpyruvate synthase

[0447] DIS-resistant mutants / Predicted Sequence- Clone 3

[0448] Evidence Position Mutation Annotation Gene

[0449] d66dis__l„5

[0450] 606,899 C— T S351S (AGC— AGT) ctpB — Carboxy-terminal processing protease CtpB 706,730 A—G M249V (ATG-*GTG) hmuU — Hemin transport system permease protein HmuU 13T (ATC— ACC) my — » Ribonuclease Y

[0451] 935,212 A—G VI 35 A (GTT— GCT) aroC — Chorismate synthase

[0452] 1,069,445 T— C V157A (GTA— GCA) virB4_l — Type IV secretion system protein virB4

[0453]

[0454] Attorney Docket No. 10063-110W01

[0455] 1,435,170 A— > G T312A (ACC^GCC) ppsA Phosphoenolpyruvate synthase

[0456] d70disl„l

[0457] 606,899 C-*T S351S (AGC-*AGT) ctpB ----> Carboxy-terminal processing protease CtpB 706,730 A— > G M249V (ATG^GTG) hmuU — > Hemin transport system permease protein HmuU 13'1' (ATC— »ACC) rny Ribonuclease Y

[0458] 935,212 A— > G V135A (GTT^GCT) aroC *— Chorismate synthase

[0459] 1,069,445 T--> C V 157 A (GTA--> GCA) virB4_l — * Type IV secretion system protein virB4 1,435,170 A— > G T312A (ACC^GCC) ppsA —>■ Phosphoenolpyruvate synthase

[0460]

[0461] Attorney Docket No. 10063-110WO1

[0462] Interestingly, AZ upregulated IpxE and DIS downregulated IpxE (lipid a-1 phosphatase). Host-pathogen interaction investigated using RNA-seq demonstrated that H. pylori infection upregulated inflammation-related genes, such as IL11, IL18, IRAK2, and IFNGR2, and oncogenic genes, such as JUNB, CREB5, LIPG, CDCP1, and PIM1 in the gastric cells, similar to previous reports. Notably, AZ and DIS inhibit FL pylori infection and decrease inflammation and oncogenic upregulation at 4xMIC (Fig. 6). The heatmap shows that critical inflammatory and oncogenes were upregulated by H. pylori infection, and AZ or DIS treatment controlled the upregulation of inflammatory genes and inflammation.

[0463] The metabolic perturbations in H. pylori under the influence of test drugs were compared to the global metabolic profile of H. pylori (Fig. 5). Amino acid and carbohydrate metabolism were regulated in AZ-treated cells, and sulfur, glutamate, and fatty acid profiling was moderated in DIS-treated cells.

[0464] It was found that AZ is inactive against Listeria monocytogens, but its analogs disclosed herein are active against L. monocytogens, and DIS is inactive against Acinetobacter baumannii, but its analogs disclosed herein are active against A. baumamii. AZ, DIS, and their analogs are also active against various gastric pathogens, including Salmonella sp. and Shigella sp. Furthermore, DIS analogs are more active than the parental molecule against the intracellular pathogen Staphylococcus aureus MW2.

[0465] Successful antimicrobial treatment against H. pylori infection is extremely challenging because the drug must penetrate thick mucus and remain active in the acidic gastric environment (Romano, M., el al., Eradication of Helicobacter pylori: a clinical update. Medscape General Medicine, 2004. 6(1): p. 19). These studies indicate that aztreonam (AZ), a monocyclic beta-lactam (Gordon, E. M., et al., Toward Orally Absorbed Prodrugs of the Antibiotic Aztreonam. Design of Novel Prodrugs of Sulfate Containing Drugs. Part 2. ACS medicinal chemistry letters, 2020. 11(2): p. 162-165) antibiotic, and disulfiram (DIS), used to treat alcohol dependence (Shirley, D.-A., et al., Drug Repurposing of the Alcohol Abuse Medication Disulfiram as an Anti-Parasitic Agent. Frontiers in Cellular and Infection Microbiology, 2021. 11), inhibit H. pylori growth and pathogenesis and remain active in acidic pH in vitro. Moreover, while II. pylori resistance to clarithromycin emerged after 20 days of continuous drug exposure, no resistance to AZ or DIS emerged even after 50 days of continuous exposure. AZ and DIS also showed synergistic anti-ZF. pylori activity with clinical antibiotics (clarithromycin or amoxicillin) and PPIs (omeprazole or pantoprazole). AZ or DIS downregulated H. pylori virulence and Attorney Docket No. 10063-110WO1

[0466] motility and hindered vacuolation in gastric epithelial ceils. Importantly, oral administration of AZ and DIS in triple therapy with amoxicillin and omeprazole decreased inflammation in the stomach and eliminated II. pylori adherence to gastric mucosal or epithelial cells after seven days of treatment. In other words, AZ and DIS show in vivo efficacy in preclinical settings.

[0467] Example 4: In vivo efficacy

[0468] AZ is an intravenous antibiotic with poor absorbance via the oral route. A formulation of ethyl alcohol (1:1) mixture with saline (1:10) was used for oral administration of AZ to combat H. pylori. The same formulation was used to deliver DIS, which is already available as an oral drug that can survive in acidic pH. The in vivo efficacy of oral AZ and DIS was tested in a C57BL / 6J mouse-H. pylori infection model. Mice were gavaged with a high dose of H. pylori SSI (2xl09 / 0.2 mL) three times a week for two weeks. Bacterial colonization was checked by H. pylori shedding at 7, 14, and 21 days after infection using an H. py / ori-specific primer in PCR. After three weeks, vehicle, AZ (25mg / kg), DIS (20 mg / kg), or control antibiotics + PPI (amoxicillin + clarithromycin + omeprazole) were administered to the infected mice for 14 days. After therapy, the stomach tissue sections were stained for H& E. The Gl-histopathologist scored the slides (blinded), and the inflammation scores suggest that AZ at 25 mg / kg or DIS at 20 mg / kg reduces inflammation (Fig. 2A) in the bacteria-colonized area, compared to control or drug treatment. The inflammation score was obtained based on leukocyte infiltration due to H. pylori infection (Fig. 2D; H& E stain).

[0469] Immunohistochemistry (IHC) was used with an H. pylori polyclonal antibody to visualize H. pylori in the stomach tissue sections. The no-treatment group showed that H. pylori adhered to the gastric epithelial cells, and AZ or DIS cleared the bacterial adherence (Fig. 2D; H. pylori IHC). Both drugs, in single therapy, were effective against H. pylori after a standard 14-day course of therapy. Based on these results, a shorter treatment course with AZ or DIS in triple therapy was hypothesized to be effective against H. pylori in vivo. A seven-day course was used to reduce the chance of resistance emergence. The same C57BL / 6J- mice-H. pylori infection model was used with a change in the bacterial strain to PMSS1. After three weeks of II. pylori infection, the mice were treated with AZ or DIS as single therapy, dual therapy (drug+PPI), or triple therapy (drug+PPI+clinical antibiotic), generating the following groups: a) AZ, b) AZ+PPI (omeprazole) (AZP), c) AZ+PPI (omeprazole)+antibiotic (amoxicillin) (AZPA), d) DIS e) DIS+PPI (omeprazole) (DISP), c) DIS+PPI (omeprazole)+antibiotic (amoxicillin) (DISPA). Seven days of triple therapy Attorney Docket No. 10063-110WO1

[0470] cleared bacterial adherence and reduced inflammation (Figs. 2B-2C). H& E stains show decreased leukocyte infiltration results in decreased inflammation and bacterial attachment (Fig. 2E). The results indicate that AZ and DIS in triple therapy can combat H. pylori in vivo.

[0471] Example 5: Gutmicrobiome analysis

[0472] Fecal samples from mice used in the preliminary studies described above were analyzed. Fecal pellets were isolated from individual mice and outsourced for genomic DNA isolation, 16s sequencing, and bioinformatic analysis (University of Minnesota Microbiome Core). It was found that AZ or DIS in single therapy had less impact on the gut microbiome after seven days of therapy, compared to triple therapy (Figs. 4A-4D).

[0473] However, AZ or DIS in triple therapy had a similar impact compared to standard antibiotic therapy (amoxicillin + clarithromycin + omeprazole). The AZ at 25 mg / kg or DIS at 20 mg / kg in triple therapy cleared bacterial adhesion and reduced inflammation score similar to the standard antibiotic control group (amoxicillin + clarithromycin + omeprazole) (Figs.

[0474] 4A-4D). Interestingly, the gut microbiome analysis with Principal Coordinate Analysis (PCoA) analysis based on Bray Curtis (abundance) and Unifrac (taxonomy) distance showed that AZPA and DISPA had microbiome profiles similar to that of the standard antibiotic therapy grouped AB (Figs. 4A-4D). The abundance is the factor differentiating keystone microorganisms from those that are dominant, and taxonomic composition helps identify the different species and strains present. A diverse range of taxa is generally associated with a healthier gut microbiome. The inclusion of amoxicillin and omeprazole explains the gut-microbiome shift as neither AZ. nor DIS induced changes when administered alone, with poor rescue from inflammation. It was also found that major beneficial bacterial populations, Akkermansia and Bacteroides, were increased after triple therapy with AZ or DIS compared to healthy, uninfected mice (Figs.4A-4D).

[0475] Example 6: AZ and DIS analog activity

[0476] Of the DIS analogs, (22 compounds tested) the following activity was observed. Twelve analogs (DIS 1-2, D1S2-2; DIS3-2, DIS4-2; DIS5-2, D1S6-2, DIS2-7, DIS9-2, DIS 10-2, DIS 11-2, DIS13-2, DIS 17-2) were active against H. pylori (Gram-negative). Three analogs (DIS9-2, DIS 10-2, DIS 11 -3) were active against A. baumannii (Gram¬ negative). Six analogs (DISl-2, DIS2-2, DIS5-2, DIS6-2, DIS7-2, DIS13-2) were active against L. monocytogenes (Gram-positive). Two (DISl-2 and DIS5-2) were active against Enterococcus faecium (Gram-positive). Eight (DISl-2, DIS2-2, DIS3-2, DIS5-2, DIS6-2, Attorney Docket No. 10063-110WO1

[0477] DIS-7, DIS-9, DIS-13) were active against Staphylococcus aureus (Gram-positive). Data is shown in Table 5 and Fig. 6. Attorney Docket No. 10063-110W01

[0478] Table 5:

[0479] Agents Minimum inhibitory concentration (mg / L)

[0480] MW2 MW2+

[0481] HP EF VRA1 AB KP EA EA PAU LM BP82 SE SF CAB CAU1 CGL 1C50 CAMFIB 10%FBS

[0482] DIS 1-2.06 2 4 8 2 >64 >64 >64 >64 >64 8 >64 >64 >64 >64 >64 16 64 DIS2-2.03 4 6 2 >64 >64 >64 >64 >64 16 >64 >64 >64 >64 >64 >64 64 DIS3-2.06 4 4 2 4 >64 >64 >64 >64 >64 32 >64 >64 >64 >64 >64 16 64 DIS4-2 4 >64 64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 64 DIS5-2.06 1 4 8 1 >64 >64 >64 >64 >64 8 >64 >64 >64 32 32 8 64 DIS6-2.03 1 4 6 1 >64 >64 >64 >64 64 8 >64 >64 >64 >64 >64 4 64 DIS7-2.06 2 2 4 >64 >64 >64 >64 64 16 >64 >64 >64 32 32 16 64 DIS8-2 2 >64 64 >64 >64 >64 >64 >64 >64 >64 4 64 8 >64 >64 >64 64 DIS9-2.06 2 4 2 8 >64 >64 >64 >64 32 4 32 32 4 4 4 64 DIS 10-2.06 16 4 16 >64 >64 >64 >64 64 4 64 16 4 8 8 64 DIS 11 -2.125 16 4 16 16 >64 >64 >64 >64 32 >64 >64 >64 8 8 16 64 DIS 12-2 2 64 64 64 32 >64 >64 >64 >64 34 >64 >64 >64 8 16 4 64 DIS 13-2.125 2 4 64 2 >64 >64 >64 >64 >64 8 >64 >64 >64 >64 >64 >64 64 DIS 14-2.25 >64 64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 64 DIS 15-2 4 >64 64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 64 DIS 17-2.125 >64 64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 64 DIS 1-3 4 >64 64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 64 DIS2-3 8 >64 64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 64 DIS3-3 8 >64 64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 64 DIS4-3 6 >64 64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 64 DIS5-3 4 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 64 DIS6-3 8 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 >64 64

[0483]

[0484] DIS.125 4-8 4-8 8 4-8 16 64 >64 >64 >64 16 16 64 32 16 16 16 16-32 Attorney Docket No. 10063-110WO1

[0485] HP- Helicobacter pylori,' MW2- S. aureus strain MW2; EF -Enterococcus f aecium; AB- Acinetobacter baumannii; KP- Klebsiella pnemoniae; EA- Enterobacter aerogenes; PA14- Pseudomonas aeruginosa LM- Listeria monocytogens; BP82- Burkholderia pseudomallei BSL2 strain 82; SE- NR-169 Salmonella enterica subsp. enterica CIP 60.62 (Serovar Typhimurium); SF- Shigella flexneri; CAB- Candida albicans strain CAN 14; CAU1- Candida auris strain CAU1; CGL- Candida glabrata; IC-50 - Cytotoxicity against HEPG2 cells.

[0486] Of the AZ Analogs, (19 compounds tested) the following activity was observed. Five analogs (AZ3-2, AZ4-2, KZH-, ALL-1, AZ- 8) showed activity against Helicobacter pylori (Gram-negative). Four analogs (AZ1-2; AZ1, AZ7, AZ8) showed activity against Acinetobacter baumannii (Gram-negative). Eight analogs (AZ1-2, AZ3-2, AZ4-2, A71-2, AZ1, AZ2, ALLI, ALLS showed activity against Enterobacteriacea sp. Klebsiela pneumoniae (Gram-negative). Eighteen (AZ1-2, AZ2-2, AZ3-2, AZ4-2, AZ5-2, AZ6-2, AZ7-2, AZ8-2 and AZ1, AZ2, AZA, AZ4, AZ5, ALLL1, AZ8, AZ10, AZ11, AZ12) showed activity against Listeria monocytogenes (Gram-positive). Data is show m in Table 6. Attorney Docket No. 10063-110W01

[0487] Table 6:

[0488] Agents Minimum inhibitory concentration (mg / L)

[0489] HP MW2 MW2 MW2+ EE VRA1 AB KP FA PAW LM BP82 SE SF CAB CAU1 CGL IC50

[0490] CAMHB 10%FBS

[0491] AZ 1-2 2 NT NT NT NT NT 32 8 >64 >64 1 >64 8 1 >64 >64 >64 64 AZ2-2 1 NT NT NT NT NT 64 4 >64 >64 8 >64 8 1 >64 >64 >64 64 AZ3-2 0.25 NT NT NT NT NT 64 2 >64 >64 32 >64 4 1 >64 >64 >64 64 AZ4-2 0.5 NT NT NT NT NT 32 2 >64 64 8 >64 4 0.5 >64 >64 >64 64 AZ5-2 1 NT NT NT NT NT 64 4 >64 >64 16 >64 8 2 >64 >64 >64 64 AZ6-2 16 NT NT NT NT NT >64 16 >64 >64 64 >64 16 4 >64 >64 >64 64 AZ7-2 0.25 NT NT NT NT NT 32 2 >64 32 4 >64 >64 2 >64 >64 >64 64 AZ8-2 2 NT NT NT NT NT 64 4 >64 64 4 4 2 2 >64 >64 >64 64 AZ1 8 NT NT NT NT NT 4 2 >64 >64 1 4 2 1 >64 >64 >64 64 AZ2 1 NT NT NT NT NT 8 1 >64 >64 1 4 1 0.5 >64 >64 >64 64 AZ4 2 NT NT NT NT NT 32 16 >64 >64 2 >64 4 2 >64 >64 >64 64 AZ5 2 NT NT NT NT NT 32 16 >64 >64 2 >64 16 4 >64 >64 >64 64 AZ6 >64 NT NT NT NT NT >64 >64 >64 >64 >64 >64 >64 30 >64 >64 >64 64 AZ7 0.25 NT NT NT NT NT 4 0.5 >64 >64 2 >64 0.5 0.5 >64 >64 >64 64 AZ8 0.5 NT NT NT NT NT 4 0.5 >64 >64 2 >64 0.5 0.5 >64 >64 >64 64 AZ9 1 NT NT NT NT NT >64 >64 >64 >64 >64 >64 >64 32 >64 >64 >64 64 AZ10 1 NT NT NT NT NT 16 16 >64 >64 2 >64 4 4 >64 >64 >64 64 AZ 11 2 NT NT NT NT NT 8 16 >64 >64 2 >64 16 4 >64 >64 >64 64 AZ12 4 NT NT NT NT NT 8 4 >64 >64 1 >64 2 1 >64 >64 >64 64

[0492]

[0493] AZ 0.25 NT NT NT NT NT 4 0.5 4 4 64 >64 0.5 0.5 >64 >64 >64 64 Attorney Docket No. 10063-110WO1

[0494] HP- Helicobacter pylori,' MW2- S. aureus strain MW2: EF -Enterococcus f aecium; AB- Acinetobacter baumannii; KP- Klebsiella pnemoniae; EA- Enterobacter aerogenes; PA14- Pseudomonas aeruginosa; LM- Listeria monocytogens; BP82- Burkholderia pseudomallei BSL2 strain 82; SE- NR- 169 Salmonella eitierica subsp. enterica CIP 60.62 (Serovar Typhimurium); SF- Shigella jlexneri; CAB- Candida albicans strain CAN14; CAU1- Candida auns strain L'AlJ 1 CGL- Candida gicibrata; IC-50 — Cytotoxicity against I'0iPG2 cells.

[0495] Notably, AZ7 and AZ8 with aliphatic chains and DIS5-2 and DIS6-2 (featuring fused pyrrole and piperidine rings) exerted enhanced antibacterial activity against H. pylori compared to their parental molecules (Figs. 2A-2C) with low cytotoxicity.

[0496] Scanning electron microscopy demonstrates H. pylori morphological change after parental compounds treatment at lOxMIC (Fig. 7).

[0497] Scanning electron microscopy demonstrates S. aureus MW2 morphological change after DIS and DIS 5-2 analog treatment at lOxMIC (Fig. 8).

[0498] A broth dilution assay at various pH show DIS analogs 1-2, 5-2, 6-2, 7-2, 9-2, 13-2 has enhanced antibacterial activity at acidic pH (Table 7). Killing kinetics show

[0499] Table 7:

[0500] Agents pH7 pH5 pH4

[0501] DIS 1-2 2 0.5 0.5-0.25

[0502] DIS 2-2 4 >64 >64

[0503] DIS 5-2 2 0.5 0.125-0.06

[0504] DIS 6-2 2 2 1-0.5

[0505] DIS 7-2 4 1 0.125-0.06

[0506] DIS 9-2 2 2 0.25

[0507] DIS 13-2 4 0.5 0.5

[0508] DIS 8 64 64

[0509]

[0510] The activity of various disulfiram analogs against Neisseria gonorrhoea is shown in Table 8.

[0511] Table 8: Antibacterial activity of DIS and its analogs against Neisseria gonorrhea reference strains, and clinical strains. Attorney Docket No. 10063-110WO1

[0512] Agents Minimum inhibitory concentration (mg / L)

[0513] Reference strain Clinical Strains BEI reference strain

[0514] NG ATCC 49226 NG 914 NG 1281 NG 1145734 DIS 1-2 0.5 0.125 0.06 0.5

[0515] DIS2-2 8 0.25 0.06 0.5

[0516] DIS3-2 1 0.125 0.06 0.25

[0517] DIS4-2 >32 >32 4 8

[0518] DIS5-2 0.5-1.0 0.125 0.03 0.25

[0519] DIS6-2 4 0.25 0.06 0.25

[0520] DIS7-2 1 0.125 0.06 0.125

[0521] DIS8-2 32 32 9 8

[0522] DIS9-2 0.25-0.5 0.25 0.06 0.25

[0523] DIS 10-2 2 0.125 0.06 0.5

[0524] DISH -2 1-2 0.125 0.06 0.25

[0525] DIS12-2 16 4 1 4

[0526] DIS 13-2 16 4 9 2

[0527] DIS 14-2 8 8 0.25 1

[0528] DIS 15 -2 >32 4 4 16

[0529] DIS17-2 8 16 0.25 4

[0530] DIS 1-3 >32 >32 >32 >32

[0531] DIS2-3 >32 >32 >32 >32

[0532] DIS3-3 >32 >32 32 >32

[0533] DIS4-3 >32 >32 32 >32

[0534] DIS5-3 >32 >32 32 32

[0535] DIS6-3 >32 >32 32 16

[0536] DIS 8 0.125 0.03 0.25

[0537] Penicillin G >16 >32 >32 NT

[0538] Ciprofloxacin 0.1 16 8 NT

[0539] Erythromycin 0.25 0.5 2 NT

[0540]

[0541] Tetracycline 0.5 2 1 NT

[0542] Table 9: Antibacterial activity of AZ and its analogs against Neisseria gonorrhea reference and clinical strains.

[0543] Agents Minimum inhibitory concentration (mg / L)

[0544] Reference strain Clinical Strains BEI reference strain NG ATCC 49226 NG 914 NG 1281 NG 1145734

[0545] AZ1-2 4 >8 >8 0.5

[0546] AZ2-2 1-2 >8 >8 0.25

[0547] AZ.3-2 4 >8 >8 1

[0548] AZ4-2 1 >8 >8 0.25

[0549] AZ5-2 2-4 >8 >8 0.25

[0550]

[0551] AZ6-2 >32 >8 >8 4 Attorney Docket No. 10063-110WO1

[0552] AZ7-2 2-4 >8 >8 0.25

[0553] AZ8-2 8 >8 >8 1

[0554] AZ1 4 >8 >8 0.5

[0555] AZ2 1-2 >8 >8 0.25

[0556] AZ4 4-8 >8 >8 2

[0557] AZ5 4-8 >8 >8 2

[0558] AZ6 >32 >8 >8 8

[0559] AZ7 0.25 4 4 2

[0560] AZ8 2 >8 >8 1

[0561] AZ9 >32 >8 >8 32

[0562] AZ10 4 >8 >8 2-1

[0563] AZ11 8 >8 >8 4-2

[0564] AZ12 8 >8 >8 2

[0565] AZ 0.125 2 4 0.01

[0566] Penicillin G >16 >32 >32 NT

[0567] Ciprofloxacin 0.1 16 8 NT

[0568] Erythromycin 0.25 0.5 2 NT

[0569]

[0570] Tetracycline 0.5 2 1 NT

[0571] Table 10: Minimal inhibitory concentration against Helicobacter pylori, methicillin resistant Staphylococcus aureus, A. baumannii

[0572] Agents H. pylori MRSA-MW2 A. baumannii

[0573] LHM-01 0.06 2 >128

[0574] LHM-02 0.03 2 >128

[0575] LHM-03 0.25 >128 >128

[0576] LHM-04 0.125 4 >128

[0577] LHM-05 0.125 4 >128

[0578] LHM-06 2 >128 >128

[0579] LHM-07 0.25 8 >128

[0580] LHM-08 0.5 16 >128

[0581] LHM-09 0.06 2 >128

[0582] LHM-10 0.06 1 >128

[0583] LHM-11 0.06 1 >128

[0584] LHM-12 0.25 8 >128

[0585] LHM-13 1 16 >128

[0586] LHM-14 1 16 >128

[0587] LHM-15 0.25 4 >128

[0588] LHM-16 0.25 16 >128

[0589] LHM-17 0.25 8 >128

[0590] LHM-18 8 >128 >128

[0591] LHM-19 0.06 165 >128

[0592] LHM-20 2 >128 >128

[0593] LHM-21 0.25 4 >128

[0594] LHM-22 0.06 8 >128

[0595] LHM-23 1 8 32

[0596]

[0597] LHM-24 4 >128 >128 Attorney Docket No. 10063-110WO1

[0598] LHM-25 0.25 39 >128

[0599]

[0600] LHM-26 1 4 >128

[0601] LHM-27 2 32 >128

[0602] LHM-28 0.125 2 >128

[0603] LHM-29 0.5 4 >128

[0604] LHM-30 0.125 4 >128

[0605] LHM-31 0.06 4 >128

[0606] LHM-32 1 16 >128

[0607] LHM-33 0.06 4 >128

[0608] LHM-34 0.125 8 >128

[0609] LHM-35 0.125 8 >128

[0610] LHM-36 1 >128 >128

[0611] LHM-37 1 >128 >128

[0612] LHM-38 2 >128 >128

[0613] LHM-39 1 16 >128

[0614] LHM-40 8 >128 >128

[0615] LHM-41 0.06 4 >128

[0616] LHM-42 0.25 16 >128

[0617] LHM-43 0.125 8 >128

[0618] LUM-44 8 >128 >128

[0619] LHM-45 4 >128 >128

[0620] LHM-46 4 >128 >128

[0621] LHM-47 2 >128 >128

[0622] LHM-48 8 >128 >128

[0623] LHM-49 16 >128 >128

[0624] LHM-50 2 >128 >128

[0625] LHM-51 0.125 4 >128

[0626] LHM-52 1 32 >128

[0627] LHM-53 2 >128 >128

[0628] LHM-55 2 >128 >128

[0629] LHM-56 0.125 8 >128

[0630] LHM-62 0.25 8 >128

[0631] LHM-63 0.125 8 >128

[0632] LHM-64 8 >128 >128

[0633] LHM-65 0.5 16 32

[0634] LHM-66 1 8 64

[0635] LHM-67 16 8 >128

[0636] LHM-68 16 >128 >128

[0637] LHM-69 16 >128 >128

[0638] LHM-70 16 >128 >128

[0639] LHM-71 8 >128 >128

[0640] LHM-78 0.5 4 >128

[0641] LHM-80 0.5 8 >128

[0642] LHM-81 2 16 >128

[0643] LHM-82 1 4 >128

[0644] LHM-83 1 4 >128

[0645] LHM-84 1 4 >128

[0646]

[0647] LHM-87 0.5 16 >128 Attorney Docket No. 10063-110WO1

[0648] Example 7: DIS analog synthesis

[0649] DIS series 2 compounds.

[0650] The di-sulfur backbone in DIS was used in new designs around the left-hand side of the molecule - carbocyclic ring (compounds DIS1-2, DIS2-2, DIS5-2, D1S6-2, DIS9-2, DIS10-2), pyrane (compounds DIS3-2, DIS7-2, DISll-2), iPr substitution (compounds DLS4-2, DIS8-2, DIS12-2), spiro ring (compound DIS13-2), bicyclic ring (compound DIS14-2, D1S15-2), and benzyl ring (compound DIS17-2).

[0651] Synthesis of octylsulfanyl pyrrolidine- 1 -carbodithioate

[0652] B(1 eq), CS2(1 eq), Et3N

[0653] (1.1 eq), CBr4(1.96 eq), dry DCM (10 mL), RT. 2 h.

[0654]

[0655] DIS1-2

[0656] To a stirred solution of pyrrolidine-(B) (0.11 mL, 1.36 mmol, 1 eq) and octane-1- thiol A (200 mg, 1.36 mmol, 1 eq) were added in anhydrous DCM (10 mL), the reaction was cooled in an ice bath. Carbon disulfide (0.082 mL, 1.36 mmol, 1 eq) was added drop wise, followed by the slow addition of triethylamine (0.2 mL, 1.49 mmol, 1.1 eq) stirred for 5 min. After solution of CB (904 mg, 2.67 mmol, 1.96 eq) in DCM (2mL) was added. The resulting reaction mixture was allowed to stir at room temperature for 2 h. Reaction was monitored by TLC. After completion, Reaction mixture was diluted with H2O (4 mL) and extracted with DCM (2 x 10 mL). The organic layers were dried over sodium sulphate and concentrated to get crude. The Crude was purified by 230-400 pre-packed Redisep column on combi flash chromatography, eluted using 20 -25 % of DCM in Heptane to afford octylsulfanyl pyrrolidine-l-carbodithioate, DIS1-2 (200 mg, 50.2 %) as pale-yellow liquid.

[0657] TLC: 50 % DCM in Heptane (Ry. 0.5).JH NMR (400 MHz, CHLOROFORM-d) 5 = 3.97 (t, J = 7.0 Hz, 2H), 3.74 (t, J = 6.9 Hz, 2H), 2.88 - 2.83 (m, 2H), 2.15 - 2.07 (m, 2H), 2.03 - 1.96 (m, 2H), 1.67 (quin, J = 7.4 Hz, 2H), 1.42 - 1.36 (m, 2H), 1.27 (br s, 8H), 0.90 - 0.85 (m, 3H) ppm. LCMS: (ES+) 97.59 %, m / z= 292.3 [M+H]+[Method:- Column:

[0658] CORTECS UPLC C18(3X30mm) Mobile Phase A: 0.05 % FA in Water, B: 0.05 % FA in ACN]; HPLC: 97.24% purity [Method:- Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile (95:05), B - Acetonitrile]. Attorney Docket No. 10063-110WO1

[0659] Synthesis of octylsulfanyl piperidine- 1 -carbodithioate

[0660] B(1 eq), CS2(1 eq), Et3N

[0661] (1.1 eq), CBr4(1.96 eq), g 1 dry DCM (10 mL), RT, 2 h. JIsHS

[0662] % - ' C Js' '

[0663]

[0664] A DIS2-2 To a stirred solution of Piperidine B (0.14 mL, 1.36 mmol, 1 eq) and octane-1- thiol A (200 mg, 1.36 mmol, 1 eq) were added in anhydrous DCM (10 mL), the reaction was cooled in an ice bath. Carbon disulfide (0.08 mL, 1.36 mmol, 1 eq) was added drop wise, followed by the slow addition of triethylamine (0.2 mL, 1.49 mmol, 1.1 eq) stirred for 5 min. After solution of CBto (904 mg, 2.67 mmol, 1.96 eq) in DCM (2mL) was added. The resulting reaction mixture was allowed to stir at room temperature for 2 h. Reaction was monitored by TLC, After completion, Reaction mixture was diluted with H? O (4 mL.) and extracted with DCM (2 x 10 mL). The organic layers were dried over sodium sulphate and concentrated to get crude. The crude was purified by 230-400 pre-packed Redisep column on combi flash chromatography, eluted using 20-25 % of DCM in Heptane to afford octylsulfanyl piperidine-l-carbodithioate, DIS2-2 (100 mg, 23.9 %) as pale-yellow liquid.

[0665] TLC: 50 % DCM in Heptane (Rf. 0.5). NMR (400 MHz, CHLOROFORM-d) 5 = 4.30 (br s, 2H), 3.98 (br s, 2H), 2.90 - 2.81 (m, 2H), 1.70 - 1.62 (m, 2H), 1.54 (s, 1H), 1.44 - 1.35 (m, 2H), 1.32 - 1.23 (m, 8H), 0.91 - 0.85 (m, 3H) ppm. LCMS: (ES+) 98.27 %, m / z= 305.87 [M+H]+[Method:- Column: CORTECS UPLC C18 (3*30)mm, 1.6um, Flow rate: 0.85 mL / min. Mobile Phase A: 0.05 % TEA in Water, Mobile Phase B: 0.05%TFA in Acetonitrile]; HPLC: 99.01% purity [Method:- Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile (95:05), B - Acetonitrile], Synthesis of octylsulfanyl morpholine-4-carbodithioate B(1 eq), CS2(1 eq), Et3N CL (1.1 eq), CBr4(1.96 eq), SHS / x - [ ] dry DCM (10 mL), RT, 2 h. ^

[0666] 'N |Sn A B DIS3-2 To a stirred solution of Morpholine B (0.12 mL, 1.36 mmol, 1 eq) and octane-1- thiol A (200 mg, 1.36 mmol, 1 eq) in anhydrous DCM (10 ml,), the reaction was cooled in

[0667]

[0668] an ice bath. Carbon disulfide (0.08 mL, 1.36 mmol, 1 eq) was added drop wise, followed by the slow addition of triethylamine (0.2 mL, 1.49 mmol, 1.1 eq) stirred for 5 min, after solution of CBr4(904 mg, 2.67 mmol, 1.96 eq) in DCM (2mL) was added. The resulting Attorney Docket No. 10063-110WO1

[0669] reaction mixture was allowed to stir at room temperature for 2 h. The progress of reaction was monitored by TLC. After completion, Reaction mixture was diluted with H? O (4 ml.,) and extracted with DCM (2 x 10 mL). The organic layers were dried over sodium sulphate and concentrated to get crude. The crude was purified by 230-400 pre-packed Redisep column on combi flash chromatography, eluted using 30 % of EtOAc in Heptane to afford octylsulfanyl moipholine-4-carbodithioate, DIS3-2 (250 mg, 59.4 %) as pale-yellow liquid.

[0670] TLC: 50 % EtOAc in Heptane

[0671]

[0672] (Rf. 0.6). NMR (400 MHz, CHLOROFORM-d) 5 = 4.21 (br s, 4H), 3.83 - 3.74 (m, 4H), 2.86 (t, 7.4 Hz, 2H), 1.66 (quin, J = 7.4 Hz, 2H), 1.44 - 1.35 (m, 2H), 1.27 (br s, 8H), 0.91 - 0.85 (m, 3H) ppm. LCMS: (ES+) 99.13 % 308.2 [M+H]+, [Method: - Column: CORTECS UPLC C18 (3X30mm), Mobile Phase A: 0.05 % FA in Water, B: 0.05% FA in ACN]; HPLC: 97.9 % purity [Method:- Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile (95:05), B - Acetonitrile).

[0673] Synthesis of octyl diisopropylcarbamo(dithioperoxo)thioate

[0674] B(1 eq), CS2(1 eq), Et3N

[0675] Y (L1 eq). CBr4(1.96 eq), I+\^NH dry DCM (10 mL), RT, 2,h.

[0676]

[0677] A B ' DIS4-2 To a stirred solution of Diisopropyl amine B (0.19 mL, 1.36 mmol, 1 eq) and octane-1- thiol A (200 mg, 1.36 mmol, 1 eq) in anhydrous DCM (10 mL), the reaction was cooled in an ice bath. Carbon disulfide (0.08 mL, 1.36 mmol, 1 eq) was added drop wise, followed by the slow addition of triethylamine (0.2 mL, 1.49 mmol, 1.1 eq) stirred for 5 min, after solution of C B: 4 (904 mg, 2.67 mmol, 1.96 eq) in DCM (2mL) was added. The resulting reaction mixture was allowed to stir at room temperature for 2 h. The progress of reaction was monitored by TLC. After completion, the reaction mixture was diluted with H2O (4 mL) and extracted with DCM (2 x 10 mL). The organic layers were dried over sodium sulphate and concentrated to get crude. The crude was purified by 230-400 pre-packed Redisep column on combi flash chromatography, eluted using 15 % of DCM in Heptane to afford octyl diisopropylcarbamo(dithioperoxo)thioate, DIS4-2 (20 mg, 4.5 %) as pale¬ yellow liquid.

[0678] TLC: 50 % DCM in Heptane

[0679]

[0680] (Ry. 0.6) NMR (400 MHz, CHLOROFORM-d) 5 = 5.62 - 4.58 (m, 2H), 2.85 (t, J = 7.3 Hz, 2H), 1.66 (td, J = 7.4, 14.9 Hz, 3H), 1.51 - 1.36 (m, UH), 1.27 (br s, 10H), 0.88 (t, J = 6.8 Hz, 3H) ppm. LCMS: (ES+) 95.01 % m / z= 321.96 [M+H]* [Method:- Column: CORTECS UPLC C18 (3*30)mm, 1.6um, Flow' rate: 0.85 Attorney Docket No. 10063-110WO1

[0681] mL / min. Mobile Phase A: 0.05 % TFA in Water, Mobile Phase B: 0.05 % TFA in Acetonitrile]; HPLC: 93.15 % purity (Column: X-Select CSH C18 (4.6*150) mm

[0682] 5 u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile (95:05), B - Acetonitrile).

[0683] Synthesis of heptylsulfanyl pyrrolidine- 1 -carbodithioate

[0684] B(1 eq), CS2(1 eq), Et3N

[0685] (1.1 eq), CBr4(1.96 eq), + dry DCM (10 mL), RT, 2^.

[0686]

[0687] A DIS5-2 To a stirred solution of Pyrrolidine B (0.19 mL, 2.26 mmol, 1 eq) and heptane-1- thiol A (300 mg, 2.26 mmol, 1 eq) in anhydrous DCM (10 mL), the reaction was cooled in an ice bath. Carbon disulfide (0.13 mL, 2.26 mmol, 1 eq) was added dropwise, followed by the slow addition of triethylamine (0.35 mL, 2.49 mmol, 1.1 eq) stirred for 5 min, after solution of CBI-4 (1.5 g, 4.53 mmol, 2 eq) in DCM (2mL) was added. The resulting reaction mixture was allowed to stir at room temperature for 2 h. The progress of reaction was monitored by TLC. After completion, the reaction mixture was diluted with H2O (4 mL) and extracted with DCM (2 x 10 mL). The organic layer was dried over sodium sulphate and concentrated to get crude. The crude was purified by 230-400 pre-packed Redisep column on combi flash chromatography, eluted using 20-25% of DCM in Heptane to afford heptylsulfanyl pyrrolidine- 1 -carbodithioate, DIS5-2 (150 mg, 23.8 %) as pale-yellow liquid.

[0688] TLC: 50 % DCM in Heptane (R

[0689]

[0690] f. 0.5). NMR (400 MHz, CHLOROFORM-d) 8 = 3.97 (t, J = 6.9 Hz, 2H), 3.74 (t, J = 6.9 Hz, 2H), 2.89 - 2.84 (m, 2H), 2.15 - 2.08 (m, 2H), 2.03 - 1.96 (m, 2H), 1.67 (quin, J = 7.4 Hz, 2H), 1.42 - 1.23 (m, 8H), 0.90 - 0.86 (m, 311) ppm. LCMS: (ES+) 96.59 %, m / z= 278 [M+H]+[Method:- Column: CORTECS UPLC C18 (3*30)mm, 1.6umFlow rate: 0.85 mL / min, Mobile Phase A: 0.05 % TFAin Water, Mobile Phase B: 0.05 % TFA in Acetonitrile]; HPLC: 96.5% purity [Method:- Column: X- Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1 % Formic acid in water:

[0691] Acetonitrile(95:05), B - Acetonitrile].

[0692] Synthesis of heptylsulfanyl piperidine-1 -carbodithioate

[0693] B(1 eq), CS2(1 eq), Et3N

[0694] (1.1 eq), CBr4(2 eq), dry DCM (10 mL), RT, 2 h.

[0695]

[0696] D! S6-2 To a stirred solution of piperidine B (0.22 mL, 2.26 mmol, 1 eq) and Heptane- 1- thiol A (300 mg, 2.26 mmol, 1 eq) in anhydrous DCM (10 mL), the reaction was cooled in Attorney Docket No. 10063-110WO1

[0697] an ice bath. Carbon disulfide (0.13 mL, 2.26 mmol, 1 eq) was added dropwise, followed by the slow addition of triethylamine (0.35 mL, 2.49 mmol, 1.1 eq) stirred for 5 min, After solution of ( Bi: (1.5 g, 4.53 mmol, 2 eq) in DCM ( 2ml. ) was added. The resulting reaction mixture was allowed to stir at room temperature for 2 h. The progress of reaction was monitored by TLC. After completion, the reaction mixture was diluted with H2O (4 mL) and extracted with DCM (2 x 10 mL). The organic layer was dried over sodium sulphate and concentrated to get crude. The crude was purified by 230-400 pre-packed Redisep column on combi flash chromatography, eluted using 20-25% of DCM in Heptane to afford heptylsulfanyl piperidine- 1 -carbodi thioate, DIS6-2 (150 mg, 22.6 %) as pale-yellow liquid.

[0698] TLC: 50 % DCM in Heptane

[0699]

[0700] (Rf. 0.5). NMR (400 MHz, CHLOROFORM-d) 5 = 4.31 (br s, 2H), 3.99 (br s, 2H), 2.88 - 2.84 (m, 2H), 1.69 - 1.62 (m, 7H), 1.42 - 1.26 (m, 8H), 0.91 - 0.86 (m, 3H) ppm. LCMS: (ES+) 98.58 %, m / z= 292 [M+H]+[Method:- Column: CORTECS UPLC C18 (3*30)mm, 1.6um Flow rate: 0.85 mL / min. Mobile Phase A: 0.05 % TFA in Water, Mobile Phase B: 0.05 % TFA in Acetonitrile]; HPLC: 99.0 % purity [Method:- Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile(95:05), B - Acetonitrile].

[0701] Synthesis of heptylsulfanyl morpholine-4-carbodithioate

[0702] B(1 eq), CS2(1 eq), Et3NsAX (1.1 eq), CBr4(1.95 eq), II | dry DCM(10 mL), RT, 2 h.

[0703]

[0704] V - * 6. J H A B DIS7-2

[0705] To a stirred solution of Morpholine B (0.2 mL, 2.26 mmol, 1 eq) and Heptane-1-thiol A (300 mg, 2.26 mmol, 1 eq) were added in anhydrous DCM (10 mL), the reaction was cooled in an ice bath. Carbon disulfide (0.13 mL, 2.26 mmol, 1 eq) was added dropwise, followed by the slow addition of tri ethylamine (0.35 mL, 2.49 mmol, 1.1 eq) Stirred for 5 min. After solution of CBri (1.5 g, 4.4 mmol, 1.95 eq) in DCM (2mL) was added. The resulting reaction mixture was allowed to stir at room temperature for 2 h. The progress of reaction was monitored by TLC. After completion, the reaction mixture was diluted with H? O (4 mL) and extracted with DCM (2 x 10 mL). The organic layers were dried over sodium sulphate and concentrated to get crude. The crude was purified by 230-400 pre-packed Redisep column on combi flash chromatography, eluted at 30% of EtOAc in Heptane to afford heptylsulfanyl morpholine-4-carbodithioate, DIS7-2 (150 mg, 22.5 %) as pale-yellow liquid. Attorney Docket No. 10063-110WO1

[0706] TLC: 50 % EtOAc in Heptane (

[0707]

[0708] J?z=0.6). NMR (400 MHz, CHLOROFORM-d) 5 = 4.20 (br s, 4H), 3.81 - 3.76 (m, 4H), 2.88 - 2.84 (m, 2H), 1.66 (quin, J = 7.4 Hz, 2H), 1.45 - 1.34 (m, 2H), 1.33 - 1.23 (m, 6H), 0.91 - 0.85 (m, 3H) ppm. LCMS: (ES+) 96.80 %, 294 [M+HJ+[Method:- Column: CORTECS UPLC C18 (3*30)mm, 1.6umFlow rate: 0.85 mL / min. Mobile Phase A: 0.05 % TFA in Water, Mobile Phase B: 0.05 % TFA in Acetonitrile]; HPLC: 95 % purity [Method:- Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile(95:05), B - Acetonitrile], Synthesis of heptyl diisopropylcarbamo( dithioperoxo)thioate

[0709] B(1 eq), CS2(1 eq), Et3N g HS+T (1.1 eq), CBr4(1.95 eq), dry I H \. NH DCM (10 mL), RT, 2 h. N S' r — — ' A.

[0710]

[0711] A B DIS8-2 To a stirred solution of Diisopropyl amine B (0.32 mL, 2.26 mmol, 1 eq) and Heptane-1- thiol A (300 mg, 2.26 mmol, 1 eq) were added in anhydrous DCM (10 mL), the reaction was cooled in an ice bath. Carbon disulphide (0.13 ml.,, 2.26 mmol, 1 eq) was added dropwise, followed by the slow addition of triethylamine (0.35 mL, 2.49 mmol, 1.1 eq), Stirred for 5 min. After solution CBr4(1.5 g, 4.4 mmol, 1.95 eq) in DCM (2niL) was added. The resulting reaction mixture was allowed to stir at room temperature for 2 h. The progress of reaction was monitored by TLC. After completion, the reaction mixture was diluted with FLO (4 mL) and extracted into DCM (2 x 10 mL). The organic layers were dried over sodium sulphate and concentrated to get crude. The crude was purified by 230-400 pre-packed Redisep column on combi flash chromatography, eluted using 15 % of DCM in Heptane to afford heptyl diisopropylcarbamo(dithioperoxo)thioate, DIS8-2 (1 0 mg, 21.5 %) as pale-yellow liquid.

[0712] TLC: 50 % DCM in Heptane (R

[0713]

[0714] / = 0.6). NMR (400 MHz, CHLOROFORM-d) 5 = 5.24 - 4.65 (m, 1H), 2.85 (t, J = 7.3 Hz, 2H), 1.66 (td, J= 7.4, 14.9 Hz, 5H), 1.50 - 1.23 (m, 18H), 0.91 - 0.85 (m, 3H) ppm. LCMS: (ES+) 94.21 %■, m / z= 308 [M+H]+[Method:-Column: CORTECS UPLC C18 (3*30)mm, 1.6um Flow rate: 0.85 mL / min. Mobile Phase A: 0.05%TFA in Water, Mobile Phase B: 0,05%TFA in Acetonitrile]; HPLC: 96.0 % purity [Method:- Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile (95:05), B - Acetonitrile], Attorney Docket No. 10063-110WO1

[0715] Synthesis of propyl sulfanyl pyrrolidine- 1 -carbodithioate

[0716] B(1 eq), CS2(1 eq), Et3N

[0717] I — \ (1.09 eq), CBr4(1.95 eq), s + < ) dry PCM (10 mL), RT, 2 h. JsHS. N * Z" N' ~'S"x

[0718] 'HVJ

[0719]

[0720] A B DIS9-2 To a stirred solution of pyrrolidine B (0.32 mL, 3.93 mmol, 1 eq) and Propane-1-thiol A (300 mg, 3.93 mmol, 1 eq) were added in anhydrous DCM (10 ml,), the reaction was cooled in an ice bath. Carbon disulphide (0.23 ml., 3.93 mmol, 1 eq) was added drop wise, followed by the slow addition of triethylamine (0.60 mL, 4.29 mmol, 1.09 eq) Stirred for 5 min. After solution of CBr4(2.6 g, 7.68 mmol, 1.95 eq) in DCM (2mL) was added. The resulting reaction mixture was allowed to stir at room temperature for 2 h. Progress of the reaction was monitored by TLC. After completion, the reaction mixture was diluted with H2O (4 mL) and extracted with DCM (2. x 10 mL). The organic layers were dried over sodium sulphate and concentrated to get crude. The crude was purified by 230-400 pre¬ packed Redisep column on combi flash chromatography, eluted using 20-25 % of DCM in Heptane to afford propylsulfanyl pyrrolidine- 1 -carbodithioate, DIS9-2 (360 mg, 41.2 %) as orange solid.

[0721] TLC: 50 % DCM in Heptane (R

[0722]

[0723] f= 0.5). NMR (400 MHz, CHLOROFORM-d) 5 = 3.97 (t, 7 = 7.0 Hz, 2H), 3.75 (t, J = 6.9 Hz, 2H), 2.86 - 2.82 (m, 2H), 2.15 - 2.07 (m, 2H), 2.03 - 1.96 (m, 2H), 1.70 (sxt, J = 7.3 Hz, 2H), 1.01 (t, J = 7.4 Hz, 3H) ppm. LCMS: (ES+) 96.01 %, m / z= 222 [M+H]+[Method:- Column: CORTECS UPLC C18 (3*30)mm, 1.6um, Flow rate: 0.85 mL / min, Mobile Phase A: 0.05 % TEA in Water, Mobile Phase B: 0.05 % 1TA in Acetonitrile]: HPLC: 95.90 % purity [Method:- Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile (95:05), B - Acetonitrile].

[0724] Synthesis of propylsulfanyl piperidine-l-carbodithioate

[0725] B(1 eq), CS2(1 eq), Et3N

[0726] C (1.1 eq), CBr4(1.96 eq), HHS\ / \. + dry DCM (10 mL), RT, 2 h.

[0727] H " A

[0728]

[0729] B

[0730]

[0731] D1S10-2 To a stirred solution of piperidine B (0.65 mL, 6.56 mmol, 1 eq) and Propane-1- thiol A (500 mg, 6.56 mmol, 1 eq) were added in anhydrous DCM (10 mL), the reaction was cooled in an ice bath. Carbon disulphide (0.40 mL, 6.56 mmol, 1 eq) was added drop wise, followed by the slow addition of triethylamine (1.01 mL, 7.21 mmol, 1.1 eq) Stirred Attorney Docket No. 10063-110WO1

[0732] for 5 min. After solution of CBr4(4.35 g, 12.9 mmol, 1.96 eq) in DCM (2mL) was added. The resulting reaction mixture was allowed to stir at room temperature for 2 h. Progress of the reaction was monitored by TLC. After completion, the reaction mixture was diluted with H2O (4 mL) and extracted with DCM (2 x 10 mL). The organic layers were dried sodium sulphate and concentrated to get crude. The crude was purified by 230-400 pre-packed Redisep column on combi flash chromatography, eluted using 30 % of EtOAc in Heptane to afford propylsulfanyl piperidine- 1 -carbodithioate, DIS10-2 (402 mg, 26.01%) as yellow liquid.

[0733] TLC: 50 % EtOAc in Heptane (Rf: 0.6).!H NMR (400 MHz, CHLOROFORM-d) 5 = 4.30 (br s, 2H), 3.99 (br s, 2H), 2.86 - 2.81 (m, 2H), 1.78 - 1.64 (m, 8H), 1.01 (t,. J = 7.4 Hz, 3H) ppm. LCMS: (ES+) 98.15 %, m / z= 236 [M+H]+[Method:- Column: CORTECS IJPLC C18 (3*30)mm, 1,6um, Flow rate: 0.85 mL / min, Mobile Phase A: 0.05 % TEA in Water, Mobile Phase B: 0.05 % TFA in Acetonitrile]; HPLC: 99.06 % purity [Metliod:- Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile (95:05), B - Acetonitrile].

[0734] Synthesis of propylsulfanyl morpholine-4-carbodithioate

[0735] B(1 eq), CS2(1 eq), Et3N

[0736] z°> (1.09 eq), CBr4(1.95 eq), S HS..+I i dry DCM (10 mL), RT, 2 h.

[0737] " "'N " ( |b

[0738]

[0739] H

[0740] A B DIS11-2 To a stirred solution of Morpholine B (0.33 mL, 3.93 mmol, 1 eq) and Propane-1-thiol A (300 mg, 3.93 mmol, 1 eq) were added in anhydrous DCM (10 mL), the reaction was cooled in an ice bath. Carbon disulphide (0.23 L, 3.93 mmol, 1 eq) was added drop wise, followed by the slow addition of triethylamine (0.60 ml,, 4.29 mmol, 1.09 eq) Stirred for 5 min. After solution of CBr4(2.6 g, 7.68 mmol, 1.95 eq) in DCM (2mL) was added. The resulting reaction mixture was allowed to stir at room temperature for 2 h. Progress of the reaction was monitored by TLC. After completion, the reaction mixture was diluted with II2O (4 mL) and extracted with DCM (2 x 10 mL). The organic layers were dried sodium sulphate and concentrated to get crude. The crude was purified by 230-400 pre-packed Redisep column on combi flash chromatography, eluted using 30 % of EtOAc in Heptane to afford propylsulfanyl morpholine-4-carbodithioate, DIS11-2 (140 mg, 14.9%) as off white solid. Attorney Docket No. 10063-110WO1

[0741] TLC: 50 % EtOAc in Heptane (

[0742]

[0743] Rf: 0.6). NMR (400 MHz, CHLOROFORM-d) 8 = 4.21 (br s, 4H), 3.81 - 3.77 (m, 4H), 2.84 (t, J = 7.3 Hz, 2H), 1.70 (sxt, J = 7.3 Hz, 2H), 1.01 (t, J = 7.3 Hz, 3H) ppm. LCMS: (ES+) 99.69 %, m / z= 238 [M+H]+[Method:- Column: CORTECS UPLC C18 (3*30)mm, 1.6uni, Flow rate: 0.85 mL / min, Mobile Phase A: 0.05%1'FAin Water, Mobile Phase B: 0.05%TFAin Acetonitrile]; HPLC: 99.53 % purity [Method:- Column: X-Select CSH Cl 8 (4.6*1 0) mm 5u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile (95:05), B - Acetonitrile].

[0744] Synthesis of propyl diisopropylcarbamo( dilhioperoxo )thioate

[0745] . B(1 eq), CS2(1 eq), Et3N g ''f'' (1.09 eq), CBr4(1.95 eq), i JJ+\ _NH dry DCM (10 mL), RT, 2 h. 'ANAS''Sx / X

[0746]

[0747] A B DIS12-2 To a stirred solution of Diisopropyl amine B (0.56 mL, 3.93 mmol, 1 eq) and Propane-1- thiol A (300 mg, 3.93 mmol, 1 eq) were added in anhydrous DCM (10 ml.,), the reaction was cooled in an ice bath. Carbon disulfide (0.23 L, 3.93 mmol, 1 eq) was added drop wise, followed by the slow addition of triethylamine (0.60 mL, 4.29 mmol, 1.09 eq) stirred for 5 min. After solution of CBt4 (2.6 g, 7.68 mmol, 1.95 eq) in DCM (2mL) was added. The resulting reaction mixture was allowed to stir at room temperature for 2 h. Progress of the reaction was monitored by TEC. After completion, Reaction mixture was diluted with H2O (4 mL) and extracted with DCM (2 x 10 mL). The organic layers were dried sodium sulphate and concentrated to get crude. The crude was purified by 230-400 pre-packed Redisep column on combi flash chromatography, eluted at 15% of DCM in Heptane to afford propyl diisopropylcarbamo(dithioperoxo)thioate, DIS12-2 (40 mg, 4.04 %) as pale-yellow liquid.

[0748] TLC: 50 % DCM in Heptane (R 0.6). Hl NMR (400 MHz, CHLOROFORM-d) 3 = 2.83 (t, J = 7.3 Hz, 2H), 1.69 (sxt, J =. Hz, 2H), 1.53 - 1.41 (m, 14H), 1.01 (t, J = 7.4 Hz, 3H) ppm. LCMS: (ES+) 91.84 %, m / z= 252 [M+H]+[Method:- Column: CORTECS UPLC C18 (3*30)mm, 1.6um Flow rate: 0.85 mL / min, Mobile Phase A: 0.05 % TFA in Water, Mobile Phase B: 0.05 % TFA in Acetonitrile]; HPLC: 95.5 % purity [Method:-Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1%’ Formic acid in water: Acetonitrile (95:05), B - Acetonitrile], Attorney Docket No. 10063-110WO1

[0749] Synthesis of octylsulfanyl 7-oxa-l -aza spiro [ 4.4 ]nonane-l -carbodithioate

[0750] B(0.89 eq), CS2(1 eq), Et3N

[0751] (1.04 eq), CBr4(1.96 eq), S dry DCM (10 mL), RT, 2 h.

[0752]

[0753] DIS13-2 To a stirred solution of 7-oxa-l -azaspiro [4.4] nonane B (154 mg, 1.2.1 mmol, 0.89 eq) and octane-1- thiol A (200 mg, 1.36 mmol, 1 eq) in anhydrous DCM (10 mL), the

[0754] reaction was cooled in an ice bath. Carbon disulphide (0.08 mL, 1.36 mmol, 1 eq) was

[0755] added dropwise, followed by the slow' addition of triethylamine (0.2 mL, 1.42 mmol, 1.04 eq) Stirred for 5 min. After solution of CBf: (904 mg. 2.67 mg, 1.96 eq) in DCM (2mL) was added. The resulting reaction mixture was allowed to stir at room temperature for 2 h.

[0756] Progress of the reaction was monitored by TLC. After completion, the reaction mass was diluted with H? O (4 mL) and extracted with DCM (2 x 10 mL). The organic layers were

[0757] dried over sodium sulphate and concentrated to get crude. The crude was purified by 230- 400 pre-packed Redisep column on combi flash chromatography, eluted using 20-25 % of DCM in Heptane to afford octylsulfanyl 7-oxa-l-aza spiro[4.4]nonane-l-carbodithioate, DIS13-2 (100 mg, 21.04 %) as pale yellow liquid.

[0758] TLC: 50 % DCM in Heptane (Ry. 0.5). Hl NMR (400 MHz, CHLOROFORM-d) 5

[0759] = 4.73 (d,.7 = 6.2 Hz, 1H), 4.68 - 4.58 (m, 3H), 4.20 (s, III), 4.01 - 3.95 (m, 2H), 3.78 (t, J

[0760] = 7.0 Hz, 1H), 2.85 (dt, J = 1.7, 7.4 Hz, 2H), 2.42 (t, J = 7.0 Hz, 1H), 2.30 (t, J = 7.3 Hz,

[0761] 1H), 1.71 - 1.61 (m, 2H), 1.43 - 1.36 (m, 2H), 1.27 (br s, 8H), 0.88 (t,.1 = 6.7 Hz, 3H) ppm.

[0762] LCMS: (ES+) 98.52 % [Method:- Column: CORTECS UPLC C18 (3*30)mm, 1.6um Flow rate: 0.85 mL / min. Mobile Phase A: 0.05 % TEA in Water, Mobile Phase B: 0.05 % TFA in Acetonitrile]; HPLC: 99.01 % purity [Method:- Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile (95:05), B - Acetonitrile], Synthesis of octyl hexahydrocyclopenta[c]pyrrole-2(lH)-carbo(difhioperoxo)thioate / x B(1 eq), CS2(1 eq), Et3Ns( / (1.04 eq), CBr4(1.96 eq), H H -e X / ~,+ / — ( dry DCM (10 mL), RT, 2 h.fN I H A B DIS14-2

[0763]

[0764] To a stirred solution of l,2,3,3-[a],4,5,6,6-[a]-octaliydrocyclopenta[c]pyrrole B (151 mg, 1.36 mmol, 1 eq) and octane-1- thiol A (200 mg, 1.36 mmol, 1 eq) were added in anhydrous DCM (10 mL), the reaction was cooled in an ice bath. Carbon disulfide (0.08 Attorney Docket No. 10063-110WO1

[0765] mL, 1.36 mmol, 1 eq) was added drop wise, followed by the slow addition of triethylamine (0.2 mL, 1.42 mmol, 1.04 eq) stirred for 5 min. After solution of CBr4(904 mg. 2.67 mg, 1.96 eq) in DCM (2mL) was added. The resulting reaction mixture was allowed to stir at room temperature for 2 h. Progress of the reaction was monitored by TLC. After completion, the reaction mixture was diluted with H O (4 mL) and extracted with DCM (2 x 10 mL). The organic layer was dried over sodium sulphate and concentrated to get crude.

[0766] The crude was purified by 230-400 pre-packed Redisep column on combi flash chromatography, eluted using 20-25% of DCM in Heptane to afford octyl hexahydrocyclopenta[c]pyrrole-2(lH)-carbo(dithioperoxo)thioate, DIS14-2 (80 mg, 17.6 % ) as pale-yellow liquid.

[0767] TLC: 50 % DCM in Heptane

[0768]

[0769] (Ry. 0.5). NMR (400 MHz, CHLOROFORM-d) 5 = 4.19 (dd, J = 8.6, 1.9 Hz, 1H), 3.95 (dd, J = 8.4, 11.8 Hz, 1H), 3.78 (dd, J = 5.3, 13.9 Hz, III), 3.54 (dd,.1= 5.0, 11.9 Hz, HI), 2.92 - 2.82 (m, 3H), 2.80 - 2.71 (m, 1H), 1.95 - 1.76 (m, 3H), 1.73 - 1.62 (m, 3H), 1.60 - 1.56 (m, 1H), 1.53 - 1.47 (m, 1H), 1.44 - 1.35 (m, 2H), 1.33 - 1.21 (m, 8H), 0.90 - 0.85 (m, 3H) ppm. LCMS: (ES+) 97.8 %, m / z= 332 [M+H]+[Method:- Column: CORTECS UPLC C18 (3*30)mm, 1.6um Flow rate: 0.85 mL / min.

[0770] Mobile Phase A: 0.05% TFA in Water, Mobile Phase B: 0.05% TFA in Acetonitrile];

[0771] HPLC: 99.1% purity [Method:- Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile (95:05), B - Acetonitrile].

[0772] Synthesis of octylsulfanyl isoindoline-2-carbodithioate B(1.3 eq), CS2(1 eq), Et3N S (1.04 eq), CBr4(1.96 eq), J] g dry DCM (10 mL), RT, 2 h. zz^~N S"

[0773]

[0774] A B DIS15-2

[0775] To a stirred solution of isoindoline B (212 mg, 1.77 mmol, 1.3 eq) and octane-1-thioi A (200 mg, 1.36 mmol, 1 eq) were added in anhydrous DCM (10 niL), the reaction was cooled in an ice bath. Carbon disulfide (0.08 mL, 1.36 mmol, 1 eq) was added drop wise, followed by the slow addition of triethylamine (0.2 mL, 1.42 mmol, 1.04 eq) stirred for 5 min. After solution of CBr4 (904 mg. 2.67 nig, 1.96 eq) in DCM (2mL) was added.

[0776] 'The resulting reaction mixture was allowed to stir at room temperature for 2 h. Progress of the reaction was monitored by TEC. After completion, the reaction mixture was diluted with H2O (4 L) and extracted with DCM (2 x 10 mL). The organic layer was dried over sodium sulphate and concentrated to get crude. The crude was purified by 230-400 pre-packed Attorney Docket No. 10063-110WO1

[0777] Redisep column on combi flash chromatography, eluted using 20-25% of DCM in Heptane to afford octylsulfanyl isoindoline-2 -carbodithioate, DIS15-2 (20 mg, 4.3 %) as brown solid.

[0778] TLC: 50 % DCM in Heptane

[0779]

[0780] (Ry. 0.5). NMR (400 MHz, CHLOROFORM-d) 5 = 7.37 - 7.27 (m, 4H), 5.25 (s, 2H), 5.09 (s, 2H), 2.91 - 2.87 (m, 2H), 1.69 (quin, J = 7.4 Hz, 2H), 1.45 - 1.37 (m, 2H), 1.28 (dd,.7 = 1.5, 4.3 Hz, 8H), 0.90 - 0.85 (m, 3H) ppm. LCMS:

[0781] (ES+) 99.14%, m / z= 340 [M+H]+[Method:- Column: CORTECS UPLC C18 (3*30)mm, 1.6um Flow rate: 0.85 mL / min. Mobile Phase A: 0.05 % TEA in Water, Mobile Phase B:

[0782] 0,05 % TFA in Acetonitrile]; HPLC: 98.3% purity [Method:- Column: X-Select CSH C18 (4.6*150) nun 5u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile (95:05), B - Acetonitrile],

[0783] Synthesis of octyl benzyll methyl )carbamo( dithioperoxo jthioate

[0784] 2 (1 eq), CS2 (1eq), CBr4 S \ (2eq), TEA (1.1 eq) DCM %s~ zr-Ty NH (10V), RT-2h _ N' 'S' # ) — i H S. J

[0785] O'

[0786]

[0787] A 8 DIS17-2 To a stirred solution of N-methyl benzylamine(A) (1 g, 8.3 mmol, 1.01 eq) and octane-1- thiol (B)(1.2 g, 8.24 mmol, 1 eq) in anhydrous DCM (10 mL), the reaction was cooled in an ice bath. Carbon disulfide (628 mg, 8.24 mmol, 1 eq) was added drop wise, followed by the slow addition of triethylamine (918 nig, 9.03 mmol, 1.1 eq) stirred for 5 min, after solution of CBr4 (5.4 g, 16 mmol, 2 eq) in DCM (2mL) was added. The reaction mixture was allowed to stir at room temperature for 2h. Progress of reaction was monitored by TLC. After completion, reaction mixture was diluted with H2O (4 mL) and extracted into DCM (2 x 10 mL). The organic layers were dried over sodium sulphate and concentrated in vacuo to obtain crude. The crude was purified by 230-400 pre-packed Redisep column on combi flash chromatography, eluted using 20-25% of DCM in Heptane to afford octyl benzyl(methyl)carbamo (dithioperoxo)thioate, DIS17-2 (800 mg, 29 % yield) as yellow liquid.

[0788] TLC: 50 % DCM: Heptane (Rf: 0.5).

[0789]

[0790] NMR (CHLOROFORM-d) 8: 7.15-7.32 (m, 5H), 5.40 (br s, 1H), 5.07 (br s, 1H), 3.52 (br s, 1H), 3.33 (br s, 2H), 2.89 (br s, 2H), 1.41 (br s, 3H), 1.28 (br s, 9H), 0.85-0.90 (m, 3H) ppm. LCMS: (ES+) 96.47 %, m / z =341.92 [Method: - Column: CORTECS UPLC C18 (3*30)mm, 1.6um, Mobile Phase A:

[0791] 0.05%FAin Water, Mobile Phase B: 0.05%FA in Acetonitrile, HPLC: 96.69 [Method:- Attorney Docket No. 10063-110WO1

[0792] Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% TFAin water, B - Acetonitrile].

[0793] DIS series 3 compounds.

[0794] Synthesis of nonyl diethylcarbamodithioate

[0795] B (0.97 eq), PPh3 (0.95 eq),

[0796] CBM (05 eq), DCM(10mL)

[0797] Acetone (6mL), RT-2h. s 40°C-4h. JL

[0798]

[0799] DIS1-3 To a stirred solution of sodium diethyl carbamodi thioate A (530 mg, 3.11 mmol, leq), nonane- 1 -thiol B (500 mg, 3.11 mmol, 0.97 eq) in DCM (10 mL) were added Triphenylphosphine (817 mg, 3.05 mmol, 0.95 eq) and CBr4 (518 mg, 1.53 mmol, 0.5 eq) at 0°C. The reaction mixture was allowed to stir at RT for 2 h. / After completion, the reaction mixture was concentrated at 5-7 °C to obtain crude. That crude dissolved with acetone (6 mL) and diethylcarbamothioylsulfanylsodium (550 mg, 3.21 mmol, 1 eq) was added. The reaction mixture was stirred at 40 °C for 4 h. Progress of the reaction was monitored by TLC & LCMS. After completion, the reaction mixture was filtered and concentrated in vacuo to obtain crude. The crude compound was purified by combi flash column chromatography using YMC-12 g cartridge eluted using 7 % EtOAc in heptane. The eluted fractions were concentrated in vacuo to afford nonyl diethylcarbamodithioate (D1S1-3) (80 mg, 9.04 % Yield) as a colorless liquid.

[0800] TLC: 20% EtOAc in Heptane (Rf: 0.4). *H NMR (CHLOROFORM-d) 5: 4.04 (q, J=6.6 Hz, 2H), 3.75 (q, J=6.5 Hz, 2H), 3.25-3.29 (m, 2H), 1.70 (quin, J=7.4 Hz, 2H), 1.36- 1.47 (m, 2H), 1.21-1.35 (m, 16H), 0.83-0.91 (m, 3H) ppm. LC-MS: (ES+) m / z = 275.5 [M+H]+, 99.80%, RT = 2.06 min (Method:- Column Name: CORTECS UPLC C18 (3X30mm, 1.6pm), Mobile Phase A: 0.05 % TEA in H ■■( ). Mobile Phase B: 0.05 % TEA in ACN). HPLC: 99.34 % purity [Method: Column: X-Select CSH C18, (4.6*150) mm 5u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile (95:05), B - Acetonitrile.

[0801] Synthesis of S-nonyl diethyicarbamothioate

[0802] O B (0.71 eq), KOH(0.29 eq), o JJ ~ Acetone(5 mL). RT,12h H N' ''ci + HS - *-

[0803]

[0804] A B DIS2-3 Attorney Docket No. 10063-110WO1

[0805] To a stirred solution of diethylcarbamic chloride A (500 nig, 3.68 mmol, 1 eq) in Acetone (5 ml.,) were added KOH (174 mg, 1.08 mmol, 0.29 eq) and nonane- 1 -thiol B (422 mg, 2.632 mmol, 0.71 eq) at 0 °C. The reaction mixture was allowed to stir at room temperature for 12 h. Progress of the reaction was monitored by TLC & LCMS. After completion, the reaction mixture was filtered and concentrated in vacuo to obtain crude. The crude compound was purified by combi flash column chromatography using YMC-12 g cartridge, and the product was eluted using 30 % EtOAc in heptane. The eluted fractions were concentrated in vacuo to afford S-nonyl diethylcarbamothioate, DIS2-3 (350 mg, 99.95 % Yield) as colorless liquid.

[0806] TLC: 50% EtOAc in Heptane (Rf: 0.6).

[0807]

[0808] NMR (CHLOROFORM-d) 5: 3.38 (br s, 4H), 2.85-2.92 (m, 2H), 1.56-1.65 (m, 2H), 1.34-1.42 (m, 2H), 1.23-1.34 (m, 10H), 1.10-1.22 (m, 6H), 0.85-0.90 (m, 3H) ppm. LC-MS: (ES+) m / z = 259.4 [M+HJ+, 99.80 %, RT = 2.06 min (Method:- Column Name: CORTECS UPLC C18 (3X30mm, 1.6um): Flow Rate: 0.85 mL / min Mobile Phase A: 0.05 % TFA in II2O, Mobile Phase B: 0.05 % TEA in ACN). HPLC: 99.80% purity [Method:- Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile(95:05), B - Acetonitrile].

[0809] Synthesis of benzyl diethylcarbamodithioate

[0810] S Br B (1 eq), Acetone S

[0811] ■' N'- SNa (3nlL)'4°°c’5 h-. ^NAS-'' y

[0812] A+U j J]

[0813] A B DIS3-3

[0814] To a stirred solution of sodium diethylcarbamodithioate A (200 nig, 1.16 mmol, leq) in acetone (3 ml,) was added (bromomethyl )benzene B (200 mg, 1.16 mmol, leq) at RT. The resulting reaction mixture was allowed to stir at 40 °C for 5 h. Reaction was monitored by TLC. After completion, the reaction mixture was concentrated in vacuo to get crude. The crude was directly purified by 100-200 silica gel column chromatography, eluted using 30% EtOAc in Heptane to afford benzyl diethylcarbamodithioate, DIS3-3 (30 mg, 10.7 %) as colorless liquid.

[0815] TLC: 50% EtOAc in Heptane (Rf. 0.6). ‘H NMR (400 MHz, DMSO-ck) 5 = 7.41 - 7.37 (m, 2H), 7.35 - 7.24 (m, 3H), 4.50 (s, 2H), 3.98 (d, J = 6.8 Hz, 2H), 3.73 (d, J = 6.8 Hz, 2H), 1.19 (d,.7= 5.9 Hz, 6H) ppm. LCMS: (ES+) 97.98 %, m / z= 239.74 [M+H]+[Method:- Column: CORTECS UPLC C18 (3*30)mm, 1.6um, Flow rate: 0.85 mL / min. Mobile Phase A: 0.05 % TFA in Water, Mobile Phase B: 0.05 % TFA in Acetonitrile]; Attorney Docket No. 10063-110WO1

[0816] HPLC: 97.74 % purity [Method:- Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile (95:05), B - Acetonitrile], Synthesis of S-benzyl diethylcarbamothioate

[0817] ? SH B (1 eq), KOH (0.35), 9

[0818] +' Acetone(5mL), RT,12 h.

[0819]

[0820] J U ' > V1

[0821] A B DIS4-3

[0822] To a stirred solution of diethylcarbamic chloride A (546 mg, 4.02 mmol, 1 eq) in Acetone (5 niL) were added KOH (226 mg, 1.40 mmol, 0.35 eq) and phenylmethanethiol B (500 mg, 4.02 mmol, 1 eq) at 0 °C. The resulting reaction mixture was allowed to stir at RT for 12 h. Reaction was monitored by TLC. After completion, the reaction mixture was concentrated in vacuo, diluted with II2O (10 mL), and extracted with EtOAc (2 x 20 mL). The organic layers were dried over sodium sulphate and concentrated in vacuo to obtain crude. The crude was purified by 100-200 silica gel column chromatography, eluted using 30% EtOAc in Heptane to afford S-benzyl diethylcarbamothioate, DIS4-3 (350 mg, 38.9 %) as colorless liquid.

[0823] TLC: 50% EtOAc in Heptane (Rf 0.6).2H NMR (400 MHz, CHLOROFORM-d) 5 = 7.37 - 7.33 (m, 2H), 7.32 - 7.20 (m, 3H), 4.16 (s, 2H), 3.49 - 3.27 (m, 4H), 1.17 (t, J= 7.1 Hz, 6H) ppm. LCMS: (ES+) 99.94 %, 223.78 [M+H]+[Method:- Column: CORTECS UPLC C18 (3*30)mm, 1.6um, Flow rate: 0.85 mL / min. Mobile Phase A: 0.05 % TFAin Water, Mobile Phase B: 0.05 % TFA in Acetonitrile]; HPLC: 99.34% purity [Method:-Column: X-Select CSH C18 (4.6*150) mm 5u Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile(95:05) B - Acetonitrile].

[0824] Synthesis of phenethyl diethylcarbamodithioate

[0825]

[0826] To a stirred solution of sodium diethylcarbamodithioate A (200 mg, 1.16 mmol, 1 eq) in acetone (3 mL) was added (2-bromoethyl)benzene B (216 mg, 1.16 mmol, 1 eq) at RT. The resulting reaction mixture was allowed to stir at 40 °C for 5 h. Reaction was monitored by TLC. After completion, the reaction mixture was concentrated in vacuo to get crude. The crude was directly purified by 100-200 silica gel column chromatography, eluted Attorney Docket No. 10063-110WO1

[0827] using 30% EtOAc in Heptane to afford phenethyl diethylcarbamodithioate, DIS5-3 (90 mg, 30.4 %) as colorless liquid.

[0828] TLC: 50% EtOAc in Heptane (Rf: 0.6). 'H NMR (400 MHz, DMSO-d6) 5 = 7.33 -7.25 (m, 4H), 7.25 - 7.19 (m, 1H), 3.97 (q, J= 6.8 Hz, 2H), 3.72 (q, J= 6.9 Hz, 2H), 3.48 - 3.43 (m, 2H), 2.95 - 2.89 (m, 2H), 1.19 (q, J = 7.2 Hz, 6H) ppm. LCMS: (ES+) 99.44 % [Method:- Column: X Select CSH-C18(3.0X50mm, 2.5pm) Mobile Phase: A: 0.05% TFA in water, Mobile Phase B: 0.05 % TFAinACN]; HPLC: 98.69 % purity [Method:- Column: X-Select CSH C18 (4.6*150) mm 5u Mobile Phase: A - 0.1% Formic acid in water:

[0829] Acetonitrile(95:05) B - Acetonitrile],

[0830] Synthesis of S-phenelhyl diethylcarbamothioate

[0831]

[0832] To a stirred solution of diethylcarbamic chloride A (490 mg, 3.61 mmol, 1 eq) and 2 -phenylethane- 1 -thiol B (500 mg, 3.61 mmol, 1 eq) in Acetone (5 mL) was added KOH (203 mg, 1.26 mmol, 0.35 eq) at RT. The reaction mixture was allowed to stir at RT for 12 h. Reach on was monitored by TLC. After completion, KOH was filtered and concentrated in vacuo to obtain crude. The crude was purified by 100-200 silica gel column chromatography eluted using 15 % EtOAc in Heptane to afford S-phenethyl diethylcarbamothioate, DIS6-3 (400 mg, 46.6 %) as colorless liquid.

[0833] TLC: 50% EtOAc in Heptane (Rf 0.6). 'H NMR (400 MHz, CHLOROFORM-d) 5 = 7.32 - 7.19 (m, 5H), 3.39 (d,.7= 6.6 Hz, 4H), 3.17 - 3.10 (m, 2H), 2.95 - 2.89 (m, 2H), 1.17 (t,.7= 6.9 Hz, 6H) ppm. LCMS: (ES+) 95.17 %, m / z= 237.85 [M+H]* [Method:-Column: CORTECS UPLC C18 (3*30)mm, 1.6um Flow rate: 0.85 mL / niin, Mobile Phase A: 0.05 % TFA in Water, Mobile Phase B: 0.05 % TFA in Acetonitrile], HPLC: 97.80 % purity [Method: Column: X-Select CSH C18 (4.6*150) mm 5u Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile (95:05), B - Acetonitrile]. Attorney Docket No. 10063-110WO1

[0834] Example 8: AZ analog synthesis

[0835] Synthesis of Common Intermediate-2, ethyl (Z)-2-(hydroxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetate

[0836] PhvPh

[0837] Ci'' Ph

[0838] 2 Ph ph

[0839] 2 i'1 eq), TEA(3 eq)

[0840] HO'NDMF(10Vol), RT-16hNX Y~-NH

[0841] HO' N

[0842] 'o Step-1

[0843] O O

[0844]

[0845] 1 Comrnon-lntermidiate-2 To a stirred solution of ethyl (2-Z)-2-(2-aminotliiazol-4-yl)-2-hydroxyimino-acetate (1) (5 g, 23.23 mmol, leq) and chloromethanetriyl)tribenzene (2) (6.47 g, 23.23 mmol, leq) in DMF (25 ml.) was added Triethylamine (9.47 ml, 69.69 mmol, 3eq) dropwise in the reaction mixture at room temperature. Further, the reaction mixture was allowed to stir at room temperature for 16 h. Progress of reaction was monitored by TLC and LCMS. After completion, the reaction mixture was diluted with ice cold water (50 mL) and extracted with ethyl acetate (3 x 40 mL). The combined organic layers were dried over anhydrous sodium sulphate and concentrated in vacuum to obtain crude. The crude was washed with DCM ( 3 x 30 mL) and collected organic layers were concentrated under vacuum to afford ethyl (Z)-2-(hydroxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetate common intermediate-2 (8 g, 75.27% Yield) as a pale yellow solid.

[0846] TLC: 70% ethyl acetate in heptane (Ry-0.3).1H NMR (DMSO-d6) 5: 7.28-7.33 (m, 6H), 7.25 (t, J= 7.6 Hz, 6H), 7.15-7.21 (m, 3H), 6.82 (s, 1H), 3.93 (q, J = 6.8 Hz, 2H), 2.88 (s, III), 2.72 (s, ITT), 1.09 (t,, Z = 7.2 Hz, 3H) ppm. LCMS: (ES+) 96.32%, 458.3 [M+H] [Method: Column: CORTECS UPLC C18 (3X30mm Instrument ID: - AMC-LCMS-15 Mobile Phase A: 0.05 % FA in Water B: 0.05% FA in ACN],

[0847] AZ series 1 compounds

[0848] In AZ, the left-hand side of the molecule at the dimethyl acetic acid chain was replaced with various hydrophobic groups. The designs are aliphatic chain length (compound AZ1, AZ2, AZ4, and AZ7), ether chain length (compound AZ8), pyrane ring (compound AZ 9), benzyl (compound AZ5 and AZ12), and cyclohexyl (compound AZ. i h. Attorney Docket No. 10063-110WO1

[0849] Synthesis of ( 2S, 3S)-3-((Zf2-( 2"€iminothiazol-4~yl)~2f (hexyloxy)imino) acetamido)-2-methyl-4-oxoazetidine--l--sulfomc acid

[0850] ncNN -o-N- N N.„ N Ox*0 Stepl O' *0

[0851] Step-2 Common-lnt-2 4 5 NH2N

[0852] 6 — NHTrit 5 oNN'

[0853] Step-3 0

[0854] N Step-4 O-S-OH

[0855]

[0856] 0

[0857] 7 AZ1

[0858] Step 1: Synthesis of ethyl (Z)-2-((hexyloxy)imino)-2-(2-(trityl amino) thiazol-4-yl) acetate

[0859] Ph Ph

[0860] X-Ph

[0861]

[0862] O' O

[0863] 4

[0864] To a stirred solution of ethyl (Z)-2-(hydroxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetate common Int-2 (1.5 g, 3.3 mmol, leq) and 1-bromohaxane (3) (1 g, 6.6 mmol, 2eq) in DMF (10 mL) was added Potassium carbonate (1.13 g, 8.2 mmol, 2.5eq) in the reaction mixture at room temperature. The reaction mixture was allowed to stir at 50°C for 16 h.

[0865] Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was diluted with ice cold water (25 mL) and extracted with ethyl acetate (3 x 50 L). The combined organic layers were dried over anhydrous sodium sulphate and concentrated in vacuo to obtain crude (1.1g). The crude was purified by silica gel flash chromatography eluted using 30% ethyl acetate in heptane to afford ethyl (2Z)-2-(cyclohexoxyimino)-2-[2-(tritylamino) thiazol-4-yl] acetate (4) (800 mg, 45% Yield) as a pale-yellow liquid. TLC: 70% ethyl acetate in heptane (Rf. 0.5) ppm. LCMS: (ES+) 76.65%, m / z = 542.26 [M+H],

[0866] Step 2: Synthesis of (Z)-2-((hexyloxy)imino)-2-(2-(tritylamino) thiazol-4-yl) acetic acid Attorney Docket No. 10063-110WO1

[0867] Ph Ph

[0868] fr\ X-Ph

[0869] / N

[0870]

[0871] HO O

[0872] 5

[0873] To a stirred solution of ethyl (Z)-2-(cyclohexoxyimino)-2-[2-(tritylamino) thiazol-4- yl] acetate (4) (750 mg, 1.4 mmol, leq) in THE: MeOH: H2O (1:1,1, 6 mL) was added LiOH (167 mg, 7 mmol, 5eq) at room temperature. The reaction mixture was allowed to stir at 45°C for 16 h. Progress of the reaction was monitored by TLC and LCMS. Reaction mixture was concentrated under vacuum and diluted with water (lOmL) and acidified with IM Aq Citric acid solution (5 mL) was obtained Precipitation which was filtered and dried under vacuum to afford the desire (Z)-2-((hexyloxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetic acid (5) (550 mg) as an off white solid. TLC: 50% methanol in DCM (Rf. 0.2). LCMS: (ES+) 53.36%, m / z = 514.15 [M+H], Step 3: Synthesis of (2S, 3S)-3-((Z)-2-((hexyloxy)imino)-2-(2-(tritylamino) thiazol-4-yl) acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid r's, N Ji NHTrit 'NHN ^O

[0874] N

[0875] O-S~OH

[0876]

[0877] 6

[0878] 7

[0879] To a stirred solution of (Z)-2-((hexyloxy)imino)-2-(2-(tritylamino)thiazol-4-yl)acetic acid Int-5 (500 mg, 0.97 mmol, 1 eq) and (2S,3S}-3-amino-2-methyl-4-oxo-azetidine-l- sulfonic acid (6) (174 mg, 0.97 mmol, 1 eq) in DME (2 mL) was added PyAOP (1.2 g, 2.2 mmol, 2.25eq) and reaction mixture at room temperature stirred for 5 min. After, DIPEA was added (2 mL). The reaction was allowed to stir at room temperature 16 h. The reaction was monitored by TLC and LCMS. After completion, the reaction mixture was diluted with water (20 mL) and extracted with ethyl acetate (3 X 10 mL). The combined organic layer was washed with brine (30 mL) dried over anhydrous Na2SO4 concentrated under vacuum to afford the crude. The crude was further purified by flash column chromatography to afford the desired (2S,3S)-3-((Z)-2-((hexvloxy)imino)-2-(2-(tritvlamino) thiazol-4-yl) Attorney Docket No. 10063-110WO1

[0880] acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid (Int-7) (200 mg, 34 % yield). LCMS:

[0881] (ES+) 50%, m / z = 676.22 [M+H].

[0882] Step 4: Synthesis of (2S S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-((hexyloxy )imino) acetamido)-2-methyl-4-oxoazetidine-l-sulfonic acid

[0883] Q N HN' ND

[0884] N

[0885] O=S ii ~OH

[0886]

[0887] o

[0888] AZ1

[0889] To a stirred solution of (2S,3S)-3-((Z)-2-((hexyloxy)imino)-2-(2-(tritylamino) thiazol-4-yl) acetamido)-2-methyl-4-oxoazetidine-l-sulfonic acid (500 mg, 0.73 mmol, leq) in Formic acid (5 mL) at room temperature. The reaction mixture was allowed to stir at 4 °C for 4 h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated under reduced pressure to obtain crude (450 mg). The crude was purified by Prep-HPLC to afford (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-((hexyloxy)imino) acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid (Target-1) (65.6 mg, 20.5%’ Yield) as an off white solid.

[0890] TLC: 15% methanol in DCM (Ry - 0.2).2H NMR (DMSO-&) 5: 0.85 - 0.89 (m, 3H), 1.28 (s, 6H), 1.36 - 1.44 (m, 4H), 1.56 - 1.64 (m, 2H), 3.65 - 3.69 (m, 1H), 4.06 (t, J = 6.50 Hz, 2H), 4.43 (dd, J= 8.00, 2.63 Hz, 1H), 6.76 (s, III), 7.50 - 8.10 (m, 1H) ppm. LCMS: (ES+) 99.67%, m / z = 434.00 [M+H]. HPLC: 99.58 % Purity [Method: Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% TFA in water, B - Acetonitrile], Attorney Docket No. 10063-110WO1

[0891] Synthesis of ( 2S, 3S)-3~((Z)-2f2-aminothiazol--4-yl)--2-( sec-butoxyimino ) acetamid )- 2-methyl-4-oxoazetidine-l -sulfonic acid *-Br3 3(2 eq), K2COs (25 eq), LiOH (25 eq), THF: MeOH: DMF. SCT'C.ie h H2O (1:1,1), RT45°C 16h JL / X-NHTrit O N

[0892] Stepl Step-2

[0893] HO

[0894] Common-lnt-2

[0895]

[0896] 7 AZ2

[0897] Step 1: Synthesis of ethyl (Z)-2-(sec-butoxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetate

[0898] Ph Ph

[0899] I X-Ph

[0900] 1,. N. A 4>-NH

[0901] o N

[0902] 0^0

[0903]

[0904] 4

[0905] To a stirred solution of ethyl (Z)-2-(hydroxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetate common Int-2 (1 g, 2.18 mmol, leq) and 2-bromobutane (3) (599 mg, 4.37 mmol, 2eq) in DMF (5 mL) was added Potassium carbonate (756 mg, 5.46 mmol, 2.5eq) at room temperature. The resulted reaction mixture was allowed to stir at 50°C for 16 h. The progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was diluted with ice cold water (20 ml. ) and extracted with ethyl acetate (3 x 50mL). The combined organic layers were dried over anhydrous sodium sulphate and concentrated under vacuum to obtain crude (1.5g). The crude was purified by flash column chromatography to afford ethyl (Z)-2.-(sec-butoxyimino)-2-(2-(tritylaniino) thiazol-4-yl) acetate (4) (800 mg, 71.26% Yield) as a pale yellow solid. TLC: 70% Ethyl acetate in heptane. (Ry: 0.5). LCMS: (ES+) 80%, m / z = 14.4 [M+H],

[0906] Step 2: Synthesis of (Z)-2-(sec-butoxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetic acid Attorney Docket No. 10063-110WO1

[0907]

[0908] To a stirred solution of ethyl (Z)-2-(sec-butoxyimino)-2-(2-(tritylamino) thiazole¬ yl) acetate (4) (800 mg, 1.55 mmol, leq) in THF: MeOH: H₂O (1:1:1, 6 ml.) was added LiOH (95 mg, 3.89 mmol, 2.5eq) in Water (2 mL) was added drop wise in the reaction mixture at room temperature and stirred at 45 °C for 16 h. The reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated under vacuum and diluted with water (10 mL) and acidified(pH~3.0) with IM aq Citric acid solution and was obtained precipitate was filtered, washed with di-ethyl ether (10 mL) and dried under vacuum to afford (Z.)-2-(sec-butoxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetic acid (5) (500 mg, 66.10% Yield) as a white solid. TLC: 15% Methanol in DCM. (Ry- 0.2). LCMS: (ES+) 58 %, m / z = 486.13 [M+H],

[0909] Step 3: Synthesis of (2S,3S)-3-((Z)-2-(sec-butoxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetamido)-2-methyl-4-oxoazetidine-l-sulfonic acid.

[0910] HN 'O

[0911]

[0912] To a stirred solution of (Z)-2-(sec-butoxyimino)-2-(2-(tritylamino)thiazol-4-yl)acetic acid (5) (300 mg, 0.61 mmol, leq) in DMF (3 mL) were added (2S,3S)-3-amino-2-methyl-4-oxoazetidine-l -sulfonic acid (6) (111.3 mg, 0.61 mmol, leq,) and N, N- Di isopropylethyl amine (403 mg, 3.08 mmol, 5eq) at room temperature stirred for 15 min followed by added PYAOP (402.6 mg, 0.74 mmol, 1,2eq) the resulted reaction mixture was allowed to stir at room temperature for 16 h. The Progress of reaction was monitored by TLC and LCMS. After completion, the reaction mixture was quenched with ice cold water and extracted with ethyl acetate (3 x 25 mL). The organic layers were dried over sodium Attorney Docket No. 10063-110WO1

[0913] sulphate and concentrated under reduced pressure to afford (2S,3S)-3-((Z)-2-(sec- butoxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetamido)-2-methyl-4-oxoazetidine- 1 -sulfonic acid (7) (450 mg) as a pale-yellow liquid. TLC: 15% methanol in DCM (Rf:0.3).

[0914] LCMS: (ES+) 31%, m / z = 648.16 [M+HJ.

[0915] Step 4: Synthesis of (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-(sec-butoxyimino) acetamido)-2-methyl-4-oxoazetidine-l-sulfonic acid

[0916] N

[0917] O-S-O

[0918]

[0919] OH

[0920] AZ2

[0921] To a stirred solution of (2S,3S)-3-((Z)-2-(sec-butoxyimino)-2-(2-(tritylamino) thiazoi-4-yl) acetamido)-2-methyl-4-oxoazetidine-l-sulfonic acid (450 mg, 0.69 mmol, leq) in Formic Acid (5 mL) at room temperature. The reaction mixture was allowed to stir at 4 °C for 4 h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated under reduced pressure to get crude (500 mg). The crude was purified by Prep-HPLC to afford (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-(sec- but oxyimino) acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid (AZ2) (8 mg, 3 % Yield) as an off white solid.

[0922] TLC: 15% methanol in DCM (Ry - 0.2).1H NMR (DMSO-cfc) 5: ppm 0.87 (td, J - 7.41, 1.81 Hz, 3H), 1.18 (dd,.7= 6.25, 1.75 Hz, 4H), 1.39 (d, J= 6.13 Hz, 3H), 1.47 - 1.53 (m, III), 1.56 - 1.64 (m, HI), 2.31 - 2.34 (m, HI), 4.05 - 4.16 (m, HI), 4.42 - 4.50 (m, HI), 6.71 (d, J = 2.00 Hz, 1H), 7.19 - 7.58 (m, 2H), 9.09 - 9.31 (m, 1H) ppm. LCMS: (ES+) 96.46 %, m / z - 406.2 [M+H]. HPLC: 91.21 % Purity [Method: Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A- 0.1% TEA in water, B - Acetonitrile]. Attorney Docket No. 10063-110WO1

[0923] Synthesis of (2S,3S)-3-((Z)-2-(2-aminothiawl-4-yl)-2-(fheptan-4-yloxy) imino) ace, tamido)-2-methyl-4-oxoazetidine-l -sulfonic acid

[0924] y3L N J / ^-NHTrit HTrit o- LiOH(2.5eq), THF: MeOH:NH2O: (1:1:1), RT-65°C 16h HO Common lnt-2 4 Step-2

[0925] O=S-OH 6 (1eq) DIC(1.2 eq) HOBt(1,2eq), TEA(3 eq), HCOOH, 50°C, 4h DMF(5V) RT,14h Step-3 * Step-4

[0926]

[0927] Step 1: Synthesis of ethyl (Z)-2-((pentan-3-yloxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetate

[0928] k

[0929] Z. 1 N A / / --NHTrit

[0930] ' V N

[0931] O'^O

[0932]

[0933] 4

[0934] To a stirred solution of ethyl (Z)-2-(hydroxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetate common Int-2 (1 g, 2.328 mmol, leq) and 3-bromopentane (3) (5 ml) in DMF (5 mL) was added Potassium carbonate (2.50 g, 5.82 mmol, 2.5eq) at room temperature. The reaction mixture was allowed to stir at 50°C for 16 h. The progress of reaction was monitored by TLC (70% of EtOAc in heptane) and LCMS. After completion, the reaction mixture was diluted with ice cold water (20 mL) and extracted with ethyl acetate (3 x 50 mL). The combined organic layers were dried over sodium sulphate and concentrated into vacuum to obtained crude (1. 1 g). The crude was purified by combi flash column chromatography, eluted using 30% ethyl acetate in heptane to afford ethyl (Z)-2-((heptan-4-yloxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetate (4) (1.2 g, 98.0%’ Yield) as a pale¬ yellow liquid. TLC: 70% ethyl acetate in heptane (R / : 0.5). Attorney Docket No. 10063-110WO1

[0935] Step 2: Synthesis of (Z)-2-((pentan-3-yloxy) imino)-2-(2-(tntylamino) thiazol-4-vl) acetic acid

[0936] k TS

[0937] X N XL X-NHTrit

[0938]

[0939] 0X

[0940] HO ON

[0941] lnt-5

[0942] To a stirred solution of ethyl (Z)-2-((pentan-3-yloxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetate (4) (1.2 g, 1.51 mmol, leq) in TIIF: MeOH: H2O (1:1:1, 6 mL) was added LiOH (452.41 nig, 3.77 mmol, 2.5eq) at room temperature. The reaction mixture was allowed to stir at 45 °C for 16 h. Progress of reaction was monitored by TLC and LCMS. After completion, the Reaction mixture was concentrated and diluted with water (10 mL) and acidified with 1 (N) Citric acid solution (5 mL) up to pH 3. Precipitate was obtained. The precipitate was filtered and dried under vacuum to afford (Z)-2-((heptan-4-yloxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetic acid (5) (500 mg, 56% pure by LCMS) as an off white solid. TLC: 50% Methanol in DCM. (Ry: 0.2). LCMS: (ES+) 56%, m / z = 528 [M+HJ.

[0943] Step 3: Synthesis of (2S, 3S)-2-methyl-4-oxo-3-((Z)-2-((pentan-3-yloxy) imino)-2-(2-(trityIamino) thiazoI-4-vl) acetamido) azetidine-l-sulfonic acid

[0944] k.,3.

[0945] A X X—NHTrit

[0946] O=S-OH

[0947]

[0948] 6

[0949] 7

[0950] To a stirred solution of (Z)-2-((heptan-4-yloxy)imino)-2-(2-(tritylamino)thiazol-4-yl)acetic acid(5) (500 mg, 0.72 mmol, leq) in DMF (5 mL) were added (2S,3S)-3-amino-2-rnethyl-4-oxoazetidine-l -sulfonic acid (130.6 mg, 0.72 mmol, leq), N, N- Di isopropylethyl amine (478 mg, 3.62 mmol, 5eq) at room temperature stirred for 15 min. Then PYAOP (472 mg, 10.86 mmol, 1.2eq) was added. The reaction mixture was allowed to stir at room temperature for 16 h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was quenched with ice cold water and Attorney Docket No. 10063-110WO1

[0951] extracted with ethyl acetate (3 x 2.5 mL). The organic layers were dried over sodium sulphate and concentrated in vacuum to afford (2S,3S)-2-methyl-4-oxo-3-((Z)-2-((pentan-3- yloxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetamido) azetidine- 1 -sulfonic acid (7) (550 mg, crude) as a pale-yellow liquid. The crude was directly taken to the next step. TLC: 15% Methanol in DCM. (Rg 0.3). LCMS: (ES+) 92.99%, m / z = 690.46 [M+HJ.

[0952] Step 4: Synthesis of (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-((pentan-3-yloxy) imino) acetamido)-2-methyl-4-oxoazetidine-l-sulfonic acid

[0953] J-"" 5,

[0954] 01

[0955] HN O

[0956] .

[0957] N

[0958] o-s-o

[0959]

[0960] OH

[0961] AZ4

[0962] To a stirred solution of (2S,3S)-2-metliyl-4-oxo-3-((Z)-2-((pentan-3-yloxy) imino)- 2-(2-(tritylamino) thiazol-4-yl) acetamido) azetidine- 1 -sulfonic acid (7) (500 mg, 0.72 mmol, leq) in Formic acid (5 mL). The reaction mixture was allowed to stir at 50°C for 4 h. The reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated under reduced pressure to get crude (450mg). Crude was purified by Prep- HPLC to afford (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-((pentan-3-yloxy) imino) acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid (Target 4) (20.6 mg, 6.35% Yield) as an off white solid.

[0963] TLC: 15% methanol in DCM. (Ry 0.2). NMR (DMSO-tfg) 8: ppm 0.84 - 0.92 (m, 6H), 1.29 - 1.37 (m, 3H), 1.40 (d, J = 6.18 Hz, 3H), 1.44 - 1.53 (m, 2H), 1.54 - 1.63 (m, 2H), 3.65 - 3.69 (m, III), 4.06 - 4.11 (m, III), 4.46 (dd,. J = 8.16, 2.63 Hz, III), 6.77 (s, 1H),

[0964]

[0965] 7.75 - 8.07 (m, 1H), 9.26 (br d, J ~ 8.16 Hz, 1H) ppm. LCMS: (ES+) 96.90%, m / z - 448.01 [M+HJ. HPLC: 99.14% Purity [Method: Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1 % TEA in water, B - Acetonitrile]. Attorney Docket No. 10063-110WO1

[0966] Synthetic scheme for (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-((benzyloxy)imino) acetam.ido)-2-m.ethyl-4-oxoazetidine-l -sulfonic acid

[0967]

[0968] Step 1: Synthesis of ethyl (Z)-2-((benzyloxy)imino)-2-(2-(tritylamino) thiazol-4-yl) acetate

[0969] r-s

[0970] / -X. / X. N A ^-NHTrit

[0971] y 'o'' vN

[0972] 0^0

[0973]

[0974] 4

[0975] To a stirred solution of ethyl (Z)-2-(hydroxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetate common intermediate-2 (1 g, 2.18 mmol, leq) in DMF (5 mL) was added Potassium carbonate (756 mg, 5.46 mmol, 2.5eq) and (bromomethyl)benzene (3) (747 mg, 4.37 mmol, 2eq) at room temperature. The resulted reaction mixture was allowed to stir at 100°C for 16 h. Progress of the reaction was monitored by TLC and I, CMS. After completion, the reaction mixture was concentrated and diluted with ice cold water (20 mL) and extracted with ethyl acetate (3 x 50 mL). The organic layers were dried over sodium sulphate and concentrated under reduced pressure to get crude (2.5g). The crude was purified by trituration with DCM (50 mL) was obtained Solid (intact common-Int-2 ) which was Filtered the and the collect the filtrate MLs were evaporated under reduced pressure to afford ethyl (Z)-2-((benzyloxy)imino)-2-(2-(tritylamiiio) thiazol-4-yl) acetate (4) (900 mg, 77.16 % Yield) as a pale-yellow liquid. TLC: 70% ethyl acetate in heptane. (Ry 0.5).

[0976] LCMS: (ES+) 72.85%, m / z = 548.14 [M+H], Attorney Docket No. 10063-110WO1

[0977] Step 2: Synthesis of (Z)-2-((benzyloxy)imino)-2-(2-(tritylamino) thiazol-4-yl) acetic acid

[0978]

[0979] 5

[0980] To a stirred solution of ethyl (Z)-2-((benzyloxy)imino)-2-(2-(tritylainino) thiazol-4-yl) acetate (4) (900 mg, 1.64 mmol, leq) inTHF: MeOH: H₂O (1:1:1, 6 mL) was added Lithium Hydroxide (100 mg, 4.10 mmol, 2.5eq) at room temperature. The reaction mixture was allowed to stir at 45 °C for 16 h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated under reduced pressure and diluted with water (10ml) and acidified (pH ~3.0) with IM aq Citric acid solution (5mL) and was obtained Precipitate, which was filtered and washed with di ethyl ether (10 ml). Solid was dried under vacuum to afford (Z)-2-((benzyloxy)imino)-2-(2-( tritylamino) thiazol-4-yl) acetic acid (5) (800 mg, 93.7% Yield) as an off white solid. TLC: 15% methanol in DCM (Ry 0.2). LCMS: (ES+) 95.55%, m / z = 520.16 [M+HJ.

[0981] Step 3: Synthesis of (2S,3S)-3-((Z)-2-((benzyloxy)imino)-2-(2-(tritylamino) thiazol-4-yl) acetamido)-2-methyl-4-oxoazetidine-l-sulfonic acid

[0982] 7-s

[0983] NJL NHTrit

[0984] 'O' %■N

[0985] HN^O

[0986]

[0987] N

[0988] O=S ii-OH

[0989] o

[0990] 6

[0991] To a stirred solution of (Z)-2-((benzyloxy)imino)-2-(2-(tritylamino) thiazol-4-yl) acetic acid (5) (200 mg, 0.38 mmol, leq) in DMF (2 ml,) was added GDI (64 mg, 0.38 mmol, leq) at room temperature for 15 mins. The reaction mixture was allowed to stir at 50°C for 2 h. Then (2S,3S)-3-amino-2-methyl-4-oxoazeti dine- 1 -sulfonic acid 6 (69 mg, 0.38 mmol, leq) was added. The reaction mixture was allowed to stir at room temperature for 16 h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated in vacuum to obtain crude. The crude compound was purified by Prep-HPLC. The pure fractions from prep HPLC were lyophilization to afford Attorney Docket No. 10063-110WO1

[0992] (2S,3S)-3-((Z)-2-((benzyloxy)imino)-2-(2-(tritylamino) thiazol-4-yl) acetamido)-2-methyl- 4-oxoazetidine-l -sulfonic acid (6) (200 mg, 76% Yield) as an off white solid. LCMS: (ES+) 65%, m / z = 682.18 [M+H],

[0993] Step 4: Synthesis of (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-((benzyloxy)imino) acetamido)-2-methyl-4-oxoazetidine-l-sulfonic acid

[0994] N

[0995] N

[0996] O“S~OH

[0997]

[0998] 6

[0999] AZ5

[1000] To a stirred solution of (2S,3S)-3-((Z)-2-((benzyloxy)imino)-2-(2-(tritylamino) thiazol-4-yl) acetamido)-2-methyl-4-oxoazetidine-l-sulfonic acid (6) (260 mg, 0.38 mmol, leq) in Formic acid (3 mL) at 0°C stirred for 15 mins. The reaction mixture was allowed to stir at 50°C for 3 h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated under reduced pressure to get crude. The crude compound was purified by Prep-HPLC. The pure fractions from prep HPLC were lyophilization to afford (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-((benzyloxy)imino) acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid (AZ5) (13.30 mg, 7.94% Yield) as an off white solid.

[1001] 'H NMR (DMSO-cfc) 5: 1.39 (d, J = 6.13 Hz, 3H), 3.17 (s, 1H), 3.73 - 3.79 (m, 1H), 4.40 (dd,.7= 8.25, 2.63 Hz, 1H), 5.29 (s, 2H), 7.32 - 7.45 (m, 6H), 7.50 - 7.54 (m, III), 9.19 - 9.24 (m, III) ppm. LCMS: (ES+) 92.98%, m / z = 439.97 [M+H], HPLC: 90.32% Purity [Method: Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% TFAin water, B - Acetonitrile], Attorney Docket No. 10063-110WO1

[1002] Synthesis of(2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-((cyclohexyloxy)imino) ace, tamido)-2-methyl-4-oxoazetidine-l -sulfonic acid

[1003] LiOH (2.25 eq), THF: MeOH: H2O(1:1:1), K2CO3(2.5 eq), HO RT to 45°C DMF, 45°C,16h 12b _ „ Stepl Step-2 Common -lnt-2

[1004] 4 5

[1005] I

[1006] O“S— OH O SM-6(1 eq) DiPEA (5 eq), PYAOP(1.2eq), DMF, HCOOH, 50cC, 4h. RT,16h N Step-3 Step-4

[1007] O“S~OH 6

[1008]

[1009] Step 1: Synthesis of ethyl (Z)-2-((cyclohexyloxy)imino)-2-(2-(tritylamino) thiazol-4-yl) acetate

[1010] O O T O

[1011]

[1012] NHTrit

[1013] 4

[1014] To a stirred solution of ethyl (Z)-2-(hydroxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetate common int-2 (2 g, 4.37 mmol, leq) and Bromo cyclohexane (718 mg, 4.37 mmol, leq) in DMF (20 mL) was added Potassium carbonate (1.53 g, 10.93 mmol, 2.5eq) at room temperature. The reaction mixture was allowed to stir at 45 °C for 16 h. The reaction was monitored by TLC and LCMS. Reaction mixture was diluted with ice-cold water (20 mL) and extracted with ethyl acetate (3 x 50 mL). The combined all organic layers and dried over sodium sulphate Concentrated under reduced pressure to get crude (2.5 g). The crude was washed with DCM (50 mL). Precipitation was observed. The solid was (Starting Attorney Docket No. 10063-110WO1

[1015] material was not soluble in DCM) collected and concentrated under vacuum to afford ethyl (Z)-2-((cyclohexyloxy)imino)-2-(2-(tritylamino) thiazol-4-yl) acetate (4) (1.9 g, 81% Yield) as a pale-yellow liquid. TLC: 70% Ethyl acetate in heptane. (Ry: 0.5). LCMS: (ES+) 70.83%, m / z = 540.24 [M+H],

[1016] Step 2: Synthesis of (Z)-2-((cyclohexyloxy)imino)-2-(2-(tritylamino) thiazol-4- yl) acetic acid

[1017]

[1018] To a stirred solution of ethyl (Z)-2-((cyclohexyloxy)imino)-2-(2-(tritylamino) thiazol-4-yl) acetate (4) (1 g, 1.85 mmol, leq) in THF: MeOH: H₂O (1:1:1, 6 mL) was added LiOH (99.6 mg, 4.16 mmol, 2.25eq) in water. The reaction mixture was allowed to stir at 4 °C for 16 h. Progress of the reaction was monitored by TLC and LCMS. After completion, the Reaction mixture was concentrated and diluted with water (10 mL) and acidified with IN Aq Citric acid solution (5 ml,) was obtained the precipitate was filtered and dried under vacuum to afford (Z)-2-((cyclohexyloxy)imino)-2-(2-(tritylamino) thiazol- 4-yl) acetic acid (5) (700 mg, 73.8% yield) as an off white solid. TLC: 50% Methanol in DCM. (Ry: 0.2). LCMS: (ES+) 72.41%, m / z = 512.18 [M+H].

[1019] Step 3: Synthesis of (2S3S)-3-((Z)-2-((cyclohexyloxy)imino)-2-(2-(tritylamino) thiazol-4-yl) acetamido)-2-methyI-4-oxoazetidine-l-sulfonic add

[1020] NHTrit HN

[1021]

[1022] 7

[1023] To a stirred solution of (Z)-2-((cyclohexyloxy)imino)-2-(2-(tritylamino)thiazol-4- yl)acetic acid (5) (500 mg, 0.97 mmol, leq) in DMF (5 ml.) were added (2S,3S)-3-amino-2- Attorney Docket No. 10063-110WO1

[1024] methyl -4-oxoazetidine-l -sulfonic acid 6 (176 nig, 0.97 mmol, leq), N, N- Diisopropylethylamine (644.5 mg, 4.88 mmol, 5eq) at room temperature stirred for 15 mins. After PYAOP (637 mg, 1.17 mmol, 1.2 eq) was added in the reaction mixture at room temperature. Further, the reaction mixture was allowed to stir at room temperature for 16 h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was quenched with ice cold water and extracted with ethyl acetate (3 x 25 mL). The combined organic layers were dried over sodium sulphate and concentrated under vacuum to afford (2S,3S)-3-((Z)-2-((cyclohexyloxy)imino)-2-(2-(tritylamino) thiazol-4-yl) acetamido)-2-methyl-4-oxoazetidine-l-sulfonic acid (7) (600 mg, 91.11% Yield) as a pale¬ yellow liquid. Without purification, crude was directly utilized for the next step. TLC: 15% methanol in DCM. (Ry 0.3). LCMS: (ES+) 87.77%, m / z = 674.44 [M+H],

[1025] Step 4: Synthesis of (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-((cyclohexyloxy)imino) acetamido)-2-methyl-4-oxoazetidine-l-sulfonic acid

[1026] N

[1027] o=s=o

[1028]

[1029] OH

[1030] AZ6

[1031] To a stirred solution of (2S,3S)-3-((Z)-2-((cyclohexyloxy)iniino)-2-(2-(tritylainiiio) thiazoi-4-yl) acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid (7) (600 mg, 0.89 mmol, leq) in Formic acid (6 mL). The reaction mixture was allowed to stir at 50°C for 4 h.

[1032] Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated under reduced pressure to obtain crude (550mg). The crude was purified by Prep-HPLC to afford (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2- ((cyclohexyloxy)imino) acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid (AZ6) (19.1 mg, 4.97% Yield) as an off white solid.

[1033] TLC: 15% Methanol in DCM. (Ry 0.2). 'H NMR (DMSO-tfc) 8: 1.22 - 1.26 (m, 1H), 1.34 - 1.42 (m, 6 II), 1.50 - 1.57 (m, 3H), 1.64 - 1.72 (m, 2H), 1.93 - 1.98 (m, 2H), 3.74 - 3.80 (m, III), 4.24 (tt, 7 = 8.24, 4.26 Hz, 1H), 4.41 (dd, J = 8.22, 2.70 Hz, 1H), 7.52 - 7.55 (m, 1H), 9.10 - 9.14 (m, 1H), ppm. LCMS: (ES+) 98.72%, m / z = 431.94 [M+H]. Attorney Docket No. 10063-110WO1

[1034] HPLC: 98.18% Purity [Method: Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile (95:05), B - Acetonitrile],

[1035] Synthetic scheme for (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-(propoxyimino) acetamido )-2-methyl-4-oxoazetidine-l -sulfonic acidw[I Stepl step-2 HO' Comrnon’int-2 NI-I2N

[1036] Step-3 Step-4

[1037]

[1038] AZ7

[1039] Step-1: Synthesis of ethyl (Z)-2-(propoxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetate

[1040]

[1041] To a stirred solution of Common-int-2 (1 g, 2.18 mmol, leq) in DMF (10 mL) were added Potassium carbonate (756 mg, 5.464 mmol, 2.5eq) and 1 -bromopropane (3) (269 mg, 2.18 mmol, leq) at room temperature. The reaction mixture was allowed to stir at 45°C for 16 h. Progress of reaction was monitored by LCMS and TLC. After completion, the reaction mixture was diluted with water (50 mL) and extracted with EtOAc (3x 50 mL). The combined organic layer was washed with brine (100 mL), dried over sodium sulphate, and concentrated in vacuo to obtain crude. The crude was purified by combi flash column chromatography, eluted using 40-45 % EtOAc in heptane resulted pure fractions were concentrated to afford ethyl (Z)-2-(propoxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetate (4) (800 mg, 73.26 % Yield) as a pale-yellow semi solid. TLC: 50% Ethyl acetate in heptane. (Ry- 0.5). LCMS: (ES+) 93.71%, m / z = 500.17 [M+H],

[1042] Step-2: Synthesis of (Z)-2-(propoxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetic acid Attorney Docket No. 10063-110WO1

[1043] NHTrit

[1044] O 'V N

[1045] HO ' O

[1046]

[1047] To a stirred solution of ethyl (Z)-2-(propoxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetate (4) (1 g, 2.0 mmol, leq) in THF: MeOH: H₂O (1:1:1, 6 mL), was added Lithium Hydroxide (122.3 mg, 5.0 mmol, 2.5eq) at room temperature. The reaction mixture was allowed to stir at 45°C for 16 h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated and diluted with water (10 mL) and acidified with IN aq Citric acid solution (5 mL) was obtained Precipitation Filtered and washed with di ethyl ether (10 mL) and dried under vacuum to afford (Z)-2- (propoxyimino)-2-(2-(tritylarnino) thiazol-4-yl) acetic acid(5) (650 mg, 68.86% Yield) as an off white solid. TLC: 50% Methanol in DCM (Rt - 0.2). LCMS: (ES+) 91.31 %, m / z = 472.10 [M+H],

[1048] Step-3: Synthesis of (2S,3S)-2-methyl-4-oxo-3-((Z)-2-(propoxyimino)-2-(2-(trityl amino) thiazol-4-yl) acetamido) azetidine-l-sulfonic acid

[1049] ,-S

[1050] N / <^~NHTrit

[1051] O' N

[1052] HN O

[1053] N

[1054] O=S ii~OH

[1055]

[1056] o

[1057] 7

[1058] To a stirred solution of (Z)-2-(propoxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetic acid(5) (200 mg, 0.42 mmol, leq) and (2S,3S)-3-amino-2-methyl-4-oxoazetidine-l-sulfonic acid 6 (76.4 mg, 0.42 mmol, leq) in Pyridine (2 mL) was added HATU (199.5 mg, 0.50 mmol, 1.2eq) at room temperature. The reaction mixture was allowed to stir at 45°C for 16 h. progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated in vacuum to obtain crude. The crude was washed with DM water (5 mL) to afford (2S,3S)-2-methyl-4-oxo-3-((Z)-2-(propoxyimino)-2-(2-(trityiamino) thiazol-4-yl) acetamido) azetidine- 1 -sulfonic acid (7) (250 mg, crude) as a Attorney Docket No. 10063-110WO1

[1059] pale-yellow semi solid. TLC: 15% Methanol in DCM (Rr: 0.3). LCMS: (ES+) 78%, m / z - 634.17 [M+H],

[1060] Step-4: Synthesis of (2S,3S)-3-((Z)-2-(2-aminothiazoI-4-yi)-2-(propoxyimiiio) acetamido)-2-methyl-4-oxoazetidine-l-sulfonic add

[1061] --S

[1062] x / x N A

[1063] °' I

[1064] HN ON

[1065] N

[1066] O-S~OH

[1067]

[1068] 6

[1069] AZ7

[1070] To a solution of (2S,3S)-2-methyl-4-oxo-3-((Z)-2-(propoxyimino)-2-(2-(triiylamino) thiazol-4-yl) acetamido) azetidine- 1 -sulfonic acid (7) (250 mg, 0.39 mmol, leq) in Formic Acid (3 mL). The reaction mixtoe was allowed to stir at 45 °C for 4 h. Progress of the reaction was monitored by TLC. Starling material was consumed in TLC and LCMS analysis. After completion, the reaction mixture was concentrated in vacuum to obtain crude. The crude was purified by Prep-HPLC to afford (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-(prop oxyimino) acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid (AZ7) (9 mg, 5.8% Yield) as an off white solid.

[1071] TLC: 1 % Methanol in DCM (Rf- 0.2). *11 NMR (DMSO-&) 5: 0.87 - 0.92 (m, 3H), 0.93 - 0.96 (m, 1H), 1.38 - 1.41 (m, 3H), 1.58 - 1.65 (m, 2H), 2.31 - 2.35 (m, 2H), 2.67 (dt, J= 3.58, 1.83 Hz, 2H), 3.17 (s, 1H), 4.02 (t, 6.58 Hz, 2H), 4.39 - 4.46 (m, 1H) ppm. LCMS: (ES+) 89.07%, m / z - 391.95 [M+H], HPLC: 89.61% Purity [Method: Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% TFA in water, B - Acetonitrile], Attorney Docket No. 10063-110WO1

[1072] Synthesis of (2S,3S)-3-((Z)-2-(2-aminothiawl-4-yl)-2-((2-ethoxyethoxy) imino) acetamido )-2 -methyl -4 -oxoaz.etidine-f -sulfonic acid

[1073] ci TritHN 3 3(1 eq), KZCO3(2.5 LiOH(2.5eq), THF: MeOH: eq), DMF,100°C,12h H2O: (1:1:1), RT-65°C 16h Stepl Step-2

[1074] Common-lnt-2

[1075] O— S— OH

[1076] 6 (1eq) DIC(1.2 eq) HOBt(1,2eq), TEA(3 eq), HCOOH, 50°C, 4h DMF(5V) RT,14h Step-3 Step-4

[1077]

[1078] AZ8 Step 1: Synthesis of ethyl (Z)-2-((2-ethoxyethoxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetate

[1079] TritHN

[1080]

[1081] To a stirred solution ethyl (Z)-2-(hydroxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetate common Int-2 (1 g, 2.18 mmol, leq) and DMF (5 mL) were added Potassium carbonate (763 mg, 5.46 mmol, 2.5eq) and (chloromethoxy)ethane (3) (204 mg, 2.18 mmol, leq) in the reaction mixture at room temperature. Further, the reaction mixture was allowed to stir at 100°C for 12 h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was diluted with ice cold water (20 mL) and extracted with ethyl acetate (3 x 50 mL). The organic layers were dried over sodium sulphate and concentrated under reduced pressure to get crude (1.1 g). The Crude was purified by combi Attorney Docket No. 10063-110WO1

[1082] flash eluted using 30% ethyl acetate in heptane to afford ethyl (Z)-2-((2-ethoxyethoxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetate (4) (900 mg) as a pale-yellow liquid. TLC: 70% ethyl acetate in heptane. (Rf: 0.5). LCMS: (ES+) 98.71%, m / z = 530.24 [M+H], Step 2: Synthesis of (Z)-2-((2-ethoxyethoxy) imino)-2-(2-(tritylamino) thiazol-4- yl) acetic acid

[1083] TritHN

[1084] CL

[1085] % N

[1086] O

[1087]

[1088] 5

[1089] To a stirred solution of (Z)-2-((2-eihoxyethoxy) imino)-2-(2-(tritylamino) thiazol-4- yl) acetic acid (4) (900 mg, 1.90 mmol, 1 eq) in TIIF: MeOII: H2O: (1:1:1, 6 mL) was added lithium hydroxide (440 mg, 3.23 mmol, 2.5eq) in the reaction mixture at room temperature. The reaction mixture was allowed to stir at 65°C for 16 h. Progress of the reaction was monitored by TLC (15% Methanol in DCM) and LCMS. After completion, the reaction mixture was concentrated under reduced pressure and diluted with water (10ml) and acidified with IN aq Citric acid solution (5 mL) was obtained precipitate Filtered, washed with di ethyl ether (10 mL) and dried under vacuum to afford (Z)-2-((2- ethoxyethoxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetic acid(5) (500 mg) as an off white solid. LCMS: (ES+) 91.86 %, m / z = 502.14 [M+H],

[1090] Step 3: Synthesis of (2S,3S)-3-((Z)-2-((2-ethoxyethoxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid

[1091] Xr-sx

[1092] JL / -NHTrit

[1093] O N HN T)

[1094] N

[1095] O=S~OH

[1096]

[1097] 6

[1098] 7

[1099] To a stirred solution of (Z)-2-((2-ethoxyethoxy)imino)-2-(2-(tritylamino)thiazol-4- yllacetic acid (5) (1 g, 0.2112 mmol, leq) in DMF (2ml) were added (2~{S },3~{S})-3- amino-2-methyl-4-oxo-azetidine-l -sulfonic acid(6) (650 mg, 0.2122, leq), N, N- Diisopropylethylamine (1.00 ml), 1.056 mmol, 5eq) and PYAOP (222 mg, 0.264 mmol, Attorney Docket No. 10063-110WO1

[1100] 1.25eq ) in the reaction mixture at room temperature. The reaction mixture was allowed to stir at room temperature for 16 h. Progress of reaction was monitored by LCMS and TLC (15% of MeOH in DCM). After completion, the reaction mixture was diluted with ice cold water (20 mL) and extracted with EtOAc (2 x 40 mL). The combined organic layer was washed with brine (20 mL), dried over sodium sulphate, and concentrated in vacuo to obtain crude. The crude was purified by flash column chromatography, eluted using 1 % MeOH in DCM. The resulted pure fractions were concentrated to afford (2S,3S)-3-((Z)-2-((2- ethoxyethoxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetamido)-2-methyl-4-oxoazetidine- 1-sulfonic acid (7) (500 mg, LCMS:58%) as an off white solid. TLC: 15 % of MeOH in DCM (R / ;0.5). LCMS: (ES+) 58 %, m / z = 664[M+H], Step 4: Synthesis of (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-((2-ethoxyethoxy) imino) acetarnido)-2-rnethyl-4-oxoazetidine-l-sulfonic acid _ g, N oxNH2 HN' X) 0=< S. N

[1101] O=S=O

[1102]

[1103] OH AZ8

[1104] To a stirred solution of (2S,3S)-3-((Z)-2-((2 -ethoxyethoxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid (7) (500 mg, 0.75 mmol, leq) in Formic acid (5 mL). The reaction mixture was allowed to stir at 50°C for 4 h.

[1105] Progress of the reaction was monitored by TLC and LCMS. After completion, Reaction mixture was concentrated under vacuum to obtain crude (450 mg). The crude was purified by Prep-HPLC to afford (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-((2-ethoxyethoxy) imino) acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid (Target-8) (9 mg, 2.8% Yield) as an off white solid.

[1106] TLC: 15% methanol in DCM (Rt: 0.2). HPLC: 91.18% Purity [Method: Column: X-Select CS II C 18 (4.6* 150) mm 5u, Mobile Phase: A - 0.1 % Formic acid in water:

[1107] Acetonitrile (95:05), B - Acetonitrile], Attorney Docket No. 10063-110WO1

[1108] Synthesis of(2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-(((tetrahydro-2H-pyran-4-yl) oxy) imino) acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid

[1109] LiOH (25 eq) THE: MeOH: 1-120(1:1,1) RT-45°C.16h Step-2 Common-int-2 4 NH2

[1110] O=S~OH 6 6 (1 eq). CDI(1 eq), DMFIRT-50°C,16h Step-3 OS-OH

[1111]

[1112] 0

[1113] 7 AZ9

[1114] Step 1: Synthesis of ethyl (Z)-2-(((tetrahydro-2H-pyran-4-yl) oxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetate

[1115] J

[1116]

[1117] 4

[1118] To a stirred solution of ethyl (Z)-2-(hydroxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetate common int-2 (1 g, 2.18 mmol, leq) in DMF (5 mL) were added Potassium carbonate (756 mg, 5.46 mmol, 2.5eq) and 4-bromotetrahydropyran (3) (721 mg, 4.37 mmol, 2eq) at room temperature. The reaction mixture was allowed to stir at 100°C for 12 h. Progress of reaction was monitored by LCMS and TLC. After completion, the reaction mixture was diluted with ice cold water (20 mL) and extracted with EtOAc (2 x 50 mL). The combined organic layer was washed with brine (30 mL), dried over sodium sulphate, and concentrated in vacuo to obtain crude product, which was purified by flash column chromatography, eluted using 60 % EtOAc in heptane. The resulted pure fractions were concentrated to afford ethyl (Z)-2-(((tetrahydro-2H-pyran-4-yl) oxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetate (4) (800 mg, 67.5% Yield) as a pale-yellow liquid. TLC: 70% ethyl acetate in heptane (Ry 0.5). LCMS: (ES+) 90.07%, m / z = 542.17 [M+H], Attorney Docket No. 10063-110WO1

[1119] Step 2: Synthesis of (Z)-2-(((tetrahydro-2H-pyran-4-yl) oxy) imino)-2-(2- (tritylamino) thiazol-4-yl) acetic acid

[1120]

[1121] 5

[1122] To a stirred solution of ethyl (Z)-2-(((tetrahydro-2H-pyran-4-yl) oxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetate (4) (700 mg, 1.29 mmol, leq) in THF: MeOH: H₂O (1:1:1, 6 ml.) was added lithium hydroxide (440 mg, 3.231 mmol, 2.5eq) at room temperature. Further, the reaction mixture was allowed to stir at 45°C for 16 h. Progress of reaction was monitored by LCMS and TLC. After completion, Reaction mixture was concentrated and diluted with water (10 mL), acidified with IN aq citric acid solution (5 ml.,) was obtained Precipitation Filtered, washed with di ethyl ether (10 mL) and dried under vacuum to afford (Z)-2-(((tetrahydro-2H-pyran-4-yl) oxy) imino)-2-(2-(tritylamino) thiazoi-4-yl) acetic acid (5) (350 mg, 52.74% yield) as an off white solid. TLC: 15% Methanol in DCM (Ry 0.2). LCMS: (ES+) 62.11%, m / z = 514.1 [M+H],

[1123] Step 3: Synthesis of (2S,3S)-2-methyl-4-oxo-3-((Z)-2-(((tetrahydro-2H-pyran-4-yl) oxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetamido) azetidine-l-sulfonic acid

[1124] O-S-OH

[1125]

[1126] 6

[1127] 7

[1128] To a stirred solution of (Z)-2-(((tetrahydro-2H-pyran-4-yl) oxy) imino)-2-(2- (tritylamino) thiazol-4-yl) acetic acid (5) (300 nig, 0.58 mmol, leq) in DMF (2 mL) was added CDI (97 mg, 0.58 mmol, leq) in the reaction mixture at room temperature stirred for 2h. After (2S,3S)-3-amino-2-metliyl-4-oxoazetidine-l -sulfonic acid (6) (105 mg, 0.58 mmol, leq) was added. The reaction mixture was allowed to stir at 50°C for 16 h. Progress of reaction was monitored by LCMS and TLC. After completion, the reaction mixture was diluted with water (50 mL) and extracted with EtOAc (2 x 50 mL). The combined organic Attorney Docket No. 10063-110WO1

[1129] layer was washed with brine (40 mL), dried over sodium sulphate, and concentrated in vacuo to obtain crude. The crude was purified by flash column chromatography, eluted using 15 % EtOAc in heptane. The pure fractions were concentrated to afford (2S,3S)-2-methyl-4-oxo-3-((Z)-2-(((tetrahydro-2H-pyran-4-yl) oxy) imino)-2-(2-(tritylamino) thiazol- 4-yl) acetamido) azetidine- 1 -sulfonic acid (7) (320 mg, 81 % Yield) as a pale-yellow semi solid. TLC: 15% Methanol in DCM (Ry 0.2). LCMS: (ES+) 55.91%, m / z = 676.22 [M+H], Step 4: Synthesis of (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-(((tetrahydro-2H-pyran-4-yl) oxy) imino) acetamido)-2-methyl-4-oxoazetidine-l-sulfonic acid

[1130] O=S~OH

[1131]

[1132] 6

[1133] AZ9

[1134] To a stirred solution of (2S,3S)-2-methyl-4-oxo-3-((Z)-2-(((tetrahydro-2H-pyran-4- yl) oxy) imino)-2-(2-(trityl amino) thiazol-4-yl) acetamido) azetidine- 1 -sulfonic acid (7) (320 mg, 0.47 mmol, leq) in Formic acid (4 mL) at room temperature. The reaction mixture was allowed to stir at 45°C for 4 h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated under vacuum to afford crude product which was purified by Prep-HPLC. The resulted pure fractions from prep HPLC were lyophilization to afford (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-(((tetrahydro-2H-pyran-4-yl) oxy) imino) acetamido)-2-methyl-4-oxoazetidine-l-sulfonicacid. (AZ9) (14.9 mg, 7.26% Yield) as awhile solid.

[1135] TLC: 15% Methanol in DCM (Ry 0.2). Hl NMR (DMSO-d6) δ:1.39 (d,.7 = 6.13 Hz, 3H), 1.62 - 1.70 (m, 2H), 1.94 - 2.01 (m, 2H), 3.17 (d, J = 5.25 Hz, 1H), 3.46 (t, J = 8.94 Hz, 2H), 3.68 - 3.72 (m, 1H), 3.76 - 3.83 (m, 2H), 4.35 - 4.42 (m, 2H), 7.09 (s, 2H), 7.47 (s, 1H), 9.09 (d, 8.25 Hz, 1H) ppm. LCMS: (ES+) 86.87%, 433.45 [M+H], HPLC: 88.97 % Purity [Method: Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1 % TFA in water, B - Acetonitrile. Attorney Docket No. 10063-110WO1

[1136] Synthesis of (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-(( 1 -phenylpropoxy) imino) acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid

[1137] o THF: MeOH: H2O TritHN— X A (1:1:1), LiOH(2.5eq), N" OH RT- 45°C, 16h Step-1 Step-2 Common lnt-2

[1138] NQ

[1139] O=S-OH 3 II \ 0 6 zx yy' N 6 (1 eq), PYAOP X. / HCOOH,45°C, 4h (1.2 eq). DIPEAu(5 eq), DMF, RT.16 h. A - >- o=C 2. Sts p-4 Step-3 N O=S~OH 7

[1140]

[1141] AZ10 Step 1: Synthesis of ethyl (Z)-2-((l-phenyIpropoxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetate

[1142]

[1143] 4

[1144] To a stirred solution of ethyl (Z)-2-(hydroxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetate common-int-2 (1 g, 2.18 mmol, leq) and 1 -bromopropylbenzene (3) (435 mg, 2.18 mmol, leq) in DMF (10 mL) was added Potassium carbonate (756 mg, 5.46 mmol, 2.5eq) in the reaction mixture at room temperature. The reaction mixture was allowed to stir at 45 °C for 16 h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was diluted with ice cold water (20 mL) and extracted with ethyl acetate (3 x 50 ml). The combined organic layers were dried over sodium sulphate and concentrated in vacuo to obtain crude (2.5g). The crude product was trituration by DCM (50 mL) and insoluble material (common Int-2) was Filtered and collected filtrate MLs were concentrated under reduced pressure to afford ethyl (Z)-2-((l -phenylpropoxy) imino)-2-(2- Attorney Docket No. 10063-110WO1

[1145] (tritylamino) thiazol-4-yl)acetate (4) (850 mg, 67.55% Yield) as a pale yellow liquid. TLC: 70% Ethyl acetate in Heptane. (R 0.5). LCMS: (ES+) 71.47%, 576.31 [M+H].

[1146] Step 2: Synthesis of (Z)-2-((l-phenylpropoxy ) imino)-2-(2-(trityIamino) thiazol-4-yl) acetic acid

[1147] ,s-n O

[1148] TritHN— % Jk U.

[1149] N Y " OH

[1150] O XQ

[1151] 5

[1152] To a stirred solution of ethyl (Z)-2-((l -phenylpropoxy imino)-2-(2-(tritylamino) thiazol-4-yl) acetate (4) (700 mg, 1.21 mmol, leq) THE: MeOH: H2O (1:1:1, 6 mL) was added LiOH (74 mg, 3.04 mmol, 2.5eq) portion wise in the reaction mixture at room temperature. The reaction mixture wras allowed to stir at 4 °C for 16 h. The reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated under reduced pressure, diluted with water (10 mL), and acidified with IN Citric acid solution (5 mL) to pH 3. Precipitation was obtained. Filtered the solid and dried under reduced pressure to afford (Z)-2-((l -phenylpropoxy) imino)-2-(2-(trityl amino) thiazol-4-yl) acetic acid (5) (350 mg, 52.5% Yield) as an off white solid. TLC: 50% of methanol in DCM. (Ry 0.2). LCMS: (ES+) 90.56%, 548.2 [M+H],

[1153] Step 3: Synthesis of (2Sr3S)-2-methyI-4-oxo-3-((Z)-2-((l-phenylpropoxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetamido) azetidine-l-sulfonic acid

[1154] O=S~OH

[1155]

[1156] 6

[1157] 7

[1158] To a stirred solution of (Z)-2-((l -phenylpropoxy )imino)-2-(2-(tritylamino)thiazol -4-yl)acetic acid (5) (400 mg, 0.73 mmol, leq) in DMF (5 ml.) were added (2S,3S)-3-amino-2-methyl-4-oxoazetidine-l -sulfonic acid (6) (131.6 mg, 0.73 mmol, leq) and N, N-Diisopropylethylamine (477 mg, 3.652 mmol, 5eq) at room temperature stirred for 15min. Attorney Docket No. 10063-110WO1

[1159] Then, PYAOP (476 mg, 0.87 mmol, 1.2eq) was added. The reaction mixture was allowed to stir at room temperature for 16 h. The reaction progress was monitored by TLC and LCMS. After completion, the reaction mixture was quenched with ice cold water and extracted with ethyl acetate (3 x 25 mL). The organic layers were dried over sodium sulphate and concentrated under reduced pressure to afford (2S,3S)-2-methyl-4-oxo-3-((Z)-2-((l- phenylpropoxy) imino)-2-(2-(tritylamino) thiazol-4-yl) acetamido) azetidine- 1 -sulfonic acid (7) (500 mg, 96.45% Yield) as a pale-yellow liquid. Which was without further purification taken for the next step. TLC: 15% methanol in DCM. (Ry. 0.3). LCMS: (ES+) 69.43%, 710.20 [M+HJ.

[1160] Step 4: Synthesis of (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-((l -phenylpropoxy) imino) acetamido)-2-methyl-4-oxoazetidine-l-sulfonic acid

[1161] N

[1162] O-S OH

[1163] 6

[1164]

[1165] AZ10

[1166] To a solution of (2S,3S)-2-methyl-4-oxo-3-((Z)-2-((l-phenylpropoxy) imino)-2-(2- (tritylamino) thiazol-4-yl) acetamido) azetidine- 1 -sulfonic acid (7) (400 mg, 0.85 mmol, leq) in Formic acid (5 mL) at room temperature and then The reaction mixture was allowed to stir at 45 °C for 4 h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated under reduced pressure to get crude (500 mg). Crude was purified by Prep-HPLC to afford (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2- ((1 -phenylpropoxy) imino) acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid (AZ10) (22 mg, 5.5% Yield) as an off white solid.

[1167] TLC: 15% methanol in DCM. (Ry 0.3). NMR (DMSO-d6) δ: 0.83 (t,.7= 7.24, 2.87 Hz, 3H), 1.41 (d,.7= 5.99 Hz, 3 H), 1.73 - 1.84 (m, 2H), 3.68 - 3.76 (m, HI), 4.46 - 4.54 (m, 1H), 4.99 - 5.05 (m, 1H), 6.65 - 6.67 (m, 1H), 6.99 - 7.10 (m, 1H), 7.15 - 7.20 (m, 2H), 7.22 - 7.35 (m, 5H), 9.25 - 9.31 (m, 1H) ppm. LCMS: (ES+) 84.45%, 468.2 [M+HJ.

[1168]

[1169] HPLC: 82.23% Purity [Method: Column: X-Bridge C18 (4.6*150) mm 5u, Mobile Phase:

[1170] A - 5mM Ammonium Bicarbonate in water, B - Acetonitrilel. Attorney Docket No. 10063-110WO1

[1171] Synthetic scheme for (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2- ((cyclohexylmethoxy)imino) acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid Brr-S N Jl NHTrit 3 LiOH(25 eq), THF: MeOH: HO' y N 3(2eq) K2CO3(2.5 eq). H2O(1:1:1), RT-45°C, L DMF.100°C.12h O ''O - ► L£tl _ » 0 Stepl Step-2 Common-int-2 4 5 NH2N

[1172] 0=4-011 I' / f~ NHTrit 0 6 ''N HC00H,45°C,4h Step-3 N Step-4

[1173] O=S-OH 0

[1174]

[1175] AZ11 Step 1: Synthesis of ethyl (Z)-2-((cyclohexylmethoxy)imino)-2-(2-(tritylamino) thiazol-4-yl) acetate

[1176] N JL <^~NHTrit

[1177] % 'V N

[1178] O^'O

[1179]

[1180] To a stirred solution of ethyl (Z)-2-(hydroxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetate common-int-2 (1 g, 2.18 mmol, leq) in DMF (5 mL) were added Potassium carbonate (756 mg, 5.46 mmol, 2.5eq) and bromomethyl cyclohexane (3) (774 mg, 4.37 mmol, 2eq) in the reaction mixture at room temperature for 15 mins. Further, the resulted reaction mixture was allowed to stir at 100°C for 12 h. Progress of the reaction was monitored by TEC (70% ethyl acetate in heptane) and LCMS. After completion, the reaction mixture was diluted with ice cold water (20 mL) and extracted with EtOAc (3 x 50 mL). The combined organic layers were washed with brine (20 mL), dried over sodium sulphate, filtered, and concentrated under reduced pressure to get crude. Which was purified by combi flash column chromatography using YMC-12 g cartridge, and the product was eluted with 70 % ethyl acetate in heptane. The eluted fractions were concentrated in vacuo to afford ethyl (Z)-2-((cyclohexylmethoxy)imino)-2-(2-(tritylamino) thiazol-4-yi) acetate (4) (900 mg, 74.36% Yield) as brown solid. LCMS: (ES+) 98.52%, m / z = 554.10 [M+H] Attorney Docket No. 10063-110WO1

[1181] Step 2: Synthesis of (Z)-2-((cyclohexylmethoxy)imino)-2-(2-(tritylamino) thiazol-4-yl) acetic acid

[1182]

[1183] To a stirred solution of ethyl (Z)-2-((cyclohexylmethoxy)imino)-2-(2-(tritylamino) thiazol-4-yl) acetate (4) (900 mg, 1.62 mmol, leq) THF: H2O: MeOH (1:1:1, 6 mL) was added 14OH (97.4 mg, 4.064 mmol, 2.5 eq) at room temperature. The reaction mixture was allowed to stir at 45 °C for 16 h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated and diluted with water (10 mL) acidified with IN aq Citric acid solution (5 mL.) was obtained Precipitate Filtered, washed with di ethyl ether (10 mL) and dried under vacuum to afford (Z)-2-((cyclohexylmethoxy)imino)-2-(2-(tritylaniino) thiazol-4-yl) acetic acid(5) (450 mg, 52.66% Yield) as an off white solid. TLC: 50% Methanol in DCM. (Ry 0.2). LCMS: (ES+) 81%, m / z = 526.21 [M+H],

[1184] Step 3: Synthesis of (2S,3S)-3-((Z)-2-((cyclohexylmethoxy)imino)-2-(2-(trityl amino) thiazol-4-yl) acetamido)-2-methyl-4-oxoazetidine-l-sulfonic acid

[1185] N

[1186] O=S~OH

[1187]

[1188] 6

[1189] 7

[1190] To a solution of (Z)-2-((cyclohexylmethoxy)imino)-2-(2-(tritylamino)thiazol-4-yl)acetic acid(5) (500 mg, 0.95 mmol, leq) in DMF (10 mL) were added (2S,3S)-3-amino-2-methyl-4-oxoazetidine-l -sulfonic acid (171 mg, 0.95 mmol, leq) and N, N- Diisopropylethylamine (621 mg, 4.75 mmol, 5eq) at room temperature stirred for 15min. Then followed by was added PYAOP (645 mg, 1.18 mmol, 1.25eq), Then the reaction mixture was allowed to at room temperature for 16 h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was quenched with ice cold water and extracted with ethyl acetate (3 x 25 mL). The Combined organic layers Attorney Docket No. 10063-110WO1

[1191] and dried over sodium sulphate and concentrated under vacuum to afford (2S,3S)-3-((Z)-2- ((cyclohexyl methoxy) imino)-2-(2-(trityl amino) thiazol-4-yl) acetamido)-2-niethyl-4-oxoazetidine- 1 -sulfonic acid (7) (600 mg, 91.72% Yield) as a pale-yellow liquid. TLC: 15% Methanol in DCM. (R 0.3). LCMS: (ES+) 60.65%, m / z = 688.19 [M+H],

[1192] Step 4: Synthesis of (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2- ((cyclohexylmethoxy)imino) acetamido)-2-methyI-4-oxoazetidine-l -sulfonic acid

[1193] N

[1194]

[1195] AZ11

[1196] To a stirred solution of (2S,3S)-3-((Z)-2-((cyclohexylmethoxy)iniino)-2-(2- (tritylamino) thiazol-4-yl) acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid (7) (600 mg, 0.87 mmol, leq) in Formic acid (10 mL). The reaction mixture was allowed to stir at 45°C for 4 h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated under vacuum to obtain Crude which was purified by Prep-HPLC to afford (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-((cyclohexylmethoxy)imino) acetamido)-2-methyl-4-oxoazetidine- 1 -sulfonic acid (AZ11) (4.2 mg, 1.1% Yield) as an off white solid.

[1197] TLC: 1 % Methanol in DCM. (Ry: 0.2). ‘H NMR (DMSO-&) 8: 0.86 - 1.03 (m, 2H), 1.09 - 1.26 (m, 3H), 1.39 (d, 6.24 Hz, 3H), 1.51 - 1.75 (m, 7H), 3.62 - 3.71 (m, 1H), 3.80 - 3.90 (m, 2H), 4.40 - 4.50 (m, 1H), 6.71 (s, 1H), 7.05 - 8.28 (m, 2H), 9.24 (d, J = 7.95 Hz, 1H) ppm. LCMS: (ES+) 99.26%, m / z= 445. 1 [M+H], HPLC: 94.09% Purity [Method: Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% TFAin water, B - Acetonitrile. Attorney Docket No. 10063-110WO1

[1198] Synthetic scheme for (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-(phenethoxyimino) acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid

[1199] 3 (2 eq), K2CO3 / TS\ THF: MeOH: H2O N JL ^--NHTrit (25 eq), DMF, 100 N JL > / —NHTi-it HO' VN‘0" N (1:1:1i LiOH (25 eq),

[1200] °C for 16 h RT-45C 16h Step) '0 Step-2 Common-Snt-2

[1201] 6 6 (1 eq), PYAOP (1 2 eq), DIPEA (5 eq), DMF, RT.16 h N

[1202] O“S ii~OH o

[1203]

[1204] AZ12 Step 1: Synthesis of ethyl (Z)-2-(phenethoxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetate

[1205]

[1206] To a stirred solution of ethyl (Z)-2-(hydroxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetate common-int-2 (1 g, 2.186 mmol, leq) in DMF (5 mL) was added Potassium carbonate (756 mg, 5.46 mmol, 2.5eq) and 2-bromoethylbenzene (3) (809 mg, 4.37 mmol, 2eq) at room temperature. The reaction mixture was allowed to stir at 100°C for 16 h.

[1207] Progress of the reaction was monitored by TEC (70% ethyl acetate in heptane) and LCMS. After completion, the reaction mixture was diluted with ice-water (70 mL) and extracted with DCM (2 x 50 mL). The combined organic layers were washed with brine (30 mL), dried over sodium sulphate, filtered, and concentrated under reduced pressure to get crude. The crude compound was purified by combi flash column chromatograph, eluted using 80 % EtOAc in heptane. The resulted pure fractions were concentrated to afford ethyl (Z)-2-(phenethoxyimino)-2-(2-(trityl amino) thiazol-4-yl) acetate (4) (800 mg, 65.57% yield) as a pale-yellow liquid. LCMS: (ES+) 36.39%, m / z- 562.17 [M+H],

[1208] Step 2: Synthesis of (Z)-2-(phenethoxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetic acid Attorney Docket No. 10063-110WO1

[1209] Njl NHTrit

[1210] O" N

[1211] HO O

[1212]

[1213] To a stirred solution of ethyl (Z)-2-(phenethoxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetate (4) (750 mg, 1.33 mmol, leq) in THF: MeOH: H₂O (1:1:1, 6 mL) was added lithium hydroxide (81 mg, 3.33 mmol, 2.5eq) at room temperature. The reaction mixture was allowed to stir at 45°C for 16 h. Progress of reaction was monitored by LCMS and TLC (15% methanol in DCM). After completion, Reaction mixture was concentrated and diluted with water (10 mL), acidified with IN aq Citric acid solution (5 mL) was obtained Precipitation filtered, washed with di ethyl ether (10 mL), dried under reduced pressure to afford (Z)-2-(phenethoxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetic acid (5) (350 mg, 49.1% yield) as an off white solid. LCMS: (ES+) 64.70%, m / z= 534.43 [M+H],

[1214] Step 3: Synthesis of (2S,3S)-2-methyl-4-oxo-3-((Z)-2-(phenethoxyimino)-2-(2-(trityl amino) thiazol-4-yl) acetamido) azetidine-l-sulfonic acid

[1215] HN O

[1216] N

[1217] O=S-OH

[1218] 6

[1219]

[1220] 7

[1221] To a stirred solution of (Z)-2-(phenethoxyimino)-2-(2-(tritylamino)thiazol-4-yl)acetic acid (5) (500 mg, 0.93 mmol, leq) in DMF (10 mL) were added (2S,3S)-3-amino-2-methyl-4-oxoazetidine-l -sulfonic acid 6 (169 mg, 0.93 mmol, leq) and N, N- Diisopropylethyl amine (618 mg, 4.685 mmol, 5eq) at room temperature for 15 mins. Then PYAOP (636 mg, 1.17 mmol, 1.2eq) was added. The resulted reaction mixture was allowed to stir at room temperature for 16 h. Progress of the reaction was monitored by TLC (15% methanol in DCM) and LCMS. After completion, the reaction mixture was diluted with water (50 mL) and extracted with EtOAc (2 x 40 mL). The combined organic layers were washed with brine (20 mL), dried over sodium sulphate, filtered, and concentrated under reduced pressure to get crude. The crude compound was purified by combi flash column chromatography using YMC-12 g cartridge, and the product was eluted with 1 % DCM in Attorney Docket No. 10063-110WO1

[1222] methanol. The eluted fractions were concentrated in vacuo to afford pure (2S,3S)-2-methyl- 4-oxo-3-((Z)-2-(phenethoxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetamido) azetidine-1-sulfonic acid (7) (550 mg, 84.36% Yield) as a pale-yellow liquid. LCMS: (ES+) 76.3%, m / z= 696.2 [M+H],

[1223] Step 4: Synthesis of (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-(phenethoxyimino) acetamido) -2-methyl-4-oxoazetidine-l -sulfonic acid

[1224]

[1225] AZ12

[1226] To a stirred solution of (2S,3S)-2-methyl-4-oxo-3-((Z)-2-(phenethoxyimino)-2-(2-(tritylamino) thiazol-4-yl) acetamido) azetidine- 1 -sulfonic acid (7) (650 mg, 0.9342 mmol, leq) in formic acid (10 mL) was added at room temperature. The reaction mixture was allowed to stir at 45 °C for 4 h. Progress of the reaction was monitored by TLC (15% methanol in DCM) and LCMS. After completion, the reaction mixture was concentrated under reduced pressure to get crude. The crude compound was purified by Prep-HPLC. The resulted pure fractions were lyophilization to afford (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-(phenethoxyimino) acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid (AZ12) (14 mg, 3.35 % Yield) as an off white solid.

[1227] NMR (DMSO-d6) δ: 1.39 (d, 7= 6.13 Hz, 3H), 2.33 (dt, 7= 3.69, 1.78 Hz, 2H) 2.67 - 2.69 (m, 1 H) 2.92 (t, 7 = 6.82 Hz, 2H), 3.65 - 3.72 (m, III), 4.24 (t, 7 = 6.88 Hz, 2H), 4.42 (dd, 7= 7.75, 2.75 Hz, III), 6.75 (s, III), 7.17 - 7.23 (m, III), 7.25 - 7.31 (m, 5H), 9.29 (d, 7= 7.75 Hz, 1H) ppm. LCMS: (ES+) 86.24%, m / z~ 454.2 [M+H]. HPLC: 90.04% Purity [Method: Column: X-Select CSH Cl 8 (4.6*150) mm 5u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile (95:05), B - Acetonitrile].

[1228] AZ series 2 compounds Attorney Docket No. 10063-110WO1

[1229] Synthesis of ( 2S, 3S)-3-[ [ (2Z)-2-( 2-aminothiazol-4-yl)-2-( 1, 1 -dimethyl-2-oxo-2-propoxy-ethoxy) imino-acetyl]amino ]-2-methyl-4-oxo-azetidine-l -sulfonic acid

[1230] 2 2 (1 eq), EDCI(1.2 eq), HOBt (1.5 eq), DMF(1.5 mL), DIPEA (3 eq),0°C-RT, 3h N

[1231] O=S OH 6

[1232]

[1233] AZ1-2 To a stirred solution of 2-((((Z)-1-(2-aminothiazol-4-yl)-2-(((2S,3S)-2-methyl-4- oxo-l-sulfoazetidin-3-yl)amino)-2-oxoethylidene)amino)oxy)-2-methylpropanoic acid [Aztreonam] 1 (300 mg, 0.68 mmol, 1 eq) in DMF (1.5 mL) were added EDCI. HC1 (160 mg, 0.817 mmol, 1.19 eq) and HOBt (139 mg, 1.01 mmol, 1.5 eq) at 0°C stirred for 5 min after DIPEA (0.35 ml, 2.02 mmol, 3 eq) and propan-l-ol, 2 (41.3 mg, 0.687 mmol, 1 eq) were added at 0°C. The reaction mixture was stirred at RT for 3h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated in vacuo to obtain of crude (430 mg). The crude was purified by prep-HPLC. The pure fraction were lyophilisation to afford (2S,3S)-3-[[(2Z)-2-(2-aminothiazol-4-yl)-2-(l,l-dimethyl-2-oxo-2-propoxy-ethoxy)imino-acetyl] amino] -2-methyl-4-oxo azetidine- 1 - sulfonic acid (AZ2-1) (77 mg, 23.40 %) as off white solid.

[1234] ¹H NMR (DMSO-d6) 5: 9.24 (d, 1=8.1 Hz, 1H), 6.85 (s, 2H), 4.47 (dd, J=2.7, 8.1 Hz, 1H), 4.04 (t, 1=6.4 Hz, 3H), 3.70 (qd, J=2.7, 6.2, Hz, 1H), 1.59 (sxt, 1=7.1 Hz, 2H), 1.47 (d, 1=4.3 Hz, 6H), 1.41 (d, 1=6.1 Hz, 3H), 0.86 (t, 1=7.5 Hz, 3H) ppm. LCMS: (ES+) 99.81%, m / z = 478.99 [M+H] [Method: Column: CORTECS UPLC Cl 8 (3*30) mm, 1.6um, Mobile Phase A: 0.05%FAin Water, Mobile Phase B: 0.05%FA in Acetonitrile]. HPLC: 99.88% Purity [Method: Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1 % TEA in water, B - Acetonitrile]. Attorney Docket No. 10063-110WO1

[1235] Synthesis of ( 2S, 3S)-3-[ [ (2Z)-2-( 2-aminothiazol-4-yl)-2-[2-( isobutylamino )-l, 1 - dimethyl-2-oxo-ethoxy imino-acetyl]amino]-2-methyl-4-oxo-azetidine-l -sulfonic acid NH22 2 (1 eq), EDCI(1.2 eq), HOBt (1.5 eq), DMF(1.5 mL), D! PEA (3 eq), 0°C-RT, 3h N

[1236] O^S-OH I I O S O OH

[1237]

[1238] AZ2-2 To a stirred solution of 2-((((Z)-l-(2-aminothiazol-4-yl)-2-(((2S,3S)-2-methyl-4-oxo-l-sulfoazetidin-3-yl)amino)-2-oxoethylidene)amino)oxy)-2-methylpropanoic acid [Aztreonam] 1 (300 mg, 0.68 mmol, 1 eq) in DMF (1.5 mL) were added EDCI. HC1 (160 mg, 0.81 mmol, 1.19 eq) and HOBt (139 mg, 1.01 mmol, 1.46 eq) at 0°C stirred for 5 mins after DIPEA (0.35 ml, 2.02 mmol, 3 eq) and 2-methylpropan-l -amine 2 (50.4 mg, 0.68 mmol, 1 eq) were added at 0°C. The reaction mixture was allowed to stir at RT for 3h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated in vacuo to obtain of crude (450 nig). The crude was purified by prep-HPLC to afford (2S,3S)-3-[[(2Z)-2-(2-aminothiazol-4-yl)-2-[2-(isobutylamino)-l,l-dimethyl-2-oxo-ethoxy]imino-acetyl]amino]-2-methyl-4-oxo-azetidine-l -sulfonic acid (AZ2-2) (111 mg, 32.84%) as an off white solid.

[1239] NMR (DMSO-d6) 5: 9.52 (d, J=8.1 Hz, 1H), 7.85-8.61 (m, 1H), 7.28 (t, J=6.1 Hz, 1H), 6.94 (s, 1H), 4.53 (dd,.1=2.6, 8.1 Hz, TH), 3.71 (qd, J=2.6, 6.1 Hz, 1H), 2.92 (id,.1=6.5, 2.9 Hz, 2H), 1.72 (dquin, J=6.8, 13.5 Hz, 1H), 1.41-1.46 (m, 9H), 0.79 (dd, J=0.9, 6.7 Hz, 6H) ppm LCMS: (ES+) 99.73%, m / z = 491.01 [M+H], [Method: Column: CORTECS UPLC C18 (3*30) mm, 1.6um, Mobile Phase A: 0.05 %FA in Water, Mobile Phase B:

[1240] 0.05%FA in Acetonitrile]. HPLC: 99.79%, [Method: Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% TFAin water, B - Acetonitrile]. Attorney Docket No. 10063-110WO1

[1241] Synthesis of(2S,3S)-3-((Z)-13-(,2-aminothiazol-4-yl)-10,10-dimetltyl-9-oxo-2,5,ll- trioxa-8, 12-diazatetradec-12-en-14-amido)-2-methyl-4-oxoaz.etidine-l-sulfonic acid

[1242] 2 (1 eq), EDC(1.2 eq), HOBt (1.5 eq), DMF(1.5 mL), DIPEA (3 eq),0°C -RT, 3h HN "0 N

[1243] o=s=o N OH O=S~OH 6

[1244]

[1245] AZ3-2 To a stirred solution of 2-((((Z)-1-(2-aminothiazol-4-yl)-2-(((2S,3S)-2-methyl-4-oxo-1-sulfoazetidin-3-yl)amino)-2-oxoethylidene)amino)oxy)-2-methylpropanoic acid [Aztreonam] 1 (100 mg, 0.22 mmol, 1 eq) in DMF (1.5 ml) were added EDCI. HC1 (53.1 mg, 0.27 mmol, 1.19 eq), HOBt (46.1 mg, 0.33 mmol, 1.46 eq) at 0°C stirred for 5 mins after DIPEA (0.12 mL, 0.69 mmol, 3eq) and 2-(2-methoxyethoxy)ethanamine 2 (27.4 mg, 0.23 mmol, 1 eq) were added at 0°C. The reaction mixture was allowed to stir at RT for 3h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated in vacuo to obtain crude compound (180 mg). The crude was purified by prep-HPLC to afford (2S,3S)-3-((Z)-13-(2-aminothiazol-4-yl)-10,10-dimethyl-9-oxo-2,5, 11 -trioxa-8, 12-diazatetradec- 12-en- 14-amido)-2-methyl-4-oxoazetidine- 1 - sulfonic acid (AZ3-2) (15 mg, 12.17 %), as off white solid.

[1246] !H NMR (DMSO-d6) 5: 9.42 (d, J=8.1 Hz, 1H), 7.46-7.92 (m, 1H), 7.35 (t, J=5.8 Hz, HI), 6.90 (s, 2H), 4.54 (dd, J=2.6, 8.1 Hz, 2H), 3.72 (qd, J=2.6, 6.2 Hz, HI), 3.45-3.49 (m, 2H), 3.39-3.43 (m, II), 3.35-3.39 (m, 2H), 3.25 (q, 1=6.1 Hz, 2H), 1.37-1.47 (m, 9H) ppm. LCMS: (ES+) 99.19% m / z = 537.2 [M+HJ Method: Column: X Select CSH-C18 (3.0X50mm, 2.5pm) Mobile Phase: A: 0.05% TEA in water, Mobile Phase: B: 0.05% TEA in ACN]. HPLC: 98.33% purity, [Method: Column: X-Select CSH C18, (4.6*150) mm 5u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile (95:05), B - Acetonitrile]. Attorney Docket No. 10063-110WO1

[1247] Synthesis of ( 2S,3S)-3-[ [ (2Z)-2-( 2-aminothiazol-4-yl)-2-[l,l -dimethyl-2-oxo-2- (propylamino) ethoxy]imi.no-acetyl]amino]-2-methyl-4-oxo-azetidine-i -sulfonic acid " NH22 HOJA. N. A / ”NH2 2(1.5 eq), DCC(0.15 mL), HOBt I o 'TN(1.36 eq), DMF(3 mL), DIPEA ° HN X) (2.5 eq), 0°C-RT, 3h N N

[1248] O=S ii~OH O-S~OH

[1249]

[1250] o o AZ4-2

[1251] To a stirred solution of 2-((((Z)-1-(2-aminothiazol-4-yl)-2-(((2S,3S)-2-methyl-4- oxo-l-sulfoazetidin-3-yl)amino)-2-oxoethylidene)amino)oxy)-2-methylpropanoic acid [Aztreonam] 1 (300 mg, 0.68 mmol, 1 eq) in DMF (3 mL) were added N, N'- Dicyclohexylcarbodiimide (0.15 mL), HOBt (129.6 nig, 0.93 mmol, 1.36 eq) at 0°C stirred for 5 mins After DIPEA (0.3 mL, 2 mmol, 2.5 eq) and propan- 1 -amine 2 (61.1 mg, 1.03 mmol, 1.5 eq) were added at 0°C. The reaction mixture was allowed to stir al RT for 3h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated in vacuo to obtain crude compound (400 mg). The crude was purified by prep-HPLC to afford (2S,3S)-3-[[(2Z)-2-(2-aminothiazol-4-yl)-2-[l,l-dimethyl- 2-oxo-2-(propylamino)ethoxy]imino-acetyl]amino]-2-methyl-4-oxo-azetidine-l -sulfonic acid (AZ4-2), (50 mg, 15.23 %), as an off white solid.

[1252] NMR (DMSO-d6) 5: 9.49 (d, J=8.1 Hz, 1H), 7.28 (t, J=5.9 Hz, 1H), 6.94 (s, 1H), 4.53 (dd, J=2.6, 8.1 Hz, 1H), 3.72 (qd, J=2.5, 6.0 Hz, 1H), 3.06 (q, J=6.6 Hz, 2H), 1.36-1.49 (m, 11H), 0.80 (t, 1=7.4 Hz, 3H) ppm. LCMS: (ES+) 96.85% m / z = 477.3 [M+H] [Method: Column: X-Select C18, (50mm*3.0mm, 2.5um), Mobile Phase A: 2.5Mm NH4CO3 In Water, Mobile Phase B: 100% Acetonitrile], HPLC: 97.22% purity [Method: Column: X-

[1253]

[1254] Select CSH Cl 8 (4.6*150) mm 5u, Mobile Phase: A - 0.1% Formic acid in water:

[1255] Acetonitrile (95:05), B - Acetonitrile]. Attorney Docket No. 10063-110WO1

[1256] Synthesis of ( 2S, 3S)-3-[ [ (2Z)-2-( 2-aminothiazol-4-yl)-2-[2-( diethylamino)-!,1 - dimethyl-2-oxo-ethoxy imino-acetyl]amino]-2-methyl-4-oxo-azetidine-l -sulfonic acid NH

[1257] — / 2 2(1 eq), EDCI (1.5 eq), HOBt (1.5 eq), DMF(3 mL), DIPEA (2.5 eq), 0°C-RT, 3h

[1258] N

[1259] O=S~OH 6

[1260]

[1261] AZ5-2 To a stirred solution of 2-((((Z)-1-(2-aminothiazol-4-yl)-2-(((2S,3S)-2-methyl-4- oxo-l-sulfoazetidin-3-yl)amino)-2-oxoethylidene)amino)oxy)-2-methylpropanoic acid [Aztreonam] 1 (300 mg, 0.68 mmol, 1 eq) in DMF (1 mL) were added EDCLHCl (160 mg, 0.817 mmol, 1.2 eq), HOBt (139 mg, 1.01 mmol, 1.5 eq) at 0°C stirred for 5 mins after DIPEA (0.35 ml, 2.02 mmol, 2.93 eq) and diethylamine 2 (50.4 mg, 0.68 mmol, 1 eq) were added 0°C. The reaction mixture was allowed to stir at RT for 3h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated in vacuo to obtain crude compound (400 mg). The crude was purified by prep-HPLC to afford the desire (2S,3S)-3-[[(2Z)-2-(2-aminothiazol-4-yl)-2-[2-(diethylamino)-l,l-dimethyl-2-oxo-ethoxy]imino-acetyl] amino] -2-methyl-4-oxo-azetidine- 1 -sulfonic acid (AZ5-2) (20 nig, 5.9% Yield), as an off white solid.

[1262]

[1263] NMR (DMSO-d6) 5: 9.32 (d, J=8.2 Hz, 1H), 6.82 (s, 1 H), 4.49 (dd, J=2.7, 8.1 Hz, 1H), 3.69 (qd, J=2.6, 6.1 Hz, 2H), 3.50-3.56 (m, 2H), 3.23 (q,.1=6.9 Hz, 2H), 1.46 (d, J=2.6 Hz, 6H), 1.41 (d, J=6.2 Hz, 3H), 1.04 (t, J=6.8 Hz, 3H), 0.98 (t, J=6.8 Hz, 3H) ppm.

[1264] LCMS: (ES+) 96.33%, m / z = 491.01 [M+H] Column: CORTECS UPLC C18 (3*30) mm, 1.6um, Mobile Phase A: 0.05%FAin Water, Mobile Phase B: 0.05%FA in Acetonitrile]. HPLC: 95.03% purity [Method: Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1 % TFA in water, B - Acetonitrile]. Attorney Docket No. 10063-110WO1

[1265] Synthesis of ( 2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-( ((!-( diisobutylamino )-2-methyl-l-oxopropan-2-yl)oxy)imino)acetamido)-2-methyl-4-oxoazetidine,-l-sulfonic acid

[1266] 2 (0.96 eq), EDCI(1.5 eq), H0Bt(1.5 eq), DMF(5 mL), DIPEA (3 eq),0°C-RT, 3h N

[1267] O=S~OH O=S-OH o o

[1268]

[1269] AZ6-2 To a stirred solution of 2-((((Z)-l-(2-aminothiazol-4-yl)-2-(((2S,3S)-2-methyl-4- oxo-l-sulfoazetidin-3-yl)aniino)-2-oxoethylidene)amino)oxy)-2-metliylpropanoic acid [Aztreonam] 1 (1 g, 2.29 mmol, 1 eq) in DMF (5 nil) were added EDO. HC1 (661 mg, 3.43 mmol, 1.5 eq), HOBt (475 mg, 3.44 mmol, 1.5 eq) at 0°C stirred for 5 min after DIPEA (1.29 mL, 6.8 mmol, 3 eq) and diisobutylamine 2 (283.8 mg, 2.19 mmol, 0.95 eq) were added 0°C. The reaction mixture was allowed to stir at RT for 3h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated in vacuo to obtain crude compound (1.4 g). The crude was purified by prep-HPLC to afford (2S,3S)-3-((Z)-2-(2-aminothiazol-4-yl)-2-(((l-(diisobutylamino)-2-methyl-l-oxopropan-2-yl)oxy)imino)acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid (AZ6-2) (222 mg, 17.68% Yield), as an off white solid.

[1270] JH NMR (DMSO-de) 8: 9.32 (br d, 1=8.0 Hz, 1H), 6.82 (s, 1H), 4.49 (dd, 1=2.6, 8.1 Hz, 1H), 3.70 (qd, 1=2.7, 6.1 Hz, 1H), 3.34 (d, 1=7.6 Hz, 2H), 3.13 (qd, 1=8.1, 13.1, Hz, 2H), 1.84-1.91 (m, 2H), 1.49 (d, 1=6.1 Hz, 6H), 1.41 (d, 1=6.1 Hz, 3H), 0.75-0.83 (m, 12H) ppm. LCMS: (ES+) 95.41%, m / z = 547.09 [M+H] [Method: Column: CORTECS UPLC C18 (3*30) mm, 1.6um, Mobile Phase A: 0.05%FA in Water, Mobile Phase B: 0.05%FAin Acetonitrile]. HPLC: 95.04% Purity [Method: Column: Bakerbond Q2100 C18, 1.8 pm, 100 X 2.1 mm, Mobile Phase A: 0.05% FA in Water, Mobile Phase B: 0.05% FA in

[1271] Acetonitrile]. Attorney Docket No. 10063-110WO1

[1272] Synthesis of (2S,3S)-3-((Z)-2-(2-aminothiaz.ol-4~yl)-2-(((l-(benzylamino)-2-methyl- l-oxopropan-2-yl)oxy)imino)acetamido)-2-methyl-4-oxoazetidine-l -sulfonic acid

[1273] N

[1274] N 2 (1 eq), EDCI (1.2 eq),

[1275] H0Bt(1.5 eq), DMF(1 mL), DIPEA(3 eq),0°C-RT, 3h N N

[1276] O^S-OH O”S~OH 6 6

[1277]

[1278] AZ7-2 To a stirred solution of 2-((((Z)-1-(2-aminothiazol-4-yl)-2-(((2S,3S)-2-methyl-4-oxo-1-sulfoazetidin-3-yl)amino)-2-oxoethylidene)amino)oxy)-2-methylpropanoic acid [Aztreonam] 1 (100 mg, 0.22 mmol, 1 eq) in DMF (1 mL) were added EDCI. HCl (53.1 mg,0.33 mmol, 1.2 eq), HOBt (46.3 mg, 0.33 mmol, 1.46 eq) at 0°C Stirred for 5 min after DIPEA (0.12 mL, 0.69 mmol, 3 eq) and phenylmethanamine 2 (24.5 mg, 0.22 mmol, 1 eq) were added at 0°C. The reaction mixture was allowed to stir at RT for 3h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated in vacuo to obtain crude compound (200 nig). The crude was purified by prep- HPLC to afford (2S,3S)-3-[[(2Z)-2-(2-aminothiazol-4-yl)-2-[2-(benzylamino)-l,l-dimethyl-2-oxo-ethoxy]imino-acetyl]amino]-2-methyl-4-oxo-azetidine-l-sulfonic acid (AZ7-2) (30 mg, 24.90%) as an off white solid.

[1279] !H NMR (DMSO-d6) 5: 9.44 (d, J=8.1 Hz, 1H), 7.90 (t, 1=6.3 Hz, 1H), 7.27-7.63 (m, III), 7.16-7.25 (m, 5H), 6.90 (s, HI), 4.53 (dd, 1=2.6, 8.1 Hz, 1H), 4.38-4.45 (m, 1H), 4.28-4.35 (m, 1H), 3.70 (qd, 1=2.6, 6.1 Hz, 1H), 1.45 (d, 1=5.6 Hz, 6H), 1.41 (d, 1=6.1 Hz, 3H) ppm. LCMS: (ES+) 99.45% m / z = 525 [M+H] [method: Column: CORTECS UPLC C18 (3*30) mm, 1.6um, Mobile Phase A: 0.05%TFA in Water, Mobile Phase B: 0.05%TFA in Acetonitrile]. HPLC: 98.36% Purity [Method: Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A - 0.1% Formic acid in water: Acetonitrile (95:05), B - Acetonitrile], Attorney Docket No. 10063-110WO1

[1280] Synthesis of ( 2S, 3S)-3-[ [ (2Z)-2-( 2-aminothiazol-4-yl)-2-(2-benzyloxy-f 1 -dimethyl- 2-oxo-ethoxy) imino-acetyl] amino] -2-methyl-4-oxo-azetidine-l- sulfonic acid

[1281] 2 (1 eq), DiPC(1.44 eq), HOBt(1.37 eq), DMF(3 mL), DIPEA(2.44 eq),0°C-RT, 3h

[1282] N

[1283] O=S-OH 6

[1284]

[1285] AZ8-2 To a stirred solution of 2-((((Z)-l-(2-aminothiazol-4-yl)-2-(((2S,3S)-2-methyl-4- oxo-l-sulfoazetidin-3-yl)amino)-2-oxoethylidene)amino)oxy)-2-methylpropanoic acid [Aztreonam] 1 (300 mg, 0.68 mmol, 1 eq) in DMF (1.5 ml) were added N, N’-Diisopropylcarbodiimide (128 mg, 0.99 mmol, 1.44 eq), HOBt (130 mg, 0.93 mmol, 1.36 eq) at 0°C stirred for 5 mins after DIPEA (0.3 ml, 2 mmol, 2.44 eq) and phenylmethanol 2 (75 mg, 0.69 mmol, 1 eq) were added at 0°C. The reaction mixture was allowed to stir at RT for 3h. Progress of the reaction was monitored by TLC and LCMS. After completion, the reaction mixture was concentrated in vacuo to obtain crude compound (400 mg). The crude was purified by prep-HPLC to afford (2S,3S)-3-[[(2Z)-2-(2-aminothiazol-4-yl)-2-(2-benzyloxy-1,1-dimethyl-2-oxo-ethoxy)imino-acetyl]amino]-2-methyl-4-oxo-azetidine-1-sulfonic acid (AZ8-2), 45 nig, 12.43%) as an off white solid.

[1286] !H NMR (DMSO-d6) 5: 9.20 (d, J=8.1 Hz, 1H), 7.26-7.39 (m, 5H), 6.78 (s, 1H), 5.18 (s, 2H), 4.46 (dd, J=2.6, 8.1 Hz, 1H), 3.69 (dd, J=2.5, 6.2 Hz, 2H), 1.48 (d, J=5.4 Hz, 6H), 1.40 (d, 1=6.1 Hz, 3H) ppm. LCMS: (ES+) 98.14%, m / z = 525.96 [M+H] [Method:

[1287] Column: CORTECS UPLC C18 (3*30) mm, 1.6um, Mobile Phase A: 0.05 %FA in Water, Mobile Phase B: 0.05 %FA in Acetonitrile], HPLC: 99.55% purity [Column: X-Select CSH C18 (4.6*150) mm 5u, Mobile Phase: A- 0.1% TEA in water, B - Acetonitrile].

[1288] Example 9: Additional DIS analog synthesis

[1289] Synthesis of Heptyl 3-azabicyclo[3.1.0]hexane-3-carbo(dithioperoxo)thioate:

[1290] CS2(1.2 eq), Et3N (1.2 eq), CBr4(2.0 eq), DCM (10 mL), 0 °C,rt, 1 h IL S NH. HCI - »-

[1291]

[1292] A B LHM-01 Attorney Docket No. 10063-110WO1

[1293] Stirred solution of 3-azabicyclo[3.1.0]hexane (B) (63 nig, 0.76 mmol, 1.0 eq) in anhydrous DCM (8 ml,) was cooled to 0 °C. Then carbon disulfide (69 mg, 0.91 mmol, 1.2 eq) and triethylamine (0.1 mL, 0.91 mmol, 1.2 eq) were added to the same temperature and stirred for 5 min. After that the solution of carbon tetrabromide (504 mg, 1.52 mmol, 2.0 eq) in DCM (2 mL) and heptane- 1 -thiol (A) (100 mg, 0.76 mmol, 1.0 eq) were added, and the resulting reaction mixture was allowed to stir at room temperature for 1 h. After completion of the reaction (reaction was monitored by TLC), the reaction mixture was diluted with H₂O (5 mL) and extracted with DCM (2 x 10 mL). The organic layers were dried over sodium sulphate and concentrated to get crude. The crude was purified by 230 - 400 mesh silica gel column on combi flash chromatography, eluted using 0 - 10% of Ethyl Acetate in Hexane to afford Heptyl 3-azabicyclo[3.1.0]hexane-3-carbo(dithioperoxo)thioate (LHM-01) (100 mg, 21%) as pale yellow liquid.

[1294] TLC: 10% EtOAc in Hexane Rf. 0.5); ¹HNMR (400 MHz, DMSO-d6) 34.10 (d,.7 = 13.20 Hz, 1H), 3.96 (d, 7 = 11.20 Hz, 1H), 3.86 - 3.77 (m, 2H), 2.80 (t, J = 7.20 Hz, 2H), 1.83 - 1.80 (m, 1H), 1.72 - 1.71 (m, 1H), 1.57 (quin, J = 7.60 Hz, 2H), 1.34 - 1.24 (m, 8H), 0.86 (q, 7 = 5.60 Hz, 4H), 0.17 (m, 1H); LCMS: (ES+) 99.05%, m / z = 290.2 [M+H]* [Method:- Flow rate: 0.8 ml / min, T / %B: 0.01 / 40,0.40 / 40,2 / 95,6 / 95,7 / 40,8 / 40, Buffer: A-5Mm NH4OAC, B-100% ACN, Column: Kinetex C18 50*4.6*5.0 Microns, Column Temp: Ambient]; HPLC: 97.76% purity [Method: Column: X-TERRA 250*4.6mm*5tun, Mobile phase A: lOmM AA IN AQ, Mobile phase B: 100% Acetonitrile].

[1295] Synthesis of Heptyl 6, 6-difluoro-3 -azabicyclo[3.1.0 ]hexane-3 - carbo( dithioperoxo )thioate:

[1296] CS2(1.2 eq), Et3N (1.2 eq), CBr4(2.0 eq) s, DCM (10 mL), 0 °C, rt, 1 h ]jcHS.NH_.. Z^N \ -X-l - / . HCI X / F

[1297]

[1298] A BFLHM-02 Stirred solution of 6,6-difluoro-3-azabicyclo[3.1.0]hexane (B) (129 mg, 0.76 mmol, 1.0 eq) in anhydrous DCM (8 mL) was cooled to 0 °C. Then carbon disulfide (69 mg, 0.91 mmol, 1.2 eq) and triethylamine (0.13 mL, 0.91 mmol, 1.2 eq) were added to the same temperature and stirred for 5 min. After that solution of carbon tetrabromide (504 mg, 1.52 mmol, 2.0 eq) in DCM (2 mL) and heptane- 1 -thiol (A) (100 mg, 0.76 mmol, 1.0 eq) were added, and the resulting reaction mixture was allowed to stir at room temperature for 1 h. After completion of the reaction (reaction was monitored by TLC), the reaction mixture was diluted with H2O (5 mL) and extracted with DCM (2 x 10 ml,). The organic layers were Attorney Docket No. 10063-110WO1

[1299] dried over sodium sulphate and concentrated to get crude. The crude was purified by 230 - 400 mesh silica gel column on combi flash chromatography, eluted using 0 - 10% of Ethyl Acetate in Hexane to afford Heptyl 6,6-difluoro-3-azabicyclo [3.1.0] hexane-3-carbo(dithioperoxo)thioate (LHM-02) (60 mg, 1%) as pale yellow liquid.

[1300] TLC: 10% EtOAc in Hexane (Ry 0.5);jHNMR (400MHz, DMSO-d6) d 4.27 - 4.07 (m, 3H), 2.87 - 2.70 (m, 4H), 1.57 (quint, J = 7.20 Hz, 2H), 1.34 - 1.24 (m, 8H), 0.86 (t, J = 7.20 Hz, 3H) ppm;19F NMR (400 MHz, DMSO-d6) <5 -130.71 (d, J = 168.80 Hz), -154.56 (d, J = 168.80 Hz) ppm; LCMS: (ES+) 99.43% m / z = 326.2 [M+H]f[Methode-Flow rate: 0.8 ml / min, T / %B: 0.01 / 5,0.1 / 5,3 / 95,5 / 95,5.10 / 5,6 / 5, Buffer: A-5Mm NH4OAC, B-100% ACN, Column: Kinetex C1850*4.6*5.0 Microns, Column Temp: 35°C]; HPLC: 98.47% purity [Method:- Column: KINETEX 250*4.6mm*5pm, Mobile phase A- lOmM A A IN AQ, Mobile phase- 100% Acetonitrile.]

[1301] Synthesis of Heptyl hexahydrocyclopentaf c]pyrrole-2(lH)-carbo(dithioperoxo)thioate:

[1302] CS2(1.0 eq), Et3N (4.0 eq), CBr4(2.0 eq) s, HCI DCM (10 mL), 0 °C, rt, 1 h II « HS,.^+ / _ - _ _ _.

[1303]

[1304] A ' rV B cy LHM-03 Stirred solution of Octahydrocyclopenta[c]pyrrole hydrochloride (B) (112 mg, 0.76 mmol, 1.0 eq) in anhydrous DCM (8 mL) was cooled to 0 °C. Then, carbon disulfide (0.04 mL, 0.76 mmol, 1.0 eq) and triethylamine (0.41 mL, 3.02 mmol, 4.0 eq) were added at the same temperature. After 5 min, a solution of carbon tetrabromide (501 mg, 1.51 mmol, 2.0 eq) in DCM (2 mL), followed by heptane- 1 -thiol (100 mg, 0.76 mmol, 1.0 eq) (A) were added. The resulting reaction mixture was allowed to stir at room temperature for 1 h. After completion of the reaction (reaction was monitored by TLC), the reaction mixture was diluted with H₂O (4 mL) and extracted with DCM (2 x 10 mL). The organic layers were dried over sodium sulphate and concentrated to get the crude compound. The crude was purified by 230 - 400 mesh silica gel column on combi flash chromatography, eluted using 0 - 10% of Ethyl Acetate in Hexane to afford Heptyl hexahydrocyclopenta[c]pyrrole-2(lH)-carbo(dithioperoxo)thioate (LHM-03) (48 mg, 20%) as pale yellow liquid.

[1305] TLC: 20% EtOAc in Hexane (R

[1306]

[1307] y 0.5); NMR (400 MHz, DMSO-d6) <54.05 (dd, J = 8.82, 13.60 Hz, 1H), 3.97 (dd, J= 8.82, 11.60 Hz, 1H), 3.64 (dd, J = 4.80, 13.60 Hz, HI), 3.52 (dd,. / = 4.80, 11.60 Hz, 1H), 2.87 - 2.79 (m, 3H), 2.73 - 2.68 (m, III), 1.82 - 1.70 (m, 3H), 1.60 - 1.54 (m, 3H), 1.51 - 1.45 (m, 2H), 1.34 - 1.25 (m, 8H), 0.86 (t, J= 7.20 Hz, Attorney Docket No. 10063-110WO1

[1308] 3H) ppm; LCMS: (ES+) 97.41%, m / z = 318.1 [M+H]+[Method:- Column: Kinetex C18 (4.6 X50 mm) 5pm. Flow rate: 0.8 ml / min. Mobile Phase A - 0.1% FA in Water, B - 100% ACN]; HPLC: 96.99% purity [Method:- Column: KINETEX, Mobile Phase: A- 0.1 % TFA in AQ, B- 100% Acetonitrile],

[1309] Synthesis of Heptyl 5, 5-difluorohexahydrocyclopenta[c jpyrrole-2( 1H)~ carbof dithioperoxo) thioate:

[1310] DAST (12.0 eq), DCM (5 mL) 4M HCI (10.0 v), DCM (5 ml) NH. HCI 0 °C, rt, 3 days 0 X, rt, 1 h Step-1 Step-2

[1311] F

[1312] 2 3

[1313] 4 (1.0 eq), CS2(1.0 eq), Et3N (4.0 eq) CBr4(2.0 eq), DCM (10 mL), 0 °C, rt, 1 h Step-3

[1314]

[1315] Step 1. Synthesis of Tert-butyl 5,5-difluorohexahydrocyclopenta[c]pyrrole- 2(lH)-carboxylate: Solution of zert-butyl 5-oxohexahydrocyclopenta[c]pyrrole-2(lH)- carboxylate (1) (150 mg, 0.66 mmol, 1.0 eq) in anhydrous DCM (5 mL) was cooled to 0 “C. Then Diethylaminosulfur trifluoride (1.06 mL, 7.99 mmol, 12.0 eq) was added dropwise at the same temperature and stirred for 5 min. The resulting reaction mixture was allowed to stir at room temperature for 3 days. After completion of the reaction (monitored by crude!H NMR), the reaction mixture was quenched with saturated ammonium chloride solution (4 ml,) and extracted with DCM (2 x 10 L). The organic layers were dried over sodium sulphate and concentrated to afford crude Tert-butyl 5,5 -Difluorohexahydro cyclopenta[c]pyrrole-2(TH)-carboxylate (2) (130 mg, 79%) as a pale yellow liquid;1H NMR (400 MHz, DMSO-d6) S 3.66 - 3.49 (m, 2H), 3.28 (brs, 2H), 2.92 - 2.80 (m, 2H), 2.39 - 2.31 (m, 2H), 2.06 - 1.93 (m, 2H), 1.46 (s, 9H), 1.43 - 1.41 (m, III), 1.30 - 1.21 (m, 3H) ppm.

[1316] Step 2. Synthesis of 5,5-Difluorooctahydrocyclopenta[c]pyrrole hydrochloride: Solution of terr-butyl 5,5-difluorohexahydrocyclopenta[c]pyrrole-2(lH)-carboxylate (2) (130 mg, 0.53 mmol, 1.0 eq) in anhydrous DCM (5 mL) was cooled to 0 C. Then, 4 M hydrochloric acid solution in 1,4-dioxane (1.3 mL, lO. Ov) was added dropwise at the same temperature and stirred for 5 minutes. The resulting reaction mixture was allowed to stir at room temperature and stirred for 30 minutes. After completion of the reaction (reaction was monitored by TLC), the reaction mixture was concentrated to get crude 5,5- Attorney Docket No. 10063-110WO1

[1317] Difluorooctahydrocyclopenta[c]pyrrole hydrochloride (3) (80 nig, 83%) as a pale yellow liquid. Obtained crude is utilized for the next step without further purification. TLC: 20% EtOAc in Hexane (Rf. 0.5).

[1318] Step 3. Synthesis of 5,5-Difluorohexahydrocyclopenta[c]pyrrole-2(lH)carbo(dithioperoxo) thioate: Stirred solution of 5,5-Difluorooctahydrocyclopenta[c]pyrrole (3) (80 mg, 0.54 mmol, 1.0 eq) in anhydrous DCM (8 mL) was cooled to 0 °C. Then carbon disulfide (0.03 mL, 0.76 mmol, 1.0 eq) and triethylamine (0.30 mL, 2.18 mmol, 4.0 eq) were added at the same temperature. After 5 min, a solution of carbon tetrabromide (361 mg, 1.08 mmol, 2.0 eq) in DCM (2 mL) followed by heptane- 1 -thiol (72 mg, 0.54 mmol, 1.0 eq) (4) were added. The resulting reaction mixture was allowed to stir at room temperature for 1 h. After completion of reaction (reaction was monitored by TLC), the reaction mixture was diluted with H2O (4 mL) and extracted with DCM (2 x 10 mL). The organic layers were dried over sodium sulphate and concentrated to get crude compound. The crude was purified by 230 - 400 mesh silica gel column on combi flash chromatography, eluted using 0 - 10% of Ethyl Acetate in Hexane to afford Heptyl 5,5-difluorohexahydrocyclopenta[c] pyrrole-2(lH)-carbo(dithioperoxo)thioate (LHM-04) (45 mg, 29%) as pale yellow liquid.

[1319] TLC: 10% EtOAc in Hexane (R

[1320]

[1321] f. 0.5); NMR (400 MHz, DMSO-d6) <54.13 -4.00 (m, 2H), 3.78 (dd, J = 4.40, 13.80 Hz, 1H), 3.69 (dd, J = 5.20, 11.40 Hz, 1H), 3.10 - 3.03 (m, III), 2.97 - 2.89 (m, 1 II), 2.82 (t, J = 7.20 Hz, 2H), 2.45 - 2.33 (m, 2H), 2.20 -2.07 (m, 2H), 1.59 (quint, J = 7.20 Hz, 2H), 1.35 - 1.25 (m, 8H), 0.86 (t, J = 6.00 Hz, 3H) ppm;i9F NMR (400 MHz, DMSO-d6) <5 -87.70 (d, J = 242.40 Hz), -89.55 (d, J = 242.40 Hz) ppm; LCMS: (ES+) 96.55%, m / z= 354.30 [M+H]+[Method:- Kinetex Cl 850*4.6*5.0 Microns. Flow rate: 0.8 ml / min. Mobile Phase A: 5 mM NH₄OAc, B: 100% Acetonitrile]; HPLC: 95.81% purity [Method:- Column: X-TERRA 250*4.6mm*5 pm, Mobile Phase: A - lOmM A A IN Aq, B - 100% Acetonitrile],

[1322] Synthesis of Heptyl 3,4-difluoropyrrolidine-l-carbo(dithioperoxo)thioate:

[1323] s p CS2(1.0 eq), Et3N (4.0 eq), CBr4(2.0 eq) oHc +. I NH.uHrC., I - DCM ( - -10 mL), - - 0 °C, -- r-t-,- - 1 h ->■cF — ( J F' y* F

[1324]

[1325] A B LHM-05 To a stirred solution of 3,4-difluoropyrrolidine hydrochloride (B) (109 mg, 0.76 mmol, 1.0 eq) in anhydrous DCM (8 mL) was cooled to 0 °C. Then carbon disulfide (56 mg, 0.76 mmol, 1.0 eq) and triethylamine (0.42 mL, 3.02 mmol, 4.0 eq) were added to the Attorney Docket No. 10063-110WO1

[1326] same temperature and stirred for 5 min. After solution of carbon tetrabromide (501 mg, 1.51 mmol, 2.0 eq) in DCM (2 mL) and heptane- 1 -thiol (100 mg, 0.76 mmol, 1.0 eq) (A) were added and the resulting reaction mixture was allowed to stir at room temperature for 1 h. After completion of reaction (reaction was monitored by TLC), the reaction mixture was diluted with H2O (4 mL) and extracted with DCM (2 x 10 mL). The organic layers were dried over sodium sulphate and concentrated to get crude. The crude was purified by 230 -400 mesh silica gel column on combi flash chromatography, eluted using 0 - 10% of Ethyl Acetate in Hexane to afford Heptyl 3,4-difluoropyrrolidine-l-carbo(dithioperoxo)thioate (LHM-05) (104 mg, 44%) as pale yellow liquid.

[1327] TLC: 10% EtOAc in Hexane (R

[1328]

[1329] f. 0.5); NMR (400 MHz, CDCI3) S 5.30 - 5.28 (m, 1H), 5.17 - 5.14 (m, 1H), 4.34 - 4.10 (m, 3H), 4.04 - 3.92 (m, 1H), 2.85 (t, J = 7.20 Hz, 2H), 1.66 (quint, J = 7.60 Hz, 2H), 1.43 - 1.28 (m, 8H), 0.88 (t, J = 5.20 Hz, 3H) ppm;i9F NMR (400 MHz, CDCI3) d -204.06 (d, J = 7.20 Hz) - 205.11 (d, J = 7.20 Hz) ppm;

[1330] LCMS: (ES+) 98.15%, m / z = 14.3 [M + H]+[Method:- Flow rate: 0.8 ml / min, T / %B:

[1331] 0.01 / 40,0.40 / 40,2 / 95,6 / 95,7 / 40, 8 / 40, Buffer: A-5Mm NH4OAC, B-100% ACN, Column: Kinetex C18 50*4.6*5.0 Microns, Column Temp: Ambient]; HPLC: 97.66% purity [Method:- Column: KINETEX 250*4.6mm*5pm, Mobile Phase- A: lOmM AA IN Aq, B: ACN],

[1332] Synthesis of Heptyl isoindoline-2-carbo(dithioperoxo)thioate:

[1333] . HCI CS2(1.20 eq), Et3N (1.20 eq), CBr4(2.0 eq) n DCM (10 mL), 0 °C, rt, 1 h

[1334]

[1335] A B LHM-06 To a stirred solution of Isoindoline (B) (180 mg, 1.13 mmol, 1.0 eq) in anhydrous DCM (8 mL) was cooled to 0 °C. Then carbon disulfide (104 mg, 1.36 mmol, 1.20 eq) and triethylamine (0.17 mL, 1.36 mmol, 1.20 eq) were added to the same temperature and stirred for 5 min. After that, the solution of carbon tetrabromide (752 mg, 2.26 mmol, 2.0 eq) in DCM (2 mL) and heptane- 1 -thiol (A) (150 mg, 1.13 mmol, 1.0 eq) were added, and the resulting reaction mixture was allowed to stir at room temperature for 1 h. After completion of the reaction (reaction was monitored by TLC), the reaction mixture was diluted with H2O (4 mL) and extracted with DCM (2 x 10 mL). The resultant organic layers were dried over sodium sulphate and concentrated to get crude. 'The crude was purified by 230 - 400 mesh silica gel column on combi flash chromatography, eluted using 0 - 10% of Attorney Docket No. 10063-110WO1

[1336] Ethyl Acetate in Hexane followed by prep HPLC purification to afford Heptyl isoindoline-2-carbo(dithioperoxo)thioate (LHM-06) (20 mg, 5%) as pale yellow solid.

[1337] TLC: 50% DCM in Hexane (Rf. 0.5); *11 NMR (400 MHz, DMSO-d6) § 7.50 -7.30 (m, 4H), 5.12 (s, 4H), 2.86 (t, J = 7.20 Hz, 2H), 1.62 (quint, 7= 6.80 Hz, 2H), 1.40 - 1.24 (m, 8H), 0.85 (t, J = 5.20 Hz, 3H) ppm; LCMS: (ES+) 98.46%, m / z = 326.3 [M + H]* [Method:- Flow rate: 0.8 ml / min, T / %B: 0.01 / 40,0.40 / 40,2 / 95,6 / 95,7 / 40,8 / 40, Buffer: A- 5Mm NH₄OAc, B-100% ACN, Column: Kinetex C18 50*4.6*5.0 Microns, Column Temp: Ambient]; HPLC: 98.20% purity [Method:- Column: X-TERRA 250*4.6mm*5um, Mobile phase A: 10 mM AA IN Aq, Mobile phase B: 100% Acetonitrile].

[1338] Synthesis of Heptyl 5,7-dihydro-6H-pyrrolo[3,4-b]pyridine-6- carbo( dithioperoxo )thioate CS2(1,0 eq), Et3N (1.2 eq), NSS (1.5 eq) o V—,„ci

[1339]

[1340] A B N LHM-07 To a stirred solution of 6,7-dihydro-5H-pyrrolo[3,4-b]pyridine (B) (142 mg, 0.76 mmol, 1.0 eq) in anhydrous DCM (8 mL), carbon disulfide (0.05 mL, 0.91 mmol, 1.2 eq), triethylamine (0.13 mL, 0.91 mmol, 1.2 eq) and heptane- 1 -thiol (A) (100 mg, 0.76 mmol, 1.0 eq) were added at room temperature. After that reaction mixture was cooled to -45 °C, and a solution of N-Iodosuccinimide (255 mg, 1.14 mmol, 1.5 eq) in DCM (2 mL) was added. The resulting reaction mixture was allowed to stir at room temperature for 1 h. After completion of the reaction (reaction was monitored by TLC), the reaction mixture was diluted with H₂O (5 mL) and extracted with DCM (2 x 10 mL). The organic layers were dried over sodium sulphate and concentrated to get crude. The crude was purified by 230-400 mesh silica gel column on combi flash chromatography, eluted using 0 - 10% of Ethyl Acetate in Hexane followed by prep HPLC purification to afford Heptyl 5,7-dihydro-6H-pyrrolo[3,4-b]pyridine-6-carbo(dithioperoxo)thioate (LHM-07) (70 mg, 17%) as pale yellow solid.

[1341] TLC: 20% EtOAc in Hexane Rf. 0.5); *HNMR (400 MHz, DMSO-d6) d 8.52 (d, J = 4.00 Hz, 1H), 7.88 (dd, 7.60, 8.00 Hz, 1H), 7.40 (dd, J = 5.20, 8.00 Hz, 1H), 5.1 (m, 4H), 2.87 (t, 7 = 7.20 Hz, 2H), 1.63 (quint, J = 7.20 Hz, 2H), 1.38 - 1.26 (m, 8H), 0.86 (t, J - 7.20 Hz, 3H) ppm; LCMS: (ES+) 98.25%, m / z = 327.1 [M+H]+[Method:- Flow rate: 0.8 ml / min, Gradient: T / % B: 0.01 / 5, 4 / 90, 6 / 90, 7 / 5, 8 / 5, Buffer: A: 0.1% FA in Water, B:

[1342] 100% ACN, Column: Kinetex C18 (4.6 X50 mm) 5pm, Column Temp: Ambient temp]; Attorney Docket No. 10063-110WO1

[1343] HPLC: 99.65% purity [Method:- Column: GEMINI NX 150*4.6MM*5MM, Mobile phase A: 0.1% TEA IN Aq, Mobile phase B: 100% Acetonitrile],

[1344] Synthesis of Heptyl benzyl(methyl )carbam.o( di.thioperoxo)thioate:

[1345] CS2(1.2 eq), Et3N (1.2 eq), CBr4(2.0 eq) 'NH DCM (10 mL), 0 °C, rt, 1 h A.3 N" S' I

[1346]

[1347] LHM-08 Stirred solution of N-methyl-1-phenylmethanamine (B) (500 mg, 3.78 mmol, 1.0 eq) in anhydrous DCM (8 mL) was cooled to 0 °C. Then carbon disulfide (345 mg, 4.54 mmol, 1.2 eq) and triethylamine (0.63 mL, 4.54 mmol, 1.2 eq) were added to the same temperature and stirred for 5 min. After that, the solution of carbon tetrabromide (2.5 g, 7.56 mmol, 2.0 eq) in DCM (2 mL) and heptane- 1 -thiol (A) (500 mg, 3.78 mmol, 1.0 eq) were added, and the resulting reaction mixture was allowed to stir at room temperature for 1 h. After completion of reaction (reaction was monitored by TLC), the reaction mixture was diluted with H₂O (15 mL) and extracted with DCM (3 x 15 mL). The organic layers were dried over sodium sulphate and concentrated to get crude. The crude was purified by 230 - 400 mesh silica gel column on combi flash chromatography, eluted using 0 - 10% of Ethyl Acetate in Hexane to afford Heptyl benzyl(methyl)carbamo(dithioperoxo)thioate (LHM- 08) (80 mg, 58%) as pale yellow liquid.

[1348] TLC: 20% EtOAc in Hexane (R 0.5);JHNMR (400MHz, DMSO-d6) b' 7.48 - 7.15 (m, 5H), 5.35 (s, 1H), 5.15 (s, 1H), 3.47 (s, 1H), 3.35 (s, 2H), 2.90 - 2.75 (m, 2H), 1.61 - 1.52 (m, 2H), 1.34 - 1.25 (m, 8H), 0.85 (d, J = 6.80 Hz, 3H) ppm: LCMS: (ES+) 96.17%, m / z = 328.3 [M+H]+[Method:- Flow rate: 0.8 ml / min, T / %B:

[1349] 0.01 / 5,0.1 / 5,3 / 95,5 / 95,5.10 / 5,6 / 5, Buffer: A-5Mm NH4OAC, B-100% ACN, Column:

[1350] Kinetex C I 8 50*4.6*5.0 Microns, Column Temp: 35°C]; HPLC: 97.79% purity [Method:- Column: X-TERRA 250*4.6mm*5pm, Mobile phase A: lOmM AA IN AQ, Mobile phase B:100% Acetonitrile],

[1351] Synthesis of 6-methylheptyl pyrrolidine- 1 -carbo( dithioperoxo )thioate:

[1352] 3ONH(3.0 eq)EtjN p Q eq( (1 2 eq)(°-3 eq)' °MF t2°mL>'rt’16 hI CBr4(1.5 eq), DCM (20 mL) J I Step-1 Step-2XJ

[1353]

[1354] 1 2 LHM-9 Step 1. Synthesis of 6-methylheptane-l -thiol (2): Stirred solution of l-bromo-6- methylheptane (1) (1.00 g, 5.18 mmol, 1.0 eq) in DMF (20 mL) was cooled to 0 °C. Then Attorney Docket No. 10063-110WO1

[1355] tetrabutylammonium bromide (501 mg, 1.55 mmol, 0.3 eq) and sodium hydrogensulfide (871 mg, 15.50 mmol, 3.0 eq) were added to the same temperature. The resultant reaction mixture was stirred to room temperature for 16 h. After completion of the reaction (reaction was monitored by TLC), the reaction mixture was diluted with H2O, and the product was extracted in EtOAc (3 x 60 mL). The combined organic layers were dried over anhydrous Na₂SO₄ and concentrated under reduced pressure to obtain 6-methylheptane-l -thiol as crude (2) (0.92 g) as a yellow liquid. TLC: 100% Hexane (Rf. 0.8).

[1356] Step 2. Synthesis of 6-methylheptyl pyrrolidine-l-carbo(dithioperoxo)thioate:

[1357] Stirred solution of pyrrolidine (3) (1.17 g, 16.40 mmol, 1.0 eq) in DCM (18 mL) was cooled to 0 °C. Then triethylamine (6.86 mL, 49.20 mmol, 3.0 eq) and carbon disulfide (974 pL, 19.70 mmol, 1.2 eq) were added dropwise to the same temperature, and stirred for 30 min.

[1358] After that carbon tetrabromide (8.16 g, 24.60 mmol, 1.5 eq) in DCM (2 ml,) and 6-methyl- 1 -heptanethiol (2) (2.40 g, 16.40 mmol, 1.0 eq) were added to the reaction mixture and allowed to stir to room temperature for 16 h. After completion of the reaction (reaction progress was monitored by TLC), the reaction mixture was diluted with H2O (40 mL) and the product was extracted in DCM (2 x 60 mL). The organic layers were dried over anhydrous Na2SO4 and concentrated under reduced pressure to obtain the crude product.

[1359] Obtained crude was purified by column chromatography followed by preparative TLC to afford 6-methylheptylthio-1-pyrrolidinecarbodithioate (LHM-09) (14.0 mg, 0.29%) as a light-yellow oil.

[1360] TLC: 10% EtOAc in Hexane Rf. 0.8).2H-NMR (400 MHz, CDCh): d 3.96 (t, J = 6.80 Hz, 2H), 3.74 (t, J = 6.80 Hz, 2H), 2.85 (t, J = 7.40 Hz, 2H), 2.14 - 1.97 (m, 4H), 1.68 - 1.63 (m, 2H), 1.54 - 1.48 (m, 1H), 1.42 - 1.12 (m, 611), 0.86 (d, J = 6.40 Hz 6H) ppm;

[1361] LCMS: (ES+) 94.95%, m / z = 292.3 [M+HJ+ [Method:- Flow rate: 0.8 ml / min, T / %B:

[1362] 0.01 / 5,0.1 / 5,3 / 95,5 / 95,5.10 / 5,6 / 5, Buffer: A-5Mm NH4OAC, B-100% ACN, Column:

[1363] Kinetex Cl 8 50*4.6*5.0 Microns, Column Temp: 35 °C], HPLC: 94.38% purity [Method:

[1364] Column: KINETEX 250*4.6mm*5|im, Mobile Phase- A: lOmM AA IN Aq, B: ACN], Synthesis of 4-fluorophenethyl pyrrolidine-1 -carbo(dithioperoxo)thioate:

[1365] " 'XSH “

[1366]

[1367] Step 1. Synthesis of 4-fluorophenethyl carbamimidothioate hydrobromide (3): l-(2-bromoethyl)-4-fluorobenzene (1g, 4.95 mmol, 1.0 eq) and thiourea (376 mg, 4.95 mmol, 1.0 eq) were dissolved in ethanol (15 mL). The resultant reaction mixture was Attorney Docket No. 10063-110WO1

[1368] refluxed for 6 h. After completion of the reaction (reaction was monitored by TLC), solvent was evaporated to get the crude compound 4-fluorophenethyl carbamimidothioate hydrobromide (3) (1.3 g) yellow gummy liquid. The obtained crude compound was used to next without further purification & characterization. TLC: 10% MeOH in DCM (Ry 0.1).

[1369] Step 2. Synthesis of 2-(4-fluorophenyl)ethane-l-thiol (4): 4-fluorophenethyl carbamimidothioate hydrobromide (3) (1.3 g, 4.68 mmol, 1.0 eq) and sodium hydroxide (281 mg, 7.02 mmol, 1.5 eq) were dissolved in water (30 mL) and heated to 100 °C for 6h. After completion of the reaction (reaction was monitored by TLC), the reaction mixture was cooled to room temperature and extracted with EtOAc (30 mL, X 2). Combined organic layer was dried over anhydrous Na₂SO₄ and concentrated under reduced pressure to get 2-(4-fluorophenyl)ethane-l -thiol (4) (400 mg) off white solid. The obtained crude compound was used to next without further purification & characterization. TLC: 10% MeOH in DCM (Rf. 0.1).

[1370] Step 3. 4-fluorophenethyl pyrrolidine-l-carbo(dithioperoxo)thioate: Stirred solution of pyrrolidine (5) (0.182 g, 2.56 mmol, 1.0 eq) in dichloromethane (8 mL) was cooled to 0 °C. Then triethylamine (1.02 mL, 7.68 mmol, 3.0 eq) and carbon disulfide (186 pL, 3.07 mmol, 1.2 eq) were added dropwise to the same temperature, and stirred for 30 min. After that carbon tetrabromide (1.27 g, 3.84 mmol, 1.5 eq) in DCM (2 mL) and 2-(4- fluorophenyl)ethane-l -thiol (4) (400 mg, 2.56 mmol, 1 eq) were added to the reaction mixture and allowed to stir to room temperature for 16 h. After completion of the reaction (reaction progress was monitored by TLC), the reaction mixture was diluted with H2O (40 mL) and the product was extracted in DCM (2 x 60 mL). The organic layers were dried over anhydrous Na2SO4 and concentrated under reduced pressure to obtain the crude product. Obtained crude was purified by column chromatography followed by prep HPLC purification to afford 4-fluorophenethyl pyrrolidine- l-carbo(dithioperoxo)thioate (LHM-10) (25 mg, 3%) as alight-yellow' oil.

[1371] TLC: 10% EtOAc in Hexane (Rf. 0.8).JH-NMR (400 MHz, CDCh) <57.32 - 7.20 (m, 2H), 7.12 (t, J = 8.80 Hz, 2H), 3.80 (t, J = 7.20 Hz, 2H), 3.73 (t, 7.60 Hz, 2H), 3.12 - 3.03 (m, 2H), 2.92 (t, J = 7.60 Hz, 2H), 2.04 (quint, J = 7.60 Hz, 2H), 1.92 (quint, J = 7.60 Hz, 2H), (m, 4H), 1.68 - 1.63 (m, 2H), 1.54 - 1.48 (m, HI), 1.42 - 1.12 (m, 6H), 0.86 (d, J = 6.40 Hz, 6H) ppm; LCMS: (ES+) 98.35%, m / z = 302.2 [M+H]+ [Method:- Flow' rate: 0.8 ml / min, T / %B: 0.01 / 40,0.40 / 40,2 / 95,6 / 95,7 / 40,8 / 40, Buffer: A-5Mm NH4OAc, B-100% ACN, Column: Kinetex C1850*4.6*5.0 Microns, Column Temp: Ambient]; HPLC: Attorney Docket No. 10063-110WO1

[1372] 99.15% purity [Method: Column: KINETEX 250*4.6mm*5µm, Mobile Phase- A: lOmM A A IN Aq, B: ACNJ.

[1373] Synthetic scheme for cyclohexylmethy I pyrrolidine-1 -carbo(dithioperoxo)thioate:

[1374] NH 3 (1.0 eq) NaSH.xHjO (3.0 eq), TBAB (0.3 eq) EtjN (3.0 eq), CS2(1.2 eq), CBr4(1. S eq) 'Vx'xBr DMF (40 mL), 0 °C, rt, 16 h DCM (10 mL), 0 °C, rt, 1h Step-1 Step-2

[1375]

[1376] LHM-11 Step 1. Synthesis of cyclohexylmethanethiol: Stirred solution of (bromomethyl)cyclohexane (1) (2.0 g, 11.29 mmol, 1.0 eq) in DMF (40 mL) was cooled to 0 °C. Tetrabutylammonium bromide (501 mg, 3.39 mmol, 0.3 eq) was added to the reaction mixture at the same temperature. After five minutes, sodium hydrogensulfide (1.09 g, 16.93 mmol, 1.5 eq) were added and stirred to room temperature for 16 h. After completion of reaction (progress was monitored by TLC) quenched with water (30 mL) and extracted with EtOAc (3 x 60 mL). The combined organic layers dried over anhydrous Na₂SO₄ and concentrated under reduced pressure to obtain the crude. Crude was purified by 230 - 400 mesh silica gel column on combi flash chromatography, eluted using 0 - 2% of Ethyl Acetate in Hexane to afford 6-methyl-l -heptanethiol (2) (0.6 g, 79.2%) as light brown oil. TLC: 100% Hexane (Rf. 0.8).JH-NMR (400 MHz, CDCh): h 3.44 (d, J = 6.40 Hz, 2H), 1.78 - 1.62 (m, 5H), 1.48 - 1.42 (m, 1H), 1.33 - 1.10 (m, 3H), 0.98 - 0.86 (m, 2H)

[1377] Step 2. Synthesis of cyclohexylmethyl pyrrolidine-l-carbo(dithioperoxo)thioate: Solution of pyrrolidine (3) (0.2 g, 2.81 mmol, 1.0 eq) in DCM (8 mL) was cooled to 0 °C then added triethylamine (1.12 mL, 8.45 mmol, 3.0 eq) and carbon disulfide (167 pL, 3.38 mmol, 1.2 eq) were added. After that, carbon tetrabromide (1.4 g, 4.21 mmol, 1.5 eq) in DCM (2 mL) and cyclohexylmethanethiol (2) (0.55 g, 4.22 mmol, 1.5 eq) were added to the same temperature, and the reaction mixture was allowed to stir for 16 h at room temperature. After completion of the reaction (progress was monitored by TLC) reaction mixture was diluted with water (20 mL) and extracted with DCM (2 x 20 mL). The organic layers were dried over anhydrous Na2SO4 and concentrated under reduced pressure to obtain the crude. The obtained crude was purified by prep TLC to afford 6-methylheptylthio-1-pyrrolidinecarbodithioate (LHM-11) (12.2 mg, 10.48%) as light-yellow oil.

[1378] TLC: 10% EtOAc in Hexane (R

[1379]

[1380] f 0.5); NMR (400 MHz, CDCh) d 3.96 (t, J = 6.80 Hz, 2H), 3.73 (t, J = 6.80 Hz, 2H), 2.76 (d, J = 6.80 Hz, 2H), 2.16 - 2.05 (m, 2H), 2.03 Attorney Docket No. 10063-110WO1

[1381] - 1.95 (m, 2H), 1.88 (d, J = 12.40 Hz, 2H), 1.72 -- 1.56 (m, 4H), 1.29 -- 1.12 (m, 3H), 1.01 - 0.92 (m, 2H) ppm; LCMS: (ES+) 96.65%, m / z = 276.2 [M+HJ+ [Method:- Flow rate: 0.8 ml / min, T / %B: 0.01 / 5,0.1 / 5,3 / 95,5 / 95,5.10 / 5,6 / 5, Buffer: A-5Mm NH4OAc, B-100% ACN, Column: Kinetex C 1850*4.6*5.0 Microns, Column Temp: 35 °CJ; HPLC: 99.17% purity [Method: Column: KINETEX 250*4.6mm*5pm, Mobile Phase- A: lOmM AA IN Aq, B: ACN],

[1382] Synthesis of Heptyl thiomorpholine-4-carbo(dithioperoxo)thioate- 1,1 -dioxide:

[1383] CS2(1.2 eq), KOH (2.0 eq), NIS (1.2 eq) DCM: DMF (3:1)(3 mL:1 mL) / " NH. HCI _ -78 °C, rt, 1 h

[1384]

[1385] LHM-12 To a stirred solution of 2-azaspiro[3.3]heptane hydrochloride (B) (20 mg, 0.15 mmol, 1.0 eq) in anhydrous DCM and DMF (3:1 mL)(3 mL:l L) was cooled to -78 °C. Then carbon disulfide (14 nig, 0.18 mmol, 1.2 eq), potassium hydroxide (18 mg, 0.30 mmol, 2.0 eq), and heptane- 1 -thiol (A) (20 mg, 0.15 mmol, 1.0 eq) were added to the same temperature and stirred for 20 min. After that, the solution of A'-lodosuccinimide (41 mg, 0.18 mmol, 1.2 eq) was added, and the resulting reaction mixture was allowed to stir at room temperature for 1 h. After completion of the reaction (reaction was monitored by TLC), reaction mixture was diluted with cold H? O (4 mL) and extracted with DCM (2 x 10 mL). The organic layers were dried over sodium sulphate and concentrated to get crude.

[1386] The crude was purified by 230 - 400 mesh silica gel column on combi flash chromatography, eluted using 0 - 10% of Ethyl acetate in Hexane to afford Heptyl 2- azaspiro[3.3]heptane-2-carbo(dithioperoxo)thioate (LHM-12) (18 mg, 39%) as pale yellow liquid.

[1387] TLC: 10% EtOAc in Hexane (

[1388]

[1389] Rf. 0.5); NMR: (400 MHz, DMS()-d6) b' 4.36 (s, 2H), 4.19 (s, 2H), 2.81 (t, J = 7.20 Hz, 2H), 2.21 (t, J = 7.60 Hz, 4H), 1.79 (quint, J = 7.60 Hz, 2H), 1.58 (quint, J = 6.80 Hz, 2H), 1.34 - 1.25 (m, 8H), 0.87 (t,. / = 6.00 Hz, 3H) ppm; LCMS: (ES+) 97.62%, m / z = 304.2 [M + H]+[Method:- Flow rate: 0.8 ml / min, T / %B:

[1390] 0.01 / 40,0.40 / 40,2 / 95,6 / 95,7 / 40,8 / 40, Buffer: A-5Mm NH4OAC, B-100% ACN, Column: Kinetex Cl 8 50*4.6*5.0 Microns, Column Temp: Ambient]; HPLC: 98.71% purity [Method:- Column: X-TERRA 250*4.6mm*5pm, Mobile Phase- A: lOmM AA IN Aq, B: ACN], Attorney Docket No. 10063-110WO1

[1391] Synthesis of Heptyl 6, 6-difluoro-2 -azaspiro [3.3]heptane-2-carbo( dithioperoxo )thioate:

[1392] s P'NH TFAcs2<1 0 eci)>Et3N<22 e% CBr4(2.0 eq) HS Lj ' DCM flO mL), 0°C, rt, 1 h "s

[1393] F

[1394]

[1395] A B F LHM-13 To a stirred solution of 6,6-difluoro-2-azaspiro[3.3]heptane 2,2,2-trifluoroacetate (B) (225 mg, 0.91 mmol, 1.2 eq) and heptane- 1 -thiol (A) (100 nig, 0.76 mmol, 1.0 eq) in anhydrous DCM (8 mL), carbon disulfide (0.01 mL, 0.76 mmol, 1.0 eq) and triethylamine (0.23 mL, 1.67 mmol, 2.2 eq) were added dropwise at 0 °C. After 5 minutes, a solution of carbon tetrabromide (250 mg, 0.36 mmol, 1.5 eq) in DCM (2 mL) was added to the same temperature. The resulting reaction mixture was allowed to stir at room temperature for 1 h. After completion of the reaction (monitored by TLC), the reaction mixture was diluted with H2O (30 mL) and extracted with DCM (2 x 30 mL). The resultant organic layers were dried over sodium sulphate and concentrated to get crude. The crude was purified by 230 - 400 mesh silica gel column on combi flash chromatography, eluted using 0 - 10% of Ethyl Acetate in Hexane to afford Heptyl 6,6-difluoro-2-azaspiro[3.3]heptane-2- carbo(dithioperoxo)thioate (LHM-13) (120 mg, 47%) as pale yellow liquid.

[1396] TLC: 10% EtOAc in Hexane (R

[1397]

[1398] f. 0.5); NMR (400 MHz, DMSO-d6) 34.53 (s, 2H), 4.34 (s, 2H), 2.93 (t, J = 12.00 Hz, 4H), 2.82 (t, J = 8.00 Hz, 2H), 1.59 (m, 2H), 1.34 - 1.25 (m, 8H), 0.87 (t, J = 7.20 Hz, 3H) ppm; LCMS: (ES+) 98.89%, m / z = 340.2 [M + H]+[Method:- Flow rate: 0.6 ml / min, T / %B: 0.01 / 40,0.20 / 40,8 / 95,12 / 95,14 / 40,16 / 40, Buffer: A-5Mm NHjOAc, B-100% ACN, Column: Kinetex C1850*4.6*5microns, Column Temp: Ambient]; HPLC: 98.00% purity [Method:- Column: KINETEX 250*4.6mm* mn, Mobile Phase A: lOmM AA IN Aq, B: ACN],

[1399] Synthesis of Heptyl 2-oxa-6-azaspiro[3.3]heptane-6-carbo(dithioperoxo)thioate:

[1400] CS2(1.0 eq), Et3N (4.0 eq), CBr4(2.0 eq) g / “OH DCM (10 ml), 0 °C, rt, 1 h II > / \ / \OH / T-J O Y NH C -J

[1401]

[1402] A B LHN1-14 To a stirred solution of 2-oxa-6-azaspiro[3.3]heptane oxalate (B) (286 mg, 1.51 mmol, 1.0 eq) anhydrous DCM (8 mL) was cooled to 0 °C. Then carbon disulfide (115 mg, 1. 1 mmol, 1.0 eq) and triethylamine (0.84 mL, 6.04 mmol, 4.0 eq) were added to the same temperature and stirred for 5 min. After that solution of carbon tetrabromide (501 mg, 2.67 mmol, 2.0 eq) in DCM (2 mL) and heptane- 1 -thiol (200 mg, 1.51 mmol, 1.0 eq) (A) were Attorney Docket No. 10063-110WO1

[1403] added, and the resulting reaction mixture was allowed to stir at room temperature for 1 h. After completion of the reaction (reaction was monitored by TLC), reaction mixture was diluted with H2O (4 mL) and extracted with DCM (2 x 10 mL). The organic layers were dried over sodium sulphate and concentrated to get crude. The crude was purified by 230 - 400 mesh silica gel column on combi flash chromatography, eluted using 0 - 10% of Ethyl Acetate in Hexane to afford Heptyl 2-oxa-6-azaspiro [3.3]heptane-6-carbo( dithioperoxo) thioate (LHM-14) (129 mg, 28%) as off white solid.

[1404] TLC: 10% EtOAc in Hexane

[1405]

[1406] (Rf. 0.5); NMR (400 MHz, CDCh) $ 4.87 - 4.85 (d, J = 4.80 Hz, 4H), 4.50 (d, J = 8.80 Hz, 4H), 2.84 (t, J = 7.60 Hz, 2H), 1.67 (quint, J = 7.20 Hz, 2H), 1.42 - 1.29 (m, 8H), 0.90 (t, J = 6.00 Hz, 3H) ppm; LCMS: (ES+) 95.70%, m / z = 306.2 [M + H]+[Method:- Flow rate: 0.6 ml / min, T / %B:

[1407] 0.01 / 40,0.20 / 40,8 / 95,12 / 95,14 / 40,16 / 40, Buffer: A-5Mm NH4OAC, B-100% ACN, Column: Kinetex Cl 8 50*4.6*5microns, Column Temp: Ambient]; HPLC: 97.14% purity [Method:- Column: X-TERRA 250*4.6mm*5pm, Mobile Phase-A: lOmM AA IN Aq, B: ACN],

[1408] Synthesis of Heptyl 2-oxa-7-azaspiro[4.4]nonane-7-carbo(dithioperoxo)thioate:

[1409] CS2(1.0 eq), Et3N (3.0 eq), CBr4(2.0 eq). / —& DCM (IO mL), 0°C, rt, 1 h ^N^S'-Sxx''\x'~'xx^HSs / \ / \ / \+“ \

[1410] HN— '' %A\

[1411]

[1412] A B LHM-15

[1413] Stirred solution of 2-Oxa-7-aza-spiro[4.4]nonane (B) (96 mg, 0.76 mmol, 1.0 eq) was dissolved in anhydrous DCM (8 mL) and cooled to 0 °C. Then, carbon disulfide (0.04 mL, 0.76 mmol, 1.0 eq) and triethylamine (0.32 mL, 2.27 mmol, 3.0 eq) were added at the same temperature. After 5 min, a solution of carbon tetrabromide (501 mg, 1. 1 mmol, 2.0 eq) in DCM (2 mL), followed by heptane- 1 -thiol (100 mg, 0.76 mmol, 1.0 eq) (A) were added. The resulting reaction mixture was allowed to stir at room temperature for 1 h. After completion of the reaction (reaction was monitored by TLC), the reaction mixture was diluted with H₂O (4 mL) and extracted with DCM (2 x 10 mL). The organic layers were dried over sodium sulphate and concentrated to get the crude compound. The crude was purified by 230 - 400 mesh silica gel column on combi flash chromatography, eluted using 0 - 10% of Ethyl Acetate in Hexane to afford Heptyl 2-oxa-7-azaspiro[4.4]nonane-7-carbo(di thioperoxo) thioate (LHM-15) (200 mg, 79%) as pale yellow liqui.

[1414] TLC: 20% EtOAc in Hexane (R

[1415]

[1416] f: 0.5); NMR (400 MHz, DMSO-d6) 33.89 -3.74 (m, 6H), 3.58 (s, 2H), 2.82 (t, J = 7.20 Hz, 2H), 2.08 (t, J = 6.80 Hz, 1H), 1.98 (t, J = Attorney Docket No. 10063-110WO1

[1417] 6.80 Hz, 1H), 1.90 (q, J = 6.80 Hz, 2H), 1.58 (quint, J = 6.80 Hz, 2H), 1.40 - 1.20 (m, 8H), 0.87 (t, J = 5.20 Hz, 3H) ppm; LCMS: (ES+) 96.70%, m / z = 334.3 [M+H]+[Method: - Kinetex C18 50*4.6*5.0 Microns. Flow rate: 0.8 ml / min. Mobile Phase A: 5 mM NH₄OAc, B: 100% Acetonitrile]; HPLC: 97.91% purity [Method:- Column: X-TERRA 250*4.6mm*5um, Mobile Phase: A- lOmM AA IN Aq, B 100% Acetonitrile].

[1418] Synthesis of Heptyl 4-(methylsulfonyl)piperazine-l-carbo(dithioperoxo)thioate:

[1419] . HCI CS2(1.0 eq), Et3N (4.0 eq), CBr4(2.0 eq) DCM (10 mL), 0 °C, rt, 1 h HS^ + O ' I _ » « [ ibx %ZS[

[1420]

[1421] A0BXO LHM-16 Stirred solution of l-(Methylsulfonyl)piperazine hydrochloride (B) (152 mg, 0.76 mmol, 1.0 eq) in anhydrous DCM (8 mL) was cooled to 0 °C. Then carbon disulfide (0.04 mL, 0.76 mmol, 1.0 eq) and triethylamine (0.41 mL, 3.02 mmol, 4.0 eq) were added at the same temperature. After 5 min, the solution of carbon tetrabromide (501 mg, 1.51 mmol, 2.0 eq) in DCM (2 mL) followed by heptane- 1 -thiol (100 mg, 0.76 mmol, 1.0 eq) (A) were added. The resulting reaction mixture was allowed to stir at room temperature for 1 h. After completion of the reaction ( reaction was monitored by TLC), the reaction mixture was diluted with H2O (4 mL) and extracted with DCM (2 x 10 mL). The organic layers were dried over sodium sulphate and concentrated to get the crude compound. The crude was purified by 230 - 400 mesh silica gel column on combi flash chromatography, eluted using 0 - 10% of Ethyl Acetate in Hexane to afford Heptyl 4-(methylsulfonyl)piperazine-l- carbo(dithioperoxo)thioate (LHM-16) (100 mg, 36%) as pale yellow liquid.

[1422] TLC: 10% EtOAc in Hexane (Rf. 0.5); Hl NMR (400 MHz, DMSO-d6) 4.36 (brs, 2H), 4.15 (brs, 2H), 3.28 (t, J = 4.80 Hz, 4H), 2.93 (s, 3H), 2.83 (t, J = 7.20 Hz, 2H), 1.59 (quint, J = 6.80 Hz, 2H), 1.35 - 1.25 (m, 8H), 0.86 (t, J - 6.00 Hz, 3H) ppm; LCMS: (ES+) 99.18%, m / z = 371.4 [M+H]+[Method:- Kinetex C1850*4.6*5.0 Microns. Flow rate: 0.8 ml / min. Mobile Phase A: 5 mM NH4OAC, B: 100%’ Acetonitrile]; HPLC: 99.27% purity [Method:- Column: KINETEX, Mobile Phase: A - 0.1% TEA IN AQ, B - 100%

[1423] Acetonitrile]. Attorney Docket No. 10063-110WO1

[1424] Synthesis of Heptyl 5-oxohexahydrocyclopenta c]pyrrole-2(l H)-carbo( dithioperoxo )thioate:

[1425]

[1426] Step 1. Synthesis of HexahydrocycIopenta[c]pyrrol-5(lH)-one hydrochloride: Solution of Stirred solution of tert-butyl 5-oxohexahydrocyclopenta|c]pyrrole-2(lH)-carboxylate (1) (150 mg, 0.88 mmol, 1.0 eq) in anhydrous DCM (5 mL) was cooled to 0 °C. Then 4M hydrochloric acid solution in 1,4-dioxane (1.3 mL, lOv) was added drop wise at same temperature and stirred for 5 min. The resulting reaction mixture was allowed to stir at room temperature and stirred for 30 minutes. After completion of reaction (reaction was monitored by TLC), the reaction mixture was concentrated to get crude Hexahydrocyclopenta[c]pyrrol-5(lH)-one hydrochloride (2) (100 mg, 93%) as a pale yellow liquid. TLC: 20% EtOAc in Hexane (R / 0.5).

[1427] Step 2. Synthesis of Heptyl 5-oxohexahydrocyclopenta[c]pyrrole-2(lH)-carbo(dithioper-oxo) thioate: Stirred solution of hexaliydrocyclopenta[c]pyrrol-5(lH)-one hydrochloride (2) (100 mg, 0.79 mmol, 1.0 eq) in anhydrous DCM (8 mL) was cooled to 0 °C. Then carbon disulfide (0.05 mL, 0.79 mmol, 1.0 eq) and triethylamine (0.45 mL, 3.19 mmol, 4.0 eq) were added at same temperature. After 5 min, solution of Carbon tetrabromide (530.23 mg, 2.67 mmol, 2.0 eq) in DCM (2 mL) followed by heptane- 1 -thiol (105.74 mg, 0.79 mmol, 1.0 eq) (3) were added. The resulting reaction mixture was allowed to stir at room temperature for 1 h. After completion of reaction (reaction was monitored by TLC), the reaction mixture was diluted with ILO (4 mL) and extracted with DCM (2 x 10 mL). The organic layers were dried over sodium sulphate and concentrated to get crude compound. The crude was purified by 230 - 400 mesh silica gel column on combi flash chromatography, eluted using 0 - 10% of Ethyl Acetate in Hexane to afford Heptyl 5-oxohexa hydrocyclopenta[c]pyrrole-2(lH)-carbo(dithioperoxo)thioate (LHM-17) (25 mg, 12%) as pale-yellow liquid.

[1428] TLC: 10% EtOAc in Hexane (Rf. 0.5): Hl NMR (400 MHz, DMSO-d6) <54.16 -4.03 (m, 2H), 3.72 - 3.61 (m, 2H), 3.16 - 3.11 (m, 1H), 3.00 (q, J = 6.00 Hz, HI), 2.81 (t, J = 7.20 Hz, 2H), 2.45 - 2.41 (m, 2H), 2.19 (dt, J = 4.80, 18.80 Hz, 2H), 1.58 (quint, J = 7.20 Hz, 2H), 1.34 - 1.24 (m, 8H), 0.85 (t, J = 6.40 Hz, 3H) ppm: LCMS: (ES+) 99.09%, m / z= Attorney Docket No. 10063-110WO1

[1429] 332.4 [M+H]+[Method:- Kinetex C1850*4.6*5.0 Microns. Flow rate: 0.8 ml / min. Mobile Phase A: 5 mM NH4OAC, B: 100% Acetonitrile]; HPLC: 96.15% purity [Method:- Column: X-TERRA 250*4.6mm*5pm, Mobile Phase: A - lOmM AA IN Aq, B - 100% Acetonitrile],

[1430] Synthesis of Cyclohexylmethyl pyrrolidine- 1 -carbodithioate:

[1431] CS2(1.1 eq), Et3N (3.0 eq) DCM (10 mL), 0 °C, rt, 1 h

[1432]

[1433] LHM-18 To a stirred solution of pyrrolidine (A) (200 mg, 2.81 mmol, 1.0 eq) in anhydrous DCM (10 mL) was cooled to 0 °C. Then carbon disulfide (235 mg, ...

Claims

Attorney Docket No. 10063-110WO1CLAIMSWhat is claimed is:

1. A compound having Formula I or II:wherein,n is 0 to 4;— - is a bond that is present or absent,X1is O or S;X2is S, CIL, or CHR4;X3and X4are, independently, CH2, O, S, NH, NR4, CHR4, CR4, or C(R4)2;R1and R2are, independently, H, branched or straight chain Ci-C4alkyl, aryl, heteroaryl, (Ci- C4alkyl)aryl, (Ci-C4alkyl)heteroaryl, Cs-Cgcycloalkyl, Cs-Cecycloheteroalkyl, (Ci- C4aIkyl)C3-C6CycloaIkyl, or (Ci-C4alkyl)C3-C6Cycloheteroalkyl, wherein R1and R2are not both H;R3is branched or straight chain Ci-Ci2alkyl, Ci-Ci2heteroalkyl, aryl, heteroaryl, (Ci- C4alkyl)aryl, (Ci-C4alkyl)heteroaryl, Cs-Cecycloalkyl, Cs-Cecycloheteroalkyl, (Ci- CralkyllCs-Cecycloalkyl, or (Ci-ChalkyliCs-Cscycloheteroalkyl, any of which are optionally substituted with oxo, sulfo, halo, Ci-Cgalkyl;R4is branched or straight chain Ci-Cialkyl, C2-C4alkenyl, Ci-Cdieteroalkyl, C2- C4heteroalkenyl, or two R4’s bond together to form a fused Cs-Cecycloalkyl, fused Cs-Cecycloheteroalkyl, fused aryl, fused heteroaryl, spiro Cs-Cecycloalkyl, or spiro C2-C6cycloheteroalkyl, any of which are optionally substituted with oxo, sulfonate, sulfonyl, halo, hydroxyl, thiol, carboxylate, carbonyl, Ci-Cealkyl, or Ci-Cghaloalkyl; or a pharmaceutically acceptable salt thereof,with the proviso that the compound is not Formula II where X1and X2are both S, R3is Csalkyl, n is 1, and X3and X4are both CH2.

2. The compound of claim 1, wherein Xsis O.

3. The compound of claim 1, wherein Xsis S.

4. The compound of claim 1, wherein X2is S.Attorney Docket No. 10063-110WO15. The compound of claim 1, wherein X2is CH2.

6. The compound of claim 1, wherein R3is straight chain C3-C10 alkyl.

7. The compound of claim 1, wherein R3is Ci-C4alkyl)aryl.

8. The compound of claim 1, wherein the compound has Formula I.

9. The compound of claim 8, wherein Rland R2are both branched or straight chain Ci - C4alkyl.

10. The compound of claim 8, wherein one of R1or R2is (Ci-C4alkyl)aryl.

11. The compound of claim 1, wherein the compound has Formula II.

12. The compound of claim 11, wherein n is 1.

13. The compound of claim 11, wherein n is 2.

14. The compound of claim 11, wherein — is a bond that is present.

15. The compound of claim 11, wherein — is a bond that is absent.

16. The compound of claim 11, wherein X3and X4are both CII2.

17. The compound of claim 11, wherein X3is ().

18. The compound of claim 11, wherein XJand X4are both CHR4and the two R4’s bond together to form a fused Cs-Cgcycloalkyl.

19. The compound of claim 11, wherein X3and X4are both CR4and the two R4’s bond together to form a fused aryl.

20. The compound of claim 11, wherein X4is C(R4)2. and the two R4’s bond together to form a spiro Cs-Cecycloheteroalkyl.

21. The compound of claim 1, wherein the compound is selected fromAttorney Docket No. 10063-110WO1DIS8-2 DIS10-2SDIS15-2DIS3-3 DIS4-3 DIS5-3 DIS6-3:00Attorney Docket No. 10063-110WO1LHM-22 LHM-23Attorney Docket No. 10063-110WO1SLHM-09 LHM-10SLHM-24LHM-25:02Attorney Docket No. 10063-110WO1 LHM-26LHM-33Attorney Docket No. 10063-110WO1 LHM-29 LHM-30SLHM-34LHM-49 LHM-51LHM-5322. A compound having Formula III:Attorney Docket No. 10063-110WO1wherein,Lis null, -NHC(O)C(CH3)2-, -NR5C(O)C(CH3)2-, -or -OC(O)C(CH3)2-;R5is branched or straight chain Ci-Cnalkyl, Ci-Cnheteroalkyl, aryl, heteroaryl, (Ci- CAalkyljaryl, (Ci-C.4alkyl)heteroaryl, C3-C6cycloalkyl, C3-C6cycloheteroalkyl, (Ci- C4alkyl)C3-C6cycloalkyl, or (Ci-C4alkyI)C3-C6cycloheteroalkyl;or a pharmaceutically acceptable salt thereof.

23. The compound of claim 22, wherein L is null.

24. The compound of claim 23, wherein R5is branched or straight chain CT-Cioalkyl or C3-Cioheteroalkyl.

25. The compound of claim 23, wherein R5is C3-Cecycloalkyl or C3- Cgcycloheteroalkyl.

26. The compound of claim 23, wherein R5is (Ci-C4alkyl)C3-C6cycloalkyl, or (Ci - C4alkyl)C3-C6cycloheteroalkyl.

27. The compound of claim 23, wherein R5is (Ci-C4alkyl)aryl.

28. The compound of claim 22, wherein L is -NHC O)C(CH3)2- or -NR3C(O)C(CH3)2.

29. The compound of claim 28, wherein R5is branched or straight chain Cz-Csalkyl.

30. The compound of claim 22, wherein the compound is selected fromAZ5 AZ6 AZ7:05Attorney Docket No. 10063-110 WO 1 AZ8 AZ9 AZ10 AZ11 AZ12 AZ1-2 AZ2-2AZ3-2 AZ4-2Attorney Docket No. 10063-110WO1AZ5-2 AZ6-2AZ7-2 AZ8-231. A method of treating a bacterial and fungal infection, comprising: administering to a subject in need thereof a composition comprising a compound of any one of claims 1-30.

32. The method of claim 31, wherein the bacterial infection is caused by a drug-resistant bacteria and fungi.

33. The method of claim 31, wherein the compound is the compound of any one of claims 22-30 and the bacterial infection is caused by Helicobacter pylori, S. aureus strain MW2, Enterococcus f aecium, Acinetobacter baumannii, Klebsiella pnemoniae, Enterobacter aerogenes, Pseudomonas aeruginosa, Listeria monocyte gens, Burkholderia. pseudomallei BSL2 strain 82, Salmonella enterica subsp. enterica CIP 60.62, or Shigella flexneri.

34. The method of claim 31, wherein the compound is the compound of any one of claims 22-30 and the fungal infection is caused by Candida albicans strain CAN14, Candida auris slrai n CAU 1, or Candida glabrata.

35. The method of claim 31, wherein the compound is the compound of any one of claims 1-21, and the bacterial infection is caused by Helicobacter pylori, S. aureus strain MW2, Enterococcus faecium, Acinetobacter baumannii, Klebsiella pnemoniae, Enterobacter aerogenes, Pseudomonas aeruginosa, Listeria, monocytogens, Burkholderia:07Attorney Docket No. 10063-110WO1pseudomallei BSL2 strain 82, Salmonella enterica subsp. enterica OP 60.62, or Shigella flexneri.

36. The method of claim 31, wherein the compound is the compound of any one of claims 1-21, and the fungal infection is caused by Candida albicans strain CAN 14, Candida auris strain CAU1, or Candida glabrata.

37. The method of claim 31, further comprising administering azetreonam and / or disulfiram.

38. The method of claim 31, further comprising administering an antibiotic.

39. The method of claim 38, wherein the antibiotic is amoxicillin, clarithromycin, metronidazole, nitrofurantoin, or any combination thereof.

40. The method of claims 31-38, further comprising administering a proton pump inhibitor.

41. The method of claim 40, wherein the proton pump inhibitor is omeprazole, lansoprazole, pantoprazole, rabeprazole, or esomeprazole.

42. The method of claim 41, wherein the bacterial infection is caused by a drug -resistant II. pylori.

43. A method of treating a bacterial infection, comprising: administering to a subject in need thereof aztreonam and / or disulfiram, an antibiotic, and a proton pump inhibitor.

44. The method of claim 43, wherein the antibiotic is amoxicillin, clarithromycin, metronidazole, nitrofurantoin, or any combination thereof.

45. The method of claim 43, wherein the proton pump inhibitor is omeprazole, lansoprazole, pantoprazole, rabeprazole or esomeprazole.

46. The method of claim 43, wherein the bacterial infection is caused by a drug -resistant H. pylori.

47. A method of treating a bacterial infection, comprising: administering to a subject in need thereofSVJDIS1-2or a pharmaceutically acceptable salt thereof.

48. A method of treating bacterial and fungal infections in single or dual therapy (DIS or DIS analogs+ PPI) or (DIS or DIS analogs+clinical antibiotic) or triple therapy (DIS or or DIS analogs+clinical antibiotic+PPI).Attorney Docket No. 10063-110WO149. A method of treating bacterial and fungal infections in dual therapy (AZ + AZ analog or DIS + DIS analog) antibiotic) or triple therapy ((AZ or DIS or AZ analog or DIS analogs+clinical antibiotic+PPI).

50. A method of composition of formulation used in polyethylene glycol fatty acid esters (Koliphor or cremophor) for oral therapy.

1. A method of formulating one pill that has AZ or AZ analogs + PPI + clinical antibiotic (amoxicillin or clarithromycin or nitrofurantoin or Bismuth Salicylate).

52. A method of delivery oral, IV, intramuscular.

53. A method of treating Neisseria gonorrhea infections using DISULFIRAM or DISULFIRAM analogs alone or combined with clinical antibiotics.

54. A method of treating Neisseria gonorrhea infections using AZTREONAM analogs alone or combined with clinical antibiotics.

55. A method of treating a bacterial and fungal infection, comprising: administering to a subject in need thereof a composition comprising a compound of any one of claims 1-30:09

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