Use of casein proteolytic enzyme p function as biomarker of drug response to IMIPRIDONE-like reagent

Abstract

The present disclosure relates to the use of casein proteolytic enzyme P function as a biomarker for the response of a drug to an IMIPRIDONE-like agent. Use of casein proteolytic enzyme P (ClpP) function and / or concentration as a biomarker for predicting the response to a compound of formula I, for a neoplastic disease, preferably cancer, or for other diseases in which enhancement of ClpP activity can provide therapeutic benefits. In other aspects, it relates to methods and kits, as well as to methods of treatment using the biomarkers.

Description

[0001] This application is a divisional application. The original application was filed on February 26, 2020, with application number 202080031455.3, and the invention title was “Use of casein hydrolysate P function as a biomarker of drug response to IMIPRIDONE-like reagent”, the entirety of which is incorporated herein by reference. Technical Field

[0002] This invention relates to the use of casein hydrolysate P (ClpP) function and / or concentration as biomarkers for predicting tumor diseases, preferably cancer, or other diseases for which enhanced ClpP activity may provide therapeutic benefit, in response to compounds of formula I. In other aspects, it relates to methods and kits, as well as therapeutic methods using biomarkers. Furthermore, chemicals for activating ClpP are described. Background Technology

[0003] Mammalian mitochondria contain a serine protease complex (ClpP), which is a proteolytic component of the ClpXP protein degradation complex. This complex plays a role in mitochondrial protein quality control (Houry, WA et al., Cell Chemical Biology 2018). 25 ClpP plays a central role in regulating cellular bioenergetic activity. Houry, WA et al. have also reported that ClpP is highly expressed in various cancers and plays an important role in cell metastasis. Furthermore, mitochondrial dysfunction is central to disease mechanisms and may be a pathogenic factor in many neurodegenerative diseases (Beal and Johri, J Pharmcol Exp Thera. 2012, 342(3), 619-630 and cited references therein). ClpP deficiency leads to an overload of misfolded / unfolded mitochondrial proteins, inhibits mitochondrial respiratory activity, increases mitochondrial oxidative damage, and causes cell death (Qi et al., Acta Neuropathologica, 2019, 137, 939-960 and cited references therein).

[0004] Agents that modulate ClpP function have been identified. Direct activation of proteases with small molecules is rare in drug discovery. Agents activating ClpP have been reported (Sieber, SA et al., Angew. Chem. Int. Ed. 2018, 57, 14, 602-14607 and references cited therein). Furthermore, agents inhibiting ClpP have been reported (Schimmer, AD et al., Cancer Cell 2015, 27, 864-876 and references cited therein). Schimmer, AD et al. and Sieber, SA et al. have both described the use of their agents in cancer treatment. Orally active agents for cancer treatment have preferred market potential due to their ease of administration during repeated dosing. However, highly potent small-molecule upregulators of ClpP activity are unknown. The larger macrocyclic activator of ClpP, namely “ADEP”, is known, but lacks structural features for oral bioavailability (Lipinski's rules, Oprea et al., Adv. Drug Deliv Rev. 2016, 101, 89-98 and the references cited therein).

[0005] Proteases highly similar to human ClpP have been found to be encoded in the genomes of bacteria and some viruses. Agents that modulate ClpP function have been shown to be useful in treating bacterial infections. Kao RYT et al. described small-molecule inhibitors of ClpP and their effects on Staphylococcus aureus (Kao, RYT et al., PNAS 2018, 115, 8003-8008 and references cited therein). Furthermore, ClpP activators have been described (Lee RE et al., ACS Infect Dis 2019, Nov 8;5(11): 1915-1925 and references cited therein). Mitochondria have many quality control systems to ensure homeostasis (protein homeostasis). Defects in these systems can lead to mitochondrial dysfunction, markers of aging, various neurodegenerative diseases, cardiovascular diseases, and cancer (Li R. et al., AnnRev Biophy, 2020, Jan 13. doi: 10.1146 / annurev-biophys-121219-081604 and references cited therein, Martins LM, J Mol Med, 2013, 91, 665-671 and references cited therein, and Jeong YY, Cells, 2020, 9(1), 150 and references cited therein. α-synuclein accumulation and mitochondrial dysfunction are associated with the pathology of Parkinson's disease and Alzheimer's disease (Qi et al., Acta Neuropathologica, 2019, 137, 939-960 and references cited therein, and Nielsen & Twohig, Mol Neurodegener, 2019, 14(1)). (23 and the references cited therein). Furthermore, α-synuclein can lead to decreased ClpP protein levels. Significantly enhanced ClpP activity in cellular systems reduces α-synuclein-related pathology.

[0006] ONC201 is a small molecule drug for treating cancer that has entered clinical trials and is being evaluated for the treatment of several cancers. Several published reports describe various aspects of the mechanism of action of ONC201. Publications describe ONC201 as acting through G protein-coupled receptors (GPCRs) (El-Deiry WS, Neoplasia 2018, 20, 80-91 and references cited therein). Furthermore, one report describes changes in cellular function, including mitochondrial function after treatment with ONC201 (Lipkowitz S., Oncotarget 2018, 9, 18, 454-18, 479 and references cited therein).

[0007] Perrault syndrome is a disorder characterized by female ovarian hypoplasia and sensorineural hearing loss. In more severe cases, additional symptoms may include ataxia, neurosis, and intellectual disability (Dougan, DA, Sci Rep 2018, 8(1), 12862 and references cited therein). Mutations in six different genes are associated with this disease, and in Perrault syndrome, the type 3 mutation of ClpP is the cause. Two mutations, Y229D and I208M, are thought to alter peptidase activity, with Y229D showing inhibition of ClpP peptidase activity.

[0008] Loss-of-function mutations in the heme biosynthesis enzyme gene can cause congenital porphyria. ClpX promotes heme biosynthesis, and mutations in ClpX (Gly298Asp) lead to the pathological accumulation of the heme biosynthesis intermediate protophyria (PPIX). (Paw BH, Proc. Natl. Acad. Sci. USA. 114:E8045-E8052 (2017) and references cited therein).

[0009] Non-dividing hepatocytes in end-stage liver disease indicate permanent growth arrest, cryptogenic cirrhosis (Ramakrishna, G. et al., Cell Mol Gastroenterol Hepatol. 2019, 8(1):73-94 and references cited therein). Fatty liver is a common cause of cryptogenic cirrhosis. The contemporary drug development process, often referred to as the translational medicine approach, focuses on identifying the right patients and targeting specific interventions for key aspects of the disease process. This requires multiple inputs, including understanding specific molecular events that are critical to an individual’s disease process and having a clear understanding of how a particular treatment will intervene in that individual’s disease process (Rossetti L., Drug Dis. Today 2016, 21, 517-526 and references cited therein). At the heart of this approach is the development and use of biomarkers and associated companion diagnostics for specific therapeutic treatments. Summary of the Invention

[0010] In this invention, we report that human ClpP (hClpP or HSClpP) is a biomarker for the chemical action of ONC201 and related chemical analogs, and that this biomarker can be used to determine whether a patient is a candidate for this drug treatment and whether the drug treatment has the expected molecular effect. Specifically, we show that these compounds directly bind to hClpP and activate the peptidase activity of hClpP. The binding and activation of hClpP occurs in a time- and dose-dependent manner and parallels the growth-inhibiting effects of these compounds on cancer cells. Therefore, our findings suggest that the biological action of ONC201 (and related compounds) depends on the physical activation of hClpP. Our findings pertain to hClpP and ClpP (ClpP) in other mammalian species. Furthermore, ONC201 and related chemical analogs directly bind to bacterial ClpP (bClpP) and activate the peptidase activity of bacterial ClpP. This pertains to Staphylococcus aureus and other bacterial species. We anticipate that the action of bClpP occurs in a time- and dose-dependent manner and is the reason for the growth-inhibiting effects of these compounds on bacterial cells. We also anticipate that the antimicrobial activity of ONC201 and structure-related compounds is due to the physical activation of bClpP. This invention also allows for the molecular evaluation of bacteria susceptible to ONC201 and chemically related compounds.

[0011] A large class of neurodegenerative disorders is characterized by the relatively selective death of neuronal subtypes. Impaired mitochondrial dysfunction can lead to many neurodegenerative diseases, such as, but not limited to, Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis (ALS), Friedreich ataxia, and Alzheimer's disease. This invention relates to the treatment of these diseases using the agents described herein, how to select patients who will benefit from such treatment, and methods for monitoring patients' responses to treatment.

[0012] We disclose fluorescence, positron emission tomography (PET) scans (F... 18-PET, near-infrared, and other small molecule probes can be chemically coupled to the compounds described in this invention as a direct method for imaging ClpP expression in tumors or other tissues (Liu HW., Chem Soc. Review, 2018, 47, 7140-7180 ​​and references cited therein; Pantel A.R., Cancer Lett, 2017, 387, 25-31 and references cited therein). This provides a basis for detecting ClpP expression in tumors as a biomarker for cancer. Secondly, the use of these probes can be used directly to measure the ClpP binding efficacy of drugs (such as ONC201 and the chemical agents or other ClpP binders described herein)—by measuring the competitive reduction of ClpP binding. Target (ClpP) binding of ONC201 or other compounds described herein can be directly measured in in vivo animal, human, or in vitro screening assays. Combined with ClpP enzymatic activity assays, the binding of small molecules to this ClpP can be directly measured. Third, the development of ClpP-dependent activity probes can be used to determine the activity of ClpP in tumors or cell lysates. Based on the principle of enzyme-activated fluorescent probes described in *Liu HW., Chem Soc. Review, 2018, 47, 7140-7180 ​​and references cited therein), for this purpose, we propose attaching responsive chemical groups to the compounds of this invention. Amine-responsive TR-compounds will be conjugated with a chemifluorescent substrate, with the aim of targeting these compounds to ClpP in intact tumors and directly measuring ClpP using this method. Fourth, the use of TR-compound probes can be used to discover novel ClpP-binding molecules by substituting unknown compounds for TR-probe compounds in high-throughput assays using time-resolved fluorescence assays or other assays. We further disclose that the use of our TR-compound probes can be similarly used to identify novel small-molecule binders of bacterial ClpP (bClpP) enzymes. Combined with bClpP activity assays, the effect of these small molecules on bClpP activity can be directly determined. This provides unique TR-probe compounds for the discovery of bClpP binders as potential antibacterial agents. In addition, new chemical substances that can be used as anticancer agents were described. Attached Figure Description

[0013] Figure 1 Kinetics of hClpP activity without pre-incubation with ONC201 and Ex. 51 (TR57) – hClpP peptide hydrolysis assay in the presence of ONC201 and Ex. 51 (TR57). Time- and dose-dependent enhancement of coumarin fluorescence release from Ac-WLA-AMC by the enzymatic activity of purified hClpP was shown. Protocol 1.

[0014] Figure 2 Kinetics of hClpP activity after 60 min pre-incubation with ONC201 and Ex. 51 (TR57) – hClpP peptide hydrolysis assay in the presence of ONC201 and TR57. The study showed a time-dependent increase in the enzymatic activity of purified hClpP from Ac-WLA-AMC for coumarin fluorescence after 60 min pre-incubation with either ONC201 or Ex. 51 (TR57). Protocol 2.

[0015] Figure 3 The dose-dependent activation of hClpP using ONC201, Ex. 51 (TR57), Ex. 14 (TR65), Ex. 57 (TR79), and Ex. 1 (D9) was studied. A dose-dependent increase in hClpP activity in response to incubation with individual compounds was shown. HClpP activity was measured as an increase in coumarin fluorescence released from Ac-WLA-AMC by the enzymatic activity of purified hClpP, as described above. For comparison, the published hClpP activator D9 was included. Activities were plotted as relative fluorescence units (RFU / ug of hClpP / h(H)). EC 50 The value represents the dose-dependent activation measured by this method.

[0016] Figure 4 HClpP is the binding protein of the compounds of this invention. HClpP is the binding protein of the compounds Ex, 2 (TR31) and Ex.59 (TR81) of this invention. The binding of hClpP to immobilized Ex.59 (TR81) agarose beads in vitro is shown. HELA cell lysates with a carrier (0.1% DMSO) or... Figure 4 The concentrations of ONC201 or Ex. 2 (TR31, ONC212) dissolved in DMSO were briefly incubated (10 min). These samples were then applied to a fixed TR-81 agarose column (50 μL) and washed to remove unbound proteins. Samples were eluted with SDS-PAGE sample buffer, applied to SDS-PAGE, and subjected to hClpP Western blotting. As shown, increasing the concentrations of ONC201 and Ex. 2 (TR31) in the lysate resulted in a dose-dependent detachment of competing hClpP from the Ex. 59 (TR81) resin. Similar results were obtained with Ex. 51 (TR57) (not shown). The study aimed to identify the protein-binding ClpP compounds of the present invention.

[0017] Figure 5Data on ClpX and TUFM concentrations, responses, and time progression in SUM159 cells using ONC201 and TR57 (Ex. 51). The study demonstrates the effects of these compounds and the reduction of proteins ClpX and TUFM (measured by Western blot) when cancer cells (SUM159) are exposed to the compounds of this invention.

[0018] Figure 6 ClpP CRISPR knockout cells are resistant to the effects of ONC201 and TR57 (Ex. 51). This study shows the effects of ONC201 and Ex. 51 (TR57) on the growth of the cancer cell line SUM159 compared to the growth of a ClpP-free cell line (SUM159, ClpP CRISPR KO).

[0019] Figure 7 Examples 81 and 82 1 HNMR.

[0020] Figure 8 LC-MS of Example 80.

[0021] Figure 9 The timeline for activating ClpP using 1 μM Ex. 60 (ONC206). Figure 2 The conditions described in Scheme 2 involved incubating purified hClpP with Ex. 60 (ONC206). This demonstrated a time-dependent increase in the release of coumarin fluorescence from Ac-WLA-AMC via the enzymatic activity of purified hClpP.

[0022] Figure 10The dose-dependent activation of ClpP using Ex. 62 (TR98), Ex. 66 (TR108), Ex. 67 (TR109), and Ex. 68 (TR122) was studied. Also, 1 μM studies were conducted on Ex. 80 (TR145), Ex. 81 (TR146), Ex. 82 (TR147), Ex. 83 (TR129), and Ex. 84 (TR130). The time-dependent increases in hClpP activity caused by TR129, TR130, TR145, TR146, and TR147 were measured. The dose-dependent increase in hClpP activity in response to incubation of purified HClpP with individual compounds was measured. HClpP activity was measured as an increase in coumarin fluorescence released from Ac-WLA-AMC by the enzymatic activity of purified hClpP (Scheme 2). The study also showed a time-dependent increase in hClpP activity measurements at 1 μM: Ex. 83 (TR129); Ex. 84 (TR130); Ex. 80 (TR145); Ex. 81 (TR146); and Ex. 82 (TR147). HClpP activity was measured as an increase in relative fluorescence units from the hydrolysis of the aforementioned substrate Ac-WLA-AMC (Scheme 2). Detailed Implementation

[0023] This invention relates to methods for determining whether an individual responds to a reagent described in Formula I and methods for determining whether an individual maintains responsiveness to a reagent described in Formula I, said methods comprising measuring the level of at least one biomarker in a biological sample. The invention further relates to kits for performing said methods. The invention further describes novel chemical substances and their uses in the treatment of cancer, various proliferative diseases, various immune diseases, various inflammatory diseases, bacterial infections, neurodegenerative diseases, viral diseases such as HIV, acquired immunodeficiency syndrome (AIDS), hereditary spastic paraplegia, cystic fibrosis (CF), and Perrault syndrome. Wong, KS, and Houry, WA (ACS Chem. Biol., 2019: DIO:10.102.1021 / acschembio.9b00347, and references cited therein) describe ClpP, its relationship with cancer and other diseases, and targeted therapy. All publications, patents, and patent applications cited in this specification are incorporated herein by reference as if each individual publication, patent, or patent application were specifically and individually indicated as incorporated herein by reference. The term following a specific citation, "and the references cited therein," whether from publications, patents, or patent applications, indicates that all citations within that specific citation are also incorporated into this text.

[0024] The reduced expression of the protein ClpP in RNA and expressed protein (ClpP) in cells from patients with hereditary spastic paraplegia can be corrected by treatment with compounds and by using the methods of the present invention (Bross, P et al., Neuroscience, 2008, 153, 474-482).

[0025] One aspect of the present invention relates to the treatment of hereditary spastic paraplegia by administering the compounds of the present invention.

[0026] One aspect of this invention relates to a novel method for detecting ClpP as a biomarker for cancer and other diseases. This is based on our initial findings that ONC201 and the chemically relevant compounds defined in Formula I are high-affinity binders and activators of ClpP enzymatic activity. Another aspect of this invention relates to using the reagents described herein as active probes to detect ClpP protein and activity levels in tumors and cells, including biological samples derived from mammals. These biological samples can be obtained from mammals before or after treatment with the compounds described in Formula I. Furthermore, these samples can also be treated with the compounds described in Formula I, and the response to the compounds can be determined by changes in ClpP activity and protein levels, or other biomarkers associated with ClpP activity.

[0027] Another aspect of the invention relates to the regulation of the complex ClpXP and its components ClpP and the AAA+ ATPase ClpX. This regulation of these components can be used to treat diseases.

[0028] One aspect of the present invention relates to a novel method for detecting ClpXP as a biomarker for cancer and other diseases.

[0029] One aspect of the present invention relates to a novel method for detecting ClpX as a biomarker for cancer and other diseases.

[0030] Another aspect of the invention relates to the identification of other chemical substances that act as ClpP binders. The compounds of the present invention can be used to screen compound libraries for the determination of new chemical substances.

[0031] A. Develop high-affinity ClpP binding probes for detecting ClpP in live animals, patients, or intact cells.

[0032] We have discovered that hClpP directly binds to the compounds TR79, TR80, and TR81 of this invention—when coupled with agarose beads. Furthermore, we have identified ONC201, ONC212 (TR31), and others (TR57) that compete with hClpP (human-ClpP) for detachment from the aforementioned functionalized agarose beads in a dose-dependent manner. Figure 4This indicates that ONC201 and other analogues and related chemicals of the present invention bind hClpP (Graves LM et al., ACS Chem Biol., 2019, 14(5), 1020-1029 and references cited therein). The present invention further describes the use of chemically reactive functions to link fluorescence, infrared, PET and other imaging motifs to a subset of compounds of Formula I. These imaging motifs disclosed herein are collectively referred to as “dyes”. Examples of compounds having these properties are TR79, TR80 and TR81. These probes are used as cell-permeable imaging probes for the detection of ClpP as a biomarker for cancer or other diseases.

[0033] B. Measure probe displacement to evaluate small molecule therapeutic binding to the biomarker protein ClpP.

[0034] As described herein, the probes of this invention will be used to measure the efficacy of therapeutic agents targeting a (ClpP) in binding to the enzyme. This will include ONC201, ONC206, ONC212, and other Formula I compounds used to treat mammalian diseases. Animals or humans will be exposed to these probes and tumors imaged by fluorescence, PET, or other imaging methods. Exposure to ONC201 or related compounds will be performed, and the amount of probe remaining bound to ClpP will be determined by imaging. Measuring the signal before and after such exposure will allow for direct measurement of how effectively this biomarker target (ClpP) binds to ONC201 or other ClpPs binding to related therapeutic agents.

[0035] C. Develop ClpP activity-based probes for detecting ClpP activity in tumors, cells, or cell lysates. A subset of Formula I compounds is used to generate activity-dependent probes selective for ClpP. Numerous cleavable fluorescent or other such chemical motifs, known to those skilled in the art, are used to generate ClpP-active probes. Suitable examples of Formula I compounds are TR79, TR80, and TR81, each possessing a chemically reactive amine suitable for conjugation with a variety of reagents (generating "coupling agents"). These coupling agents will be used to 1) guide these molecules to bind directly to ClpP, and 2) measure ClpP activity via the hydrolysis of the fluorescent molecule. These reagents will also be used to image ClpP activity in tumor, tissue, or cell lysates.

[0036] D. Develop ClpP probes for high-throughput screening of ClpP binding and regulation. The various probe / conjugated reagents described in this invention are diagnostic reagents for evaluating the binding of compounds to ClpP from mammalian and bacterial sources. This assay is based on the displacement of ClpP by a fluorescent (or other) probe. Time-resolved fluorescence anisotropy (or a similar assay) will be used to measure the displacement of the probe compound from ClpP by said compound. This will form the basis of an HTS screening procedure to discover novel small molecule interactants of ClpP from human or bacterial sources.

[0037] definition The terms used herein have their ordinary meanings, and the meaning of these terms is independent each time they appear. Nevertheless, and unless otherwise stated, the following definitions apply throughout the specification and claims.

[0038] a) Biologically relevant definitions Cancer: Tumor formation It is the abnormal growth and proliferation of abnormal cells or an abnormal number of cells caused by benign or malignant processes.

[0039] Biological samples. The term "sample" in relation to an individual encompasses blood and other liquid samples of biological origin, solid tissue samples such as biopsy samples and their progeny. The definition also includes samples that have been processed in any way since their acquisition, such as by reagent treatment; washing; or enrichment for certain cell populations (e.g., cancer cells). The definition also includes samples rich in specific types of molecules (e.g., nucleic acids, peptides, etc.).

[0040] The term "biological sample" encompasses clinical samples. Types of "biological samples" include, but are not limited to: tissue obtained through surgical resection, tissue obtained through biopsy, cultured cells, cell supernatant, cell lysate, tissue samples, organs, bone marrow, blood, plasma, serum, fine needle aspirate, lymph node aspirate, cyst aspirate, puncture samples, thoracentesis samples, etc. A "biological sample" may include cells (e.g., target cells, normal cells, blood cells, tissue cells, etc.) that may or may not contain such cells. A biological sample may include a biological fluid derived from cells (e.g., cancer cells, infected cells, etc.), for example, a sample containing polynucleotides and / or polypeptides obtained from such cells (e.g., cell lysate or other cell extracts containing polynucleotides and / or polypeptides). A biological sample containing infected cells from a patient may also contain uninfected cells. In some embodiments, the biological sample is blood or a derivative thereof, such as plasma, serum, etc.

[0041] Obtaining and Measuring Samples. The term “methodization” is used herein to refer to the physical steps involved in manipulating a biological sample to produce data related to the sample. As will be readily understood by one of ordinary skill in the art, a biological sample must be “obtained” before it can be measured. In this respect, the term “methodization” implies that the sample has been obtained. As used herein, the term “obtained” or “acquired” includes the act of receiving an extracted or isolated biological sample. For example, a testing institution may “obtain” a biological sample by mail (or by delivery, etc.) before it can be measured. In some cases, a biological sample is “extracted” or “isolated” from an individual by another party before being mailed (i.e., delivered, transferred, etc.) and then “obtained” by the testing institution upon the sample’s arrival. In this respect, the testing institution can obtain the sample and then measure it, thereby generating data related to the sample.

[0042] As used herein, the terms "obtained" or "acquired" may also include the physical extraction or separation of a biological sample from an object. Thus, a biological sample may be separated from the object (and therefore "obtained") by the same person or entity from which the sample is subsequently determined. When a biological sample is "extracted" or "separated" from a first party or entity and then transferred (e.g., delivered, mailed, etc.) to a second party, the sample is obtained by the first party (and also "separated" by the first party), and then subsequently "obtained" by the second party (but not "separated"). Therefore, in some embodiments, the obtaining step does not include the step of separating the biological sample.

[0043] In some embodiments, the obtained steps include the step of separating biological samples (e.g., pre-processed biological samples, post-processed biological samples, etc.). Methods and procedures for separating various biological samples (e.g., blood samples, serum samples, plasma samples, biopsy samples, extracts, etc.) will be known to those skilled in the art, and any convenient method can be used to separate biological samples.

[0044] Those skilled in the art will understand that in some cases, it is convenient to wait until multiple samples (e.g., pre-treatment biological samples and post-treatment biological samples) are available before determining the samples. Therefore, in some cases, isolated biological samples (e.g., pre-treatment biological samples, post-treatment biological samples, etc.) are stored until all suitable samples are available. Those skilled in the art will understand how to properly store multiple different types of biological samples and that any convenient storage method suitable for a particular biological sample (e.g., refrigeration) can be used. In some embodiments, pre-treatment and post-treatment biological samples are determined in parallel. In some cases, multiple different post-treatment biological samples and / or pre-treatment biological samples are determined in parallel. In some cases, biological samples are processed immediately or as soon as possible after they are obtained.

[0045] In the subject-matter approach, the concentration (i.e., “level”) or expression level of a gene product—which may be RNA, protein, etc. (hereinafter referred to as a biomarker)—is measured (i.e., “as determined”) in a biological sample. With regard to “expression level” (or “level”), it refers to the level of the gene product (e.g., the absolute and / or normalized value determined for the RNA expression level of a biomarker in a biological sample, or for the expression level of an encoded polypeptide, or for the concentration of a protein). The terms “gene product” or “expression product” are used herein to refer to the RNA transcript of a gene (RNA transcripts, such as mRNA, unspliced ​​RNA, spliced ​​variant mRNA, and / or fragmented RNA), including mRNA, and the polypeptide translation products of such RNA transcripts. Gene products can be, for example, unspliced ​​RNA, mRNA, spliced ​​variant mRNA, microRNA, fragmented RNA, polypeptides, post-translational modified polypeptides, spliced ​​variant polypeptides, etc.

[0046] The terms “determine,” “measure,” “assess,” “evaluate,” “assay,” and “analyze” are used interchangeably herein to refer to any form of measurement and include determining the presence of an element. These terms include quantitative and / or qualitative determinations. An assay can be relative or absolute. For example, an “assay” can determine whether an expression level is less than or “greater than or equal to” a specific threshold (which may be predetermined or determined by measuring a control sample). On the other hand, an “assay to determine an expression level” can mean determining a quantitative value (using any convenient measure) representing the expression level (i.e., the amount of expression, e.g., protein and / or RNA (e.g., mRNA)). Expression levels can be expressed in any unit associated with a particular assay (e.g., fluorescence units, e.g., mean fluorescence intensity (MFI)) or can be expressed as an absolute value with defined units (e.g., the number of mRNA transcripts, the number of protein molecules, the concentration of protein, etc.). Furthermore, the expression level of a biomarker can be compared with the expression levels of one or more other genes (e.g., nucleic acids and / or proteins they encode) to derive a standardized value representing the standardized expression level. The specific metric (or unit) chosen is not critical, as long as the same unit (or conversion to the same unit) is used when evaluating multiple biological samples from the same individual (e.g., biological samples collected from the same individual at different time points). This is because units are eliminated when calculating the fold change in the expression level of one biological sample relative to the next (e.g., biological samples collected from the same individual at different time points) (i.e., determining the ratio).

[0047] To measure RNA levels, the amount or level of RNA in a sample is determined, for example, the level of mRNA. In some cases, the expression levels of one or more additional RNAs may also be measured, and the biomarker expression level is compared to the levels of said one or more additional RNAs to provide a standardized value for the biomarker expression level. Any convenient protocol for assessing RNA levels can be used, in which the levels of one or more RNAs in the sample being measured are determined.

[0048] Many exemplary methods for measuring the expression level of RNA (e.g., mRNA) (e.g., the expression level of nucleic acid biomarkers) in a sample are known to those skilled in the art, and any convenient method may be used. Exemplary methods include, but are not limited to: hybridization-based methods (e.g., RNA blotting, array hybridization (e.g., microarrays); in situ hybridization; in situ hybridization followed by FACS; etc.) (Parker & Barnes, Methods in Molecular Biology 106:247-283 (1999)); RNA protection assays (Hod et al., Biotechniques, 1992, 13 852-854 and references cited therein); PCR-based methods (e.g., reverse transcription PCR (RT-PCR), quantitative RT-PCR (qRT-PCR), real-time RT-PCR, etc.) (Weis et al., Trends in Genetics 1992, 8 263-264 and references cited therein); nucleic acid sequencing methods (e.g., Sanger sequencing, next-generation sequencing (i.e., massively parallel high-throughput sequencing, such as Illumina's reversible terminator method, Roche pyrosequencing (454), Life Technologies' ligation sequencing (SOLiD platform), Life...). Technologies' ion flow platform, single-molecule sequencing, etc.); and so on.

[0049] In some implementations, biological samples can be directly measured. In some implementations, the nucleic acids of the biological sample are amplified (e.g., by PCR) before measurement. Therefore, techniques such as PCR (polymerase chain reaction), RT-PCR (reverse transcriptase PCR), and qRT-PCR (quantitative RT-PCR) can be used before the hybridization and / or sequencing methods described above.

[0050] To measure mRNA levels, the starting material is typically total RNA or poly(A+) RNA isolated from biological samples (e.g., cell suspensions—from peripheral blood, bone marrow, etc., or homogenized tissues, such as homogenized biopsy samples, extracts, paraffin-embedded homogenates, or OCT-embedded samples). General methods for mRNA extraction are well-known in the art and disclosed in standard textbooks of molecular biology, including Ausubel et al., Current Protocols of Molecular Biology, John Wiley and Sons (1997). RNA isolation can also be performed according to the manufacturer's instructions using purification kits, buffer kits, and proteases from commercial manufacturers. For example, RNA can be isolated from cell suspensions using the Qiagen RNeasy mini column, and from TRIzol-based reagent kits (Invitrogen), MasterPure, etc. TM Complete DNA and RNA Purification Kit (EPICENTRE) TM RNA can be isolated from cell suspensions or homogenized tissue samples using the Paraffin Block RNA Isolation Kit (Ambion, Inc.) or the RNA Stat-60 Kit (Tel-Test).

[0051] Various methods for measuring mRNA levels are known in the art, such as those used in differential gene expression analysis. One representative and convenient type of protocol for measuring mRNA levels is array-based gene expression mapping. Such protocols are hybridization assays, in which nucleic acids are used as “probe” nucleic acids that display the genes to be measured / mapped in the generated map. In these assays, a target nucleic acid sample is first prepared from the initial nucleic acid sample to be measured, where preparation may include labeling the target nucleic acid with a marker—e.g., a member of a signal generation system. After the target nucleic acid sample preparation, the sample is contacted with the array under hybridization conditions, thereby forming a complex between the target nucleic acids that is complementary to the probe sequence attached to the array surface. The presence of the hybridization complex is then detected qualitatively or quantitatively.

[0052] Specific hybridization techniques that can be implemented to generate expression maps used in the subject method include U.S. Patent Nos. 5,143,854; 5,288,644; 5,324,633; 5,432,049; 5,470,710; 5,492,806; 5,503,980; 5,510,270; 5,525,464; 5,547,839; 5,580,732; 5,661,028; 5,800,992, the disclosures of which are incorporated herein by reference; and WO 95 / 21265; WO 96 / 31622; WO 97 / 10365; WO 97 / 27317; EP373 203; and EP 785280. In these methods, an array of "probe" nucleic acids is contacted with target nucleic acids as described above, the array comprising probes expressing the phenotypic determinants being measured. Contact is performed under hybridization conditions, such as stringent hybridization conditions, followed by removal of unbound nucleic acids. As used herein, the term "stringent assay conditions" refers to conditions that produce sufficient complementary nucleic acid (e.g., surface-bound and solution-phase nucleic acid) binding pairs to provide the desired level of specificity in the assay, while having less compatibility with binding members with insufficient complementarity to form binding pairs to provide the desired specificity. Stringent assay conditions are the sum or combination (overall) of hybridization and washing conditions.

[0053] The resulting patterns of hybrid nucleic acids provide information about the expression of each gene that has been detected. This expression information is based on data on whether a gene is expressed and at what level it is typically expressed, i.e., expression profiles (e.g., in the form of transcripts), which can be both qualitative and quantitative.

[0054] Alternatively, non-array-based methods can be used to quantify the levels of one or more nucleic acids in a sample. These include methods based on amplification protocols, such as polymerase chain reaction (PCR)-based assays including quantitative PCR, reverse transcription PCR (RT-PCR), real-time PCR, etc., such as TaqMan® RT-PCR, MassARRAY® system, BeadArray® technology, and Luminex® technology; and those that rely on probe-filter hybridization, such as RNA blotting. In situ examples of some of the nucleic acid sequencing methods listed above are described in the following references: Margulies et al., Nature 2005, 437, 376-80 and cited references therein; Ronaghi et al., Analytical Biochemistry 1996, 242, 84-89 and cited references therein; Shendure et al., Science 2005, 309 1728 and cited references therein; Imelfort et al., Brief Bioinform. 2009, 10, 609-618 and cited references therein; Fox et al., Methods Mol Biol. 2009, 553, 79-108 and its cited references; Appleby et al., Methods Mol Biol. 2009; 513, 19-39 and its cited references; and Morozova et al., Genomics 2008, 92, 255-264 and its cited references, which are incorporated herein by reference for a general description of these methods and specific steps of the methods, including all starting products, reagents and final products of each step.

[0055] For measuring protein levels, the amount or level of peptides in a biological sample is determined. In some embodiments, extracellular protein levels are measured. For example, in some cases, the protein being measured (i.e., the peptide) is a secreted protein (e.g., a cytokine or chemokine), and therefore its concentration can be measured in the extracellular fluid of the biological sample (e.g., the concentration of a protein can be measured in serum). In some embodiments, the concentration is a relative value measured by comparing the level of one protein to another. In other embodiments, the concentration is an absolute measurement of weight / volume or weight / weight hybridization.

[0056] In some cases, cells are removed from a biological sample (e.g., by centrifugation, by adhering cells to a culture dish or plastic, etc.), and then the concentration is measured. In some cases, intracellular protein levels are measured by lysing the cells removed from the biological sample to measure the protein levels in the cellular contents. In some cases, both extracellular and intracellular protein levels are measured by separating the cellular and fluid portions of the biological sample (e.g., by centrifugation), measuring the extracellular protein levels by measuring the protein levels in the fluid portion of the biological sample, and measuring the intracellular protein levels by measuring the protein levels in the cellular portion of the biological sample (e.g., after cell lysis). In some cases, the total protein level (i.e., combined extracellular and intracellular proteins) is measured by lysing the cells of the biological sample to include the intracellular contents as part of the sample.

[0057] In some cases, the concentration of one or more additional proteins may also be measured, and the biomarker concentration may be compared to the level of said one or more additional proteins to provide a standardized value for the biomarker concentration. Any convenient protocol for assessing protein levels may be used, wherein the level of one or more proteins in the sample being measured is determined.

[0058] While a variety of different methods for determining protein levels are known to those skilled in the art and any convenient method can be used, a representative and convenient type of protocol for determining protein levels is ELISA—an antibody-based method. In ELISA and ELISA-based assays, one or more antibodies specific to the protein of interest are immobilized on a selected solid surface, preferably a surface exhibiting protein affinity, such as the wells of a polystyrene microtiter plate. After washing away incompletely adsorbed material, the assay plate wells are coated with a nonspecific “blocking” protein known to be antigen-neutral to the test sample, such as bovine serum albumin (BSA), casein, or a solution of powdered milk. This allows the nonspecific adsorption sites on the immobilized surface to be blocked, thereby reducing background caused by nonspecific binding of antigens to the surface. After washing away unbound blocking protein, the immobilized surface is contacted with the test sample under conditions favorable to the formation of immune complexes (antigen / antibody). After incubation, the antiserum-contacted surface is washed to remove the non-immune composite material. The occurrence and amount of immune complex formation can then be determined by subjecting the bound immune complexes to a second antibody that is specific to the target and different from the first antibody, and detecting the binding of the second antibody. In some embodiments, the second antibody will have an associated enzyme, such as urease, peroxidase, or alkaline phosphatase, which produces a colored precipitate upon incubation with a suitable chromogenic substrate. After incubation with the second antibody and washing to remove unbound material, the amount of the label is quantified, for example by incubation with a chromogenic substrate such as urea and bromocresol purple in the case of a peroxidase label, or by incubation with 2,2'-azino-di-(3-ethylbenzthiazoline)-6-sulfonic acid (ABTS) and H₂O₂ in the case of a peroxidase label. Quantification is then achieved by measuring the degree of color formation, for example, using a visible spectrophotometer.

[0059] The aforementioned approach can be modified by first binding the sample to the assay plate. Then, the primary antibody is incubated with the assay plate, and the bound primary antibody is detected using a labeled secondary antibody specific to the primary antibody. The solid substrate immobilizing one or more antibodies can be made from a variety of materials and in over 30 different shapes, such as microtiter plates, microbeads, dipsticks, resin particles, etc. The substrate can be selected to maximize the signal-to-noise ratio, minimize background binding, facilitate separation, and reduce costs. Washing can be performed in a manner most suitable for the substrate used, for example, by removing beads or dipsticks from the reservoir, emptying or diluting the reservoir (e.g., microtiter plate wells), or rinsing the beads, particles, column, or filter with a washing solution or solvent.

[0060] Alternatively, non-ELISA-based methods can be used to measure the levels of one or more proteins in a sample. Representative exemplary methods include, but are not limited to, antibody-based methods (e.g., Western blotting, proteomics arrays, xMAP™ microsphere technology (e.g., LuMinex® technology), immunohistochemistry, flow cytometry, etc.) and non-antibody-based methods (e.g., mass spectrometry).

[0061] Biomarkers. As used herein, the term “biomarker” refers to a gene product, i.e., a protein or RNA, whose concentration (i.e., “level”) and enzymatic activity (function) report the activity (level and / or function) of an administered ClpP modulator. This ClpP modulator is also called a ClpP agent. Because some individuals may not respond to treatment with a ClpP agent, biomarkers can be used to determine whether a ClpP agent has the desired effect on an individual (e.g., determining whether an individual has responded to a ClpP agent, determining whether an individual maintains a response to a ClpP agent, and whether an individual is a candidate for treatment with a ClpP agent, etc.). For example, a biomarker that increases in the level of a ClpP agent after administration when an individual responds to it is a “positive biomarker”; a biomarker that decreases in the level of a ClpP agent after administration when an individual responds to it is a “negative biomarker”; and a biomarker that does not change in the level of a ClpP agent after administration when an individual responds to it is a “neutral biomarker”.

[0062] In some implementations, the concentration or level of the biomarker is measured before and after administration of the ClpP agent, and the degree of change or absence is interpreted as whether the administered ClpP agent actually affects the function and / or level of ClpP, and / or whether such blockade has the desired effect (i.e., whether the immune system has been activated in response to contact with or administration of the ClpP agent). In summary, the concentration or level of the biomarker is measured before and after administration of the ClpP agent to the individual, and the degree of change or absence of its level and / or enzyme function (taken in relation to the duration of exposure to the ClpP agent) is interpreted as an indication of whether the individual will “respond” to the ClpP agent, whether the individual has “responded” to the ClpP agent, and / or whether the individual has “maintained a response” to the ClpP agent.

[0063] A “positive biomarker” is a biomarker whose level increases in response to exposure to and / or treatment with a ClpP agent when an individual and / or cells respond to the ClpP agent. Therefore, if the ClpP agent has the desired effect, biological samples isolated from individuals who have been given the ClpP agent show an increased level of the positive biomarker (relative to the level of the same biomarker measured from the same type of biological sample from the same individual prior to administration of the ClpP agent). In some embodiments, when an individual and / or cells respond to the ClpP agent, the level of the positive biomarker increases by approximately 1.5 times or more (e.g., 2 times or more, 2.5 times or more, 3 times or more, 3.5 times or more, 4 times or more, 4.5 times or more, or 5 times or more, 8 times or more, 10 times or more, 15 times or more) in response to exposure to and / or treatment with the ClpP agent.

[0064] Positive biomarkers include, but are not limited to: ClpP, ClpX, ClpXP, H3K27M, LONP, and malic acid 1 (ME1). Other positive biomarkers established by treating cancer cells with compounds of formula I (>2X increase) include: The levels of any combination of the above-mentioned positive biomarkers can be measured and utilized in the subject approach.

[0065] A “negative biomarker” is a biomarker whose level decreases in response to exposure to and / or treatment with a ClpP agent when an individual and / or cells respond to a ClpP agent. Therefore, if a ClpP agent has the desired effect, biological samples isolated from individuals who have been given a ClpP agent show a reduction in the level of a negative biomarker (relative to the level of the same biomarker measured from the same type of biological sample from the same individual prior to administration of the ClpP agent). In some embodiments, when an individual and / or cells respond to an anti-CD47 agent, the level of a negative biomarker decreases by approximately 1.5 times or more (e.g., 2 times or more, 2.5 times or more, 3 times or more, 3.5 times or more, 4 times or more, 4.5 times or more, or 5 times or more, 8 times or more, 10 times or more, 15 times or more) in response to exposure to and / or treatment with a ClpP agent. Negative biomarkers include, but are not limited to: ClpP, ClpX, ClpXP, H3K27M, LONP, and malic acid 1 (ME1). Other negative biomarkers established by treating cancer cells with compounds of formula I (>2X reduction) include: A "neutral biomarker" is a biomarker whose level does not significantly increase or decrease in response to exposure to and / or treatment with a ClpP agent when an individual and / or cells respond to a ClpP agent. The term "neutral biomarker" is used to refer to a protein or RNA whose level is expected to change (e.g., because gene levels change in other environments that alter an individual's immune status, such as during an inflammatory response), but which experiments show does not change when ClpP levels and / or function are modulated by the use of a ClpP agent. Therefore, if a ClpP agent has the desired effect, biological samples isolated from individuals who have been given a ClpP agent will exhibit similar levels of neutral biomarkers (relative to the levels of the same biomarker measured from the same type of biological sample from the same individual before or before a standardized control). In some implementations, when an individual and / or cell responds to a ClpP agent, the level of the neutral biomarker changes by less than about 5-fold (e.g., less than about 4.5-fold, less than about 4-fold, less than about 3.5-fold, less than about 3-fold, less than about 2.5-fold, less than about 2-fold, or less than about 1.5-fold) in response to contact with and / or treatment with the ClpP agent. Neutral biomarkers include, but are not limited to, ClpP, ClpXP, ClpX, H3K27M, LONP, and malicase 1 (ME1). Furthermore, for neurodegenerative diseases, α-synuclein and α-synuclein A53T (mutant) may be used. Levels of any combination of the above-described neutral biomarkers may be measured and utilized in the subject method.

[0066] Chemical related definitions Structures can be described interchangeably using chemical names, common names, and chemical structures. If there is ambiguity between a chemical structure and a chemical name, and between the structure and the name, the structure shall prevail. Unless otherwise stated, these definitions apply whether the terms are used alone or in combination with other terms. Therefore, the definition of "alkyl" applies to the "alkyl" portion of "hydroxyalkyl," "fluoroalkyl," "-O-alkyl," etc.

[0067] Unless otherwise stated, the following terms used herein and throughout this disclosure shall be understood to have the following meanings: As used herein, the term "therapeutic effective amount" means the amount by which a compound of formula (I) and / or other therapeutic agents, or combinations thereof, effectively produce the desired therapeutic, ameliorative, inhibitory, or preventative effect when administered to a patient with cancer or other disease or disorder of unwanted cell proliferation. In the combination therapy of the present invention, the therapeutic effective amount may refer to each individual agent or a combination as a whole, wherein the amounts of all agents administered together are effective, but wherein the component agents of the combination may be present individually without being in an effective amount. With respect to the treatment of cancer, the therapeutic effective amount means an amount that has the following effects: (1) reducing tumor size, (2) inhibiting (i.e., to some extent slowing down, preferably stopping) tumor metastasis, (3) to some extent inhibiting (preferably stopping) tumor growth or tumor invasion and / or (4) to some extent alleviating (or preferably eliminating) one or more signs or symptoms associated with cancer.

[0068] As used in this article, the term "prevention" in relation to cancer or unwanted cell proliferation refers to reducing the likelihood or rate of disease or disorder progression.

[0069] The use of dashed or dotted lines indicates single bonds between the stated molecular segment and other defined molecular segments. For example, choosing Q1 in equation (I) yields the following structure: .

[0070] In another instance, choosing Q2 in equation (I) yields the following structure: .

[0071] In another instance, choosing Q3 in equation (I) yields the following structure: .

[0072] In another instance, choosing Q4 in equation (I) yields the following structure: .

[0073] In another instance, choosing Q5 in equation (I) yields the following structure: .

[0074] In another instance, choosing Q6 in equation (I) yields the following structure: .

[0075] In another instance, choosing Q7 in equation (I) yields the following structure: .

[0076] In another instance, choosing Q8 in equation (I) yields the following structure: .

[0077] In another instance, choosing Q9 in equation (I) yields the following structure: .

[0078] In another instance, choosing Q10 in equation (I) yields the following structure: .

[0079] In another instance, choosing Q11 in equation (I) yields the following structure: .

[0080] In another instance, choosing Q12 in equation (I) yields the following structure: .

[0081] As used herein, the term "alkyl" refers to an aliphatic hydrocarbon group in which one of its hydrogen atoms is replaced by a bond having a specified number of carbon atoms. Alkyl groups can be straight-chain or branched. In addition to the term "alkyl," alkyl groups can also be defined by the number of carbon atoms. Alkyl substituents typically comprise 1 to 20 carbon atoms ("(C1-C20)alkyl"), preferably 1 to 12 carbon atoms ("(C1-C12)alkyl"), more preferably 1 to 8 carbon atoms ("(C1-C8)alkyl"), or 1 to 6 carbon atoms ("(C1-C6)alkyl"), or 1 to 4 carbon atoms ("(C1-C4)alkyl". In various embodiments, alkyl groups comprise 7 to 12 carbon atoms ("(C7-C12)alkyl") or 7 to 20 carbon atoms ("(C7-C20)alkyl"). Non-limiting examples of alkyl groups include methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, n-pentyl, neopentyl, isopentyl, n-hexyl, isohexyl, and neohexyl. Unless otherwise stated, all alkyl groups described herein may optionally be substituted with one or more substituent groups, which are independently chosen. Unless otherwise stated, alkyl groups described herein as substituted alkyl groups (“substituted alkyl”) will be substituted with one or more substituent groups, which are independently chosen. The total number of substituent groups may be equal to the total number of hydrogen atoms on the alkyl moiety, provided that such substitution is chemically meaningful. Optionally substituted alkyl groups (“optionally substituted alkyl”) typically contain 1 to 6 optional substituents, preferably 1 to 4 optional substituents, and more preferably 1 to 3 optional substituents. For example, the optionally substituted ethyl group is "optionally substituted (C2) alkyl" or "(C2) optionally substituted alkyl", and the substituted ethyl group is "substituted (C2) alkyl" or "(C2) substituted alkyl".

[0082] Suitable substituents for alkyl, "alkyl", "optionally substituted alkyl", and "substituted alkyl" include, but are not limited to, (C3-C8) cycloalkyl, 3-12-membered heterocyclic, (C6-C12) aryl, 5-12-membered heteroaryl, halo, =O (oxo), =S (thiocarbonyl), =N-CN, =N-OR X =NR X -CN, -C(O)R X -CO2R X -C(O)NR X R Y -SR X -SOR X -SO2R X -SO2NR X R Y -NO2, -NR X R Y -NRX C(O)R y -NR X C(O)NR X R Y -NR X C(O)OR X -NR X SO2R Y -NR X SO2NR X R Y -OR X -OC(O)R X and -OC(O)NR X R Y ; where each R X With R Y Independently, it is hydrogen, (C1-C6)alkyl, (C2-C6)alkenyl, (C2-C6)ynyl, (C3-C6)cycloalkyl, 3-12-membered heterocyclic, (C6-C12)aryl, or 5-12-membered heteroaryl, or R X With R Y They can form 3-12 membered heterocyclic groups or 5-12 membered heteroaryl systems together with the nitrogen atoms to which they are attached, each optionally containing 0, 1, or 2 additional heteroatoms; each R X With R Y Optionally substituted with one to three substituents independently selected from the following: halogen, =O, -CN, -C(O)R', -CO2R', -C(O)NR'2, -SO2R', -NR'2, -OR', wherein each R' is independently hydrogen, (C1-C6)alkyl, (C3-C6)cycloalkyl, or a 3-12 membered heterocyclic group. However, suitable substituents for "substituted alkyl" do not include hydrogen.

[0083] "Alkenyl" means an alkyl group as defined herein, consisting of at least two carbon atoms and at least one carbon-carbon bond. Typically, an alkenyl group has 2 to 20 carbon atoms ("C2-C20"), preferably 2 to 12 carbon atoms ("C2-C12"), more preferably 2 to 8 carbon atoms ("C2-C8"), or 2 to 6 carbon atoms ("C2-C6"), or 2 to 4 carbon atoms ("C2-C4"). Representative examples include, but are not limited to, vinyl, 1-propenyl, 2-propenyl, 1-, 2-, or 3-butenyl. The alkenyl group may be optionally substituted ("optionally substituted alkenyl"). Suitable substituents for the alkenyl group are described herein with respect to "optionally substituted alkyl", "substituted alkyl", and alkyl.

[0084] "Alynyl" means an alkyl group as defined herein, consisting of at least two carbon atoms and at least one carbon-carbon triple bond. An alkynyl group has 2 to 20 carbon atoms ("C2-C20"), preferably 2 to 12 carbon atoms ("C2-C12"), more preferably 2 to 8 carbon atoms ("C2-C8"), or 2 to 6 carbon atoms ("C2-C6"), or 2 to 4 carbon atoms ("C2-C4"). Representative examples include, but are not limited to, ethynyl, 1-propynyl, 2-propynyl, 1-, 2-, or 3-butynyl. Any alkynyl group may be optionally substituted. Suitable substituents for the alkynyl group are described herein with respect to "optionally substituted alkyl", "substituted alkyl", and alkyl.

[0085] As used herein, the term "fluoroalkyl" refers to an alkyl group as defined above, wherein one or more of the hydrogen atoms of the alkyl group are substituted with fluorine. In one embodiment, the fluoroalkyl group has 1 to 6 carbon atoms. In another embodiment, the fluoroalkyl group has 1 to 3 carbon atoms. In yet another embodiment, the fluoroalkyl group is substituted with 1 to 3 fluorine atoms. Non-limiting examples of fluoroalkyl groups include -CH2F, -CHF2, and -CF3. The term "(C1-C3)fluoroalkyl" refers to a fluoroalkyl group having 1 to 3 carbon atoms. The term "(C1)fluoroalkyl" refers to -CH2F, -CHF2, and -CF3.

[0086] As used herein, the term "aryl" refers to an aromatic monocyclic or polycyclic system comprising 6 to about 14 carbon atoms. In one embodiment, the aryl group comprises about 6 to 10 carbon atoms (C6-C10) aryl. In another embodiment, the aryl group is phenyl. Non-limiting examples of aryl groups include phenyl and naphthyl. The aryl group may be optionally substituted. Suitable substituents for aryl groups are described herein with respect to "optionally substituted alkyl", "substituted alkyl", and alkyl.

[0087] As used herein, the term "cycloalkyl" refers to a saturated ring containing a specified number of cyclic carbon atoms and free of heteroatoms. Cycloalkyl substituents typically contain 3 to 8 carbon atoms ("C3-C8")cycloalkyl, preferably 3 to 7 carbon atoms ("C3-C7")cycloalkyl, more preferably 3 to 6 carbon atoms ("C3-C6")cycloalkyl, or 3 to 5 carbon atoms ("C3-C5")cycloalkyl. Non-limiting examples of monocyclic cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl. Unless otherwise stated, all cycloalkyl groups described herein may optionally be substituted with one or more substituent groups, which are independently selected. Unless otherwise stated, cycloalkyl groups described herein as optionally substituted ("optionally substituted cycloalkyl") may be substituted with one or more substituent groups, which are independently selected. Unless otherwise stated, the cycloalkyl group of a substituted cycloalkyl group described herein ("substituted cycloalkyl") will be substituted with one or more substituent groups, which are independently selected. The total number of substituent groups may be equal to the total number of hydrogen atoms on the cycloalkyl moiety, provided that such substitution is chemically meaningful. Optionally substituted cycloalkyl groups typically contain 1 to 6 optional substituents, preferably 1 to 4 optional substituents, and more preferably 1 to 3 optional substituents. For example, an optionally substituted cyclopropyl group is an “optionally substituted (C3)cycloalkyl”, while a substituted cyclopropyl group is a “substituted (C2)cycloalkyl”. In one embodiment, the cycloalkyl group contains 3 to 9 carbon atoms, “(C3-C9)cycloalkyl”. In another embodiment, the substituted cycloalkyl group contains 3 to 9 carbon atoms, “substituted (C3-C9)cycloalkyl”. Suitable substituent groups for cycloalkyl groups are as described herein with respect to “optionally substituted alkyl”, “substituted alkyl”, and alkyl.

[0088] As used herein, the term "cycloalkenyl" refers to a partially unsaturated carbocyclic system containing a specified number of carbon atoms. Cycloalkenyl substituents typically contain 4 to 8 carbon atoms ("(C4-C8)cycloalkenyl"), and preferably 5 to 6 carbon atoms ("(C5-C6)cycloalkenyl"). Non-limiting examples of monocyclic cycloalkenyl groups include cyclobutenyl, cyclopentenyl, cyclohexenyl, and cycloheptenyl. Unless otherwise stated, the cycloalkenyl groups described herein may optionally be substituted with one or more substituent groups, which are independently chosen. The total number of substituent groups may be equal to the total number of hydrogen atoms on the cycloalkenyl moiety, provided that such substitution is chemically meaningful. Optionally substituted cycloalkenyl groups typically contain 1 to 6 optional substituents, preferably 1 to 4 optional substituents, and more preferably 1 to 3 optional substituents. For example, a cyclopentenyl group is "(C5)cycloalkenyl," while an optionally substituted cyclopentenyl is "optionally substituted (C5)cycloalkenyl." In one embodiment, the cycloalkenyl group comprises 4 to 8 carbon atoms, "(C4-C8)cycloalkenyl". Suitable substituent groups for the cycloalkenyl group are as described herein with respect to "optionally substituted alkyl", "substituted alkyl", and alkyl.

[0089] As used herein, the term "cycloalkylalkyl" refers to a cycloalkyl ring, typically (C3-C9)cycloalkyl, which is linked to a base molecule via an alkylene linker "(C1-C6)alkylene" of 1 to 6 carbon atoms. The cycloalkylalkyl group is described by the number of carbon atoms in the carbon ring and the number of carbon atoms in the linker. Unless otherwise stated, the cycloalkylalkyl group described herein may optionally be substituted with one or more substituent groups, which are independently chosen. Unless otherwise stated, a cycloalkylalkyl group described herein as optionally substituted ("optionally substituted cycloalkylalkyl") may be substituted with one or more substituent groups, which are independently chosen. Unless otherwise stated, the cycloalkylalkyl group described herein as substituted ("substituted cycloalkylalkyl") will be substituted with one or more substituent groups, which are independently chosen. The total number of substituent groups may be equal to the total number of hydrogen atoms on the cycloalkylalkyl moiety, provided that such substitution is chemically meaningful. The optionally substituted cycloalkyl group typically contains 1 to 6 optional substituents, preferably 1 to 4 optional substituents, and more preferably 1 to 3 optional substituents. In one embodiment, the cycloalkyl group contains 3 to 9 carbon atoms and the linker alkyl group contains 1 to 6 carbon atoms, "(C3-C9)cycloalkyl(C1-C6)alkyl". For example, the cyclopropylethyl group is "(C3)cycloalkyl(C2)alkyl", while the optionally substituted cyclopropylethyl group is "optionally substituted (C3)cycloalkyl(C2)alkyl". Furthermore, the substituted cyclopropylethyl group is "substituted (C3)cycloalkyl(C2)alkyl". Suitable substituent groups for cycloalkyl alkyl groups are as described herein with respect to "optionally substituted alkyl", "substituted alkyl", and alkyl.

[0090] As used herein, the term "cycloalkenylalkyl" refers to a cycloalkenyl ring, typically (C4-C8)cycloalkenyl, which is linked to a base molecule via an alkylene linker, "(C1-C6)alkylene," of 1 to 6 carbon atoms. The cycloalkenylalkyl group is described by the number of carbon atoms in the carbon ring and the number of carbon atoms in the linker. Thus, the "(C5)cycloalkenyl(C1)alkyl" group is a cyclopentenyl group of the base molecule linked by a methylene group (-CH2-). Unless otherwise stated, the cycloalkenylalkyl group described herein may optionally be substituted with one or more substituent groups, which are independently chosen. The total number of substituent groups may be equal to the total number of hydrogen atoms on the cycloalkenylalkyl moiety, provided that such substitution is chemically meaningful. Optionally substituted cycloalkenylalkyl groups typically contain 1 to 6 optional substituents, preferably 1 to 4 optional substituents, and more preferably 1 to 3 optional substituents. In one embodiment, the cycloalkenyl group comprises 4 to 8 carbon atoms, while the linker alkyl group comprises 1 to 6 carbon atoms, and is a “(C4-C8)cycloalkenyl(C1-C6)alkyl”. For example, the cyclopentenylethyl group is a “(C5)cycloalkenyl(C2)alkyl”, and the optionally substituted cyclopentenylethyl group is an “optionally substituted (C5)cycloalkenyl(C2)alkyl”. Suitable substituents for cycloalkenylalkyl groups are as described herein with respect to “optionally substituted alkyl”, “substituted alkyl”, and alkyl.

[0091] In some cases, the substituted alkyl group may be specifically named by reference to the substituent group. For example, "haloalkyl" refers to an alkyl group in which a specified number of carbon atoms are replaced by one or more halogen substituents and typically contains 1 to 6 carbon atoms and 1, 2, or 3 halogen atoms (i.e., "(C1-C6)haloalkyl"). Thus, (C1-C4)haloalkyl groups include trifluoromethyl (-CF3) and difluoromethyl (-CF2H). Unless otherwise stated, the haloalkyl groups described herein may optionally be replaced by one or more substituent groups, which are chosen independently. The total number of substituent groups (the sum of the number of halogens and any other substituents defined herein) may be equal to the total number of hydrogen atoms on the unsubstituted parent alkyl moiety, provided that such substitution is chemically meaningful. For example, for -CH2CH2CH(OH)CH2CF3, the parent alkyl moiety is N-pentyl (-(CH2)4CH3), with 11 possible substitution positions. This example is not intended to be limiting. Unless otherwise stated, a halogenated alkyl group described herein as optionally substituted (“optionally substituted alkyl halogroup”) may be substituted by one or more substituent groups, which are independently chosen. Unless otherwise stated, a halogenated alkyl group described herein as substituted (“substituted alkyl halogroup”) may be substituted by one or more substituent groups, which are independently chosen. The total number of substituent groups may be equal to the total number of hydrogen atoms on the alkyl halogroup, provided that such substitution is chemically meaningful. Optionally substituted alkyl halogroups typically contain 1 to 6 optional substituents, preferably 1 to 4 optional substituents, more preferably 1 to 3 optional substituents. For example, an optionally substituted propyl halogroup is an “optionally substituted (C3) alkyl halogroup”, while a substituted propyl halogroup is a “substituted (C3) alkyl halogroup”. In one embodiment, the cycloalkyl group contains 1 to 6 carbon atoms, “(C1-C6) alkyl halogroup”. In another embodiment, the substituted haloalkyl group comprises 1 to 6 carbon atoms, and is referred to as "substituted (C1-C6) haloalkyl". Suitable substituents for haloalkyl groups are as described herein with respect to "optionally substituted alkyl" and "substituted alkyl".

[0092] "Alkoxy" refers to a monovalent -O-alkyl group, wherein the alkyl moiety has a specified number of carbon atoms. The alkyl moiety of an alkoxy group can be a straight-chain or branched group. Alkoxy groups typically contain 1 to 8 carbon atoms ("C1-C8")alkoxy", 1 to 6 carbon atoms ("C1-C6")alkoxy", or 1 to 4 carbon atoms ("C1-C4")alkoxy". Non-limiting examples of alkoxy groups include methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, and tert-butoxy. Unless otherwise stated, all alkoxy groups described herein may optionally be substituted with one or more substituent groups, which are independently chosen. Unless otherwise stated, alkoxy groups described herein as optionally substituted ("optionally substituted alkoxy") may be substituted with one or more substituent groups, which are independently chosen. Unless otherwise stated, alkoxy groups described herein as substituted ("substituted alkoxy") will be substituted with one or more substituent groups, which are independently chosen. The total number of substituent groups may be equal to the total number of hydrogen atoms on the alkoxy moiety, provided that such substitution is chemically meaningful. Optionally substituted alkoxy groups typically contain 1 to 6 optional substituents, preferably 1 to 4 optional substituents, and more preferably 1 to 3 optional substituents. For example, an optionally substituted ethoxy group is an “optionally substituted (C2)alkoxy”, while a substituted butoxy group is a “substituted (C4)alkoxy”. In one embodiment, the alkoxy group contains 1 to 6 carbon atoms, “(C1-C6)alkoxy”. In another embodiment, the substituted alkoxy group contains 1 to 6 carbon atoms, “substituted (C1-C6)alkoxy”. Suitable substituent groups for alkoxy groups are as described herein with respect to “optionally substituted alkyl”, “substituted alkyl”, and alkyl.

[0093] "Cycloalkoxy" refers to a monovalent -O-cycloalkyl group, wherein the cycloalkyl moiety has a specified number of carbon atoms. The cycloalkyl moiety of an alkoxy group typically comprises a "(C3-C9)cycloalkoxy" with 3 to 9 carbon atoms, or a "(C3-C6)cycloalkoxy" with 3 to 6 carbon atoms. Non-limiting examples of cycloalkoxy groups include cyclopropoxy, cyclobutoxy, and cyclopentoxy. Unless otherwise stated, all cycloalkoxy groups described herein may optionally be substituted with one or more substituent groups, which are independently chosen. The total number of substituent groups may be equal to the total number of hydrogen atoms on the cycloalkoxy moiety, provided that such substitution is chemically meaningful. Optionally substituted cycloalkoxy groups typically comprise 1 to 6 optional substituents, preferably 1 to 4 optional substituents, more preferably 1 to 3 optional substituents. Suitable substituent groups for cycloalkoxy groups are as described herein with respect to "optionally substituted alkyl," "substituted alkyl," and alkyl.

[0094] The term "haloalkoxy" refers to a monovalent -O-haloalkyl group, wherein the alkyl moiety has a specified number of carbon atoms substituted by one or more halogen substituents, and typically comprises 1 to 6 carbon atoms and 1, 2, or 3 halogen atoms (i.e., "(C1-C6)haloalkoxy"). In some cases, the substituted alkyl group may be specifically named with reference to the substituent group. For example, "haloalkoxy" refers to an alkyl group having a specified number of carbon atoms. Thus, (C1-C4)haloalkoxy groups include trifluoromethoxy (-OCF3). Unless otherwise stated, the haloalkoxy group described herein may be substituted by one or more substituent groups, which are independently chosen. The total number of substituent groups may be equal to the total number of hydrogen atoms on the haloalkyl moiety, provided that such substitution is chemically meaningful. Optionally substituted haloalkoxy groups typically comprise 1 to 3 optional substituents, preferably 1 to 2 optional substituents. In one embodiment, the haloalkoxy group comprises 1 to 6 carbon atoms, "(C1-C6)haloalkoxy". Examples of substituted haloalkoxy groups include 1 to 6 carbon atoms, "(C1-C6) haloalkoxy". Suitable substituent groups for haloalkoxy groups are described herein with respect to "optionally substituted alkyl" and "substituted alkyl".

[0095] As used herein, the term "halo" refers to -F, -Cl, -Br, or -I. In one embodiment, the halogen group is -Cl. In another embodiment, the halogen group is -Br.

[0096] As used herein, the term "halogen" refers to -F, -Cl, -Br, or -I. In one embodiment, the halogen group is -Cl. In another embodiment, the halogen group is -Br.

[0097] As used herein, the term "acyl" refers to a -C(O) alkyl or a -C(O) cycloalkyl group. The alkyl group can be straight-chain or branched. The alkyl substituent of the acyl group typically contains 1 to 20 carbon atoms, preferably 1 to 12 carbon atoms, more preferably 1 to 8 carbon atoms, 1 to 6 carbon atoms, or 1 to 4 carbon atoms. The cycloalkyl substituent of the acyl group typically contains 3 to 8 carbon atoms, preferably 3 to 7 carbon atoms, more preferably 3 to 6 carbon atoms, or 3 to 5 carbon atoms. The alkyl and cycloalkyl moieties of the acyl group can be substituted. Suitable substituent groups are described herein with respect to "optionally substituted alkyl," "substituted alkyl," and alkyl.

[0098] The term "aryl" or "aromatic" refers to an optionally substituted monocyclic biaryl or fused bicyclic system with well-known aromatic characteristics, wherein at least one ring comprises a fully conjugated π-electron system. Typically, the aryl group contains 6 to 20 carbon atoms, with "(C6-20)aryl" as the ring member, preferably "(C6-C14)aryl" with 6 to 14 carbon atoms, or more preferably "(C6-C12)aryl" with 6 to 12 carbon atoms. Fused aryl groups may include aryl rings fused to another aryl ring or fused to a saturated or partially unsaturated carbon ring or heterocyclic ring (e.g., a benzene ring). The connection point to the base molecule in such a fused aryl ring system can be a carbon atom of the aromatic portion of the ring system or a carbon or nitrogen atom of the non-aromatic portion. Examples of aryl groups, without limitation, include phenyl, biphenyl, naphthyl, anthracenyl, phenanthrenyl, indenyl, indenyl, and tetrahydronaphthyl. Unless otherwise stated, the aryl groups described herein may optionally be replaced by one or more substituent groups, which are independently chosen. Suitable substituent groups for the aryl groups are further described herein.

[0099] The terms “heteroaryl” or “heteroaromatic” are used interchangeably herein to refer to an aromatic monocyclic or polycyclic ring system comprising about 5 to about 14 ring atoms, wherein 1 to 4 of the ring atoms are independently N, O, or S, and the remaining ring atoms are carbon atoms. These systems have well-known aromatic characteristics. The aromaticity is maintained by the connection of the heteroaryl ring to the base molecule via the ring atoms of the heteroaryl ring. The inclusion of heteroatoms allows for aromaticity in 5-membered and 6-membered rings. In one embodiment, the heteroaryl group has 5 to 10 ring atoms. In another embodiment, the heteroaryl is a monocyclic system and has 5 to 6 ring atoms. In yet another embodiment, the heteroaryl group is a bicyclic system. The term “heteroaryl” also includes a heteroaryl group as defined above fused with a heterocyclic group as defined below. The term “heteroaryl” also includes a heteroaryl group as defined above fused with a benzene, cyclohexadiene, or cyclohexane ring. Non-limiting examples of heteroaryl groups include pyridyl, pyrazinyl, furanyl, thiophene, pyrimidinyl, pyridine (including N-substituted pyridine), and isoaryl. azole group, isothiazol group, azole group, Diazolyl, thiazolyl, pyrazolyl, furyl, pyrroleyl, triazolyl, 1,2,4-thiadiazolyl, pyrazinyl, pyridazinyl, indolyl, quinoxalinyl, phthalazinyl, oxindolyl, imidazo[1,2-a]pyridyl, imidazo[2,1-b]thiazolyl, etc. Unless otherwise stated, the heteroaryl or heteroaromatic groups described herein may optionally be substituted with one or more substituent groups, which are independently chosen. Suitable substituent groups for heteroaryl or heteroaromatic groups are further described herein.

[0100] The terms “heterocyclic group,” “heterocyclic,” or “heterocyclic atom” are used interchangeably herein to refer to a non-aromatic saturated or partially saturated monocyclic or polycyclic ring system comprising 3 to 11 ring atoms, wherein 1 to 4 ring atoms are independently O, S, or N, and the remaining ring atoms are carbon atoms. In one embodiment, the heterocyclic group is monocyclic and has 6 ring atoms, “6-membered heterocycle.” In another embodiment, the heterocyclic group is monocyclic and has 6 ring atoms, wherein 1 or 2 ring atoms are heteroatoms, “6-membered heterocycle containing 1 or 2 heteroatoms.” In another embodiment, the heterocyclic group is monocyclic and has 4 or 5 ring atoms, “4- or 5-membered heterocycle.” In another embodiment, the heterocyclic group has 7, 8, or 9 ring atoms, “7-, 8-, or 9-membered heterocycle.” In another embodiment, the heterocyclic group is bicyclic. The heterocyclic group may be attached to the rest of the molecule via a ring carbon or ring nitrogen atom. The nitrogen or sulfur atom of the heterocyclic group can optionally be oxidized to the corresponding N-oxide, S-oxide, or S,S-dioxide. Any carbon atom with two hydrogen atoms can optionally be oxidized to the corresponding carbonyl group. Non-limiting examples of monocyclic heterocycles include oxetanyl, piperidinyl, pyrrolyl, piperazine, morpholinyl, thiomorpholinyl, thiazolyl, dihydropyranyl, pyran, and 1,4-dioxane. Alkyl, tetrahydrofuranyl, tetrahydrothiophenyl, δ-lactam, δ-lactone, etc. Unless otherwise stated, the heterocyclic groups described herein may optionally be substituted with one or more substituent groups, which are independently chosen. Suitable substituent groups for heterocyclic groups are further described herein. Heterocyclic groups may be unsubstituted or substituted with the same groups applicable to alkyl, aryl, or heteroaryl groups. In one embodiment, the heterocycle comprises 6 atoms and is substituted with 1 to 4 groups as defined herein, “a 6-membered heterocycle substituted with one to four groups.” Furthermore, when specified, the cyclic nitrogen atom may optionally be substituted with groups applicable to amines, such as alkyl, acyl, carbamoyl, sulfonyl, etc., and the cyclic S atom may optionally be substituted with 1 or 2 oxo groups (i.e., S(O)). q (where q is 0, 1, or 2). In one embodiment, a 4- or 5-membered heterocycle is optionally substituted, as given above, “Optionally substituted 4- or 5-membered heterocycles”. In another embodiment, a 7-, 8-, or 9-membered heterocycle is optionally substituted, as given above, “Optionally substituted 7-, 8-, or 9-membered heterocycles”.

[0101] Unless otherwise stated, aryl, heteroaryl, and heterocyclic moieties described herein as optionally substituted (“Optionally Substituted”) may be substituted by one or more substituent groups, which are independently chosen. Optionally substituted aryl, heteroaryl, or heterocyclic moieties described herein as substituted (“Substituted”) may be substituted by one or more substituent groups, which are independently chosen. Optionally substituted aryl, heteroaryl, or heterocyclic groups typically contain 1 to 5 optional substituents, sometimes 1 to 4 optional substituents, preferably 1 to 3 optional substituents, or more preferably 1 to 2 optional substituents. Substituted aryl, heteroaryl, or heterocyclic groups contain at least one substituent as described herein and may optionally contain up to 5 independently chosen substituents in total. The substituent groups used are those suitable for the purposes described herein.

[0102] Substituents applicable to aryl, heteroaryl, and heterocyclic groups include, but are not limited to: (C1-C8)alkyl, (C2-C8)alkenyl, (C2-C8)ynyl, (C3-C8)cycloalkyl, 3-12 membered heterocyclic groups, (C6-C12)aryl, 5-12 membered heteroaryl, halogen, =O (oxo), =S (thiocarbonyl), =N-CN, =N-OR X =NR X -CN, -C(O)R X -CO2R X -C(O)NR X R Y -SR X -SOR X -SO2R X -SO2NR X R Y -NO2, -NR X R Y -NR X C(O)R y -NR X C(O)NR X R Y -NR X C(O)OR X -NR X SO2R Y -NR X SO2NR X R Y -OR X -OC(O)R X and -OC(O)NR X R Y ; where each R X With R YIndependently, it is hydrogen, (C1-C6)alkyl, (C2-C6)alkenyl, (C2-C6)ynyl, (C3-C6)cycloalkyl, 3-12-membered heterocyclic, (C6-C12)aryl, or 5-12-membered heteroaryl, or R X With R Y They can form 3-12 membered heterocyclic groups or 5-12 membered heteroaryl systems together with the nitrogen atoms to which they are attached, each optionally containing 0, 1, or 2 additional heteroatoms; each R X With R Y Optionally substituted with one to three substituents independently selected from the following: halogen, =O, -CN, -C(O)R', -CO2R', -C(O)NR'2, -SO2R', -NR'2, -OR', wherein each R' is independently hydrogen, (C1-C6)alkyl, (C3-C6)cycloalkyl, or a 3-12 membered heterocyclic group. However, suitable substituents for "substituted alkyl" do not include hydrogen.

[0103] "Unsubstituted amino" refers to the group -NH2. When the amino group is described as substituted or optionally substituted, the term includes -NR. X R Y Groups in the form of R, wherein each R X With R Y The alkyl substituent is independently selected from hydrogen, (C1-C8)alkyl, (C3-C9)cycloalkyl, alkynyl, heterocyclic, acyl, aryl, heteroaryl, thioacyl, cycloalkylalkyl, arylalkyl, or heteroalkylalkyl, and in each case has a specified number of atoms and is optionally substituted as described herein. Typically, the alkyl substituent on the amine comprises 1 to 8 carbon atoms, preferably 1 to 6 carbon atoms, or more preferably 1 to 4 carbon atoms. The term also includes R wherein... X With R Y Together with the nitrogen atoms to which they are attached, they form 3-12 membered heterocyclic groups or 5-12 membered heteroaryl rings, each of which may optionally be substituted, as described herein with respect to heterocyclic groups or heteroaryl rings, and may contain 1 to 3 additional heteroatoms selected from N, O, and S as ring members, provided that such a ring does not contain consecutive oxygen atoms or consecutive sulfur atoms. As mentioned above, the term is extended to amino residues of another functional group (e.g., -C(O)NR). X R Y -S(O)2NR X R Y (etc.). In one implementation, -NR X R Y R X and R Y ;-C(O)NR X R Y R X and R YThese can form rings (3-12 membered heterocyclic groups or 5-12 membered heteroaromatic rings) together with the nitrogen to which they are attached (“together with the nitrogen to which they are attached”), each ring may optionally be substituted as described herein with respect to heterocyclic groups or heteroaromatic rings, and may contain 1 to 3 additional heteroatoms selected from N, O and S as ring members, provided that such ring does not contain consecutive oxygen atoms or consecutive sulfur atoms). In another embodiment, -NR X R Y R X and R Y ;-S(O)2NR X R Y R X and R Y They can form rings (3-12-membered heterocyclic groups or 5-12-membered heteroaryl rings) together with the nitrogen to which they are attached, each ring may optionally be substituted as described herein with respect to heterocyclic groups or heteroaryl rings, and may contain 1 to 3 additional heteroatoms selected from N, O and S as ring members, provided that such ring does not contain consecutive oxygen atoms or consecutive sulfur atoms.

[0104] Two adjacent substituents on a ring can form a ring together with the atoms they are attached to. The term "can form a ring together with the carbon atoms they are attached to" is defined herein as meaning that two adjacent residues on a ring can combine with the carbon atoms they are attached to form a 4-6 membered heterocyclic group, a 4-6 membered carbocyclic group, or a 4-6 membered heteroaryl ring, each of which can optionally be substituted as described herein with respect to heterocyclic groups or heteroaryl rings. The resulting heterocyclic group or heteroaryl ring can thus contain 1 to 3 additional heteroatoms selected from N, O, and S as ring members (provided that such a ring does not contain consecutive oxygen atoms or consecutive sulfur atoms). Representative examples derived from the phenyl moiety include, but are not limited to, benzofuranyl, benzothiophenyl, indolyl, benzimidazolyl, indazolyl, benzotriazolyl, indazolyl, quinolinyl, isoquinolinyl, cinnolinyl, azaquinazoline, quinoxalinyl, 2,3-dihydro-1H-indenyl, phthalanyl, 2,3-dihydrobenzofuranyl, benzodioxoyl, and benzodioxoyl. Alkyl groups (benzodioxanyl), etc. Representative examples of the resulting heterocyclic rings include, but are not limited to: And so on. Therefore, representative examples of the resulting carbocyclic base rings include, but are not limited to: , , etc.

[0105] Two substituents bonded to a common carbon atom can form a ring together with the carbon atoms to which they are attached. The term "can form a non-aromatic ring with two oxygen atoms together with the carbon atoms to which they are attached" is defined herein as meaning that two alkoxy or two oxygen-substituted alkyl groups can combine with the carbon atoms to which they are attached to form a ring of 4 to 7 atoms containing two oxygen atoms. Representative examples of the resulting heterocycles include, but are not limited to: , etc.

[0106] Two substituents bonded to a common nitrogen atom can form a ring together with the nitrogen atom to which they are attached. The term “can form a ring together with the nitrogen atom” is defined herein as meaning that two residues residing on the nitrogen atom can combine to form a 3-12 membered heterocyclic group, a 3-7 membered carbocyclic group, or a 5-12 membered heteroaryl ring, each of which may optionally be substituted, as described herein with respect to heterocyclic groups or heteroaryl rings. The resulting heterocyclic group or heteroaryl ring may contain 1 to 3 additional heteroatoms selected from N, O, and S as ring members (provided that such a ring does not contain consecutive oxygen or sulfur atoms). Non-limiting examples derived from the nitrogen atom include the following moieties: azetidinyl, pyrrolidinyl, piperidinyl, morpholinyl, 1,4-azathianyl, 1,3,4-triazolyl, tetrazolyl, imidazolyl, etc.

[0107] Two substituents can together form an oxo residue (=O). "R5 and R6 can together form =O" means that the oxygen atom is bonded to a carbon atom with both R5 and R6 residues in a double bond. For A1, which will result in the following substructure, see 1AA. Furthermore, for "R7 and R8 can together form =O" means that the oxygen atom is bonded to a carbon atom with both R7 and R8 residues in a double bond, see 1AB. The term "substituted" refers to the substitution of one or more specified hydrogen atoms by a selection from a specified group, provided that the substitution does not exceed the normal valence of the atoms under normal conditions, and that the substitution produces a stable compound. A "stable compound" or "stable structure" refers to a compound that is stable enough to be isolated from the reaction mixture with useful purity and formulated into an effective therapeutic agent.

[0108] When any substituent or variable appears more than once in any ingredient or compound of formula (I), its definition for each occurrence is independent of its definition for each subsequent occurrence, unless otherwise stated.

[0109] As used herein, the term "purified form" refers to the physical state of a compound after it has been isolated from a synthetic process (e.g., from a reaction mixture), a natural source, or a combination thereof. The term "purified form" also refers to the physical state of a compound after it has been obtained from one or more purification processes (e.g., chromatography, recrystallization, etc.) described herein or known to those skilled in the art.

[0110] The term "alkyl optionally substituted with dye" ("alkyl optionally substituted with dye") means that the alkyl residue can be substituted by a substituent as defined herein with respect to the optionally substituted alkyl residue, and the carbon of the alkyl residue or a suitable substituent substitution can be modified with dye. As part of the dye residue, there may be a linker portion, such as an alkyl chain or a polyether chain. The compounds described when Q is Q2 or Q3 can be coupled with various infrared, fluorescence, phosphorescence, radioactivity, or infrared fluorescence, as shown in Synthesis Scheme 3. The compound shown as SS10 is a valuable intermediate for preparing other diagnostic agents from the compounds of the present invention. The length of the carbon linker, determined by n, can be 1-30, but n=1-5 is preferred. These analogues are prepared as described above using appropriately protecting groups with terminal functionality. The amine terminus of the alkyl chain has particular value as a reactive substance and can readily form many common functional groups, such as amides, carbamates, secondary amines, etc., using acyl chlorides, ketenes, carboxylic acids (with coupling agents). Other terminal residues besides amines can be used to form linkers, such as -SH, -OH, -Cl, -Br, and -I. These terminal residues can be linked to a variety of dyes and imaging agents. Commercially available fluorescent dyes (BroadPharm, Inc., 6625 Top Gun Street, Suite 103, San Diego, CA 92121) contain a variety of easily coupled functional groups and PEG spacers of varying lengths to increase water solubility. These enable efficient biolabeling in imaging and diagnostic research. BroadPharm, Inc. sells reagents including BDP, Cyanide Blue 3, Cyanide Blue 5, Cyanide Blue 5.5, Cyanide Blue 7, fluorescein, and pyrene. This example is not intended to be limiting.

[0111] It should be noted that any carbon and heteroatom with an unsatisfied valence in the text, schemes, examples and tables of this article are assumed to have a sufficient number of hydrogen atoms to satisfy that valence.

[0112] Compounds can be known by one or more names. For example, ONC201 is also TIC10. Other compounds may be referred to by names beginning with "TR". The following examples illustrate the nomenclature for the same compound. For instance, the following names refer to the same compound: TR57, TR-57, Tr57, Tr-57, tr-57, and tr57.

[0113] One or more compounds of the present invention may exist in a non-solventized form as well as in a solvated form with pharmaceutically acceptable solvents such as water, ethanol, etc., and the present invention is intended to include both solvated and non-solventized forms.

[0114] Compounds of formula (I) may contain one or more stereocenters and thus may exist as racemates, racemic mixtures, single enantiomers, diastereomer mixtures, and single diastereomers. Each such asymmetric center independently produces two optical isomers, and it is intended that all possible optical isomers and diastereomers in the mixture, as well as compounds as pure or partially purified compounds, be included within the scope of this invention.

[0115] As used herein, the term "composition" is intended to cover products containing specified amounts of specified ingredients, and any product produced directly or indirectly from a combination of specified amounts of specific specified ingredients.

[0116] In compounds of formula (I) and of formulas 1A, 2A, 3A, 4A, 5A, 6A, 7A, 8A, 9A, 10A, 11A, 12A, 13A, 14A, 15A, and 16A, atoms may exhibit their natural isotopic abundance, or one or more of these atoms may be artificially enriched with specific isotopes having the same number of atoms but different atomic masses or mass numbers from those predominantly found in nature. This invention aims to include all suitable isotopic variants of compounds of formula (I) and of formulas 1A, 2A, 3A, 4A, 5A, 6A, 7A, 8A, 9A, 10A, 11A, 12A, 13A, 14A, 15A, and 16A. Enrichment with specific isotopes can provide advantageous characteristics (one or more), for example, enrichment of deuterium can provide certain therapeutic advantages, such as increased in vivo half-life or reduced dosage. Furthermore, isotope enrichment can enhance the usefulness of compounds in the characterization of biological samples. Compounds enriched with specific isotopes can be prepared using the synthetic methods described herein and methods known to those skilled in the art, by using reagents and starting materials enriched with that specific isotope.

[0117] This document considers prodrugs of the compounds of the present invention. As used herein, the term "prodrug" means a compound that, upon administration to a subject, undergoes a chemical transformation via metabolism or a chemical process to produce a compound of formula (I). Prodrugs may have beneficial properties, such as, but not limited to, enhanced absorption and / or oral bioavailability.

[0118] Compounds of formula (I) can form salts in some cases, and such salts are also within the scope of this invention. Unless otherwise stated, references to compounds of formula (I) herein should be understood to include references to their salts. As used herein, the term "salt (one or more)" means an acidic and / or basic salt formed from inorganic and / or organic acids and bases. Zwitterions (internal salts) are included in the term "salt (one or more)" as used herein (and may be formed, for example, where the R substituent contains an acid moiety, such as a carboxyl group). This document also includes quaternary ammonium salts, such as alkyl ammonium salts. Pharmaceutically acceptable (i.e., non-toxic and physiologically acceptable) salts are preferred, but other salts may also be used, for example, in separation or purification steps that may be employed during the preparation process. Salts of compounds of formula (I) may be formed, for example, by reacting a compound of formula (I) with an equivalent amount of acid or base in a medium, such as a medium that allows the salt to precipitate (e.g., diethyl ether), or by reacting in an aqueous medium and then freeze-drying.

[0119] Exemplary acid addition salts include acetates, ascorbic acid salts, benzoates, benzenesulfonates, bisulfates, borates, butyrates, citrates, camphorates, camphorsulfonates, fumarates, hydrochlorides, hydrobroms, hydroiodates, lactates, maleates, methanesulfonates, naphthalenesulfonates, nitrates, oxalates, phosphates, propionates, salicylates, succinates, sulfates, tartrates, thiocyanates, and toluenesulfonates (also known as tosylates). Additionally, acids generally considered suitable for forming pharmaceutically usable salts from basic pharmaceutical compounds are discussed, for example, by P. Stahl et al., Camille G. (eds.) Handbook of Pharmaceutical Salts. Properties, Selection and Use. (2002) Zurich: Wiley-VCH. This disclosure is incorporated herein by reference.

[0120] Exemplary basic salts include ammonium salts, alkali metal salts such as sodium, lithium, and potassium salts, alkaline earth metal salts such as calcium and magnesium salts, salts formed with organic bases (e.g., organic amines) such as dicyclohexylamine and tert-butylamine, and salts formed with amino acids such as arginine and lysine. The basic nitrogen-containing group can be quaternized with reagents such as lower alkyl halides (e.g., methyl, ethyl, and butyl chlorides, bromides, and iodides), dialkyl sulfates (e.g., dimethyl, diethyl, and dibutyl sulfates), long-chain halides (e.g., decyl, lauryl, and stearyl chlorides, bromides, and iodides), and aralkyl halides (e.g., benzyl and phenethyl bromides).

[0121] The present invention also includes compounds of formula (I) in all their discrete forms.

[0122] This invention provides a method for determining whether a mammal responds to a compound of general formula I, the method comprising: After separating pre-processed biological samples and before separating post-processed biological samples of the same type, an individual is given a compound of formula I, wherein the biological samples are selected from blood samples, serum samples, plasma samples, bone samples, biopsy samples, fine needle aspirates, lymph node aspirates, cyst aspirates, puncture samples, and thoracentesis samples. Determine the levels of the biomarker ClpP in biological samples before and after treatment; When the level of the pre-treatment biomarker is 1.5 times or higher than the normal level, the individual is identified as a candidate for treatment with a compound of formula I, or when the level of either biomarker ClpP is reduced by more than 50% of the pre-treatment biomarker level, the individual is identified as a candidate for treatment with a compound of formula I. Z1-Q Formula I Z1 is: ; Z2 is: ; Q is selected independently from: Q1 Q2 Q3 Q4 Q5 Q6 Q7 Q8 Q9 Q10 Q11 and Q12; Ar1 and Ar2 are independently selected from aryl, heteroaryl, phenylthio, and phenyl groups; Ar1 can be optionally substituted with 1 to 5 J groups; Ar2 is optionally substituted with 1 to 5 JJ groups; J is independently selected from halogen, -CN, (C1-C6) optionally substituted alkyl, (C3-C9) optionally substituted cycloalkyl, (C3-C9) cycloalkyl(C1-C6) alkyl, (C1-C6) haloalkyl, -CF3, -NH2, -NO2, -SH, -SR15, -OH, (C1-C6) optionally substituted alkoxy, -NR17R18, substituted (C3-C9) cycloalkyl(C1-C6) alkyl, (C3-C9) cycloalkyl(C2-C6) alkynyl, (C4-C8) cycloalkenyl, (C4-C8) cycloalkenyl Alkyl (C1-C6), aryl, heteroaryl, heterocyclic, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted heterocyclic, -C(O)OH, -C(O)OR15, -OC(O)OR15, (C2-C6) alkynyl, (C2-C8) alkenyl, (C1-C6) haloalkoxy, -S(O)2OR15, -SO2NR17R18, -S(O)2R15, -NR15S(O)2R16, -C(O)NR17R18, -C(O)R15, and -NR15C(O)R16; JJ is independently selected from hydrogen, halogen, -CN, (C1-C6) optionally substituted alkyl, (C3-C9) optionally substituted cycloalkyl, (C3-C9) cycloalkyl(C1-C6) alkyl, (C1-C6) haloalkyl, -CF3, -NH2, -NO2, -SH, -SR15, -OH, (C1-C6) optionally substituted alkoxy, -NR17R18, substituted (C3-C9) cycloalkyl(C1-C6) alkyl, (C3-C9) cycloalkyl(C2-C6) alkynyl, (C 4-C8)cycloalkenyl, (C4-C8)cycloalkenyl(C1-C6)alkyl, aryl, heteroaryl, -C(O)OH, -C(O)OR15, -OC(O)OR15, (C2-C6)alkynyl, (C2-C8)alkenyl, (C1-C6)haloalkoxy, -S(O)2OR15, -SO2NR17R18, -S(O)2R15, -NR15S(O)2R16, -C(O)NR17R18, -C(O)R15, and -NR15C(O)R16; R1, R2, R3, R4, R5, R6, R7 and R8 are each independently selected from hydrogen, halogens and (C1-C3) optionally substituted alkyl groups; R9, R10, R11 and R12 are each independently selected from hydrogen, halogen, (C3-C6) cycloalkyl and (C1-C6) optionally substituted alkyl; R10 and R11, together with the carbon atoms they are attached to, can form non-aromatic rings with 3 to 6 carbon atoms; R13 is independently selected from hydrogen, (C1-C6) optionally substituted alkyl, (C3-C6) optionally substituted cycloalkyl, (C1-C6) haloalkyl, (C2-C6) optionally substituted alkenyl, (C2-C6) optionally substituted alkynyl, -CN, -S(O)2R15, -NR17R18, -S(O)2R15, -C(NH)NH2, -C(O)R15, ZW, and -C(O)OR15; R14 is independently selected from hydrogen, (C1-C6) optionally substituted alkyl, (C3-C6) cycloalkyl, (C1-C6) haloalkyl, (C2-C6) optionally substituted alkenyl, (C2-C6) optionally substituted alkynyl, -CN, -S(O)2R15, -NR17R18, -S(O)2R15, -C(NH)NH2, -C(O)R15, and -C(O)OR15; R15, R16, R17, R18, R19, R28 and R29 are independently selected from hydrogen and (C1-C6) optionally substituted alkyl groups; R17 and R18, together with the nitrogen atoms they are attached to, can form rings of 3 to 6 atoms; ZW is an alkyl group (C1-C6) optionally substituted with a dye; W1 and W2 are selected independently from: Nitrogen and ; W3 is independently selected from oxygen, -N(R15)-, and sulfur; W4 is independently selected from =C(R14)- and nitrogen; W5 can be independently selected from single key, SS and ; W6 is independently selected from oxygen, sulfur, and -NR. 14 ; A is independently selected from SS and ; G is independently selected from SS and ; M is independently selected from SS and ; E is independently selected from single bonds, SS, and ; SS is selected independently from: , , , , , , , and ; R20, R21, R26 and R27 are each independently selected from hydrogen, halogens and (C1-C6) optionally substituted alkyl groups; R22, R23, R24, and R25 are each independently selected from hydrogen, halogen, -CN, (C1-C6) optionally substituted alkyl, (C3-C9) optionally substituted cycloalkyl, (C3-C9)cycloalkyl(C1-C6)alkyl, (C1-C6) haloalkyl, -NH2, -NO2, -SH, -SR15, -OH, (C1-C6) optionally substituted alkoxy, -NR17R18, substituted (C3-C9)cycloalkyl(C1-C6)alkyl, (C3-C9)cycloalkyl(C2-C6) Alkynyl, (C4-C8)cycloalkenyl, (C4-C8)cycloalkenyl(C1-C6)alkyl, aryl, heteroaryl, -C(O)OH, -C(O)OR15, -OC(O)OR15, (C2-C6)alkynyl, (C2-C8)alkenyl, (C1-C6)haloalkoxy, -S(O)2OR15, -SO2NR17R18, -S(O)2R15, -NR15S(O)2R16, -C(O)NR17R18, -C(O)R15, and -NR15C(O)R16; R22 and R23, together with the carbon atoms they are attached to, can form non-aromatic rings with 3 to 6 carbon atoms; R22 and R23, together with the carbon atoms they are attached to, can form non-aromatic rings with 1-2 oxygen atoms; R24 and R25, together with the carbon atoms they are attached to, can form non-aromatic rings with 1-2 oxygen atoms; R24 and R25, together with the carbon atoms they are attached to, can form non-aromatic rings with 3 to 6 carbon atoms; R30 and R31 are each independently selected from hydrogen and (C1-C6) optionally substituted alkyl groups.

[0123] Embodiments of the present invention include testing the level of in vitro ClpP in samples taken from mammals.

[0124] The embodiments of the present invention include test samples derived from normal tissue, tumor tissue, circulating tumor cells, plasma or whole blood.

[0125] Embodiments of the present invention include samples to be tested derived from tumor tissue or circulating tumor cells.

[0126] Embodiments of the present invention include predicting an effective response of the disease to treatment with a compound of formula I or a pharmaceutically acceptable formulation thereof by a higher level of ClpP in the native sample relative to a standard value or a set of standard values.

[0127] Embodiments of the present invention include predicting an effective response by a lower level of ClpP in a sample relative to a standard value or a set of standard values ​​following treatment with a compound of formula I or a pharmaceutically acceptable formulation thereof.

[0128] Embodiments of this invention include other biomarkers described herein. These include the use of positive biomarkers described herein. Additionally, negative biomarkers may be used in this invention. Furthermore, negative biomarkers described herein may be used.

[0129] This invention provides a compound of formula 1A: Or its pharmaceutically acceptable salt.

[0130] This invention provides a compound of formula 2A: Or its pharmaceutically acceptable salt.

[0131] This invention provides a compound of formula 3A: Or its pharmaceutically acceptable salt.

[0132] This invention provides a compound of formula 4A: Or its pharmaceutically acceptable salt.

[0133] This invention provides a compound of formula 5A: Or its pharmaceutically acceptable salt.

[0134] This invention provides a compound of formula 6A: Or its pharmaceutically acceptable salt.

[0135] This invention provides a compound of formula 7A: Or its pharmaceutically acceptable salt.

[0136] This invention provides a compound of formula 8A: Or its pharmaceutically acceptable salt.

[0137] This invention provides a compound of formula 9A: Or its pharmaceutically acceptable salt.

[0138] This invention provides a compound of formula 10A: Or its pharmaceutically acceptable salt.

[0139] This invention provides a compound of formula 11A: Or its pharmaceutically acceptable salt.

[0140] This invention provides a compound of formula 12A: Or its pharmaceutically acceptable salt.

[0141] This invention provides a compound of formula 13A: Or its pharmaceutically acceptable salt.

[0142] This invention provides a compound of formula 14A: Or its pharmaceutically acceptable salt.

[0143] This invention provides a compound of formula 15A: Or its pharmaceutically acceptable salt.

[0144] This invention provides a compound of formula 16A: Or its pharmaceutically acceptable salt.

[0145] The various free radicals and / or variables of A1, A2, A3, A4, A5, A6, A7, A8, A9, A10, A11, A12, A13, A14, A15 and A16 are as defined herein with respect to equation (I).

[0146] In another embodiment, the present invention provides compounds and pharmaceutically acceptable salts of formulas A1, A2, A3, A4, A5, A6, A7, A8, A9, A10, A11, A12, A13, A14, A15 and A16.

[0147] In another embodiment, the present invention provides compounds and pharmaceutically acceptable salts of formulas A1, A2, A3, A4, A5, A6, A7, A8, A9, A10, A11, A12, A13, A14, A15, and A16, wherein: Z1 is: ; Z2 is: ; Ar1 and Ar2 are independently selected from aryl, heteroaryl, phenylthio, and phenyl groups; Ar1 can be optionally substituted with 1 to 5 J groups; Ar2 is optionally substituted with 1 to 5 JJ groups; J is independently selected from halogen, -CN, (C1-C6) optionally substituted alkyl, (C3-C9) optionally substituted cycloalkyl, (C3-C9) cycloalkyl(C1-C6) alkyl, (C1-C6) haloalkyl, -CF3, -NH2, -NO2, -SH, -SR15, -OH, (C1-C6) optionally substituted alkoxy, -NR17R18, substituted (C3-C9) cycloalkyl(C1-C6) alkyl, (C3-C9) cycloalkyl(C2-C6) alkynyl, (C4-C8) cycloalkenyl, (C4-C8) cycloalkenyl Alkyl (C1-C6), aryl, heteroaryl, heterocyclic, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted heterocyclic, -C(O)OH, -C(O)OR15, -OC(O)OR15, (C2-C6) alkynyl, (C2)-C8) alkenyl, (C1-C6) haloalkoxy, -S(O)2OR15, -SO2NR17R18, -S(O)2R15, -NR15S(O)2R16, -C(O)NR17R18, -C(O)R15, and -NR15C(O)R16; JJ is independently selected from hydrogen, halogen, -CN, (C1-C6) optionally substituted alkyl, (C3-C9) optionally substituted cycloalkyl, (C3-C9) cycloalkyl(C1-C6) alkyl, (C1-C6) haloalkyl, -CF3, -NH2, -NO2, -SH, -SR15, -OH, (C1-C6) optionally substituted alkoxy, -NR17R18, substituted (C3-C9) cycloalkyl(C1-C6) alkyl, (C3-C9) cycloalkyl(C2-C6) alkynyl, (C 4-C8)cycloalkenyl, (C4-C8)cycloalkenyl(C1-C6)alkyl, aryl, heteroaryl, -C(O)OH, -C(O)OR15, -OC(O)OR15, (C2-C6)alkynyl, (C2-C8)alkenyl, (C1-C6)haloalkoxy, -S(O)2OR15, -SO2NR17R18, -S(O)2R15, -NR15S(O)2R16, -C(O)NR17R18, -C(O)R15, and -NR15C(O)R16; R1, R2, R3, R4, R5, R6, R7 and R8 are each independently selected from hydrogen, halogens and (C1-C3) optionally substituted alkyl groups; R9, R10, R11 and R12 are each independently selected from hydrogen, halogen, (C3-C6) cycloalkyl and (C1-C6) optionally substituted alkyl; R10 and R11, together with the carbon atoms they are attached to, can form non-aromatic rings with 3 to 6 carbon atoms; R13 is independently selected from hydrogen, (C1-C6) optionally substituted alkyl, (C3-C6) optionally substituted cycloalkyl, (C1-C6) haloalkyl, (C2-C6) optionally substituted alkenyl, (C2-C6) optionally substituted alkynyl, -CN, -S(O)2R15, -NR17R18, -S(O)2R15, -C(NH)NH2, -C(O)R15, ZW, and -C(O)OR15; R14 is independently selected from hydrogen, (C1-C6) optionally substituted alkyl, (C3-C6) cycloalkyl, (C1-C6) haloalkyl, (C2-C6) optionally substituted alkenyl, (C2-C6) optionally substituted alkynyl, -CN, -S(O)2R15, -NR17R18, -S(O)2R15, -C(NH)NH2, -C(O)R15, and -C(O)OR15; R15, R16, R17, R18, R19, R28 and R29 are independently selected from hydrogen and (C1-C6) optionally substituted alkyl groups; R17 and R18, together with the nitrogen atoms they are attached to, can form rings of 3 to 6 atoms; ZW is an alkyl group (C1-C6) optionally substituted with dye; W1 and W2 are selected independently from: Nitrogen and ; W3 is independently selected from oxygen, -N(R15)-, and sulfur; W4 is independently selected from =C(R14)- and nitrogen; W5 allows independent selection of free single keys, SS, and ; W6 is independently selected from oxygen, sulfur, and -NR. 14 ; A is independently selected from SS and ; G is independently selected from SS and ; M is independently selected from SS and ; E is independently selected from single bonds, SS, and ; SS is selected independently from: , , , , , , , and ; R20, R21, R26 and R27 are each independently selected from hydrogen, halogens and (C1-C6) optionally substituted alkyl groups; R22, R23, R24, and R25 are each independently selected from hydrogen, halogen, -CN, (C1-C6) optionally substituted alkyl, (C3-C9) optionally substituted cycloalkyl, (C3-C9)cycloalkyl(C1-C6)alkyl, (C1-C6) haloalkyl, -NH2, -NO2, -SH, -SR15, -OH, (C1-C6) optionally substituted alkoxy, -NR17R18, substituted (C3-C9)cycloalkyl(C1-C6)alkyl, (C3-C9)cycloalkyl(C2-C6) Alkynyl, (C4-C8)cycloalkenyl, (C4-C8)cycloalkenyl(C1-C6)alkyl, aryl, heteroaryl, -C(O)OH, -C(O)OR15, -OC(O)OR15, (C2-C6)alkynyl, (C2-C8)alkenyl, (C1-C6)haloalkoxy, -S(O)2OR15, -SO2NR17R18, -S(O)2R15, -NR15S(O)2R16, -C(O)NR17R18, -C(O)R15, and -NR15C(O)R16; R22 and R23, together with the carbon atoms they are attached to, can form non-aromatic rings with 3 to 6 carbon atoms; R22 and R23, together with the carbon atoms they are attached to, can form non-aromatic rings with 1-2 oxygen atoms; R24 and R25, together with the carbon atoms they are attached to, can form non-aromatic rings with 1-2 oxygen atoms; R24 and R25, together with the carbon atoms they are attached to, can form non-aromatic rings with 3 to 6 carbon atoms; R30 and R31 are each independently selected from hydrogen and (C1-C6) optionally substituted alkyl groups.

[0148] In another embodiment, the present invention provides compounds and pharmaceutically acceptable salts of formulas A1, A2, A3, A4, A5, A6, A7, A8, A9, A10, A11, A12, A13, A14, A15, and A16, wherein: Z1 is substituted with 0-5 J groups. ; Z2 is replaced by 1-5 JJ groups. .

[0149] In another embodiment, the present invention provides compounds and pharmaceutically acceptable salts of formulas A1, A2, A3, A4, A5, A6, A7, A8, A9, A10, A11, A12, A13, A14, A15, and A16, wherein: Z1 is substituted with one J group. ; Z2 is replaced by 1-5 JJ groups. .

[0150] In another embodiment, the present invention provides compounds and pharmaceutically acceptable salts of formulas A1, A2, A3, A4, A5, A6, A7, A8, A9, A10, A11, A12, A13, A14, A15, and A16, wherein: Z1 is substituted with one J group. ; Z2 is replaced by one JJ group. .

[0151] In another embodiment, the present invention provides compounds and pharmaceutically acceptable salts of formulas A1, A2, A3, A4, A5, A6, A7, A8, A9, A10, A11, A12, A13, A14, A15, and A16, wherein: Z1 is ; Z2 is replaced by 1-5 JJ groups. .

[0152] In another embodiment, the present invention provides compounds and pharmaceutically acceptable salts of formulas A1, A2, A3, A4, A5, A6, A7, A8, A9, A10, A11, A12, A13, A14, A15, and A16, wherein: Z1 is ; Z2 is replaced by one JJ group. .

[0153] In another embodiment, the present invention provides compounds and pharmaceutically acceptable salts of formulas A1, A2, A3, A4, A5, A6, A7, A8, A9, A10, A11, A12, A13, A14, A15, and A16, wherein: Z1 is substituted with one J group. ; Z2 is replaced by one JJ group. ; R5, R6, R7, R8, R9, R10, R11, and R12 are hydrogen; R14 is independently selected from hydrogen, (C1-C6) alkyl, and -NH2; W1 and W2 are nitrogen; W3 is independently selected from oxygen and sulfur; W4 is independently selected from nitrogen and carbon; W5 is independently selected from single key, and ; W6 is independently selected from oxygen, sulfur, and NH2; R13 is independently selected from hydrogen and (C1-C6) alkyl groups; A is ; G is selected independently and ; M is selected independently and ; E is independently selected from single bonds, and ; R14 is independently selected from hydrogen, (C1-C6) alkyl, and NH2; R19 is independently selected from hydrogen and (C1-C6) alkyl groups.

[0154] The methods for treating cancer described herein include methods for treating cancer in a subject, including administering effective amounts of compounds of formula 1A, 2A, 3A, 4A, 5A, 6A, 7A, 8A, 9A, 10A, 11A, 12A, 13A, 14A, 15A, and 16A, or pharmaceutically acceptable salts thereof.

[0155] The pharmaceutical compositions described herein comprise compounds of formulas 1A, 2A, 3A, 4A, 5A, 6A, 7A, 8A, 9A, 10A, 11A, 12A, 13A, 14A, 15A, and 16A, or pharmaceutically acceptable salts thereof, and pharmaceutically acceptable carriers or excipients.

[0156] This invention also provides treatment for diseases in which activation of ClpP is effective. The methods described herein for treating such diseases will include administering compounds of the formulas 1A, 2A, 3A, 4A, 5A, 6A, 7A, 8A, 9A, 10A, 11A, 12A, 13A, 14A, 15A, and 16A, or pharmaceutically acceptable salts thereof. Additionally, the compounds described herein can be used to treat various neurodegenerative diseases. The methods described herein for treating various neurodegenerative diseases will include administering compounds of the formulas 1A, 2A, 3A, 4A, 5A, 6A, 7A, 8A, 9A, 10A, 11A, 12A, 13A, 14A, 15A, and 16A, or pharmaceutically acceptable salts thereof. Furthermore, the methods described herein for treating Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis, and Alzheimer's disease will include administering compounds of the following formulas: 1A, 2A, 3A, 4A, 5A, 6A, 7A, 8A, 9A, 10A, 11A, 12A, 13A, 14A, 15A, and 16A, or pharmaceutically acceptable salts thereof.

[0157] This invention also provides a treatment for diseases in which a reduction in the concentration and / or activity of ClpX will be effective. The methods described herein for treating such diseases will comprise administering compounds of the following formulas: 1A, 2A, 3A, 4A, 5A, 6A, 7A, 8A, 9A, 10A, 11A, 12A, 13A, 14A, 15A, and 16A, or pharmaceutically acceptable salts thereof. This invention also provides a treatment for diseases in which a reduction in the concentration and / or activity of TUFM will be effective. The methods described herein for treating such diseases will comprise administering compounds of the following formulas: 1A, 2A, 3A, 4A, 5A, 6A, 7A, 8A, 9A, 10A, 11A, 12A, 13A, 14A, 15A, and 16A, or pharmaceutically acceptable salts thereof.

[0158] In one embodiment of the invention, the following compounds are disclosed as activators of the protein ClpP. These compounds are formed by selecting the FA2 fragment and independently selecting fragments FA1 and FA3 to form a single molecule. For FA1, Ar1 is a phenyl group optionally substituted with 1-5 J groups.

[0159] FA1: FA3: FA2: In another embodiment, the compound is FA1-FA2-FA3.

[0160] In another embodiment, preferred compounds of the present invention are Examples 66, 76 and 77.

[0161] This invention provides compounds: , , , , , , , , , , , , , , , , , , , , , , , , , , , Or its pharmaceutically acceptable salt.

[0162] The methods for treating cancer described in this article include methods for treating cancer in the target population, including administering an effective amount of a compound: , , , , , , , , , , , , , , , , , , , , , , , , , , , Or its pharmaceutically acceptable salt.

[0163] This invention discloses the following compounds: , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , Or its pharmaceutically acceptable salt.

[0164] The methods for treating cancer described herein include methods for treating cancer in subjects, including administering an effective amount of a compound: , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , Or its pharmaceutically acceptable salt.

[0165] Another implementation method is to determine whether mammals are responsive to the compound: Or a pharmaceutically acceptable salt thereof that has a responsive method.

[0166] Another implementation method is to determine whether mammals are responsive to the compound: Or a pharmaceutically acceptable salt thereof that has a responsive method.

[0167] Another implementation is a method for treating bacterial infections in a subject, comprising administering an effective amount of a compound of formula I or a pharmaceutically acceptable salt thereof.

[0168] Another implementation is a method for treating bacterial infections in a subject, comprising administering an effective amount of a compound of formula I or a pharmaceutically acceptable salt thereof, wherein: Q is independently selected from Q2, Q3, Q4, Q5, Q6, Q7, Q8, Q9, Q10, Q11, and Q12.

[0169] The compounds of the present invention Dosage form and regimen The administration of the compounds of the present invention can be affected by any method capable of delivering the compound to the site of action. These methods include oral administration, duodenal administration, parenteral administration (including intravenous, subcutaneous, intramuscular, or infusion), local administration, and rectal administration.

[0170] Dosing regimens can be adjusted to provide the optimal desired response. For example, a single bolus can be administered, several separate doses can be administered over time, or the dose can be reduced or increased proportionally according to the urgency of the treatment situation. Formulating parenteral compositions in unit dosage form for ease of administration and uniform dosage is particularly advantageous. As used herein, unit dosage form refers to physically discrete units suitable as a unit dose for a mammalian subject to be treated; each unit contains a predetermined amount of active compound calculated to produce the desired therapeutic effect along with the required drug carrier. The specifications of the unit dosage form of the present invention may be defined by and directly depend on: (a) the unique characteristics of the chemotherapeutic agent and the specific therapeutic or preventative effect to be achieved, and (b) the inherent limitations in the art regarding the sensitivity of such active compounds in the treatment of individuals.

[0171] Therefore, based on the disclosure provided herein, those skilled in the art will understand that dosage and administration regimens can be adjusted according to methods known in the therapeutic field. That is, a maximum tolerated dose can be readily established, and an effective amount to provide a detectable therapeutic benefit to the patient can be determined, as can the time required to administer each agent to provide a detectable therapeutic benefit to the patient. Therefore, while certain dosages and administration regimens are exemplified herein, these examples are by no means limiting to the dosages and administration regimens that can be provided to patients in the practice of this invention. It should be noted that dosage values ​​may vary depending on the type and severity of the condition to be alleviated and may include single or multiple doses. It should be further understood that, for any particular subject, the specific dosage regimen should be adjusted over time according to individual needs and the professional judgment of the person administering or supervising the administration of the composition, and the dosage ranges set forth herein are merely exemplary and not intended to limit the scope or practice of the claimed compositions. For example, dosages may be adjusted based on pharmacokinetic or pharmacodynamic parameters—which may include clinical effects such as toxic effects and / or laboratory values. Therefore, this invention covers intra-patient dose escalation determined by those skilled in the art. Determining appropriate dosages and administration regimens of chemotherapeutic agents is well known in the relevant field and will be understood to be covered by those skilled in the art—following the teachings disclosed herein.

[0172] The amount of the compound of the present invention administered will depend on the patient being treated, the severity of the impairment or condition, the rate of administration, the disposal of the compound, and the prescribing physician's judgment. However, the effective dose is in the range of about 0.001 to about 100 mg / kg body weight / day in a single or divided dose, preferably about 1 to about 35 mg / kg / day. For a 70 kg person, this would equate to about 0.05 to about 7 g / day, preferably about 0.1 to about 2.5 g / day. In some cases, dose levels below the lower limit of the above range may be sufficient, while in others, even larger doses may be used without causing any harmful side effects, provided that such larger doses are first divided into several smaller doses administered throughout the day.

[0173] Formulation and route of administration As used in this article, "pharmaceutically acceptable carrier" refers to a carrier or diluent that does not cause significant irritation to organisms and does not eliminate the biological activity and properties of the active compound.

[0174] Pharmaceutically acceptable carriers can include any conventional drug carrier or excipient. The choice of carrier and / or excipient depends largely on factors such as the specific route of administration, the effect of the excipient on solubility and stability, and the nature of the dosage form.

[0175] Suitable drug carriers include inert diluents or fillers, water, and various organic solvents (such as hydrates and solvates). If desired, the drug composition may contain additional ingredients such as flavoring agents, binders, excipients, etc. Therefore, for oral administration, tablets containing various excipients (such as citric acid) can be used with various disintegrants (such as starch, alginate, and certain complex silicates) and binders (such as sucrose, gelatin, and gum arabic). Examples of excipients, without limitation, include calcium carbonate, calcium phosphate, various sugars and starches, cellulose derivatives, gelatin, vegetable oils, and polyethylene glycol. Furthermore, lubricants such as magnesium stearate, sodium dodecyl sulfate, and talc are commonly used in tablet formulations. Similar types of solid compositions can also be used in soft and hard-filled gelatin capsules. Therefore, non-limiting examples of materials include lactose or milk sugar and high molecular weight polyethylene glycol. When oral administration requires an aqueous suspension or elixir, the active compound therein may be combined with various sweeteners or flavorings, colorants or dyes, and, if necessary, emulsifiers or suspending agents together with diluents such as water, ethanol, propylene glycol, glycerin, or combinations thereof.

[0176] The pharmaceutical composition may be in the following forms, for example, as tablets, capsules, pills, powders, sustained-release preparations, solutions, suspensions or emulsions suitable for oral administration, as ointments or creases suitable for topical administration, or as suppositories suitable for rectal administration.

[0177] Exemplary parenteral administration forms include solutions or suspensions of the active compound in sterile aqueous solutions, such as propylene glycol or dextrose. This dosage form may also be appropriately buffered if desired.

[0178] The pharmaceutical composition may be a single-dose unit dosage form suitable for precise dosage.

[0179] Pharmaceutical compositions suitable for delivering active agents and methods for their preparation are readily apparent to those skilled in the art. Such compositions and methods of preparation can be found, for example, in "Remington's Pharmaceutical Sciences," 19... th The contents of the book are found in Edition (Mack Publishing Company, 1995), and are incorporated herein by reference in their entirety.

[0180] The compounds of the present invention can be administered orally. Oral administration may include swallowing, thereby allowing the compound to enter the gastrointestinal tract, or it may be administered orally or sublingually, allowing the compound to enter the bloodstream directly from the mouth.

[0181] Oral formulations include solid dosage forms such as tablets, capsules containing granules, liquids, or powders. Other formulations include lozenges (including liquid-filled), chewable tablets, multi-particle and nanoparticle formulations, gel solid solutions, liposomes, membranes, ovules, sprays, and liquid formulations.

[0182] Liquid formulations include suspensions, solutions, syrups, and elixirs. These formulations can be used as fillers in soft or hard capsules and typically include a carrier such as water, ethanol, polyethylene glycol, propylene glycol, methylcellulose, or a suitable oil, as well as one or more emulsifiers and / or suspending agents. Liquid formulations can also be prepared by reconstructing solids, such as from sachets.

[0183] The compounds of the present invention can also be used in rapidly soluble and rapidly disintegrating dosage forms, such as those described by Liang and Chen (2001) in Expert Opinion in Therapeutic Patents, 11 (6), 981-986, the disclosure of which is incorporated herein by reference in its entirety.

[0184] For tablet dosage forms, the active agent can comprise 1 wt% to 80 wt% of the dosage form, more typically 5 wt% to 60 wt%. In addition to the active agent, tablets usually contain a disintegrant. Examples of disintegrants include sodium starch glycolate, sodium carboxymethyl cellulose, calcium carboxymethyl cellulose, croscarmellose sodium, crospovidone, polyvinylpyrrolidone, methylcellulose, microcrystalline cellulose, lower alkyl-substituted hydroxypropyl cellulose, starch, pregelatinized starch, and sodium alginate. Typically, the disintegrant can comprise 1 wt% to 25 wt% of the dosage form, preferably 5 wt% to 20 wt%.

[0185] Binders are commonly used to impart cohesive qualities to tablet formulations. Suitable binders include microcrystalline cellulose, gelatin, sugars, polyethylene glycol, natural and synthetic gums, polyvinylpyrrolidone, pregelatinized starch, hydroxypropyl cellulose, and hydroxypropyl methylcellulose. Tablets may also contain diluents such as lactose, mannitol, xylitol, dextrose, sucrose, sorbitol, microcrystalline cellulose, starch, and calcium hydrogen phosphate dihydrate.

[0186] The tablets may optionally include surfactants such as sodium lauryl sulfate and polysorbate 80, and glidants such as silica and talc. When present, the amount of surfactant is typically 0.2 wt% to 5 wt% of the tablet, while the amount of glidant is typically 0.2 wt% to 1 wt% of the tablet.

[0187] Tablets typically also contain lubricants such as magnesium stearate, calcium stearate, zinc stearate, sodium stearate fumarate, and mixtures of magnesium stearate and sodium dodecyl sulfate. The lubricant is typically present in an amount of 0.25 wt% to 10 wt%, preferably 0.5 wt% to 3 wt%, of the tablet.

[0188] An exemplary tablet may contain up to about 80 wt% of an active agent, about 10 wt% to about 90 wt% of a binder, about 0 wt% to about 85 wt% of a diluent, about 2 wt% to about 10 wt% of a disintegrant, and about 0.25 wt% to about 10 wt% of a lubricant.

[0189] Tablet formulations are discussed in detail in “pharmaceutical dosage forms: tablets, Vol. 1” by H. Lieberman and L. Lachman, Marcel Dekker, NY, NY, 1980 (ISBN 0-8247-6918-X), the disclosure of which is incorporated herein by reference in its entirety.

[0190] Suitable improved release formulations are described in U.S. Patent No. 6,106,864. Details of other suitable release technologies, such as high-energy dispersion and permeation, and coated particles, can be found in Verma et al., Pharmaceutical Technology Online 25(2), 1-14 (2001). The disclosure of this reference is incorporated herein by reference in its entirety.

[0191] It should be understood that the compound of formula (I) can be formulated as a disalt.

[0192] Parenteral administration The compounds of this invention can also be administered directly into the bloodstream, muscles, or internal organs. Suitable methods of parenteral administration include intravenous, intra-arterial, intraperitoneal, intrathecal, intraventricular, intraurethral, ​​intracranial, intramuscular, and subcutaneous administration. Devices suitable for parenteral administration include needle injectors, needle-free injectors, and infusion techniques.

[0193] Parenteral preparations are typically aqueous solutions that may contain excipients such as salts, carbohydrates, and buffers (preferably at pH 3 to 9), but for some applications they may be better formulated as sterile non-aqueous solutions or used as a dry form in combination with a suitable medium, such as sterile pyrogen-free water.

[0194] The preparation of parenteral preparations under aseptic conditions, for example by lyophilization, can be easily accomplished using standard pharmaceutical techniques known to those skilled in the art.

[0195] The solubility of the compounds of the present invention used for preparing parenteral solutions can potentially be increased by using appropriate formulation techniques, such as adding solubility enhancers.

[0196] Formulations intended for parenteral administration can be formulated for immediate and / or improved release. Improved release formulations include delayed release, sustained release, pulsatile release, controlled release, targeted release, and programmed release. Therefore, the compounds of the present invention can potentially be formulated as solids, semi-solids, or thixotropic liquids for administration as implantable depots providing improved release of the active compound. Examples of such formulations include drug-coated scaffolds and PGLA microspheres.

[0197] The compounds of the present invention can also potentially be administered topically to the skin or mucous membranes, i.e., dermal or transdermal. Typical formulations for this purpose include gels, hydrogels, lotions, solutions, creams, ointments, powders, dressings, foams, films, skin patches, wafers, implants, sponges, fibrous bandages, and microemulsions.

[0198] dose The amount of active compound administered will depend on the patient being treated, the severity of the impairment or condition, the rate of administration, the disposal of the compound, and the prescribing physician's judgment. However, the effective dose is in the range of about 0.001 to about 100 mg / kg body weight / day in a single or divided dose, preferably about 0.01 to about 35 mg / kg / day. For a human, this would equate to about 0.07 to about 700 mg / day, preferably about 0.7 to about 2500 mg / day. In some cases, dose levels below the lower limit of the above range may be sufficient, while in others, even larger doses may be used without causing any harmful side effects, where such larger doses are typically divided into several smaller doses administered throughout the day.

[0199] Combination therapy As used herein, the term "combination therapy" refers to the sequential or simultaneous administration of the compound of the present invention together with at least one other drug or agent (e.g., an anticancer agent).

[0200] As described above, the compounds of the present invention can potentially be used in combination with one or more other anticancer agents as described below. When using combination therapy, one or more other anticancer agents may be administered sequentially or simultaneously with the compounds of the present invention. In one embodiment, the other anticancer agent is administered to a mammal (subject, patient) prior to administration of the compounds of the present invention. In another embodiment, the other anticancer agent is administered to a mammal after administration of the compounds of the present invention. In yet another embodiment, the other anticancer agent is administered to a mammal concurrently with administration of the compounds of the present invention.

[0201] The present invention also relates to a pharmaceutical composition for treating abnormal cell growth in mammals, including humans, comprising an amount of the compound of the present invention as defined herein, combined with one or more (preferably one to three) anticancer agents selected from antiangiogenic agents and signal transduction inhibitors, and a pharmaceutically acceptable carrier, wherein, when considered as a whole, the amount of the active agent and the combined anticancer agent is therapeutically effective for treating said abnormal cell growth.

[0202] In one embodiment of the invention, the anticancer agent used in combination with the compounds and pharmaceutical compositions of the invention described herein is an anti-angiogenic agent (e.g., an agent that prevents tumors from developing new blood vessels). Examples of anti-angiogenic agents include, for example, VEGF inhibitors, VEGFR inhibitors, TIE-2 inhibitors, PDGFR inhibitors, angiopoietin inhibitors, PKCβ inhibitors, COX-2 inhibitors, integrin, MMP-2 (matrix metalloproteinase 2) inhibitors, and MMP-9 (matrix metalloproteinase 9) inhibitors.

[0203] Preferred anti-angiogenic agents include sunitinib (Sutent®), bevacizumab (Avastin®), axitinib (AG 13736), SU 14813 (Pfizer), and AG13958 (Pfizer).

[0204] Other anti-angiogenic agents include vtalanib (CGP 79787), sorafenib (Nexavar®), pegaptanib octasodium (Macugen®), vandetanib (Zactima®), PF-0337210 (Pfizer), SU 14843 (Pfizer), AZD 2171 (AstraZeneca), ranibizumab (Lucentis®), Neovastat® (AE 941), tetrathiomolyb-data (Coprexa®), AMG 706 (Amgen), VEGF Trap (AVE 0005), CEP 7055 (Sanofi-Aventis), XL880 (Exelixis), telatinib (BAY 57-9352), and CP-868,596 (Pfizer).

[0205] Other anti-angiogenic agents include enzastaurin (LY 317615), midostaurin (CGP 41251), perifosine (KRX 0401), teprenone (Selbex®), and UCN 01 (Kyowa Hakko).

[0206] Other examples of antiangiogenic agents that can be used in combination with the compounds and pharmaceutical compositions of the present invention described herein include celecoxib (Celebrex®), parecoxib (Dynastat®), deracoxib (SC 59046), lumiracoxib (Preige™), valdecoxic (Bextra™), rofecoxib (Vioxx™), iguratimod (Careram®), IP 751 (Invedus), SC-58125 (Pharmacia), and etoricoxib (Arcoxia®).

[0207] Other anti-angiogenic agents include exisulind (Aptosyn®), amigesic®, diflunisal (Dolobid®), ibuprofen (Motrin®), ketoprofen (Orudis®), nabumetone (Relafen®), piroxicam (Feldene®), naproxen (Aleve®, Naprosyn®), diclofenac (Voltarn®), indomethacin (Indocin®), sulindac (Clinoril®), tolectin®, lodine®, toradol®, and Promethazine (Day-pro®).

[0208] Other anti-angiogenic agents include ABT 510 (abbott), apratastat (TMI 005), AZD 8955 (AstraZeneca), incyclinide (Metastat®), and PCK 3145 (Procyon).

[0209] Other anti-angiogenic agents include acitretin (Neotigason®), plitidepsin (Aplidine®), cilengtide (EMD 121974), compretastatin A4 (CA4P), fenvitamin A (4 HPR), tempostatin®, Panzem®, rebimastat (BMS 275291), catumaxomab (Removab®), lenalidomide (Revlimid®), squalene (EVIZON®), thalidomide (Thalomid®), Ukrain® (NSC 631570), Vitaxin® (MEDI 522), and zoledronic acid (Zomata®).

[0210] In another embodiment, the anticancer agent is a so-called signal transduction inhibitor (e.g., inhibiting the way regulatory molecules that govern the fundamental processes of cell growth, differentiation, and survival communicate within the cell). Signal transduction inhibitors include small molecules, antibodies, and antisense molecules. Signal transduction inhibitors include, for example, kinase inhibitors (e.g., tyrosine kinase inhibitors or serine / threonine kinase inhibitors) and cell cycle inhibitors. More specifically, signal transduction inhibitors include, for example, farnesyltransferase inhibitors, EgF inhibitors, ErbB-1 (EGFR) inhibitors, ErbB-2 inhibitors, pan-erb inhibitors, IGF1R inhibitors, MEK(1,2) inhibitors, c-Kit inhibitors, FLT-3 inhibitors, K-Ras inhibitors, PI3 kinase inhibitors, JAK inhibitors, STAT inhibitors, Raf kinase inhibitors, Akt inhibitors, mTOR inhibitors, P70S6 kinase inhibitors, CDK inhibitors, CDK4 / 6 inhibitors, BTK inhibitors of the WNT pathway, and so-called multi-target kinase inhibitors.

[0211] Preferred signal transduction inhibitors include gefitinib (Iressa®), cetuximab (Erbitux®), erlotinib (Tarceva®), trastuzumab (Herceptin®), sunitinib (Sutent®), imatinib (Gleevec®), Trametinib® (GSK1120212), abemaciclib (Verzenio®), palbociclib (Ibrance®), ibrutinib (IMBRUVICA®), acalabrutinib (CALQUENCE®, LOXO-305, and Cobimetinib® (XL518)).

[0212] Further examples of signal transduction inhibitors that can be used in combination with the compounds and pharmaceutical compositions of the present invention described herein include BMS 214662, lonafarnib (Sarasar®), pelitrexol (AG 2037), matuzumab (EMD 7200), nimotuzumab (TheraCIMh-R3®), panitumumab (Vectibix®), vandetanib (Zactima®), pazopanib (SB 786034), BIBW 2992 (Boehringer Ingelheim), and Cervene® (TP38).

[0213] Other examples of signal transduction inhibitors include canertinib (CI 1033), pertuzumab (Omnitarg®), lapatinib (Tycerb®), pelitinib (EKB 569), miltefosine (Miltefosin®), BMS 599626, Lapuleucel-T (Neuvenge®), NeuVax®, Osidem® (IDM 1), mubritinib (TAK-165), panitumumab (Vectibix®), lapatinib (Tycerb®), pelitinib (EKB 569), erbafitinib (Balversa), and pertuzumab (Omnitarg®).

[0214] Other examples of signal transduction inhibitors include ARRY142886, everolimus (Certican®), zotarolimus (Endeavor®), temsirolimus (Torisel®), and VX 680 (Vertex).

[0215] This invention is intended for use with antitumor agents. Antitumor agents include, but are not limited to, hormones, anti-estrogenic agents, histone deacetylase (HDAC) inhibitors, gene silencing or gene activating agents, ribonucleases, proteomics, topoisomerase I inhibitors, camptothecin derivatives, topoisomerase II inhibitors, alkylating agents, antimetabolites, poly(ADP-ribose), polymerase-1 (PARP-1) inhibitors, tubulin inhibitors, antibiotics, spindle inhibitors, platinum coordination compounds, gene therapy agents, antisense oligonucleotides, vascular targeting agents (VTAs), and inhibin derivatives.

[0216] Examples of antitumor agents used in combination therapies employing the compounds of the present invention include, but are not limited to, glucocorticoids such as dexamethasone, prednisone, prednisolone, methylprednisolone, hydrocortisone, and progesters such as medroxyprogesterone acetate, megestrol acetate (Megace), mifepristone (RU-486), selective estrogen receptor modulators (SERMs) such as tamoxifen, raloxifene, lasofoxifene, afimoxifene, azoxifene, bazedoxifene, fispemifene, ormeloxifene, ospemifene, tesmilifene, toremifene, trilostance, and CHF. 4227 (Cheisi), selective estrogen receptor downregulators (SERDs, such as fulvestrant), exemestane (Aromasin®), anastrozole (Arimidex®), atamestane, fadrozole, letrozole (Femara), gonadotropin-releasing hormone (GnRH, often also called luteinizing hormone-releasing hormone [LHRH]) agonists, such as buserelin (Suprefact), goserelin (Zoladex), leuprorelin (… Lupron, triptorelin (Trelstar®), abalexix (Plenaxis®), bicalutamide (Casodex®), cyproterone acetate, flutamide (Eulexin®), medroxyprogesterone acetate, Nilandron, osaterone, dutasteride, epristeride, finasteride, abalexix, goserelin, leuprorelin, triptorelin, bicalutamide, tamoxifen, exemestane, anastrozole, fazodazole, fromestane, letrozole, and combinations thereof.

[0217] Other examples of antitumor agents used in combination with the compounds of this invention include, but are not limited to, suberolanilide (SAHA®, Merck), ester peptide (FR901228), G2M-777, MS-275, pivaloyloxymethyl butyrate, PXD-101 / Onconase® (ranpimase), PS-341, Valcade® (bortezomib), 9-aminocamptothecin, belotecone, BN-80915, camptothecin, diflomotecan, edotecarin, exatecan, gimatecan, 10-hydroxycamptothecin, irinotecan HCl (Camptosar®), lurtotecan, and Orathecin® (rubitecan). Supergen®, SN-38, Topotecan, Camptothecin, 10-Hydroxycamptothecin, 9-Aminocamptothecin, Irinotecan, Aromarubicin, Doxorubicin, Aminofibrate, Aminofibrate, Annamycin, Daunorubicin, Doxorubicin, Exarucin, Epirubicin, Etoposide, Idarubicin, Galarubicin, Hydroxyurea, Nemorubicin, Novantrone, Pirarubicin, Pixantrone, Procarbazine, Rebeccamycin, Sobuzoxane, Taflupoglycine afluposide, valrubicin, Zinecard® (dexrazoxane), nitrogen mustard N-oxide, cyclophosphamide, AMD-473, altretamine, Ap-5280, apaziquone, brostallicin, bendamustine, busulfan, carboquinone, carmustine, chlorambucil, dacarbazine, estradiol, formustine, glufosfamide, ifosfamide, KW-2170, lomustine, malphosphamide, nitrogen mustard, melphalan, dibromomannitol, dibromoeusol, mitomycin C, mitoxantrone, nimustine, ramustine, temozolomide, thiotepa, and platinum-coordinating alkylating agents such as cisplatin.Paraplatin, eptaplatin, lobaplatin, nedaplatin, Eloxatin®, oxaliplatin, satrplatin, and combinations thereof.

[0218] The present invention also contemplates the use of the compounds of the invention in conjunction with: dihydrofolate reductase inhibitors (e.g., methotrexate and NeuTrexin® (trimetresate glucoronate)), purine antagonists (e.g., 6-mercaptopurine nucleoside, mercaptopurine, 6-thioguanine, cladribine, clofarabine) Clolar®, fludarabine, nelarabine, and raltitrexed), and pyrimidine antagonists (e.g., 5-fluorouracil (5-FU), Alimta® (premetrexed disodium), capecitabine (Xeloda®), cytosine, arabinoside, Gemzar® (gemcitabine), Tegafur® (UFT) Orzel® or UForal®, and including the TS-1 combination of tegafur, gimestat and otostat), doxifluridine, carmofluridine, cytarabine (including octadecyl phosphate, phosphate stearate, sustained-release and liposomal forms), enocitabine, 5-azacitidine (Vidaza®), decitabine and etynyl-cytidine), and other antimetabolites such as eflomithine, hydroxyurea, leucovorin, nolatrexed, triapine, trimetrexate, ABT-472, Ino-1001, KU-0687 and GPI18180 and combinations thereof.

[0219] Further examples of antitumor agents used in combination therapies employing the compounds of the present invention, optionally with one or more other agents, include, but are not limited to, Advexin. ®Genasense (oblimersen, Genta®), Combretastatin A4P (CA4P), Oxi4503, AVE-8062, ZD-6126, TZT 1027, Atorvastatin (Lipitor) ® Pravastatin (Pravachol®), lovastatin (Mevacor®), simvastatin (Zocor®), fluvastatin (Lescol®), cerivastatin (Baycol®), rosuvastatin (Crestor®), niacin (Advicor®), caduet, and combinations thereof.

[0220] The present invention also contemplates the use of the compounds of the present invention in conjunction with agents that modulate the immune system, including but not limited to pembrolizumab (Keytruda®), nivolumab (Opdivo®), cimiplimab (Liptayo®), atezolizumab (Tecentrig®), avelumab (Bavencio®), durvalumab (Imfinzi®), ipilimumab (Yervoy®), rituximab (RITUXAN®, Thor-707, and dexamethasone).

[0221] The present invention also contemplates the use of the compounds of the present invention in conjunction with agents that regulate the BCL-2 protein family, including but not limited to venetoclax. ® , ABT-199) and AMG176.

[0222] The present invention also contemplates the use of the compounds of the present invention in conjunction with agents that inhibit androgen receptors, including but not limited to apalutamide (Erleada®, flutamide (Eulexin®), nilandron®, diicalutamide (Casodex®)) and enzalutamide (Xtandi®).

[0223] The present invention also contemplates the use of the compounds of the present invention in conjunction with agents that regulate the PARP protein family, including but not limited to niraparib (Zejula®), olaparib (Lynparza®), rucaparib (Rubraca®), and talazoparib (Talzenna®).

[0224] Another embodiment of the invention of particular interest relates to a method for treating breast cancer in a person requiring such treatment, comprising administering to the person an amount of the compound of the invention in combination with one or more (preferably one to three) anticancer agents selected from trastuzumab, tamoxifen, docetaxel, paclitaxel, capecitabine, gemcitabine, vinorelbine, exmestane, letrozole, and anastrozole.

[0225] Another embodiment of the invention relates to a method of treating neurodegenerative diseases in persons requiring such treatment, comprising administering to the person an amount of the compound of the invention, in combination with one or more agents selected from: anti-taumAb, anti-β-amyloid mAb, BIIB067 (Tofersen), BAN2401, BIIB054 (anti-α-synuclein), BIIB074, BIIB092, BIIB092 (gosuranemab), BIIB104, natalizumab, BIIB076 (anti-tau mAb), BIIB078 (IONIS-C9RX), BIIB080 (IONIS-MAPTRX), BIIB095 (NAV) 1.7), BIIB (XPO1 inhibitor), BIB110, cholinesterase inhibitors (Aricept®, Exelon®, Razadyne®), memantine (Namenda®), levodopa, Lodosyn, dopamine agonists (pramipexole, ropinirole, rotigotine, and apomorphine), MAO B inhibitors (selegiline, rasagiline, safinamide), catechol O-methyltransferase (COMT) inhibitors (entacapone and tolcapone), anticholinergics (benzatropine and trihexphenidyl), amantadine, riluzole, edavarone, xenazine, antipsychotics, and benzodiazepines.

[0226] Treatment methods and uses The present invention further provides treatment methods and uses, including administering the compounds of the present invention, or pharmaceutically acceptable salts thereof, alone or in combination with one or more other therapeutic agents or sedatives. The compositions and methods described herein can be used to treat a variety of disease conditions, including cancer.

[0227] Cancers treated with the methods, compositions, and / or reagents described herein are characterized by abnormal cell proliferation, including but not limited to hyper-proliferation, carcinoma in situ, tumors, and metastases. The methods and compositions described herein can be used to prevent and improve signs and / or symptoms of cancer.

[0228] On the one hand, the compositions and methods described herein are used to treat diseases such as ocular melanoma, desmoplastic round cell tumor, chondrosarcoma, leptomengial disease, diffuse large B-cell lymphoma, acute lymphoblastic leukemia, acute myeloid leukemia, adrenocortical carcinoma, AIDS-related cancer, AIDS-related lymphoma, anal or rectal cancer, appendix cancer, astrocytoma, and atypical teratoid / rhabdoid tumor.

[0229] On the one hand, the compositions and methods described herein are used to treat diseases such as basal cell carcinoma, basal cell nevus syndrome, Gorlin-Nevus Syndrome, cholangiocarcinoma, bladder cancer, bone cancer, osteosarcoma, and malignant fibrous histiocytoma, brain tumors, breast cancer, bronchial tumors, Burkitt lymphoma, and spinal cord tumors.

[0230] On the one hand, the compositions and methods described herein are used to treat diseases such as carcinoid tumors and carcinomas of unknown primary origin. Primary), atypical teratoma / band tumor of the central nervous system, meningeal diseases, embryonal tumor of the central nervous system, lymphoma of the central nervous system, cervical cancer, chordoma, chronic lymphocytic leukemia, chronic myeloid leukemia, chronic myelodysplastic syndrome, colon cancer, colorectal cancer, craniopharyngioma, cutaneous T-cell lymphoma, embryonal tumor of the central nervous system, endometrial cancer, ependymoma, ependymoma, esophageal cancer, Ewing sarcoma, extracranial germ cell tumor, gonadal germ cell tumor, extrahepatic bile duct cancer, eye cancer, gallbladder cancer, gastric cancer, gastrointestinal cancer, gastrointestinal stromal tumor, germ cell tumor, gestational trophoblastic tumor, glioma, hairy cell leukemia, head and neck cancer, hepatocellular carcinoma, histiocytosis, Hodgkin's lymphoma, hypopharyngeal cancer, Kaposi's sarcoma, renal cancer, Langhans cell histiocytosis, laryngeal cancer, lip cancer and oral cavity cancer. Cancer), liver cancer, lung cancer, non-Hodgkin's lymphoma, macroglobulinemia, malignant fibrous histiocytoma and osteosarcoma of bone, medulloblastoma, medullary epithelioma, melanoma, Meckel cell carcinoma, mesothelioma, metastatic squamous neck cancer with occult primary, multiple neoplasia syndrome, mouth cancer, multiple / plasma cell tumors, mycosis fungoides, myelodysplastic syndrome, tumors, multiple myeloma and myelodysplastic disorders.

[0231] On the one hand, the compositions and methods described herein are used to treat cancer.

[0232] The present invention further provides treatment methods and uses, including administering the compounds of the present invention, or pharmaceutically acceptable salts thereof, alone or in combination with one or more therapeutic agents or sedatives.

[0233] On one hand, the present invention provides a method for treating a disease state in which an abnormally high concentration of a ClpP substrate protein is present in the subject, the method comprising administering to the subject a therapeutically effective amount of the compound of the present invention or a pharmaceutically acceptable salt thereof.

[0234] On one hand, the present invention provides a method for treating disease states, including cancer, wherein a decrease in the concentration of a ClpP substrate protein in the subject leads to improvement of the disease, the method comprising administering to the subject a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof.

[0235] On one hand, the present invention provides a method for treating disease states, including cancer, wherein an abnormally high concentration of the protein ClpP is present in the subject, the method comprising administering to the subject a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof.

[0236] On one hand, the present invention provides a method for treating a disease state in which an abnormally low concentration of the protein ClpP is present in the subject, the method comprising administering to the subject a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof.

[0237] On one hand, the present invention provides a method for treating abnormal cell growth in a subject, comprising administering to the subject a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof.

[0238] On the other hand, the present invention provides a method for treating abnormal cell growth in a subject, comprising administering to the subject a quantity of a compound of the present invention, or a pharmaceutically acceptable salt thereof, in combination with a quantity of an antitumor agent, the amounts of which together effectively treat the abnormal growth. In some embodiments, the antitumor agent is selected from mitosis inhibitors, alkylating agents, antimetabolites, intercalating antibiotics, growth factor inhibitors, radiation, cell cycle inhibitors, enzymes, topoisomerase inhibitors, biological response modulators, antibodies, cytotoxics, antihormones, and antiandrogens.

[0239] On the other hand, the present invention provides a method for inhibiting the proliferation of cancer cells in a target, comprising administering to the target a compound of the present invention, or a pharmaceutically acceptable salt thereof, in an amount that effectively inhibits cell proliferation.

[0240] On the other hand, the present invention provides a method for treating cancers selected from solid tumors, liquid tumors, lymphomas, leukemia, or myeloma. In some embodiments, the treatment of cancer includes preventing tumor growth in the cancer subject, including administering the compound of the present invention, or a pharmaceutically acceptable salt thereof, to the subject in an amount that effectively inhibits cell proliferation.

[0241] On the other hand, the present invention provides a method for inhibiting the invasiveness of cancer cells in a target, comprising administering to the target a compound of the present invention, or a pharmaceutically acceptable salt thereof, in an amount that effectively inhibits cell proliferation.

[0242] On the other hand, the present invention provides a method for inducing apoptosis in cancer cells in a subject, comprising administering to the subject a compound of the present invention, or a pharmaceutically acceptable salt thereof, in an amount that effectively inhibits cell proliferation.

[0243] On the other hand, the present invention provides a method for inducing apoptosis in a subject, comprising administering to the subject a compound of the present invention, or a pharmaceutically acceptable salt thereof, in an amount that effectively inhibits cell proliferation.

[0244] In a common implementation of the method provided herein, the abnormal cell growth is cancer, wherein the cancer is selected from basal cell carcinoma, medulloblastoma, liver cancer, rhabdomyosarcoma, lung cancer, bone cancer, pancreatic cancer, skin cancer, head and neck cancer, melanoma of the skin or eye, uterine cancer, ovarian cancer, rectal cancer, cancer of the anal region, stomach cancer, colon cancer, breast cancer, uterine cancer, fallopian tube cancer, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, Hodgkin's disease, esophageal cancer, small bowel cancer, endocrine system cancer, thyroid cancer, parathyroid cancer, adrenal cancer, soft tissue sarcoma, urethral cancer, penile cancer, prostate cancer, chronic or acute leukemia, lymphocytic lymphoma, bladder cancer, kidney cancer or ureteral cancer, renal cell carcinoma, renal pelvis cancer, tumors of the central nervous system (CNS), primary CNS lymphoma, spinal axis tumor, brainstem glioma, pituitary adenoma, or a combination of one or more of the above cancers. In some implementations, the cells are in tissue or tumor, and the tissue or tumor can be in a subject, including a human.

[0245] Cancers treated with the methods and compositions described herein are characterized by abnormal cell proliferation, including but not limited to metastasis, pretumor overgrowth, carcinoma in situ, and tumors. In addition to improving the signs and / or symptoms of cancer, the compounds of the present invention can also be used for prevention. Examples of cancers treated with the compounds of the present invention include, but are not limited to, breast cancer, CNS cancer, colon cancer, prostate cancer, leukemia, lung cancer, and lymphoma.

[0246] On the other hand, the present invention provides a method for treating leukemia selected from the following: acute lymphoblastic leukemia (ALL), chronic lymphocytic leukemia (CLL), chronic myelodysplastic syndrome, hairy cell leukemia, acute myeloid leukemia (AML), chronic myeloid leukemia (CML) and Langerhans cell histiocytosis.

[0247] On the other hand, the present invention provides a method for treating lymphomas selected from the following: diffuse large B-cell lymphoma, AIDS-related lymphoma, cutaneous T-cell lymphoma, Seseli syndrome, mycosis fungoides (MF), histiocytosis, Burkitt lymphoma, central nervous system lymphoma, non-Hodgkin lymphoma, primary central nervous system lymphoma, Hodgkin lymphoma, macroglobulinemia, mycosis fungoides, and lymphoplasmacytic lymphoma.

[0248] On the other hand, the present invention provides a method for treating cancer in a subject, comprising administering to the subject a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof.

[0249] On the other hand, the present invention provides a method for treating cancers selected from the following: vaginal cancer, vulvar cancer, endometrial cancer, cancer of unknown primary origin, and cancer of unknown primary origin.

[0250] On the other hand, the present invention provides a method for treating bacterial infections in a subject, comprising administering to the subject a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof.

[0251] On the other hand, the present invention provides a method for treating Staphylococcus aureus infection in a subject, comprising administering to the subject a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof.

[0252] On the other hand, the present invention provides a method for treating the following neurodegenerative diseases in subjects, including but not limited to Alzheimer's disease, Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis, spinocerebellar ataxia, spinal muscular atrophy, and motor neuron disease, the method comprising administering to the subject a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof.

[0253] On the other hand, the present invention provides a method for treating hematopoietic protoporphyrin (EPP) in a subject, comprising administering to the subject a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof.

[0254] On the other hand, the present invention provides a method for treating subjects with a dominant mutant (ClpX: p.Gly298Asp) of hematopoietic protoporphyrin (EPP), comprising administering to the subject a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof.

[0255] On the other hand, other conditions for which the methods described herein may be applied include, but are not limited to, attention deficit disorder; addiction; epilepsy; viral infection; inflammation; neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, Huntington's disease, and amyotrophic lateral sclerosis; cardiovascular diseases such as coronary artery disease, cardiomyopathy, hypertensive heart disease, heart failure, pulmonary heart disease, cardiac arrhythmia, inflammatory heart disease, endocarditis, inflammatory hypertrophic heart, myocarditis, valvular heart disease, cerebrovascular disease, peripheral artery disease, congenital heart disease, and rheumatic heart disease; diabetes and light chain amyloidosis.

[0256] On the other hand, the present invention provides a method for treating cystic fibrosis.

[0257] On the other hand, the present invention provides a method for treating Perrault syndrome.

[0258] On the other hand, the present invention provides a method for treating type 3 Perrault syndrome.

[0259] On the other hand, the present invention provides a method for treating autoimmune diseases. Autoimmune diseases include, but are not limited to, alopecia areata, antiphospholipids, autoimmune hepatitis, abdominal diseases, type 1 diabetes, Graves' disease, Graves-Barré syndrome, Hashimoto's disease, hemolytic anemia, idiopathic thrombocytopenic purpura, inflammatory bowel disease, inflammatory myopathy, multiple sclerosis, primary biliary cirrhosis, psoriasis, rheumatoid arthritis, scleroderma, Sjögren's syndrome, systemic lupus erythematosus, psoriatic arthritis, Crohn's disease, and vitiligo.

[0260] On the other hand, the present invention provides a method for treating allogeneic transplant rejection. On the other hand, the present invention provides a method for treating hereditary spastic paraplegia.

[0261] On the other hand, the present invention provides a method for treating a condition—acquired immunodeficiency syndrome (AIDS).

[0262] On the other hand, the present invention provides a method for treating HIV and the condition—Acquired Immunodeficiency Syndrome (AIDS).

[0263] On the other hand, the present invention provides a method for treating a condition—pneumonia.

[0264] On the other hand, the present invention provides a method for treating a condition called sepsis.

[0265] On the other hand, the present invention provides a method for treating a condition—a viral infection.

[0266] On the other hand, the present invention provides a method for treating hepatitis in a subject, comprising administering to the subject a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof.

[0267] On the other hand, the present invention provides a method for treating cryptogenic cirrhosis in subjects, comprising administering to the subject a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof.

[0268] On the other hand, the present invention provides a method for treating hepatocyte senescence in a subject, comprising administering to the subject a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof.

[0269] On the other hand, the present invention provides a method for treating non-alcoholic fatty liver disease (NAFLD) in a subject, comprising administering to the subject a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof.

[0270] On the other hand, the present invention provides a method for treating non-alcoholic steatohepatitis (NASH) in a subject, comprising administering to the subject a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof.

[0271] Preparation methods, chemical compounds The compounds of this invention can be prepared by various methods, including standard chemistry. Unless otherwise stated, any previously defined variables will continue to have the meaning they were previously defined for. Illustrative general synthetic methods are listed below, specific compounds of formula (I) are prepared in the examples, and further information regarding the synthesis of these compounds is described in the following citations: Sun H. et al. ACS Med. Chem. Lett. 2019, 10, 191-195 and references cited therein, WO2018 031990 and references cited therein, WO 2018 031987 and references cited therein, CN 1048600948 and references cited therein, and US 8,318,751 and references cited therein.

[0272] There are currently many suppliers of chemical reagents. Examples of chemical suppliers include: Sigma Aldrich, Saint Louis, MO; Alfa Aesar, Tewksbury, MA; TCI America, Portland, OR; BroadPharm, San Diego, CA; and Cambridge BioSciences, Cambridge, UK. This list is by no means limiting. BroadPharm also offers custom services, providing reagents for the synthesis of the compounds of this invention. ONC201 (CAS1616632-77-9) is commercially available from many suppliers, including: MEDCHEM Express, 1 Deer Park Drive, Suite Q, Monmouth Junction, NJ, 08852. 2-(3-iodopropyl)isoindoline-1,3-dione is available from several distributors, including Sigma-Aldrich (Aldrich CPR-R465674). In addition, 2-(4-iodobutyl)isoindoline-1,3-dione is also available from several suppliers, including Sigma-Aldrich (Aldrich CPR-R260312). ONC201 and ONC206 are available from commercial suppliers, including SelleckChem, Houston, TX77014, MedKoo BioSciences, Inc., and Matrix Scientific, Columbia, SC 29224.

[0273] Compounds of general formula (I) can be prepared by methods known in the field of organic synthesis, as listed in the following synthetic schemes section. In all the schemes described below, it is well known that protecting groups are employed, in accordance with general principles of chemistry, for sensitive or responsive groups when necessary. Protecting groups are manipulated according to standard methods of organic synthesis (TW Green and PGMWuts (1991) Protecting Groups in Organic Synthesis, John Wiley & Sons). Those skilled in the art will recognize the presence or absence of a stereocenter in compounds of formula (I). Therefore, the present invention includes all possible stereoisomers and not only mixtures of stereoisomers (such as racemic compounds) but also individual stereoisomers. When a single isomer of the compound is desired, it can be obtained by various methods of isolating the final product or key intermediate, or by stereospecific synthesis using isomerically pure intermediates or methods that impart isomeric purity. These are known to those skilled in the art.

[0274] The compounds were analyzed using methods known to those skilled in the art. NMR, HPLC, and LCMS were used to evaluate the separated compounds and the reaction mixture. LCMS conditions used water and MeCN as two solvents, employing a Symmetry C18, 5 μm, 4.6 x 50 mm column. A linear gradient was used from time 0 (90% H₂O, 10% MeCN, 0.1% TFA) to time 4.5 min (5% H₂O, 95% MeCN, 0.1% TFA). The flow rate was 1.7 mL / min. Evaluation was performed at 254 nm.

[0275] The following solvent, reagent, protecting group, part, and other names can be referred to by their abbreviations: Me: Methyl group; Et: Ethyl; Pr: Propyl; i-Pr: Isopropyl; Bu: Butyl; t-Bu: tert-butyl; Ac: Acetyl group ACN: Acetonitrile AcOH: Acetic acid Aq: water based AUC: Area under the curve BOC or Boc: tert-butoxycarbonyl Conc.: concentrated DMF: Dimethylformamide DMSO: Dimethyl sulfoxide EDCI or EDC: 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide EtOAc: Ethyl acetate EtOH: Ethanol Example: Implementation g: grams h: hours HPLC: High Performance Liquid Chromatography LCMS: Liquid Chromatography-Mass Spectrometry MeOH: Methanol MeI: Methyl iodide MS: Mass spectrometry NA: Not applicable ND: No data report NMR: Nuclear Magnetic Resonance Spectroscopy NT: Not tested Ph: phenyl Ret time: Retention time RT or rt: room temperature Satd, Sat'd, sat'd, and satd.: saturated TFA: Trifluoroacetic acid THF: Tetrahydrofuran Of particular note is the use of toluene analogs as reagents and synthetic intermediates. Numerous commercial sources of toluene analogs are available, which can be used directly or converted into useful reagents or intermediates for the synthesis of the compounds of this invention. Many methods for interconverting toluene analogs to provide reagents and intermediates that can be used to synthesize the compounds of this invention are known to those skilled in the art. The examples described herein include bromination of methyl residues (Ex. 64) and the conversion of functionalized benzyl alcohols to their corresponding bromides (Ex. 79). Furthermore, benzyl alcohols can be converted to the corresponding benzylamines by oxidation to an aldehyde followed by reductive amination. These examples are not limiting.

[0276] Aromatic residues with a single J substituent are intended to represent various J residues (one or more) as described herein and at different positions of the aromatic residues to which they are connected.

[0277] When Q is Q3, the compound described can be prepared as shown in Scheme 1. Furthermore, the scheme used to prepare Ex. 61 can be used to prepare the compounds of the present invention. Those skilled in the art can infer this preparation method to prepare the reagent using the information contained in the references cited herein and common synthetic chemistry knowledge. Particular attention is paid to the information regarding the synthesis of chemically relevant substances in US 8,318,751 and the references cited therein. Furthermore, further synthetic details for preparing the compound when Q is Q3 can be found in WO 2008 / 130584 and the references contained therein. Similarly, the compounds of the present invention when Q is Q10 can be prepared in a similar manner to the compounds when Q is Q3. The chemistry described for the synthesis of the compound when Q is Q3 uses various functionalized piperidine compounds as synthetic intermediates; similarly, the compounds when Q is Q10 can use the same or similar synthetic routes, using various functionalized pyrrolidine compounds as synthetic intermediates.

[0278] Option 1 When Q is Q4, the compound described can be prepared as shown in Scheme 2. Those skilled in the art can infer this preparation method to produce the reagent using the information contained in the references cited herein (Stahl M., et al., Angew. Chem. Int. Ed. 2018, 57, 14,602-14,607 and references cited therein) and common synthetic chemistry knowledge.

[0279] Option 2 When Q is Q2, the compounds described can be prepared as described in WO 2018031990 and the references cited therein. Furthermore, the synthetic methods and schemes described by Ma, Z. (Ma, Z. et al., ACS Med. Chem. Lett. 2019, 10, 191-195 and the references cited therein) and Furrer (US 5,556,854 and the references cited therein) are suitable for preparing the reagents of this invention. Those skilled in the art can infer this preparation method to produce the reagents using the information contained in the references cited herein and common synthetic chemistry knowledge.

[0280] When Q is Q1, the compounds described can be prepared as described in WO 2018031987 and the references cited therein. Numerous other publications describe the synthesis of these reagents, such as: El-Deiry, WS et al., CellCycle 2017, 16, 1790-1799 and the references cited therein. Those skilled in the art can infer this preparation method to produce the reagents using the information contained in the references cited herein and common synthetic chemistry knowledge.

[0281] When Q is Q2, the compounds described can be coupled with various infrared, fluorescent, phosphorescent, radioactive, or infrared fluorescent agents, as shown in Scheme 3. The compound shown as SS10 is a valuable intermediate for preparing other diagnostic agents from the compounds of this invention. The length of the carbon linker, determined by n, can be 1-30, but n = 1-5 is preferred. These analogues are prepared as described above using appropriately protecting groups with terminal functionality. The amine terminus of the alkyl chain has particular value as a reactive substance and can readily form many common functional groups, such as amides, carbamates, and secondary amines, using acyl chlorides, ketenes, carboxylic acids (with coupling agents), etc. Other terminal residues besides amines can be used to form linkers, such as -SH, -OH, -Cl, -Br, and -I. These terminal residues can be coupled with various dyes and imaging agents. Commercially available fluorescent dyes (BroadPharm, Inc., 6625 Top Gun Street, Suite 103, San Diego, CA 92121) contain a variety of easily conjugated functional groups and PEG spacers of varying lengths to increase water solubility. These enable highly efficient biolabeling in imaging and diagnostic research. BroadPharm, Inc. sells reagents including BDP, Cyanide 3, Cyanide 5, Cyanide 5.5, Cyanide 7, fluorescein, and pyrene. This example is not intended to be limiting.

[0282] Further experimental information on the synthesis of coupling dyes can be found in the following references: Wang L. et al., AngewChem Int Ed. 2019 Mar 7. Doi: 10.1002 / anie.201901061 and the references cited therein; Gomes da Costa, S. et al., Morphologie 2019, Mar;103(341):11-16 and the references cited therein; Wei H. et al., Future Med Chem 2018, Dec 6. doi: 10.4155 / fmc-2018-0198 and the references cited therein; Alamudi, SH et al., Chem Commun 2018 Dec 4;54(97): 13641-13653 and the references cited therein; Iliopoulos-Tsoutsouvas C. et al., Expert Opin DrugDiscov 2018 Oct; 13(10):933-947 and the references cited therein, Vernall AJ et al., Br JPharmacol 2014 Mar;171(5):1073-84 and the references cited therein, and Broyles CN et al., Cells 2018 May 31;7(6) and the references cited therein.

[0283] Option 3 The general synthetic route shown in Scheme 4 represents a series of responses that can be used by those skilled in the art to prepare the compounds of the present invention. Substituents X and Y represent various substituents that can be used in this reaction sequence, and their positions on their respective aromatic residues are not limited. Furthermore, more than one substituent may be present on a single aromatic residue. At the heart of this chemical synthetic route is the use of an isocyanate shown herein as SS15. In the case that J is a single chlorine atom and the remaining positions that can be substituted are hydrogen, the desired isocyanate has the chemical formula: C8H6ClNO. Furthermore, it is envisioned that the final step (d) allows for the linkage of various residues identified herein by R. Optional methods of N-alkylation are known to those skilled in the art. For example, SS13 can be prepared from SS11 using the corresponding benzaldehyde and reducing agent. This example is not limited in terms of the number and type of substituents that can be used. Optional reaction conditions known to those skilled in the art can be used for various transformations in Scheme 4.

[0284] Option 4 Compounds were synthesized by scheme 4: (a) DMF, Et3N; (b) sodium carbonate, NH3, ethanol, 70℃ for 5 h; (c) Et3N, toluene, reflux, 80℃ for 8 h; (d) RBr, K2CO3, DMF, 100℃ for 12 h.

[0285] The general synthetic route shown in Scheme 5A is a series of reactions that can be used by those skilled in the art to prepare the compounds of the present invention. Substituents J are independently chosen Y, and their positions in the aromatic system are unrestricted. At the heart of this chemical synthetic route is the use of a two-step synthetic sequence to form a ring. A carbon-nitrogen bond is formed at SS16 to give SS19. Crucially, the SS18 reagent has a protected nucleophile (nitrogen), which, once deprotected, yields SS21, which is now ready to self-condense to form the ring in SS23. SS23 is an example when Q is Q5. These examples are unrestricted in terms of the number and type of substituents that can be used. Optional reaction conditions known to those skilled in the art can be used for various transformations in Scheme 5A.

[0286] Option 5A Compounds were synthesized by scheme 5a: (a) sodium carbonate, DMF, 85℃ for 5 h; (b) CH3NH2, EtOH, reflux, 80℃ for 4 h; (c) pTSA, DMF, iPrOH, 80℃ for 12 h.

[0287] Scheme 5B shows an alternative to the synthesis scheme shown in Scheme 5A for the preparation of the compound of the present invention.

[0288] Option 5B Compounds were synthesized by scheme 5b: (a) 4-Cl-benzylamine, DMF, 85 °C for 5 h; (b) SS25, cat p-TSA, EtOH, reflux, 80 °C for 4 h.

[0289] Scheme 6 illustrates the preparation of an amine-protected alkylating agent. Optional reaction conditions known to those skilled in the art can be used for various transformations in Scheme 6.

[0290] Option 6 Compounds were synthesized via scheme 6: (a) SS29, K2CO3, DMF.

[0291] Scheme 7 illustrates the preparation of various compounds of the present invention using the following key reagents: SS33, SS35, SS37, and SS39. Using the chemistry disclosed herein—with particular attention to transformations such as those shown in Schemes 5A, 5B, and 6—a series of reactions that can be used by those skilled in the art to prepare the compounds of the present invention are shown. Of particular note are the reaction conditions that promote alkylation, such as sodium carbonate, DMF, 85°C for 12 h. J-substituents indicate various substituents that can be used in this reaction sequence, and their positions on the molecule are not limited. This example is not limited in terms of the number and type of substituents that can be used. Optional reaction conditions known to those skilled in the art can be used for various transformations in Scheme 7 to prepare the compounds.

[0292] Option 7 Scheme 8 illustrates the preparation of various compounds of the present invention using the following key reagents: SS41, SS42, SS43, and SS44. Using the chemistry disclosed herein—with particular attention to the reaction sequences shown in Schemes 5a and 5b—a series of reactions that can be used by those skilled in the art to prepare the compounds of the present invention are shown. J-substituents represent various substituents that can be used in this reaction sequence, and their positions on their aromatic systems are not limited. This example is not limited in terms of the number and type of substituents that can be used. Optional reaction conditions known to those skilled in the art can be used in various transformations in Scheme 8 to prepare the compounds.

[0293] Option 8 Scheme 9 illustrates the preparation of various compounds of the present invention, and in particular demonstrates the use of the key synthetic intermediates SS40 and SS45. The terminal olefin of SS40 and the ketone residue of SS45 can be converted into many new analogs under reaction conditions known to those skilled in the art.

[0294] Option 9 Compounds were synthesized by scheme 9: (a) iodomethylzinc iodide, Et2O (Simmons-Smith reaction) or (CH3)2S(O)CH2, DMSO, THF 50℃; (b) O3, CH2Cl2, -78℃, followed by Me2S, and (C) pTSA, DMF, ROH (or HOCH2CH2OH), 80℃ 12 h.

[0295] Scheme 10 is a general synthetic scheme for preparing the compounds of the present invention. This scheme, together with other chemistry disclosed herein and known to those skilled in the art, can be used to prepare compounds with Q=Q6. In particular, the chemistry of schemes 4, 5a, and 5b is applicable to this synthetic route.

[0296] Option 10 Compounds were synthesized by scheme 10: (a) Z2-N=C=O, Et3N, toluene, reflux, 80 °C, 8 h and (b) RBr, K2CO3, DMF, 100 °C, 12 h.

[0297] Scheme 11 is a general synthetic scheme for preparing the compounds of the present invention. This scheme, together with other chemistry disclosed herein and known to those skilled in the art, can be used to prepare compounds with Q=Q6. Note that SS51 is prepared as shown in Scheme 10, using the chemistry described herein, particularly Schemes 5a, 5b, 6, 7 and 8.

[0298] Option 11 Compounds were synthesized by scheme 11: (a) pTSA, DMF, iPrOH, 80℃ for 12 h.

[0299] Schemes 12A and 12B are general synthetic schemes for preparing the compounds of the present invention. These schemes, together with other chemistry disclosed herein and known to those skilled in the art, can be used to prepare compounds with Q=Q8. In particular, the chemistry of Scheme 4 is applicable to this synthetic route. Furthermore, the chemistry described in CN 104860948 and WO 2016 / 184437 can be used. SS53 can be prepared from the corresponding secondary amine by a reductive amination process using the corresponding aldehyde and reducing agent to form the Z1 residue.

[0300] Option 12A Compounds were synthesized by scheme 12A: (a) Z2-NH(CO)Cl, Et3N, toluene, reflux, 80 °C, 8 h and (b) RBr, K2CO3, DMF, 100 °C, 12 h.

[0301] Option 12B Compounds synthesized by scheme 12B are: (a) NH3, t-BuOH, (b) O=N=CH(Ph-JJ), TNF, (c) toluene, Et3N, 100℃ or toluene, cat p-TSA, 100℃, (d) R13-Br, K2CO3, DMF, 100℃, (e) TFA, CH2Cl2 and (f) CH3CN, BrCH2Ph-J, Et3N.

[0302] Schemes 13A and 13B are general synthetic schemes for preparing the compounds of the present invention. These schemes, together with other chemistry disclosed herein and known to those skilled in the art, can be used to prepare compounds with Q=Q9. Note that SS56 is prepared using the chemistry described herein, particularly schemes 5a, 5b, 6, 7, and 8, as shown in scheme 12. Alternatively, SS57 can be prepared using the chemical sequence given in scheme 5b.

[0303] Option 13A According to scheme 13A, the following compounds were synthesized: (a) pTSA, DMF, iPrOFI, 80℃ for 12 h.

[0304] Option 13B Compounds were synthesized by scheme 13B: a) MeI, Et3N, THF, 50 °C; b) J-benzylamine, THF reflux; c) Et3N, toluene reflux; d) TFA, CH2Cl2; e) J-benzyl bromide, Cs2CO3.

[0305] Scheme 14 is a general synthetic scheme for preparing the compounds of the present invention. This scheme, together with other chemistry disclosed herein and known to those skilled in the art, can be used to prepare compounds of formula 8A.

[0306] Option 14 Compounds were synthesized by scheme 14: (a) MeI, Et3N, THF, 50 °C; (b) 4-Cl-benzylamine, THF reflux; Cl(CO)OEt, NaOEt, EtOH, 60 °C; (d) SS60, Et3N, toluene reflux.

[0307] Scheme 15 is a general synthetic scheme for preparing the compounds of the present invention. This scheme, together with other chemistry disclosed herein and known to those skilled in the art, can be used to prepare compounds of formula 9A.

[0308] Option 15 Compounds were synthesized by scheme 15: (a) NH3, cat. NH4Cl, EtOH, reflux; (b) HN=C=O or equivalent, Et3N, toluene reflux; (c) 4-Cl-benzyl bromide, Et3N, DMF heated; (d) K2CO3, MeI, DMF heated.

[0309] Example Chemical Examples Examples of these chemical compounds are shown below. This is in no way intended to be limiting.

[0310] Example 1 D9 is prepared as described in Sieber SA et al., Angew. Chem. Int. Ed. 2008, 57, 14,602-14,607.

[0311] Example 2-27 Examples 2-27 were prepared as described in WO 2018 031987. Examples 28-58 Examples 28-58 were prepared as described in WO 2018 031990 and the references cited therein. Example 57 3-((1-(3-aminopropyl)-2,4-dioxo-3-(4-(trifluoromethyl)benzyl)-1,2,3,4,7,8-hexahydropyridino[4,3-d]pyrimidin-6(5H)-yl)methyl)benzylnitrile Step 1: A mixture of methyl 1-(3-cyanobenzen)-4-oxopiperidin-3-carboxylic acid ester SS26 (8.55 g, 31.4 mmol) and an ammonia solution (7 ml, 25%) in ethanol (110 ml) was heated at 70 °C for 5 h. The solution was concentrated, extracted with DCM (2 x 300 ml), and washed with brine. The extract was dried over Na2SO4 and evaporated under reduced pressure to give 8 g of 2-((4-amino-3-(methoxycarbonyl)-5,6-dihydropyridin-1(2H)-yl)methyl)-4-cyanobenzen-1-ide)INT2 (oil), which was used directly in the next step.

[0312] Step 2: To a solution of INT2 (2 g, 7.4 mmol) in 20 mL of toluene, add 1-(isocyanomethyl)-4-(trifluoromethyl)benzene (1.6 g, 7.5 mmol) and triethylamine (1.1 g, 10.4 mmol). Heat the solution to 80 °C for 8 h. Cool the reaction solution to rt and concentrate under vacuum. Filter the resulting white solid and dissolve it in MeOH (20 mL). Add NaOMe (350 mg) and reflux the mixture overnight. Then remove about 10-15 mL of methanol and filter the precipitate. The desired product, 3-((2,4-dioxo-3-(4-(trifluoromethyl)benzyl)-1,2,3,4,7,8-hexahydropyridino[4,3-d]pyrimidin-6(5H)-yl)methyl)benzyl nitrile, INT2, is given as a pale yellow solid (0.8 g, 25%).

[0313] Step 3: Add potassium carbonate (150 mg) and 2-(3-iodopropyl)isoindoline-1,3-dione (150 mg) to a solution of INT2 (200 mg) in DMF (2 ml). Heat the mixture at 100 °C for 12 h. Add water (approximately 3 ml) and extract the solution with EtOAc (3 x 5 ml). Wash the combined extracts three times with brine (approximately 5 ml), dry over Na2SO4, filter, and concentrate under vacuum to produce the crude product. The purified product INT3, 100 mg, 35% yield, was obtained by preparative TLC.

[0314] Step 4: Add methylamine solution (0.25 ml, 30%) to a solution of product INT3 (100 mg) in EtOH (3 ml). Heat the mixture at 80 °C for 4 h. Add water and extract the solution with DCM (3 x 3 ml). Dry the combined organic extracts with Na2SO4, filter, and concentrate under vacuum to produce the crude product, Example 57. The final product, Example 57, 15 mg, yielded 19%, was obtained by preparative HPLC.

[0315] 1 HNMR (400MHz, CD3OD) δ 2.03 (t, J = 7.2Hz, 2H), 2.99 (t, J = 6.8Hz,2H), 3.18 (s, 2H), 3.67 (s, 2H), 4.01 (t, J = 6.8Hz, 2H), 4.07 (s, 2H), 4.62(s, 2H), 5.17 (s, 2H), 7.5-7.57 (m, 4H), 7.69 (t, J = 8Hz, 1H), 7.86-7.93 (m,2H), 7.99 (s, 1H); LC-MS: m / z = 498.1(M+1). Example 58 3-((1-(4-aminobutyl)-2,4-dioxo-3-(4-(trifluoromethyl)benzyl)-1,2,3,4,7,8-hexahydropyrido[4,3-d]pyrimidin-6(5H)-yl)methyl)benzylnitrile Example 58 was prepared in a manner similar to that of Example 57.

[0316] 1 HNMR (400MHz, CD3OD) δ 1.7 (s, 4H), 2.95 (s, 2H), 3.16 (s, 2H), 3.64(s, 2H), 3.9 (s, 2H), 4.03 (s, 2H), 4.59 (s, 2H), 5.15 (s, 2H), 7.49-7.57 (m,4H), 7.67-7.7 (m, 1H), 7.88 (t, J = 8Hz, 2H), 7.98 (s, 1H); LC-MS: m / z =512.2(M+1). Example 59 3-((1-(4-aminobutyl)-3-(4-chlorobenzyl)-2,4-dioxo-1,2,3,4,7,8-hexahydropyrido[4,3-d]pyrimidin-6(5H)-yl)methyl)benzylnitrile Example 59 was prepared in a manner similar to that of Example 57.

[0317] 1 HNMR (400MHz, CD3OD) δ 1.72 (s, 4H), 2.98-2.99 (d, 2H), 3.15-3.17 (d,2H), 3.61 (t, J=5.6Hz, 2H), 3.91-3.93(d, 2H), 4.01 (s, 2H), 4.57 (s, 2H),5.08 (s, 2H), 7.28-7.3 (d, 2H), 7.35-7.37 (d, 2H), 7.71 (t, J=7.6Hz, 1H),7.9-7.92 (d, 2H), 7.99 (s, 1H). Example 60 11-Benzyl-7-[(2,4-difluorophenyl)methyl]-2,5,7,11-tetraazatricyclo[7.4.0.0] 2,6 [Deca-1(9),5-dien-8-one] Example 60 was prepared as described in WO 2018 031987.

[0318] Example 61 3-({3-[(4-chlorophenyl)methyl]-2-methyl-4-oxo-3H,4H,5H,6H,7H,8H-pyrimidin-6-yl}methyl)benzylnitrile The synthesis of Example 61 was carried out through the following scheme: To a 10 mL three-necked flask, add SS26 (0.4 mmol), acetamidine hydrochloride (0.4 mmol), methanol (3 mL), and K2CO3 (1.2 mmol). Reflux the mixture for 12–15 h. Confirm the reaction is complete by LC-MS. Cool the reaction to room temperature and remove half of the solvent under vacuum. Add water (2 mL) dropwise. A white solid precipitate forms; filter and wash with water. Dry the solid under vacuum to provide INT4 (72% yield).

[0319] To a 10 mL three-necked flask, add INT4 (0.4 mmol), 1-(bromomethyl)-4-chlorobenzene (0.4 mmol), THF (3 mL), and Cs₂CO₃ (1.2 mmol). Reflux the mixture for 12–15 h. Confirm the reaction was complete by LC-MS. Wash the solution with water (100 mL x 2) and brine (100 mL x 1). Dry the combined organic layers in Na₂SO₄ and purify by silica gel column chromatography to provide Example 61 (30% yield).

[0320] 1 HNMR (400MHz, CDOD3) δ 7.78 (s, 1H), 7.72-7.74 (d, J = 8Hz, 1H), 7.65-7.67 (d, J=8Hz, 1H), 7.54 (t, J=8Hz, 1H), 7.34-7.36 (d, J=8Hz, 2H),7.17-7.19 (d, J=8Hz, 2H), 5.32 (s, 2H), 3.81 (s, 2H), 3.41 (s, 2H), 2.81 (t,J = 6Hz, 2H), 2.74 (t, J = 5.2Hz, 2H), 2.46 (s, 3H); LC-MS: m / z = 404.9 (M). Example 62 (TR98) 3-[(8-O-9-{[4-(trifluoromethyl)phenyl]methyl}-1,5,9,11-tetraazatricyclo[8.4.0.0] 2,7 [Tetradecyl-2(7),10-dien-5-yl)methylbenzylnitrile] Example 62 was prepared using the following method: Tetrahydroimidazole 2-thione (59.8 mmol) INT5 was dissolved in methanol (70 ml), and CH3I (89.7 mmol) was added dropwise at 25 °C. After reflux for 30 min, the solvent was removed under vacuum. The residue was suspended in MTBE (50 ml) and filtered. The solid was dried under vacuum to provide a white solid INT6 (yield 83%).

[0321] Compound INT6 (2 mmol) and ((4-trifluoromethyl)phenyl)methylamine (4.2 mmol) were dissolved in a distillate solution. In alkyl (5 ml). The mixture was refluxed for 12 hours. LC-MS confirmed the completion of the reaction. The solvent was removed, and the residue was suspended in toluene for 12 hours. The suspension was filtered and the filter cake was dried under vacuum to provide compound INT7.

[0322] Compound INT7 (0.4 mmol), SS26 (0.4 mmol), methanol (3 mL), and MeONa (1.2 mmol) were charged into a 10 mL three-necked flask. The mixture was refluxed for 12–15 h. The reaction was confirmed to be complete by LC-MS. The reaction was cooled to room temperature. Half of the solvent was removed under vacuum. Water (2 mL) was added dropwise. A white solid precipitate was formed, filtered, and washed with water. The solid was dried under vacuum to provide Example 62 (yield 25%).

[0323] 1 H-NMR (400 MHz, CD3OD): δ 7.64-7.77 (m, 4H), 7.52-7.57 (m, 2H), 7.38-7.45 (m, 2H), 5.25 (s, 1H), 5.20 (s, 1H), 3.72-3.88 (m, 4H), 3.42 (s, 2H), 3.26 (s, 2H), 2.57-2.76 (m, 4H), 1.86-1.91 (m, 2H). LCMS [Mobile phase: from 20% water (0.05% NH3·H2O) and 80% CH3CN (0.05% NH3·H2O) to 5% water (0.05% NH3·H2O) and 95% CH3CN (0.05% NH3·H2O) (linear gradient, C18 column (50 mm, 5 μm, 1 μm injection)) under these conditions, sustained at 0.5 ml / min.] Purity 97.5%, Rt = 3.6 min; MS Calcd.: 479.5. MS measured value: 480.1 [M+1] + ). Example 63 N-[(4-chlorophenyl]-5-[(3-cyanophenyl)methyl]-1,3,4- diazole-2-formamide Example 63 was prepared using the following synthesis scheme: 1 HNMR (400 MHz, DMSO) d6): δ 9.83 (s, 1H), 7.72-7.87 (m, 3H), 7.6 (t, J=8Hz, 1H), 7.38 (t, J=7.2Hz, 4H), 4.44 (t, J=4.8Hz, 4H); LC-MS: m / z = 352.9 (M+) Example 64 7-[(4-chlorophenyl)methyl]-11-[(3-oxo-2,3-dihydro-1H-inden-5-yl)methyl]-2,5,7,11-tetraazatricyclo[7.4.0.0] 2,6 [Deca-1(9),5-dien-8-one] Example 64 was prepared by the following synthesis scheme: 1 HNMR (400MHz, CDCl3) δ 2.46 (s, 3H), 2.75-7.92 (m, 5H), 3.05 (s, 1H), 3.43-3.46 (d, J=12Hz, 1H), 3.62-3.66 (d, J=16Hz, 1H), 4.07 (s, 2H), 4.21 (s,2H), 4.99 (s, 1H), 5.21 (s, 2H), 7.29 (s, 2H), 7.33-7.35 (d, J=8Hz, 2H), 7.53-7.55 (d, J=8Hz, 1H), 7.63-7.64 (d, J=8Hz, 2H); LC-MS: m / z = 460.9 (M+1). Example 65 3-({3-[(4-chlorophenyl)methyl]-4-oxo-3H,4H,5H,6H,7H,8H-pyrido[4,3-d]pyrimidin-6-yl}methylbenzylnitrile Example 65 was prepared by the following synthesis scheme: 1HNMR (400MHz, CDCl3) δ 3.06 (s, 2H), 3.42 (s, 2H), 3.92 (s, 2H), 4.35(s, 2H), 5.03 (s, 2H), 7.24 (s, 2H), 7.33-7.35 (d, J=8Hz, 2H), 7.6 (t, J=8Hz,1H), 7.72-7.81 (m, 3H), 8.14 (s, 1H); LC-MS: m / z = 390.9(M+1) Example 66 (TR108) 3-({8-[(4-chlorophenyl)methyl]-7-oxo-1,4,8,10-tetraazatricyclo[7.3.0.0]) 2,6 Dodeca-2(6),9-dien-4-ylmethylbenzylnitrile Example 66 was prepared by the following synthesis scheme: 1 LC-MS: m / z = 418 (M+1). Example 67 (TR109) 3-[(5-oxo-4-{[4-(trifluoromethyl)phenyl]methyl}-1H,2H,4H,5H,6H,7H,8H,9H-imidazo[1,2-a]quinazolin-7-yl)methyl]benzylnitrile Example 67 was prepared by the following synthesis scheme: 1 HNMR (400MHz, CDCl3) δ 1.36-1.4 (m, 1H), 1.8-1.95 (m, 3H), 2.37-2.75(m, 5H), 3.87-3.97 (m, 4H), 5.1 (s, 2H), 7.39-7.57 (m, 8H); LC-MS: m / z = 465(M+1). Example 68 (TR122) 3-({4-[(4-chlorophenyl)methyl]-5-oxo-1H,2H,4H,5H,6H,7H,8H,9H-imidazo[1,2-a]quinazolin-7-yl}methyl)benzylnitrile Example 68 was prepared using the synthetic sequence described in reference to Example 67.

[0324] 1 HNMR (400MHz, CDCl3) δ 1.32-1.42 (m, 1H), 1.81-1.94 (m, 3H), 2.31-2.74 (m, 5H), 3.86-3.96 (m, 4H), 5.01 (s, 2H), 7.25 (t, J = 5.6Hz, 2H), 7.37-7.45 (m, 5H), 7.51 (t, J = 4Hz, 1H); LC-MS: m / z = 431 (M+1). Example 69 3-({3-[(4-chlorophenyl)methyl]-2-methyl-4-oxo-3H,4H,5H,6H,7H-pyrrolo[3,4-d]pyrimidin-6-yl}methyl)benzylnitrile Example 69 was prepared by the following synthesis scheme: 1 HNMR (400MHz, CDCl3) δ 2.51 (s, 3H), 4.46-4.48 (ss, 6H), 5.26(s, 2H), 7.11-7.13 (d, J=8Hz, 2H), 7.33-7.35 (d, J=8Hz, 2H), 7.63(t, J=8Hz, 1H), 7.74-7.79(m, 2H), 7.85-7.87 (d, J=8Hz, 1H); LC-MS: m / z = 390.9 (M+1). Example 70 3-({9-[(4-chlorophenyl)methyl]-13,13-dimethyl-8-oxo-1,5,9,11-tetraazatricyclo[8.4.0.0]) 2,7 [Tetradecyl-2(7),10-diene-5-yl)methylbenzylnitrile] Example 70 was prepared by the following synthesis scheme: 1 HNMR (400MHz, CDCl3) δ 1.03 (s, 6H), 2.98 (s, 2H), 3.1-3.17 (m, 4H), 3.59-3.68 (m, 4H), 3.75 (s, 2H), 4.15 (s, 2H), 5.25 (s, 2H), 7.28-7.3 (d, J=8Hz, 2H), 7.40-7.42 (d, J=8Hz, 2H), 7.65 (t, J=8Hz, 1H), 7.8-7.82 (d, J=8Hz,1H), 7.86-7.88 (d, J=8Hz, 1H), 7.93 (s 1H); LC-MS: m / z = 473.9 (M+1). Example 71 3-({9-[(4-chlorophenyl)methyl]-13,13-difluoro-8-oxo-1,5,9,11-tetraazatricyclo[8.4.0.0]) 2 ,7 [Tetradecyl-2(7),10-diene-5-yl)methylbenzylnitrile] Example 71 was prepared by the following synthesis scheme: 1 HNMR (400MHz, CDCl3) δ 2.94 (s, 2H), 3.58-3.75 (m, 8H), 4.31 (s, 2H), 5.17(s, 2H), 7.05-7.07 (d, J=8Hz, 1H), 7.26-7.33(m, 3H), 7.59-7.79(m, 4H);LC-MS: m / z = 481.9 (M+1). Example 72 3-({3-[(4-bromophenyl)methyl]-2-methyl-4-oxo-3H,4H,5H,6H,7H,8H-pyrido[4,3-d]pyrimidin-6-yl}methyl)benzylnitrile Example 72 was prepared by the following synthesis scheme: 1HNMR (400MHz, CDCl3) δ 2.04 (s, 3H), 2.43 (s, 4H), 3.46 (s, 2H), 3.75 (s, 2H), 5.21 (s, 2H), 7.05-7.07 (d, J=8Hz, 2H), 7.42-7.47 (m, 3H), 7.56-7.61(m, 2H), 7.7 (s, 1H); LC-MS: m / z = 450.9 (M+1). Example 73 3-[(2-methyl-4-oxo-3-{[4-(trifluoromethyl)phenyl]methyl}-3H,4H,5H,6H,7H,8H-pyrido[4,3-d]pyrimidin-6-yl)methyl]benzylnitrile Example 73 was prepared using the same synthetic route as described in Example 72: LC-MS: m / z = 439.0 (M+1) and retention time 1.743 min. Example 74 3-({3-[(4-bromophenyl)methyl]-4-oxo-3H,4H,5H,6H,7H,8H-pyrido[4,3-d]pyrimidin-6-yl}methyl)benzylnitrile Example 74 was prepared using the method described in Example 65.

[0325] 1 HNMR (400MHz, CDCl3) δ 2.73-2.79 (m, 4H), 3.46 (s, 2H), 3.75 (s, 2H), 5.03 (s, 2H), 7.21-7.23 (d, 2H), 7.43-7.5 (m, 3H), 7.59 (t, J=8.8Hz, 2H), 7.7(s, 1H), 8.06 (s, 1H); LC-MS: m / z = 434.1(M+2). Example 75 3-[(4-oxo-3-{[4-(trifluoromethyl)phenyl]methyl}-3H,4H,5H,6H,7H,8H-pyrido[4,3-d]pyrimidin-6-yl)methyl]benzylnitrile Example 75 was prepared using the method described in Example 65.

[0326] 1 HNMR (400MHz, CDCl3) δ 3.09 (s, 2H), 3.45 (s, 2H), 3.95 (s, 2H), 4.37(s, 2H), 5.13 (s, 2H), 7.43-7.75 (d, 2H), 7.59-7.65 (m, 3H), 7.75-7.82 (m,3H), 8.18 (s, 1H); LC-MS: m / z = 424.2(M). Example 76 3-({8-[(4-bromophenyl)methyl]-7-oxo-1,4,8,10-tetraazatricyclo[7.3.0.0]) 2,6 Dodecyl-2(6),9-dien-4-yl)methylbenzylnitrile Example 76 was prepared by the following synthesis scheme: 1 HNMR (400MHz, CDCl3) δ 4.1-4.3 (m, 10H), 5.19 (s, 2H), 7.25 (s, 1H), 7.27 (s, 1H), 7.43-7.45 (d, 2H), 7.53 (t, J=7.6Hz, 1H), 7.66-7.72 (m, 3H); LC-MS: m / z = 463.8 (M +2). Example 77 3-[(7-O-8-{[4-(trifluoromethyl)phenyl]methyl}-1,4,8,10-tetraazatricyclo[7.3.0.0] 2,6 [dodecyl-2(6),9-dien-4-yl)methylbenzylnitrile] Example 77 was prepared using the synthesis scheme described in respect of Example 76.

[0327] 1 HNMR (400MHz, CDCl3) δ 4.06-4.15 (m, 8H), 4.28 (t, J=8.4Hz, 2H), 5.33(s, 2H), 7.52-7.61 (m, 5H), 7.66-7.68 (d, 2H), 7.72 (s, 1H); LC-MS: m / z =451.9(M). Example 78 2-[(4-(bromophenyl)methyl]-7-{[3-(prop-1-yn-1-yl)phenyl]methyl}-1,2,5,6,7,8-hexahydro-2,7-naphthidium-1-one Example 78 was prepared by the following synthesis scheme: 1 HNMR (400MHz, DMSO_d6) δ 1.97 (s, 3H), 2.91 (s, 2H), 3.32-3.36 (m,1H), 3.62-3.65 (m, 1H), 3.91 (s, 2H), 4.46 (s, 2H), 5.08 (s, 2H), 7.29-7.6(m, 8H), 8.71 (s, 1H); LC-MS: m / z = 449.8 (M+2). Example 79 7-{[3-(prop-1-yn-1-yl)phenyl]methyl}-2-{[4-(trifluoromethyl)phenyl]methyl}-1,2,5,6,7,8-hexahydro-2,7-naphthidium-1-one Example 79 was prepared using the synthesis scheme described in respect of Example 78.

[0328] 1 HNMR (400MHz, DMSO_d6) δ 2.06 (s, 3H), 2.92 (s, 2H), 3.29-3.36 (m,1H), 3.62-3.65 (m, 1H), 3.93 (s, 2H), 4.46 (s, 2H), 5.2 (s, 2H), 7.44-7.81(m, 8H), 8.75 (s, 1H); LC-MS: m / z = 437.9 (M). Example 80 4-Phenylacetyl-8-[(4-chlorophenyl)methyl]1,4,8,10-tetraazatricyclo[7.3.0.0] 2,6 Dodeca-2(6),9-dien-7-one Example 80 was prepared by the following synthesis scheme: LC-MS: Retention time: 1.546 min, m / z = 393.1(M+1). See conditions in the attached table. Figure 8 And Example 62.

[0329] Example 81 4-Phenylacetyl-8-[(4-bromophenyl)methyl]1,4,8,10-tetraazatricyclo[7.3.0.0] 2,6 Dodeca-2(6),9-dien-7-one Example 81 was prepared using the synthesis scheme described with respect to Example 80.

[0330] 1 HNMR (400MHz, CDCl3) δ 4.0 (s, 2H), 4.2-4.24 (d, 6H), 4.4 (s, 2H), 5.15 (s, 2H), 7.23-7.24 (d, 2H), 7.42 (s, 7H); LC-MS: m / z = 439.1(M+2). Example 82 4-Phenylacetyl-8-{[4-(trifluoromethyl)phenyl]methyl}1,4,8,10-tetraazatricyclic[7.3.0.0] 2,6 Dodeca-2(6),9-dien-7-one Example 82 was prepared using the synthesis scheme described with respect to Example 80.

[0331] 1 HNMR (400MHz, CDCl3) δ 4.01 (s, 2H), 4.21-4.25 (d, 6H), 4.41 (s,2H), 5.26 (s, 2H), 7.37-7.46 (m, 7H), 7.54-7.56 (d, 2H); LC-MS: m / z = 426.9(M). Example 83 3-({9-[(4-chlorophenyl)methyl]-8-oxo-1,5,9,11-tetraazatricyclo[8.4.0.0]) 2,7 [Tetradecyl-2(7),10-dien-5-yl)methylbenzylnitrile] Example 83 was prepared using the synthesis scheme described in connection with Example 62.

[0332] 1HNMR (400MHz, DMSO & CDCl3) 2.13 (s, 2H), 2.86 (s, 4H), 3.38 (s,2H), 3.5 (s, 2H), 3.84 (s, 2H), 4.05 (s, 2H), 5.28 (s, 2H), 7.27-7.34 (m,3H), 7.53 (t, J=8Hz, 1H), 7.65-7.67 (d, 2H), 7.74 (s, 1H), 8.0 (s, 1H); LC-MS: m / z = 446.1(M+1). Example 84 3-({9-[(4-bromophenyl)methyl]-8-oxo-1,5,9,11-tetraazatricyclo[8.4.0.0]) 2,7 [Tetradecyl-2(7),10-dien-5-yl)methylbenzylnitrile] Example 84 was prepared using the synthesis scheme described in connection with Example 62.

[0333] 1 HNMR (400MHz, DMSO) 2.05 (s, 2H), 2.87 (s, 4H), 3.36-3.43 (m, 4H), 3.89 (s, 2H), 3.99(t, J=5.6Hz, 2H), 5.16 (s, 2H), 7.22-7.24 (d, 2H), 7.54-7.62 (m, 3H), 7.71-7.73 (d, 1H), 7.8-7.83 (d, 2H); LC-MS: m / z = 492.1(M+2). Biological Examples and Experiments The following embodiments are provided to provide a complete disclosure and description of how to prepare and utilize the invention to those skilled in the art, and are not intended to limit the scope of the invention. Efforts have been made to ensure the accuracy of the figures used (e.g., quantities, temperatures, etc.), but some experimental errors and biases should be taken into account.

[0334] a) Experimental procedures / materials and methods Measurement of human ClpP activity 。As previously described (Maurizi, MR et al., Methods Enzymol. 1994, 244, 314-331 and references cited therein, and Woo, KM et al., Biol. Chem. 1989, 264, 2088-2091 and references cited therein), with slight modifications, the activity of recombinant human casein hydrolase hClpP (Cat # MBS204060, MyBioSource, Boston USA) was measured in vitro based on monitoring the release of fluorescent coumarin from the fluorescent substrate Ac-WLA-AMC (Cat # S330, Boston Biochem, Inc., Cambridge, MA). In summary, the activity of the recombinant hClpP protein hydrolytic subunit (1 µg / mL) was measured using 10 µM of the fluorescent Ac-WLA-AMC substrate as described in the references above, in an assay buffer consisting of 50 mM Tris, 10 mM MgCl2, 100 mM KCl, 1 mM DTT, 4 mM ATP, 0.02% Triton X-100, and 5% glycerol—pH 8.0 (HCl). Two different protocols were used to investigate the effects of ONC201 and the compounds of this invention on ClpP activity. The first protocol (Protocol 1) was used, initiating the reaction by immediately mixing the enzyme and substrate in the presence of an indicated dose of the compound. The second protocol (Protocol 2) was used, initiating the reaction by mixing the enzyme and compound and incubating in the assay buffer for 60 min, followed by adding the Ac-WLA-ACM substrate. The kinetics of free coumarin fluorescence were monitored using a black µ-CLEAR 96-well flat-bottom plate (Cat # 655090, Greiner Germany), and the fluorescence of released coumarin was recorded at 350 nm excitation and 460 nm emission using a Pherastar reader equipped with a suitable FI module (BMG LABTECH, Durham NC). The slope of the linear portion of the fluorescence signal over time is a measure of hClpP activity. Measurements were performed three times and expressed as the rate of change in fluorescence at a given concentration of hClpP and substrate in the presence or absence of ONC201 or the compounds of the present invention. The dose dependence of hClpP activation for different compounds was used for the determination of substances (relative IC50). 50 ), and subtract the activity of the DMSO (medium) treated sample as background from the experimental data, and the activity of ClpP is expressed as ClpP / h RFU / µg See also Greer, YE et al., Oncotarget, 2018, 9, 18, 454-18479 and the references cited therein.

[0335] Cancer cell line The cell data described in Tables 1 and 2 were determined as described in CN104860948 and US 10,526,332. Other information for cell testing is as follows: HCT116 (human colon cancer) or MDA-MB-231 (MDA231, human breast adenocarcinoma) was partitioned into 100 μL of cell suspension in 96-well plates. The plates were incubated in a humidified incubator (37°C, 5% CO2) for 24 h. An appropriate test concentration of the compound of the present invention was added to the culture medium of the plates. The plates were incubated for 48 h. CCK-8 (10 μL, see below) was added to each well. The plates were incubated under the above conditions for 1–4 h, and the absorbance at 450 nm and 650 nm was measured using a plate reader.

[0336] Cell Counting Kit-8 (CCK-8) allows for sensitive colorimetric determination of viable cell counts in proliferation and cytotoxicity assays. Cell counting is performed using WST-8 (2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfonylphenyl)-2H-tetrazole, monosodium salt) in the presence of the electron carrier 1-methoxyPMS, which, upon bioreduction, produces a water-soluble formazan dye. The CCK-8 solution is added directly to the cells. WST-8 is bioreduced by cellular dehydrogenases to an orange formazan product soluble in tissue culture medium. The amount of formazan produced is directly proportional to the number of viable cells.

[0337] Measurement of antibacterial activity. Several publications describe the testing of the antibacterial activity of ClpP modifiers (Kao, YT et al., PNAS 2018, 115, 8003-8008 and references contained therein, and Quellette SP et al., J. Bacteriol 2018, 201(2) pii: e00635-18, doi:10.1128 / JB.00635-18 and references cited therein). The experimental conditions described by Kao, YT et al. and Quellette SP et al. can be used to measure the antibacterial activity of the compounds of the present invention, including activity against Staphylococcus aureus.

[0338] b) Results QNC201 and TR compounds activate CLPP peptidase activity.To investigate the effects of ONC201 and the compounds of this invention on ClpP activity, we tested their effects on the enzymatic activity of isolated human hClpP. Using purified recombinant human mitochondrial ClpP proteolytic subunits (Cat # MBS204060, MyBioSource, Cambridge, MA) and the selectively fluorescent 7-aminomethylcoumarin-conjugated tripeptide Ac-WLA-AMC (Cat # S330, MyBioSource, Cambridge, MA), we measured the activity of hClpP peptidase in the presence or absence of ONC201 and TR-compounds. The enzymatic activity of hClpP was measured in assay buffer (as described in Experimental Procedures / Materials and Methods), and the fluorescence level of the released coumarin was continuously monitored. Figure 1 As shown, we observed that incubation of hClpP with ONC201 or a selected TR compound (TR-57) resulted in a time-dependent and exponential increase in the fluorescence of coumarin AMC released due to hClpP peptidase activity. However, pre-incubation of the recombinant hClpP proteolytic subunit with the selected compound in standard assay buffer for 60 min resulted in a permanent increase in enzyme activity and a linearization of the coumarin release rate over time. Figure 2 Examples illustrating the kinetic and dose-dependent activity variations of hClpP with respect to ONC201 and TR57 are shown. Plotting the dose dependence of hClpP activity relative to compound concentration on a semi-logarithmic scale allows for analysis of IC50 values. 50 The assay showed that the concentration of the reagent resulted in a 50% increase in the activity of pre-incubated hClpP. Figure 3 ).

[0339] Tables 1 and 2 provide the bioactivity of the selected examples against human cancer cells.

[0340] Table 1: Bioactivity data of selected analogues in human cancer cells Table 2: Bioactivity data of selected analogues in human cancer cells List of abbreviations A549: Human Non-Small Cell Lung Cancer Cell Line BSA: Bovine serum albumin ClpP: Casein hydrolysate P DMSO: Dimethyl sulfoxide DNA: deoxyribonucleic acid EDTA: Ethylenediaminetetraacetic acid ELISA: Enzyme-linked immunosorbent assay FACS: Fluorescence-activated cell scanning / sorting HEPES: 4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid HsClpP: Human mitochondrial ClpP HsClpX: AAA+ protein unblocking enzyme HsClpXP: An ATP-dependent protease complex found in the mitochondrial matrix. IHC: Immunohistochemistry MAB: Monoclonal antibody mRNA: messenger ribonucleic acid PBS: Phosphate-buffered saline RPMI-1640: A cell culture medium for culturing transformed and non-transformed eukaryotic cells and cell lines. siRNA: Small inhibitory ribonucleic acid TR compounds or multiple TR compounds: Any compound or group of compounds whose names begin with TR as described herein. For example: TR57.

[0341] amino acid sequence Protein: ClpP Organism: Homo sapiens (sp|Q16740|CLPP_HUMAN ATP-dependent Clp protease proteolytic subunit, mitochondrial OS=Homo sapiens OX=9606 GN=CLPP PE=1 SV=1) (SEQ ID NO:1) MWPGILVGGARVASCRYPALGPRLAAHFPAQRPPQRTLQNGLALQRCLHATATRALPLIP IVVEQTGRGERAYDIYSRLLRERIVCVMGPIDDSVASLVIAQLLFLQSESNKKPIHMYIN SPGGVVTAGLAIYDTMQYILNPICTWCVGQAASMGSLLLAAGTPGMRHSLPNSRIMIHQP SGGARGQATDIAIQAEEIMKLKKQLYNIYAKHTKQSLQVIESAMERDRYMSPMEAQEFGI LDKVLVHPPQDGEDEPTLVQKEPVEAAPAAEPVPAST.

Claims

1. Compounds of general formula I: Z1-Q (Formula I) Or its pharmaceutically acceptable salt, wherein: Z1 is: ； Z2 is: ； Q is: Q3； Ar1 is independently selected from aryl, heteroaryl, phenylthio, and phenyl groups; Ar1 can be optionally substituted with 1 to 3 J groups; Ar2 is independently selected from phenylthio and phenyl, and is substituted by 1 to 3 JJ groups; J is independently selected from halogen, -CN, (C1-C6) optionally substituted alkyl, (C1-C6) haloalkyl, -CF3, -SR15, (C1-C6) optionally substituted alkoxy, -NR17R18 and (C2-C6) ynyl; JJ is independently selected from halogen, -CN, (C1-C6) haloalkyl, (C1-C6) optionally substituted alkyl, -CF3, -SR15, (C1-C6) optionally substituted alkoxy and (C2-C6) alkynyl; R1, R2, R3, R4, R5, R6, R7 and R8 are each independently selected from hydrogen, halogen, -OH and (C1-C3) optionally substituted alkyl groups; R5 and R6 can be combined to form =O; R7 and R8 can be combined to form =O; R14 is independently selected from hydrogen and (C1-C6) optionally substituted alkyl groups; R15, R16, R17 and R18 are independently selected from hydrogen and (C1-C6) optionally substituted alkyl groups; W4 is independently selected from =C(R14)- and nitrogen.

2. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein: R1, R2, R3, and R4 are each hydrogen.

3. The compound according to claim 2 or a pharmaceutically acceptable salt thereof, wherein: R5, R6, R7, and R8 are each hydrogen.

4. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein: J is independently selected from halogens, -CN, and (C2-C6) ynyl groups; JJ is independently selected from halogens, -CF3, and (C1-C6) haloalkyl groups.

5. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, wherein: R1, R2, R3, R4, R5, R6, R7, and R8 are hydrogen; W4 is nitrogen.

6. The compound of claim 5 or a pharmaceutically acceptable salt thereof, wherein: J is independently selected from halogens, -CN, and (C2-C6) ynyl groups; JJ is independently selected from halogens, -CF3, and (C1-C6) haloalkyl groups; R14 is independently selected from hydrogen, -NH2, and optionally substituted (C1-C6) alkyl groups.

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