Cyclic pyridine derivatives as cGAS inhibitors

Abstract

The invention relates to compounds of formula I # imgabs0, wherein R1, R2, R3, R4, A, D, E, G, J, K and L are as defined in claim 1; as well as prodrugs, deuterated analogs and pharmaceutically acceptable salts thereof; the compounds are useful in the treatment of diseases such as systemic lupus erythematosus, systemic sclerosis (SSc), interferon lesions, non-alcoholic steatohepatitis (NASH), interstitial lung disease (ILD) and idiopathic pulmonary fibrosis (IPF).

Description

Background Art

[0001] 1.1 cGAS inhibitor

[0002] Innate immunity is regarded as the first-line cellular stress response, which protects host cells from pathogen invasion and initiates signal transduction to the adaptive immune system. These processes are triggered by the sensing of conserved pathogen-associated molecular patterns (PAMPs) via different pattern recognition receptors (PRRs) and subsequent activation of cytokine and type I interferon gene expression. Major antigen-presenting cells such as monocytes, macrophages, and dendritic cells produce type I interferons and are crucial for initiating adaptive T-cell and B-cell immune system responses. Major PRRs detect abnormal nucleic acids on the cell surface, inside lysosomal membranes, or in other cellular compartments, i.e., mislocalized, immature, or unmodified nucleic acids (Barbalat et al., Annu. Rev. Immunol. 29, 185-214 (2011)).

[0003] "Cyclic GMP-AMP synthase" (cGAS; UniProtKB-Q8N884)) is the major sensor for mislocalized or mishandled abnormal double-stranded DNA (dsDNA) from pathogens or nuclear or mitochondrial cell dsDNA (Sun et al., Science 339, 786-791 (2013); Wu et al., Science 339, 826-830 (2013); Ablasser et al., Nature 498, 380-384 (2013)). Binding of dsDNA to cGAS activates the reaction of GTP and ATP to form the cyclic dinucleotide GMP-AMP (called cGAMP). Subsequently, cGAMP travels to and activates the endoplasmic reticulum membrane-anchored adaptor protein, "Stimulator of Interferon Gene" (STING). The activated STING recruits and activates TANK-binding kinase 1 (TBK1), which in turn phosphorylates the transcription factor family of interferon regulatory factor (IRF), inducing cytokine and type I interferon mRNA expression.

[0004] The key role of cGAS in dsDNA sensing has been established in diverse pathogenic bacteria (Hansen et al., EMBO J. 33, 1654 (2014)), viruses (Ma et al., PNAS 112, E4306 (2015)), and retroviruses (Gao et al., Science 341, 903 - 906 (2013)). Additionally, cGAS is crucial in various other biological processes, such as cellular senescence (Yang et al., PNAS 114, E4612 (2017), Glück et al., Nat. Cell Biol. 19, 1061 - 1070 (2017)) and the recognition of broken micronuclei during potential cancer cell surveillance (Mackenzie et al., Nature 548, 461 - 465 (2017); Harding et al., Nature 548, 466 - 470 (2017)).

[0005] Although the cGAS pathway is crucial for host defense against pathogen invasion, cellular stress and genetic factors can also lead to the production of abnormal cellular dsDNA, for example, through nuclear or mitochondrial leakage, and thereby trigger an autoinflammatory response. Aicardi - Goutieres syndrome (AGS; Crow et al., Nat. Genet. 38, 917 - 920 (2006)), a lupus - like severe autoinflammatory immune - mediated disorder, is caused by loss - of - function mutations in TREX1, a major DNA exonuclease responsible for degrading abnormal DNA in the cytosol. Deletion of cGAS in TREX1 - deficient mice prevents the otherwise lethal autoimmune response, thus supporting cGAS as a driver of interferonopathies (Gray et al., J. Immunol. 195, 1939 - 1943 (2015); Gao et al., PNAS 112, E5699 - E5705 (2015)). Similarly, the embryonic lethality caused by the deficiency of DNAse2, an endonuclease responsible for degrading excess DNA in lysosomes during endocytosis, is fully rescued by additional gene deletion of cGAS (Gao et al., PNAS 112, E5699 - E5705 (2015)) or STING (Ahn et al., PNAS 109, 19386 - 19391 (2012)). These observations support cGAS as a drug target, and inhibition of cGAS could provide therapeutic strategies for preventing autoinflammation and treating diseases involving anti - dsDNA antibodies such as systemic lupus erythematosus (SLE) (Pisetsky et al., Nat. Rev. Rheumatol. 12, 102 - 110 (2016)). 1.2 Background Art

[0007] Since it has been observed that inhibiting the cGAS pathway can provide therapeutic strategies for preventing autoinflammation and treating, for example, autoimmune diseases, many efforts have been made to develop cGAS inhibitors.

[0008] In WO 2019 / 241787, compounds such as methyl 4-amino-6-(phenylamino)-1,3,5-triazine-2-carboxylate, such as CU-32 and CU-76, have been disclosed as cGAS inhibitors, where the "in vitro hcGAS IC 50 value" is slightly below 1 μM (IC 50 (CU-32)=0.66 μM and IC 50 (CU-76 = 0.27 μM).

[0009] In Hall et al., PLoS ONE 12(9); e0184843 (2017), the compound PF-06928215 has been disclosed as a cGAS inhibitor, where the "in vitro hcGAS IC 50 value" measured by fluorescence polarization assay is 0.049 μM. However, the compound PF-06928215 does not exhibit acceptable cellular activity as a cGAS inhibitor.

[0010] In WO 2020 / 142729 and WO2022 / 174012, (benzofuro[3,2-d]pyrimidin-4-yl)pyrrolidine-2-carboxylic acid derivatives have been disclosed as cGAS inhibitors for the treatment of autoimmune disorders such as Aicardi-Goutières syndrome (AGS), lupus erythematosus, scleroderma, inflammatory bowel disease, and non-alcoholic steatohepatitis (NASH). However, the compounds of the present invention are different from the (benzofuro[3,2-d]pyrimidin-4-yl)pyrrolidine-2-carboxylic acid derivatives of WO 2020 / 142729, which have a completely different substitution pattern at the 4-position of the pyrrolidine ring.

[0011] Recently provided cGAS inhibitors, such as the cGAS inhibitors in WO 2020 / 142729 or WO 2022 / 174012, generally exhibit insufficient cellular cGAS inhibitory potency (where the IC 50 value measured in cell assays for inhibiting the cGAS / STING pathway is generally greater than 1 μM, often greater than 5 μM). However, it is crucial to provide therapeutic cGAS inhibitors that not only exhibit satisfactory biochemical (in vitro) inhibitory potency ("hcGAS IC 50 "), but also exhibit satisfactory cellular inhibitory potency (e.g., by demonstrating inhibition of IFN induction in virus-stimulated THP-1 cells (THP1 (vir) IC 50)) to ensure that the compound can exhibit a therapeutic effect in patients. Other important properties that may be predictive of the successful development of a cGAS inhibitor as a therapeutic agent are satisfactory cGAS selectivity (versus off-target activity) and acceptable inhibitory potency in human whole blood.

[0012] Surprisingly, it has now been found that the compounds of formula I or II simultaneously exhibit the following three properties:

[0013] · Satisfactory "biochemical (in vitro) IC 50 value for cGAS inhibition" (where hcGAS IC 50 ≤ 100 nM, preferably ≤ 50 nM, particularly ≤ 10 nM),

[0014] · Satisfactory "inhibition of IFN induction in virus-stimulated THP-1 cells" (where THP1 IC 50(vir) ≤ 1 μM, preferably ≤ 500 nM, more preferably ≤ 100 nM, particularly ≤ 50 nM), and

[0015] · Satisfactory selectivity for cGAS inhibition

[0016] (where the ratio THP1 IC 50(cGAMP) / THP1 IC 50(vir) ≥ 10, more preferably ≥ 50, more preferably ≥ 500, particularly ≥ 1000).

[0017] In addition, the compounds of formula I or II also exhibit an acceptable IC 50 value for the inhibition of IFN induction in a dsDNA-stimulated human whole blood assay, preferably where the human whole blood IC 50 value for cGAS inhibition (hWB IC 50 ) ≤ 5000 nM, more preferably ≤ 1000 nM, particularly ≤ 100 nM.

[0018] The cGAS inhibitors of the present invention having this specific pharmacological profile, combined with excellent in vitro inhibitory potency and excellent cellular inhibitory potency as well as high selectivity for cGAS inhibition, are very likely to also exhibit good therapeutic effects in patients. Due to their high cellular inhibitory potency, the compounds having this specific pharmacological profile should be able to cross the cell membrane barrier and thus reach their intracellular target location, and due to their selectivity for specifically inhibiting cGAS activity, these compounds should not exhibit unwanted off-target effects, such as producing side effects or cytotoxic effects somewhere downstream of the cGAS signaling pathway. SUMMARY OF THE INVENTION

[0019] The present invention relates to compounds of formula I

[0020]

[0021] wherein

[0022] R 1 is selected from the group consisting of hydrogen, halogen, methyl, ethyl, -CF3, -CHF2, -CFH2, and methoxy;

[0023] R 2 is selected from the group consisting of hydrogen and methyl;

[0024] R 3 is selected from the group consisting of hydrogen, methyl, halogen, ethynyl, propargyl, -CO-(C 1-3 -alkyl), -CO-NH2, -CO-NHCH3, -CO-N(CH3)2, and a 5- or 6-membered heteroaryl ring having 1 or 2 heteroatoms each independently selected from N, S, or O, wherein the heteroaryl ring may optionally be further substituted with one or two other substituents each independently selected from the group consisting of F, Cl, Br, -O-CH3, methyl, -CF3, -CHF2, and CH2F;

[0025] R 4 is selected from the group consisting of hydrogen, -OH, and F;

[0026] and wherein

[0027] A is selected from the group consisting of -CH2-, -O-, -CF2-, and -CHCH3-;

[0028] D is selected from the group consisting of -CH2-, -O-, -CF2-, and -CHCH3-;

[0029] E is selected from the group consisting of -CH2-, -CO-, -O-, -CF2-, and -CHCH3-;

[0030] G is selected from the group consisting of -NH-, -NCH3-, -CH2-, -O-, -CF2-, and -CHCH3-;

[0031] J is selected from the group consisting of -CO-, -CH2-, -O-, -CF2-, and -CHCH3-;

[0032] K is selected from the group consisting of -CH2-, -CF2-, and -O- or is absent;

[0033] L is selected from the group consisting of -CH2-, -CHCH3-, and -CF2- or is absent;

[0034] and their prodrugs, deuterated analogs, and pharmaceutically acceptable salts.

[0035] Accordingly, variables A, D, E, G, J, K, and L are preferably selected in such a way that two or more heteroatoms do not directly follow each other.

[0036] In a preferred embodiment, the present invention relates to a compound of formula II, wherein

[0037]

[0038] R 1 is hydrogen and / or is a halogen preferably selected from the group consisting of F and Cl;

[0039] R 2 is selected from the group consisting of hydrogen and methyl;

[0040] R 3 is selected from the group consisting of: hydrogen, Cl, Br, ethynyl, propargyl-CO-(CH3), and a 5- or 6-membered heteroaryl ring having 1 or 2 heteroatoms each independently selected from N, S, or O, wherein this heteroaryl ring may optionally be further substituted with one or two additional substituents each independently selected from the group consisting of F, -O-CH3, and methyl;

[0041] R 4 is F;

[0042] and wherein

[0043] A is selected from the group consisting of: -CH2-, -O-, -CF2-, and -CHCH3-;

[0044] D is selected from the group consisting of -CH2- and -O-;

[0045] E is selected from the group consisting of: -CH2-, -CO-, and -O-;

[0046] G is selected from the group consisting of: -NH-, -NCH3-, -CH2-, and -O-;

[0047] J is selected from the group consisting of: -CO-, -CH2-, and -O-;

[0048] K is selected from the group consisting of -CH2-, -CF2-, and -O- or is absent;

[0049] L is -CH2- or is absent;

[0050] as well as its prodrugs, deuterated analogs, and pharmaceutically acceptable salts.

[0051] Accordingly, variables A, D, E, G, J, K, and L are preferably selected in such a way that two or more heteroatoms do not directly follow each other.

[0052] In another preferred embodiment, the present invention relates to a compound of formula I or a compound of formula II as mentioned above,

[0053] wherein L is absent,

[0054] and its prodrugs, deuterated analogs and pharmaceutically acceptable salts.

[0055] In another preferred embodiment, the present invention relates to a compound of formula I or a compound of formula II as mentioned above,

[0056] wherein L and K are absent,

[0057] and its prodrugs, deuterated analogs and pharmaceutically acceptable salts.

[0058] In another preferred embodiment, the present invention relates to a compound of formula I or a compound of formula II as mentioned above,

[0059] wherein L is absent and wherein K is -CF2-,

[0060] and its prodrugs, deuterated analogs and pharmaceutically acceptable salts.

[0061] In another preferred embodiment, the present invention relates to a compound of formula I or a compound of formula II as mentioned above,

[0062] wherein L is absent and wherein A is selected from the group consisting of -CH2- and -CF2-, and its prodrugs, deuterated analogs and pharmaceutically acceptable salts.

[0063] In another preferred embodiment, the present invention relates to a compound of formula I or a compound of formula II as mentioned above,

[0064] wherein L is absent and wherein A is -O-,

[0065] and its prodrugs, deuterated analogs and pharmaceutically acceptable salts.

[0066] In another preferred embodiment, the present invention relates to a compound of formula I or a compound of formula II as mentioned above,

[0067] wherein L is absent and wherein D is -O-,

[0068] and its prodrugs, deuterated analogs and pharmaceutically acceptable salts.

[0069] In another preferred embodiment, the present invention relates to a compound of formula I or a compound of formula II as mentioned above,

[0070] wherein L is absent, and

[0071] wherein R 3 is selected from the group consisting of Cl, Br, ethynyl, propargyl and a 5- or 6-membered heteroaryl ring selected from the group consisting of pyridyl and pyrazolyl, wherein said heteroaryl ring may optionally be further substituted with one or two additional substituents each independently selected from the group consisting of F, -O-CH3 and methyl;

[0072] and prodrugs, deuterated analogs and pharmaceutically acceptable salts thereof.

[0073] In another preferred embodiment, the present invention relates to a compound of formula I or a compound of formula II as mentioned above,

[0074] wherein R 1 is selected from the group consisting of F and Cl,

[0075] and prodrugs, deuterated analogs and pharmaceutically acceptable salts thereof.

[0076] In another preferred embodiment, the present invention relates to a compound of formula I or a compound of formula II as mentioned above,

[0077] wherein R 1 is hydrogen,

[0078] and prodrugs, deuterated analogs and pharmaceutically acceptable salts thereof.

[0079] In another particularly preferred embodiment, the present invention relates to a compound of formula I or a compound of formula II as mentioned above, which is selected from the group consisting of:

[0080]

[0081]

[0082]

[0083] and prodrugs, deuterated analogs and pharmaceutically acceptable salts thereof.

[0084] In another preferred embodiment, the present invention relates to a compound of formula I or a compound of formula II as mentioned above, wherein

[0085] A is selected from the group consisting of -CH2- and -O-;

[0086] D is selected from the group consisting of -CH2- and -O-;

[0087] E is selected from the group consisting of -CH2- and -O-;

[0088] G is selected from the group consisting of -CH2- and -O-;

[0089] J is selected from the group consisting of -CH2- and -O-;

[0090] K is selected from the group consisting of -CH2- and -CF2-;

[0091] L is absent;

[0092] and their prodrugs, deuterated analogs, and pharmaceutically acceptable salts.

[0093] In another particularly preferred embodiment, the present invention relates to the compound of formula II mentioned above, which is selected from the group consisting of:

[0094]

[0095]

[0096] and their prodrugs, deuterated analogs, and pharmaceutically acceptable salts.

[0097] In another particularly preferred embodiment, the present invention relates to the compound of formula I or the compound of formula II mentioned above, wherein

[0098] A is selected from the group consisting of -CH2- and -O-;

[0099] D is selected from the group consisting of -CH2- and -O-;

[0100] E is -CH2-;

[0101] G is selected from the group consisting of -CH2- and -O-;

[0102] J is -CH2-;

[0103] K is selected from the group consisting of -CH2- and -CF2-;

[0104] L is absent;

[0105] and their prodrugs, deuterated analogs, and pharmaceutically acceptable salts.

[0106] In another particularly preferred embodiment, the present invention relates to the compound of formula II mentioned above, which is selected from the group consisting of:

[0107]

[0108] and their prodrugs, deuterated analogs, and pharmaceutically acceptable salts.

[0109] The prodrug of the compound of formula I is preferably the compound of formula Ia

[0110] wherein the variable R 1 、R2 , R 3 , R 4 , A, D, E, G, J, K, and L are as defined above, and wherein R 5 is C 1-4 -alkyl, aryl, -CH2-aryl, NH-SO2-C 1-3 -alkyl.

[0111] Particularly preferred are prodrugs of formula Ia, wherein the variables R 1 , R 2 , R 3 , R 4 , A, D, E, G, J, K, and L are as defined above and wherein R 5 is methyl.

[0112] The prodrug of the compound of formula II is preferably a compound of formula IIa

[0113]

[0114] wherein the variables R 1 , R 2 , R 3 , R 4 , A, D, E, G, J, K, and L are as defined above and wherein R 5 is C 1-4 -alkyl, aryl, -CH2-aryl, NH-SO2-C 1-3 -alkyl.

[0115] Particularly preferred are prodrugs of formula Ia, wherein the variables R 1 , R 2 , R 3 , R 4 , A, D, E, G, J, K, and L are as defined above and wherein R 5 is methyl.

[0116] In another preferred embodiment, the present invention relates to

[0117] a) an intermediate compound of formula (A-I)

[0118]

[0119] wherein R 1 , R 2 , R 3 , R 4 , A, D, E, G, J, K, and L are as defined above, and wherein R 13 is selected from the group consisting of hydrogen, methyl, ethyl, and tert-butyl,

[0120] b) an intermediate compound of formula (A-II)

[0121]

[0122] wherein R 1 、R 2 、R 3 、R 4 、A, D, E, G, J, K and L are as defined above, wherein R 13 is selected from the group consisting of hydrogen, methyl, ethyl and tert-butyl, and wherein R is hydrogen or a protecting group selected from the group consisting of tert-butyl, methyl, ethyl and benzyl,

[0123] c) Intermediate compound of formula (B-I)

[0124]

[0125] wherein R 1 、R 2 、R 3 、R 4 、A, D, E, G, J, K and L are as defined above, and wherein R 13 is selected from the group consisting of hydrogen, methyl, ethyl and tert-butyl,

[0126] d) Intermediate compound of formula (C-I)

[0127]

[0128] wherein R 1 、R 2 、R 3 、R 4 、A, D, E, G, J, K and L are as defined above, and wherein R 13 is selected from the group consisting of hydrogen, methyl, ethyl and tert-butyl, or

[0129] e) Formula (C-II)

[0130]

[0131] wherein R 1 、R 2 、R 3 、R 4 、A, D, E, G, J, K and L are as defined above, and wherein R 13 is selected from the group consisting of hydrogen, methyl, ethyl and tert-butyl, and wherein PG is selected from the group consisting of tert-butoxycarbonyl (Boc), allyloxycarbonyl (Alloc), benzyloxycarbonyl (Cbz) and fluorenylmethoxycarbonyl (Fmoc).

[0132] In another preferred embodiment, the present invention relates to the above-mentioned compound of formula I or the above-mentioned compound of formula II or the prodrug of formula Ia or IIa, its deuterated analogs and pharmaceutically acceptable salts for use in treating diseases that can be treated by inhibiting cGAS.

[0133] In another preferred embodiment, the present invention relates to the above-mentioned compounds of formula I or II or prodrugs of formula Ia or IIa, deuterated analogs and pharmaceutically acceptable salts thereof, for use in treating a disease selected from the group consisting of: systemic lupus erythematosus (SLE); interferonopathy; Acardi-Guterres syndrome (AGS); COPA syndrome; familial pernio lupus; age-related macular degeneration (AMD); retinopathy; glaucoma; amyotrophic lateral sclerosis (ALS); diabetes; obesity; inflammatory bowel disease (IBD); chronic obstructive pulmonary disease (COPD); Bloom's syndrome; Sjogren's syndrome; Parkinson's disease ( disease); heart failure and cancer; systemic sclerosis (SSc); dermatomyositis; nonalcoholic steatohepatitis (NASH); interstitial lung disease (ILD), preferably progressive fibrosing interstitial lung disease (PF-ILD), in particular idiopathic pulmonary fibrosis (IPF); aging; muscle disorders; sepsis; rheumatoid arthritis; osteoarthritis; and COVID-19.

[0134] In another preferred embodiment, the present invention relates to the above-mentioned compound of formula I or II or prodrug of formula Ia or IIa, its deuterated analog and pharmaceutically acceptable salt, which is used for treating a disease selected from the group consisting of systemic lupus erythematosus (SLE), interferonopathy, Acardi-Guterres syndrome (AGS), COPA syndrome, familial pernio lupus, dermatomyositis, age-related macular degeneration (AMD), amyotrophic lateral sclerosis (ALS), inflammatory bowel disease (IBD), chronic obstructive pulmonary disease (COPD), Bloom's syndrome, Sjögren's syndrome, rheumatoid arthritis and Parkinson's disease.

[0135] In another preferred embodiment, the present invention relates to the compounds of formula I or II or prodrugs of formula Ia or IIa, deuterated analogs thereof and pharmaceutically acceptable salts thereof, for use in treating a disease selected from the group consisting of: systemic sclerosis (SSc); non-alcoholic steatohepatitis (NASH); interferonopathy; interstitial lung disease (ILD), preferably progressive fibrosing interstitial lung disease (PF-ILD), especially idiopathic pulmonary fibrosis (IPF).

[0136] In another preferred embodiment, the present invention relates to a compound of formula I or II or a prodrug of formula Ia or IIa, a deuterated analogue thereof, and a pharmaceutically acceptable salt thereof, which are used for treating diseases selected from the group consisting of age-related macular degeneration (AMD), retinopathy, glaucoma, diabetes, obesity, aging, muscle disorders, sepsis, osteoarthritis, heart failure, COVID19 / SARS-CoV-2 infection, kidney inflammation, renal fibrosis, metabolic disorders, vascular diseases, cardiovascular diseases, and cancer.

[0137] In another preferred embodiment, the present invention relates to a pharmaceutical composition comprising a compound of formula I or II or a prodrug of formula Ia or IIa, a deuterated analogue thereof, and a pharmaceutically acceptable salt thereof, and optionally one or more pharmaceutically acceptable carriers and / or excipients.

[0138] In another preferred embodiment, the present invention relates to a pharmaceutical composition comprising a compound of formula I or II or a prodrug of formula Ia or IIa, a deuterated analogue thereof, and a pharmaceutically acceptable salt thereof, and one or more active agents selected from the group consisting of anti-inflammatory agents; anti-fibrotic agents; anti-allergic agents / anti-histamines; bronchodilators; β2 agonists / β mimetics; adrenergic agonists; anticholinergics; methotrexate; mycophenolate mofetil; leukotriene regulators; JAK inhibitors; anti-interleukin antibodies; non-specific immunotherapeutic agents such as interferons or other cytokines / chemokines; cytokine / chemokine receptor regulators; Toll-like receptor agonists; immune checkpoint regulators; anti-TNF antibodies such as Humira TM ; anti-BAFF antibodies such as belimumab and etanercept; and optionally one or more pharmaceutically acceptable carriers and / or excipients.

[0139] In another preferred embodiment, the present invention relates to a pharmaceutical composition comprising a compound of formula I or II or a prodrug of formula Ia or IIa, a deuterated analogue thereof, and a pharmaceutically acceptable salt thereof, and one or more anti-fibrotic agents selected from the group consisting of pirfenidone and nintedanib, and optionally one or more pharmaceutically acceptable carriers and / or excipients.

[0140] In another preferred embodiment, the present invention relates to a pharmaceutical composition comprising a compound of formula I or II or a prodrug of formula Ia or IIa, a deuterated analogue thereof, and a pharmaceutically acceptable salt thereof, and one or more anti-inflammatory agents selected from the group consisting of NSAIDs and corticosteroids, and optionally one or more pharmaceutically acceptable carriers and / or excipients.

[0141] In another preferred embodiment, the present invention relates to a pharmaceutical composition comprising a compound of formula I or II or a prodrug of formula Ia or IIa, a deuterated analogue thereof, and a pharmaceutically acceptable salt, as mentioned above, and one or more active agents selected from the group consisting of bronchodilators, β2 agonists / β mimetics, adrenergic agonists, and anticholinergics, and optionally one or more pharmaceutically acceptable carriers and / or excipients.

[0142] In another preferred embodiment, the present invention relates to a pharmaceutical combination comprising a compound of formula I or II or a prodrug of formula Ia or IIa, a deuterated analogue thereof, and a pharmaceutically acceptable salt, as mentioned above, and one or more anti-interleukin antibodies selected from the group consisting of: anti-IL-23 antibodies such as Risankizumab; anti-IL-17 antibodies; anti-IL-1 antibodies; anti-IL-4 antibodies; anti-IL-13 antibodies; anti-IL-5 antibodies; anti-IL-6 antibodies such as Actemra TM ; anti-IL-12 antibodies; and anti-IL-15 antibodies.

[0143] 3. Terms and Definitions Used

[0144] Unless otherwise specified, all substituents are independent of each other. For example, if multiple C 1-6 alkyl groups are possible substituents on a group, then in the case of, for example, three substituents, the C 1-6 alkyl groups can independently represent methyl, n-propyl, and tert-butyl.

[0145] Cross bonds, such as the middle bond in the following butyl molecule,

[0146]

[0147] represent a double bond of unknown configuration (cis, trans, or a mixture thereof).

[0148] The term "C 1-6 alkyl" (including those alkyl groups that are part of other groups) means branched-chain and unbranched alkyl groups having 1 to 6 carbon atoms, and the term "C 1-3 alkyl" means branched-chain and unbranched alkyl groups having 1 to 3 carbon atoms. "C 1-4"Alkyl" thus denotes branched and unbranched alkyl groups having from 1 to 4 carbon atoms. Alkyl groups having from 1 to 4 carbon atoms are preferred. Examples of such alkyl groups include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl, neopentyl and hexyl. The groups mentioned above may optionally also be used in abbreviated form, such as Me, Et, n-Pr, i-Pr, n-Bu, i-Bu, t-Bu, etc. Unless otherwise stated, the definitions of propyl, butyl, pentyl and hexyl include all possible isomeric forms of these groups. Thus, for example, propyl includes n-propyl and isopropyl, and butyl includes isobutyl, sec-butyl and tert-butyl, etc.

[0149] The term "C 1-6 alkylene" (including those alkylene groups which are part of other groups) means branched and unbranched alkylene groups having from 1 to 6 carbon atoms, and the term "C 1-4 alkylene" means branched and unbranched alkylene groups having from 1 to 4 carbon atoms. Alkylene groups having from 1 to 4 carbon atoms are preferred. Examples of such alkylene groups include methylene, ethylene, propylene, 1-methylethylene, butylene, 1-methylpropylene, 1,1-dimethylethylene, 1,2-dimethylethylene, pentylene, 1,1-dimethylpropylene, 2,2-dimethylpropylene, 1,2-dimethylpropylene, 1,3-dimethylpropylene and hexylene. Unless otherwise stated, the definitions of propylene, butylene, pentylene and hexylene include all possible isomeric forms of the groups having the same number of carbon atoms. Thus, for example, propylene also includes 1-methylethylene and butylene includes 1-methylpropylene, 1,1-dimethylethylene, 1,2-dimethylethylene, etc.

[0150] If the carbon chain is substituted by a group which together with one or two carbon atoms of the alkylene chain forms a carbocyclic group having 3, 5 or 6 carbon atoms, it particularly includes the following ring examples:

[0151]

[0152] The term "C 2-6 alkenyl" (including those alkenyl groups which are part of other groups) means branched and unbranched alkenyl groups having from 2 to 6 carbon atoms, and the term "C 2-4"Alkenyl" means a branched and unbranched alkenyl having 2 to 4 carbon atoms, provided that it has at least one double bond. Alkenyls having 2 to 4 carbon atoms are preferred. Examples include: ethenyl or vinyl, propenyl, butenyl, pentenyl or hexenyl. Unless otherwise stated, the definitions of propenyl, butenyl, pentenyl and hexenyl include all possible isomeric forms of said groups. Thus, for example, propenyl includes 1-propenyl and 2-propenyl, and butenyl includes 1-butenyl, 2-butenyl and 3-butenyl, 1-methyl-1-propenyl, 1-methyl-2-propenyl, etc.

[0153] The term "C 2-5 "Alkynyl" (including alkynyl as part of other groups) means a branched and unbranched alkynyl having 2 to 5 carbon atoms, and the term "C 2-4 "Alkynyl" means a branched and unbranched alkynyl having 2 to 4 carbon atoms, provided that it has at least one triple bond. Alkynyls having 2 to 4 carbon atoms are preferred.

[0154] The term "C 2-6 "Alkenylene" (including those alkenylene as part of other groups) means a branched and unbranched alkenylene having 2 to 6 carbon atoms, and the term "C 2-4 "Alkenylene" means a branched and unbranched alkenylene having 2 to 4 carbon atoms. Alkenylenes having 2 to 4 carbon atoms are preferred. Examples of these alkenylenes include: vinylene, propenylene, 1-methylvinylene, butenylene, 1-methylpropenylene, 1,1-dimethylvinylene, 1,2-dimethylvinylene, pentenylene, 1,1-dimethylpropenylene, 2,2-dimethylpropenylene, 1,2-dimethylpropenylene, 1,3-dimethylpropenylene and hexenylene. Unless otherwise stated, the definitions of propenylene, butenylene, pentenylene and hexenylene include all possible isomeric forms of said groups having the same number of carbon atoms. Thus, for example, propenylene also includes 1-methylvinylene and butenylene includes 1-methylpropenylene, 1,1-dimethylvinylene, 1,2-dimethylvinylene.

[0155] The term "aryl" (including aryl as part of other groups) means an aromatic ring system having 6 or 10 carbon atoms. Examples include phenyl or naphthyl, and the preferred aryl is phenyl. Unless otherwise stated, the aromatic group may be substituted with one or more groups selected from: methyl, ethyl, isopropyl, tert-butyl, hydroxy, fluoro, chloro, bromo and iodo.

[0156] The term "aryl-C 1-6"Alkylene" (including those groups which are part of other groups) means a branched or unbranched alkylene having 1 to 6 carbon atoms which is substituted by an aromatic ring system having 6 or 10 carbon atoms. Examples include benzyl, 1-phenylethyl or 2-phenylethyl and 1-naphthylethyl or 2-naphthylethyl. Unless otherwise specified, the aromatic group may be substituted by one or more groups selected from: methyl, ethyl, isopropyl, tert-butyl, hydroxy, fluoro, chloro, bromo and iodo.

[0157] The term "heteroaryl-C 1-6 "Alkylene" (including those groups which are part of other groups), even if it is already included under "aryl-C 1-6 "Alkylene", means a branched or unbranched alkylene having 1 to 6 carbon atoms which is substituted by a heteroaryl group.

[0158] If not otherwise specifically defined, such heteroaryl groups include five- or six-membered heteroaromatic groups or 5- to 10-membered bicyclic heteroaryl rings which may contain one, two, three or four heteroatoms selected from oxygen, sulfur and nitrogen and contain a number of conjugated double bonds forming an aromatic system. The following are examples of five- or six-membered heteroaromatic groups and bicyclic heteroaryl rings:

[0159]

[0160] Unless otherwise specified, these heteroaryl groups may be substituted by one or more groups selected from: methyl, ethyl, isopropyl, tert-butyl, hydroxy, amino, nitro, alkoxy, fluoro, chloro, bromo and iodo.

[0161] The following are examples of heteroaryl-C 1-6 "Alkylene":

[0162]

[0163] The term "C 1-6 "Haloalkyl" (including those haloalkyls which are part of other groups) means a branched or unbranched alkyl having 1 to 6 carbon atoms which is substituted by one or more halogen atoms. The term "C 1-4 "Haloalkyl" means a branched or unbranched alkyl having 1 to 4 carbon atoms which is substituted by one or more halogen atoms. Preferably it is an alkyl having 1 to 4 carbon atoms. Examples include: CF3, CHF2, CH2F, CH2CF3.

[0164] If not otherwise specifically defined, the term "C 3-7"Cycloalkyl" (including those cycloalkyls that are part of other groups) means a cycloalkyl having 3 to 7 carbon atoms. Examples include: cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and cycloheptyl. Unless otherwise stated, the cycloalkyl may be substituted by one or more groups selected from: methyl, ethyl, isopropyl, tert-butyl, hydroxy, fluoro, chloro, bromo and iodo.

[0165] Unless otherwise specifically defined, the term "C" 3-10 "cycloalkyl" also means a monocycloalkyl having 3 to 7 carbon atoms and also means a bicycloalkyl having 7 to 10 carbon atoms, or a monocycloalkyl bridged by at least one C 1-3 carbon bridge.

[0166] Unless otherwise stated, the term "heterocyclic ring" or "heterocycle" means a five-, six- or seven-membered saturated, partially saturated or unsaturated heterocycle which may contain one, two or three heteroatoms selected from oxygen, sulfur and nitrogen, and the ring may be linked to the molecule via a carbon atom or via a nitrogen atom (if there is one). Although included by the term "heterocyclic ring" or "heterocycle", the term "saturated heterocycle" refers to a five-, six- or seven-membered saturated ring. Examples include:

[0167]

[0168] Although included by the term "heterocycle" or "heterocyclic group", the term "partially saturated heterocyclic group" refers to a five-, six- or seven-membered partially saturated ring which contains one or two double bonds but not so many as to form an aromatic system, unless otherwise specifically defined. Examples include:

[0169]

[0170]

[0171] Although included by the term "heterocyclic ring" or "heterocycle", unless otherwise specifically defined, the terms "heteroaromatic ring", "unsaturated heterocyclic group" or "heteroaryl" mean a five- or six-membered heteroaromatic group or a 5- to 10-membered bicyclic heteroaryl ring which may contain one, two, three or four heteroatoms selected from oxygen, sulfur and nitrogen, and contains many conjugated double bonds forming an aromatic system. Examples of five- or six-membered heteroaromatic groups include:

[0172]

[0173] Unless otherwise mentioned, a heterocyclic ring / heterocycle may have a ketone group. Examples include:

[0174]

[0175] Although covered by the term "cycloalkyl", the term "bicycloalkyl" generally refers to an eight-, nine- or ten-membered bicyclic carbocyclic ring. Examples include:

[0176]

[0177] Although included by the term "heterocycle", unless otherwise specifically defined, the term "bicyclic heterocycle" generally refers to an eight-, nine- or ten-membered bicyclic ring that may contain one or more heteroatoms, preferably 1 to 4, more preferably 1 to 3, even more preferably 1 to 2, and especially one heteroatom selected from oxygen, sulfur and nitrogen. The ring may be linked to the molecule via a carbon atom in the ring or via a nitrogen atom in the ring (if there is one). Examples include:

[0178]

[0179] Although included by the term "aryl", the term "bicyclic aryl" refers to a 5- to 10-membered bicyclic aryl ring containing conjugated double bonds sufficient to form an aromatic system. An example of a bicyclic aryl is naphthyl.

[0180] Although included under "heteroaryl", unless otherwise specifically defined, the term "bicyclic heteroaryl" refers to a 5-10 membered bicyclic heteroaryl ring that may contain one, two, three or four heteroatoms selected from oxygen, sulfur and nitrogen and contains conjugated double bonds sufficient to form an aromatic system.

[0181] Although included by the term "bicycloalkyl" or "bicyclic aryl", the term "fused cycloalkyl" or "fused aryl" refers to a bicyclic ring where the bridge separating the rings represents a direct single bond. The following are examples of fused bicycloalkyls:

[0182]

[0183] Although included by the term "bicyclic heterocycle" or "bicyclic heteroaryl", the term "fused bicyclic heterocycle" or "fused bicyclic heteroaryl" refers to a 5- to 10-membered bicyclic heterocycle containing one, two, three or four heteroatoms selected from oxygen, sulfur and nitrogen, and where the bridge separating the rings represents a direct single bond. "Fused bicyclic heteroaryl" additionally contains conjugated double bonds sufficient to form an aromatic system. Examples include pyrrolozine, indole, indolizine, isoindole, indazole, purine, quinoline, isoquinoline, benzimidazole, benzofuran, benzopyran, benzothiazole, benzothiazole, benzisothiazole, pyridopyrimidine, pteridine, pyrimidinopyrimidine,

[0184]

[0185] "Halogen" within the scope of the present invention means fluorine, chlorine, bromine or iodine. Unless otherwise stated, fluorine, chlorine and bromine are regarded as preferred halogens.

[0186] As previously mentioned, the compounds of formula I or II can be converted into their salts, especially into their physiologically and pharmacologically acceptable salts for pharmaceutical use. The phrase "pharmaceutically acceptable" is used herein to refer to those compounds, substances, compositions and / or dosage forms that are suitable for contact with the tissues of humans and animals within the scope of reasonable medical judgment without excessive toxicity, irritation, allergic reaction or other problems or complications, and that match a reasonable benefit / risk ratio. On the one hand, these salts can exist in the form of physiologically and pharmacologically acceptable acid addition salts formed by the compounds of formula I or II with inorganic acids or organic acids. On the other hand, the compounds of formula I or II can be converted into physiologically and pharmacologically acceptable salts by reaction with inorganic bases, where alkali metal or alkaline earth metal cations act as counterions. Acid addition salts can be prepared, for example, using hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, methanesulfonic acid, p-toluenesulfonic acid, acetic acid, fumaric acid, succinic acid, lactic acid, citric acid, tartaric acid or maleic acid. Mixtures of the acids mentioned above can also be used. For the preparation of alkali metal salts and alkaline earth metal salts of the compounds of formula I or II, alkali metal and alkaline earth metal hydroxides and hydrides are preferably used, among which the hydroxides and hydrides of alkali metals (especially sodium, potassium), magnesium, calcium, zinc and diethanolamine are preferred, and sodium hydroxide and potassium hydroxide are particularly preferred.

[0187] The present invention relates to said compounds, which can optionally be in the form of individual optical isomers, diastereoisomers, mixtures of diastereoisomers, individual enantiomers or mixtures of racemates; in the form of tautomers; and in the form of the free base or the corresponding acid addition salts formed with pharmacologically acceptable acids, such as acid addition salts formed with hydrohalic acids (e.g., hydrochloric acid or hydrobromic acid) or organic acids (such as oxalic acid, fumaric acid, glycolic acid or methanoic acid).

[0188] The compounds of formula I or II according to the present invention can optionally exist in the form of a mixture of diastereoisomers, but can also be obtained in the form of pure diastereoisomers. Preferred are compounds having a specific stereochemistry of formula II or III, especially compounds of formula II.

[0189] 4. Synthetic methods

[0190] General procedure

[0191] The following methods are applicable to the preparation of compounds of general formula I or II. Synthetic methods known to those skilled in the art and described in the organic synthesis literature can be used to obtain the compounds according to the present invention. General methods for functional group protection and deprotection steps are described, for example, in Greene, T.W. and Wuts, P.G.M. (eds.): Protective Groups in Organic Synthesis, 3rd Edition 1999; John Wiley and Sons, Inc. Preferably, the compounds are obtained analogously to the preparation methods more fully explained hereinafter, especially those described in the experimental section. Compounds of general formula (I) can be prepared using several alternative synthetic routes, of which the following routes should serve as examples. Route A:

[0192] Compounds of general formula I and II, especially those of general formula I and II in which A represents -CH2- or substituted -CH2-, can be obtained from compounds of formula (A-I) via standard amidation procedures. R13 can thus represent hydrogen or a protecting group, such as tert-butyl or methyl, which can be removed by standard deprotection methods. R can represent hydrogen or a protecting group, such as tert-butyl, benzyl, methyl or ethyl, which can be removed by standard deprotection methods. Compounds of formula (A-I) can be prepared from compounds of formula (A-II) using standard deprotection methods. Compounds of formula (A-II) can be prepared by reacting compounds of formula (A-III) with compounds of formula (A-IV) in the presence of a strong base such as sodium hydride. The methods applicable to the preparation of compound (A-III), according to the search of Routes C and D described below and the examples described in the experimental section, will be obvious to those skilled in the art. Compounds of formula (A-IV) can be prepared via the methods described hereinafter for synthetic intermediate P.

[0193] Route A

[0194]

[0195] Route B:

[0196] Compounds of general formula I and II can be prepared by reacting a compound of general formula (B-I) in the presence of a strong base such as sodium hydride. R13 can thus represent hydrogen or a protecting group, such as tert-butyl, which can be removed by standard deprotection methods. Compound (B-I) can be prepared by reacting a compound of general formula (B-II) with a compound of general formula (B-III) under standard amidation conditions. According to the synthesis of intermediate P described in the experimental section of the query, the method applicable to the preparation of compound (B-III) becomes obvious to those skilled in the art. Alternatively, a compound of general formula (B-I) can be prepared by reacting a compound of general formula (B-IV) with a compound of general formula (B-V) under standard reaction conditions for transition metal-catalyzed coupling reactions (such as the Heck reaction), followed by hydrogenating the resulting olefin under standard hydrogenation conditions. Compound (B-V) can be prepared via the method described below for the synthesis of intermediate P.

[0197] Route B

[0198]

[0199] Route C:

[0200] Compounds of general formula I and II can be prepared by reacting a compound of general formula (C-I) with an activating reagent such as BOP ((1H-benzo[d][1,2,3]triazol-1-yl)oxytris(dimethylamino)phosphonium (V) hexafluorophosphate) in the presence of a base, namely DBU. (C-I) is generated from (C-II) by removing the protecting group PG under standard deprotection conditions known to those skilled in the art. PG in formula (C-II) can represent, for example, tert-butoxycarbonyl (Boc), allyloxycarbonyl (Alloc), benzyloxycarbonyl (Cbz). Compounds of general formula (C-II) can be prepared from compounds of general formula (C-III) and the respective enantiopure hydroxyproline (optionally carrying the protecting group R13), optionally in the presence of a base. Compounds of general formula (C-III) can be obtained from compounds of general formula (C-IV) under high temperature and basic conditions, such as triethylamine / TMSCl. Compounds of general formula (C-IV) can be synthesized by reacting a carboxylic acid of general formula (C-VI) with an amino-benzofuran of general formula (C-V) under standard amidation conditions.

[0201] Route C

[0202]

[0203] Route D:

[0204] Compounds of general formula I and II can be prepared from compounds of general formula (D-I) by various types of cyclization reactions, such as (but not limited to): Heck-type coupling reactions (wherein R11 represents a bromine or iodine atom and R12 contains a terminal olefin), metathesis reactions (wherein both R11 and R12 contain terminal olefins); followed by hydrogenation of the resulting olefin; amidation (wherein R11 carries a carboxylic acid and R12 carries a primary or secondary amino group, or vice versa). R13 can thus represent hydrogen or a protecting group, such as a tert-butyl group, which can be removed by standard deprotection methods. The method for synthesizing compound (D-I) becomes obvious to those skilled in the art according to the above-described Routes A and B and the examples described in the experimental section.

[0205] Route D

[0206]

[0207] Synthesis of intermediates

[0208] The synthesis described below is carried out partly according to the general procedures indicated below.

[0209] General procedure Int-A: Hydrogenation with Pd / C catalyst (see Intermediate N-09, step 4)

[0210] General procedure Int-B: Cyclization under basic conditions (see: Intermediate N-09, step 5)

[0211] General procedure Int-C: Chlorination using phosphorus oxychloride (see: Intermediate N-09, step 7)

[0212] General procedure Int-D: Use of S of hydroxyproline ester N Ar (see: Intermediate N-09, step 8)

[0213] General procedure Int-E: Ester cleavage using lithium hydroxide (see: Intermediate N-04, step 2)

[0214] General procedure Int-F: Amidation using PFTU (see: Intermediate N-04, step 3) General procedure Int-G: Heck-type coupling (see: Intermediate N-17, step 1)

[0215] General procedure A: Amidation (TBTU / NMP) (see: EX-01, step 1)

[0216] General procedure B: S N Ar (NaH / DMA) (see: EX-01, step 2)

[0217] General Procedure C: Ester Hydrolysis (LiOH / THF) (See: EX-01, Step 4)

[0218] General Procedure D: Amidation (HATU / DMF) (See: EX-01, Step 5)

[0219] General Procedure E: Amidation (HATU / DMA) (See: EX-02, Step 5)

[0220] General Procedure F: Removal of BOC Protecting Group with PTSA (See: EX-03, Step 2) General Procedure G: Macrocyclization (HATU / DMA) (See: EX-03, Step 3)

[0221] General Procedure H: Removal of tBu Protecting Group (TFA / DCM) (See: EX-04, Step 6) General Procedure I: Cyclization to Pyridine (NaH / NMP) (See: EX-05, Step 2)

[0222] General Procedure J: Amidation with DCC (See: EX-08, Step 3)

[0223] General Procedure K: Olefin Metathesis (See: EX-08, Step 4)

[0224] General Procedure L: Hydrogenation with Raney-Ni (See: EX-08, Step 5) General Procedure M: Heck Coupling (See: EX-22, Step 1)

[0225] General Procedure N: Hydrogenation with Pd / C (See: EX15, Step 3)

[0226] General Procedure O: Suzuki Coupling (See: EX-20.01, Step 1)

[0227] Intermediate N-01

[0228] (2S,4S)-1-[4-(2-Ethoxy-2-oxoethyl)-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-6-yl]-4-hydroxypyrrolidine-2-carboxylic acid tert-butyl ester

[0229] Step 1: Ethyl 2-{6-oxo-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,10,12-pentaen-4-yl}acetate

[0230] At room temperature, ethyl cyanoacetate (5.19 mL; 48.7 mmol) was added to a mixture of ethyl 3-aminobenzofuran-2-carboxylate (5.00 g; 24.4 mmol) in 4 M HCl (50.00 mL; 200 mmol). The mixture was heated at 100 °C for 4 h. After cooling to room temperature, 2.6 mL (24.4 mmol) of ethyl cyanoacetate was added again and the heating was continued at 100 °C for 48 h. The solvent was evaporated and the crude residue was diluted with 100 mL of MeOH, filtered and dried. The crude product was used directly for the next step.

[0231] ESI-MS: 273.0 [M+H] + ; R t (HPLC): 0.63 min (Method B)

[0232] Step 2: Ethyl 2-{6-chloro-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-4-yl}acetate

[0233] A mixture of ethyl 2-{6-oxo-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,10,12-pentaen-4-yl}acetate (3.35 g; 12.3 mmol) in POCl3 (50.0 mL; 547 mmol) was heated at 110 °C for 1.5 h. The reaction mixture was cooled to room temperature and added dropwise with stirring over 30 min to an ice bath (500 mL). Ethyl acetate was added and the layers were separated. Saturated NaHCO3 solution was slowly added to the organic layer and the phases were separated. The organic layer was washed with water and brine, dried over sodium sulfate, filtered and concentrated in vacuo.

[0234] ESI-MS: 291.0 [M+H] + ; R t (HPLC): 0.43 min (Method A)

[0235] Step 3: (2S,4S)-1-[4-(2-Ethoxy-2-oxoethyl)-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-6-yl]-4-hydroxypyrrolidine-2-carboxylic acid tert-butyl ester

[0236] To a solution containing 2-{6-chloro-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7Ethyl (8.17 mmol; 1.00 eq., 2.50 g) {4-bromo-8-oxa-3,5-diazatricyclo[7.4.0.0

[0237] ESI-MS: 442.0 [M+H] + ; R t (HPLC): 0.67 min (Method B)

[0238] Intermediate N-02

[0239] (2S,4S)-1-{4-bromo-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(13),2(7),3,5,9,11-hexaen-6-yl}-4-hydroxypyrrolidine-2-carboxylic acid tert-butyl ester

[0240] Step 1: 4,6-Dibromo-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(13),2(7),3,5,9,11-hexaene

[0241] A mixture of 1H-benzo[4,5]furo[3,2-d]pyrimidine-2,4-dione (11.6 g; 0.0573 mol, preparation described in KR2015 / 84657) and POBr3 (40.8 mL, 0.402 mol) was heated at 150 °C for 3 h. The mixture was cooled to room temperature, the pH was adjusted to pH = 7 using saturated aqueous NaHCO3 while cooling at 0 °C, and the mixture was extracted with EtOAc (3 × 50 mL). The organic layers were combined, dried over sodium sulfate, and evaporated under reduced pressure. The remaining residue was purified by dissolving in a mixture of DCM (3V relative to the weight of the crude material) and EtOAc (3V relative to the weight of the crude material) with stirring. After stirring at room temperature for 30 min, the mixture was filtered, and the supernatant was evaporated under reduced pressure.

[0242] ESI-MS: 327 / 329 / 331 [M+H] + (2Br); R t (HPLC): 0.67 min (Method A)

[0243] Step 2: (2S,4S)-1-{4-bromo-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(13),2(7),3,5,9,11-hexaen-6-yl}-4-hydroxypyrrolidine-2-carboxylic acid tert-butyl ester

[0244] To a mixture of 4,6-dibromo-8-oxa-3,5-diazatricyclo[7.4.0.02,7]trideca-1(13),2(7),3,5,9,11-hexaene (5.56 g, 16.9 mmol) in 93 mL of DMF was added (2S,4S)-4-hydroxypyrrolidine-2-carboxylic acid tert-butyl ester hydrochloride (4.17 g, 16.6 mmol, described in WO2005 / 35525) and K2CO3 (7.03 g, 50.8 mmol). After stirring overnight at room temperature, the reaction mixture was poured into water and neutralized with 4 M aqueous HCl. The precipitate was collected by filtration and dried in vacuo.

[0245] ESI-MS: 434 / 436 [M+H] + (2Br); R t (HPLC): 0.64 min (Method A)

[0246] Intermediate N-03:

[0247]

[0248] N-03 Step 1: (2S,4S)-1-[4-(2-ethoxy-1,1-difluoro-2-oxoethyl)-8-oxa-3,5-diazatricyclo-[7.4.0.0 2,7 trideca-1(13),2(7),3,5,9,11-hexaen-6-yl]-4-hydroxypyrrolidine-2-carboxylic acid tert-butyl ester

[0249] Copper bronze powder (508 mg, 8.00 mmol) was added to a mixture of ethyl bromodifluoroacetate (533 μL, 4.00 mmol) and Intermediate N-02 (914 mg, 2.00 mmol) in 19.4 mL of DMSO. The mixture was heated to 70 °C and stirred for 3 h. Subsequently, an additional amount of ethyl bromodifluoroacetate (533 μL, 2.00 equiv) and copper bronze powder (508 mg, 4.00 equiv) were added and stirring was continued at 70 °C for 1 h. The reaction mixture was diluted with ACN / H2O, acidified with TFA, filtered through a Celite pad and purified by preparative HPLC (P09; XBridge C18; ACN / H2O / TFA).

[0250] ESI-MS: 478 [M+H] + ; Rt (HPLC): 0.67 min (Method A)

[0251] Step 2 of N-03: 2-{6-[(2S,4S)-2-[(tert-Butoxy)carbonyl]-4-hydroxypyrrolidin-1-yl]-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-4-yl}-2,2-difluoroacetic acid

[0252] 4.34 mL of THF containing the product (ethyl ester) obtained in the previous step (217 mg, 0.432 mmol) was treated with 2 M saturated aqueous LiOH (432 μL; 2.00 equivalents, 0.864 mmol) at room temperature until the starting material was completely consumed (1 h). The reaction mixture was diluted with 10 mL of water, acidified with 1 mL of 1 N HCl and extracted with DCM. The combined organic phases were dried over sodium sulfate, filtered and evaporated. The residue was dissolved in ACN / H2O and lyophilized.

[0253] ESI-MS: 450 [M+H] + ; R t (HPLC): 0.53 min (Method A)

[0254] Step 3 of N-03: (2S,4S)-1-(4-{Difluoro[(4-methoxy-4-oxobutyl)(methyl)carbamoyl]methyl}-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-6-yl)-4-hydroxypyrrolidine-2-carboxylic acid tert-butyl ester

[0255] At room temperature, HATU (250 mg, 0.624 mmol) was added to a mixture of 2-{6-[(2S,4S)-2-[(tert-Butoxy)carbonyl]-4-hydroxypyrrolidin-1-yl]-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-4-yl}-2,2-difluoroacetic acid (422 mg, 0.939 mmol), methyl 4-(methylamino)butanoate hydrochloride (110 mg; 0.624 mmol, 1.0 equivalent, prepared as described in CN114173824) and DIPEA (231 μL, 1.34 mmol) in 3.17 mL of DMF. The reaction mixture was stirred at room temperature until the starting material was completely consumed (2.5 h), then diluted with ACN / H2O, acidified with TFA, and purified by preparative HPLC (XBridge C18; ACN / H2O / TFA).

[0256] ESI-MS: 563 [M+H] + ; R t (HPLC): 0.61 min (Method A)

[0257] N-03 Step 4: 4-(2-{6-[(2S,4S)-2-[(tert-butoxy)carbonyl]-4-hydroxypyrrolidin-1-yl]-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-4-yl}-2,2-difluoro-N-methylacetamido)-butyric acid

[0258] React the product obtained in the previous step according to the general procedure Int-E to obtain Intermediate N-03 as the final product.

[0259] ESI-MS: 450 [M+H] + ; R t (HPLC): 0.53 min (Method A)

[0260] Intermediate N-04:

[0261]

[0262] N-04 Step 1:

[0263] The zinc powder used was washed with 2% aqueous hydrochloric acid, water and acetone before use. The purified powder was dried under high vacuum and stored under argon. Charge a flask with zinc powder (2.44 g; 36.9 mmol), nickel(II) chloride hexahydrate (0.754 g; 3.14 mmol), THF (35.0 mL) and 3 drops of water, and stir the mixture at room temperature for 10 min. Subsequently, add tert-butyl hex-5-enoate (4.63 g; 18.5 mmol; prepared as described in WO2010 / 15447) in one portion, and add ethyl iodo fluoroacetate (2.80 mL; 18.5 mmol) dropwise (an exothermic reaction occurs during the addition), while maintaining the temperature below 30 °C. After the addition is complete, stir the reaction mixture at 60 °C for 4 h. Pour the reaction mixture into a mixture of saturated ammonium chloride solution (100 mL) and diethyl ether (100 mL) and stir for 10 min. Subsequently, filter the mixture through a pad of diatomaceous earth, and after phase separation, extract the aqueous phase with diethyl ether. Wash the combined organic layers with water, dry over sodium sulfate, filter and evaporate. Purify the crude product by FC (silica gel; CH / DCM 10% -> 100%).

[0264] ESI-MS: 312 [M+NH4] + Rt (HPLC): 0.78 min (Method A)

[0265] Step 2 of N-04:

[0266] General Procedure Int-E:

[0267] Lithium hydroxide (276 mg; 11.0 mmol) was added to a solution of the product of Step 1 in 2:1 THF / H2O and the reaction mixture was stirred at room temperature until the reaction control by RP HPLC indicated depletion of the starting material (here: 2.5 h). Volatiles were removed in vacuo; the residue was acidified to pH = 1 by addition of 5.50 mL of 1 N aqueous hydrochloric acid and the mixture was extracted three times with EtOAc. The combined organic phases were dried over sodium sulfate, filtered and evaporated.

[0268] ESI-MS: 211 [M - isobutene + H] + R t (HPLC): 0.59 min (Method A)

[0269] N-04 Step 3:

[0270] General Procedure Int-F:

[0271] At room temperature, PFTU (2.40 g; 5.60 mmol) was added to a stirred solution of the product of Step 2 (1.40 g; 5.09 mmol) and DIPEA (970 μL; 5.60 mmol) in DMF (21.0 mL) and the mixture was stirred for 30 min. Subsequently, 3-aminobenzofuran-2-carboxamide (1.01 g; 5.60 mmol) and an additional DIPEA (970 μL; 5.60 mmol) were added and the mixture was stirred at room temperature for 10 min. Subsequently, the reaction mixture was heated to 50 °C and stirred at this temperature for 16 h. Since the reaction control by RP HPLC indicated incomplete depletion of the starting material, additional DIPEA (441 μL; 2.80 mmol) and PFTU (0.86 g; 2.04 mmol) were added and stirring was continued at 50 °C for 2.5 h. The reaction mixture was diluted with water, acidified with TFA, filtered, and purified by means of RP HPLC (XBridge C18; ACN / water; modifier: TFA).

[0272] ESI-MS: 425 [M + H] + R t (HPLC): 0.72 min (Method A)

[0273] N-04 Step 4:

[0274] At room temperature, chlorotrimethylsilane (4.05 mL; 30.3 mmol) was slowly added to a solution of the product of step 3 (950 mg; 2.13 mmol) and triethylamine (13.0 mL; 92.4 mmol) in 1,2-dichloroethane (28.5 mL). After completion of the addition, the reaction mixture was heated to 85 °C and stirred at this temperature for 24 h. The reaction mixture was poured into 30 mL of 4 M hydrochloric acid (pH = 1) and extracted twice with DCM. The combined organic phases were washed with water, dried over sodium sulfate, filtered and evaporated to give tert-butyl 7,7-difluoro-7-{6-oxo-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(13),2(7),3,9,11-pentien-4-yl}heptanoate.

[0275] ESI-MS: 405 [M-H] - R t (HPLC): 0.44 min (method D)

[0276] N-04 Step 5:

[0277] Applying the general procedure Int-C, the product of step 4 was converted into a chlorinated product, which was then purified by RP-HPLC (XBridge C18; ACN / water; modifier: TFA).

[0278] ESI-MS: 367 [M-H] - R t (HPLC): 0.64 min (method A)

[0279] N-04 Step 6:

[0280] The product of step 5 was reacted according to the general procedure Int-D to give the title compound.

[0281] ESI-MS: 520 [M+H] + R t (HPLC): 0.61 min (method A)

[0282] Intermediate N-05

[0283]

[0284] N-05 Step 1:

[0285] To a solution of magnesium powder (45.0 g; 1.85 mol) in THF (285 mL) was added iodine (1.00 g; 3.94 mmol), and then a solution of 1-bromo-3-butene (111 g; 821 mmol) in THF (850 mL) was added dropwise. The temperature was thereby maintained below 50 °C. The resulting mixture was cooled to -75 °C and a solution of diethyl oxalate (100 g; 684 mmol; 93.5 mL) in THF (1.89 L) was added dropwise. The mixture was stirred at -75 °C for an additional 4 h and then quenched by the addition of saturated aqueous ammonium chloride (900 mL) at 0 °C. The pH was adjusted to pH 3 by the addition of aqueous hydrochloric acid (1 M). The mixture was extracted three times with EtOAc (500 mL). The combined organic layers were washed with brine (900 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure to give a residue, which was purified by FC (silica gel; petroleum ether / ethyl acetate 0% -> 100%).

[0286] 1 H NMR: (400 MHz, CDCl3) δ = 5.85 - 5.72 (m, 1H), 5.11 - 4.92 (m, 2H), 4.37 - 4.24 (m, 2H), 2.92 (t, J = 7.3 Hz, 2H), 2.36 (q, J = 7.1 Hz, 2H), 1.24 - 1.21 (m, 3H)

[0287] N-05 Step 2:

[0288] At 0 °C, bis-(2-methoxyethyl)aminosulfur trifluoride (Deoxofluor) (120 g, 544 mmol, 119 mL) and ethanol (2.95 g, 64.0 mmol) were added to a solution of the product of Step 1 (50.0 g, 320 mmol) in DCM (1000 mL). The mixture was stirred at 25 °C for 12 h. The reaction mixture was quenched by the addition of 500 mL of saturated aqueous sodium bicarbonate, and then extracted three times with DCM (500 mL). The combined organic layers were washed with aqueous hydrochloric acid (1 M; 200 mL) and brine (200 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure to give a residue. The crude product was vacuum distilled (30 °C, 0.09 MPa / oil pump).

[0289] 1 H NMR: (400 MHz, CDCl3): δ = 5.80 (br dd, J = 10.3, 16.9 Hz, 1H), 5.13 - 5.00 (m, 2H), 4.39 - 4.28 (m, 2H), 2.30 - 2.12 (m, 4H), 1.43 - 1.34 (m, 3H)

[0290] N-05 Step 3:

[0291] To a mixture of zinc powder (8.79 g, 134 mmol) and THF (30.0 mL) was added nickel(II) chloride hexahydrate (804 mg, 3.38 mmol). The mixture was stirred at -65 °C for 5 min. Subsequently, ethyl difluoroiodoacetate (12.0 g, 48.0 mmol) and the product of Step 2 (6.00 g, 33.7 mmol) were added dropwise at -65 °C. The mixture was stirred at 25 °C for 12 h. The reaction mixture was quenched by adding saturated aqueous ammonium chloride solution (60.0 mL) at 0 °C, and then extracted three times with DCM (60.0 mL). The combined organic layers were washed with brine (60.0 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure.

[0292] 1 1H NMR: (400 MHz, chloroform-d): δ = 4.33 (q, J = 7.2 Hz, 4H), 2.18 - 1.98 (m, 4H), 1.56 (td, J = 3.8, 8.0 Hz, 4H), 1.36 (t, J = 7.2 Hz, 6H) N-05 Step 4:

[0293] To a solution of the product of Step 3 (11.0 g, 36.4 mmol) in dioxane (30.0 mL) was added 1,5,7-triazabicyclo[4.4.0]dec-5-ene (14.2 g, 102 mmol) and 3-amino-1-benzofuran-2-carboxamide (4.50 g, 25.5 mmol). The mixture was stirred at 110 °C for 2 h, and then diluted with water (30.0 mL) and extracted three times with EtOAc (30.0 mL). The combined organic layers were washed with brine (30.0 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure. The crude product was purified by RP HPLC (ACN / water, modifier: TFA).

[0294] 1 1H NMR: (400 MHz, DMSO-d): δ = 8.09 (d, J = 7.7 Hz, 1H), 7.88 (d, J = 8.4 Hz, 1H), 7.72 (t, J = 7.8 Hz, 1H), 7.53 (t, J = 7.5 Hz, 1H), 2.40 - 2.37 (m, 2H), 2.12 - 2.07 (m, 2H), 1.54 - 1.48 (m, 4H).

[0295] N-05 Step 5:

[0296] Chlorinate the product of Step 4 using the general procedure Int-C.

[0297] N-05 Step 6:

[0298] The product of Step 5 was subjected to the general procedure Int-D reaction to give the title compound.

[0299] ESI-MS: 556 [M+H] +

[0300] R t (HPLC): 0.64 min (Method A)

[0301] Intermediate N-06

[0302]

[0303] N-06 Step 1:

[0304] 2-Allyloxyacetic acid (9.02 g; 73.8 mmol) was dissolved in DCM with a drop of DMF and cooled to 0 °C. Oxalyl chloride (23.4 g; 184 mmol) was added dropwise. The reaction mixture was stirred at 0 °C for 2 h. Then the solvent was evaporated under reduced pressure. The residue was dissolved in DMF and added to a stirred solution of 3-aminobenzofuran-2-carboxamide (13.0 g; 73.8 mmol) in DMF. After 2 h, the mixture was poured into 100 mL of water and stirred for 5 min. The solid formed was collected, dissolved in DCM and dried over magnesium sulfate. The volatiles were evaporated and the solid was triturated with tert-butyl methyl ether to give 3-[2-(prop-2-en-1-yloxy)acetamido]-1-benzofuran-2-carboxamide.

[0305] N-06 Step 2:

[0306] A suspension of the product of Step 1 in 4 M aqueous sodium hydroxide was stirred at 70 °C for 2 h. The mixture was acidified by adding aqueous hydrochloric acid and the precipitate formed was collected and dried to give 4-[(prop-2-en-1-yloxy)methyl]-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,10,12-pentaen-6-one.

[0307] The product of Step 2 was further reacted according to the general procedures Int-C and then Int-D in a 2-step sequence to give the title compound.

[0308] ESI-MS: 426 [M+H] + R t (HPLC): 0.49 min (Method A)

[0309] Intermediate N-07:

[0310]

[0311] Step 1 of N-07:

[0312] A mixture of 3-aminobenzofuran-2-carboxamide (2.00 g; 11.4 mmol), ethyl 2-[2-(2-ethoxy-2-oxoethoxy)ethoxy]acetate (5.32 g; 22.7 mmol) and 1H,2H,3H,4H,6H,7H,8H-[1,3]diazino[1,2-a]pyrimidine (6.45 g; 45.4 mmol) was heated to 120 °C for 150 min. After cooling to room temperature, aqueous hydrochloric acid solution (1 M; 70 mL) was added. The precipitate was filtered off with suction, washed with water and dried in vacuo at 60 °C to give a mixture of ethyl 2-[2-({6-oxo-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,10,12-pentaen-4-yl}methoxy)ethoxy]acetate and the corresponding free acid, and the mixture was used in the next step.

[0313] Step 2 of N-07:

[0314] For re-esterification of the acid, the mixture from Step 1 (2.8 g) was dissolved in DCM (200 mL). 2 drops of DMF were added, followed by oxalyl chloride (392 μL, 4.57 mmol). The mixture was stirred overnight, then ethanol (10 mL) was added and the mixture was stirred for an additional 2 h. The mixture was concentrated under reduced pressure. Methyl tert-butyl ether was added and the resulting precipitate was washed with methyl tert-butyl ether and dried at 50 °C to give ethyl 2-[2-({6-oxo-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,10,12-pentaen-4-yl}methoxy)ethoxy]acetate.

[0315] The product of Step 2 was further reacted according to the first general procedure Int-C and then Int-D in a 2-step sequence to give the title compound.

[0316] ESI-MS: 516 [M+H] + R t (HPLC): 0.80 min (Method E)

[0317] Intermediate N-08

[0318]

[0319] Step 1 of N-08:

[0320] Sodium hydride (60% in mineral oil; 1.73 g, 44.2 mmol) was added portionwise to a solution of sodium 2-chloro-2,2-difluoroacetate (4.5 g, 29.5 mmol) and prop-2-en-1-ol (2.14 g, 36.8 mmol) in THF (30 mL) under nitrogen at 0 °C over a period of 2 minutes. The resulting suspension was stirred at 65 °C for 16 h. The reaction mixture was cooled and diluted with aqueous hydrochloric acid (2 M) until the pH reached 5 to 6, and the aqueous layer was then extracted twice with DCM (30 mL). The combined organic layers were washed with brine, dried over sodium sulfate, filtered and evaporated to give the crude product. The crude product was purified by FC (silica gel; petroleum ether / EtOAc 0% -> 30%) to give 2-(allyloxy)-2,2-difluoroacetic acid.

[0321] 1 1H NMR (400 MHz, CDCl3) δ 9.34 (s, 1H), 6.01 - 5.91 (m, 1H), 5.45 - 5.36 (m, 1H), 5.33 - 5.26 (m, 1H), 4.50 (dt, J = 5.8, 1.2 Hz, 2H)

[0322] Step 2 of N-08:

[0323] At 0 °C under a nitrogen atmosphere, 3-amino-1-benzofuran-2-carboxamide (4.63 g, 26 mmol) was added to a solution of the product of Step 1 (5 g, 33 mmol) in pyridine (100 mL), followed by the dropwise addition of phosphorus oxychloride (15.3 g, 0.1 mol). The resulting mixture was stirred at room temperature overnight. The mixture was diluted with water (200 mL) and extracted three times with EtOAc (200 mL). The combined organic layers were washed with brine, dried over sodium sulfate and concentrated. The residue was purified by FC (silica gel; petroleum ether / EtOAc 15%) to give 3-(2-(allyloxy)-2,2-difluoroacetamido)benzofuran-2-carboxamide.

[0324] ESI-MS: 311 [M+H] +

[0325] Step 3 of N-08:

[0326] The product of step 2 (3 g, 9.7 mmol) was added to an aqueous sodium hydroxide solution (14.5 mL, 4 M, 58.2 mmol), and then THF (1.5 mL) was added. The reaction mixture was stirred at 70 °C for 4 h. The cooled (RT) reaction mixture was acidified to pH = 5 to 6 with aqueous hydrochloric acid (2 M), and the formed precipitate was collected and dried to give crude 2-((allyloxy)difluoromethyl)benzofuro[3,2-d]pyrimidin-4(3H)-one, which was used in the next step without further purification.

[0327] N-08 Step 4:

[0328] DMF (1.36 g, 18.6 mmol) and DCM (250 mL) were charged into a 0.5 L three-necked flask equipped with a thermometer and a nitrogen balloon. The flask was cooled to 0 °C, and then a solution of oxalyl chloride (3.54 g, 27.9 mmol) in DCM (5 mL) was added dropwise over 5 min while maintaining the temperature at 0 °C to 5 °C. The mixture was stirred at ambient temperature for 0.5 h. The reaction mixture was cooled to 0 °C in an ice-water bath and the product of step 3 (1.8 g, 6.2 mmol) was added portionwise. The reaction mixture was stirred at room temperature for 15 min and then at 40 °C for 2 h. After cooling to room temperature, the reaction mixture was poured into ice, neutralized with aqueous sodium bicarbonate, and extracted twice with DCM (100 mL). The combined organic phases were washed with water, dried over sodium sulfate and concentrated. The crude product was purified by FC (silica gel; petroleum ether / EtOAc 2% -> 10%) to give the title compound 2-((allyloxy)difluoromethyl)-4-chlorobenzofuro[3,2-d]pyrimidine.

[0329] 1 H NMR (400 MHz, DMSO-d6) δ 8.41 - 8.31 (m, 1H), 7.79 (m, 2H), 7.60 - 7.54 (m, 1H), 7.26 (s, 1H), 6.06 (dq, J = 10.8, 6.0 Hz, 1H), 5.45 (dd, J = 17.2, 1.2 Hz, 1H), 5.30 (dd, J = 10.4, 1.0 Hz, 1H), 4.70 (d, J = 6.0 Hz, 2H) N-08 Step 5:

[0330] The product of step 4 was reacted according to the general procedure Int-D to give the title compound. ESI-MS: 462 [M+H] +

[0331] R t (HPLC): 0.68 min (Method A)

[0332] Intermediate N-09:

[0333]

[0334] Step 1 of N-09:

[0335] A mixture of tert-butyl 3-(2-oxoethoxy)propionate (7.25 g; 38.5 mmol) prepared as described in EP2409977 and methyl (triphenylphosphoranylidene)acetate (13.1 g; 38.5 mmol) in DCM (200 mL) was stirred overnight at room temperature. The mixture was evaporated under reduced pressure and dissolved in CH / EtOAc (3:1). The insoluble material was removed by filtration and the filtrate was evaporated. The crude product was purified by FC (silica gel; CH / EtOAc 10% -> 45%) to give the product as a mixture of cis and trans isomers.

[0336] Step 2 of N-09:

[0337] The product of Step 1 (1.50 g; 6.14 mmol) was stirred into a mixture of DCM (15 mL) and TFA (10 mL) overnight. The mixture was evaporated and dissolved in methanol (10 mL). Polymer-bound tetraalkylammonium carbonate (2 weight equivalents) was added and the mixture was stirred for 90 min. The insoluble material was filtered off and the filtrate was evaporated.

[0338] Step 3 of N-09:

[0339] 1-Chloro-N,N,2-trimethylpropenylamine (4.45 mL; 33.6 mmol) was added to a solution of the product of Step 2 (5.00 g; 21.3 mmol) in ACN (140 mL). The mixture was stirred at room temperature for 10 min, then pyridine (5.10 mL; 63.0 mmol) and 3-aminobenzofuran-2-carboxamide (3.70 g; 21.0 mmol) were added. The mixture was stirred overnight at room temperature, then water was added. The mixture was extracted with DCM, and the organic layer was separated and evaporated. The crude product was purified first by FC (silica gel; petroleum ether / EtOAc 40% -> 80%) and secondly by RP HPLC (Sunfire C18, ACN / water, modifier: TFA).

[0340] ESI-MS: 347 [M+H] + R t (HPLC): 0.80 min (Method C)

[0341] Step 4 of N-09: General Procedure Int-A:

[0342] A mixture of the product from Step 3 (3.40 g; 9.73 mmol), palladium / carbon (10%; 350 mg), and ethanol (500 mL) was shaken under a hydrogen pressure of 50 psi until RP HPLC indicated that the starting material had been converted (here: 90 min). The catalyst was filtered off and the filtrate was evaporated to dryness.

[0343] ESI-MS: 349 [M+H] + R t (HPLC): 0.81 min (Method C)

[0344] N-09 Step 5:

[0345] General Procedure Int-B:

[0346] A mixture of the product from Step 4 (4.55 g; 13.1 mmol) and aqueous sodium hydroxide (4 M; 100 mL; 400 mmol) was stirred at 60 °C until RP HPLC reaction control indicated that the starting material had been converted (here: 60 min). The mixture was cooled to room temperature and then acidified by addition of aqueous hydrochloric acid (4 M). The precipitate was collected and dried at 60 °C.

[0347] ESI-MS: 317 [M+H] + R t (HPLC): 0.73 min (Method C)

[0348] N-09 Step 6:

[0349] At room temperature, oxalyl chloride (1.24 mL; 14.5 mmol) was added to a mixture of the product from Step 5 (3.53 g; 11.2 mmol), DCM (60 mL), and a few drops of DMF. The mixture was stirred at room temperature for 3 h, then methanol was added and stirring was continued for 60 min. The mixture was extracted with water, and the organic layer was separated and evaporated to dryness.

[0350] ESI-MS: 331 [M+H] + R t (HPLC): 0.82 min (Method C)

[0351] N-09 Step 7:

[0352] General Procedure Int-C:

[0353] A mixture of the product from Step 6 (3.70 g; 11.2 mmol) and phosphorus oxychloride (70 mL) was stirred at 90 °C for 4 h. The excess phosphorus oxychloride was removed by distillation, and water was carefully added. The resulting mixture was extracted with EtOAc, the organic layer was separated and evaporated. The crude product was used in the next step.

[0354] ESI-MS: 349 [M+H] + R t (HPLC): 1.02 min (Method C)

[0355] Step 8 of N-09:

[0356] General Procedure Int-D:

[0357] Stir a mixture of the product of Step 7 (300 mg; 0.896 mmol), (2S,4S)-tert-butyl 4-hydroxypyrrolidine-2-carboxylate hydrochloride (253 mg; 1.08 mmol), potassium carbonate (300 mg; 2.06 mmol), and DMF (7.0 mL) at room temperature overnight. Add water and acidify the mixture by adding aqueous hydrochloric acid (1 M). Extract the mixture with EtOAc, evaporate the organic layer, and purify the crude product by FC (silica gel; petroleum ether / EtOAc 40% -> 75%).

[0358] ESI-MS: 500 [M+H] + R t (HPLC): 0.75 min (Method C)

[0359] Step 9 of N-09:

[0360] React the product of Step 8 according to General Procedure Int-E to give the title compound with ester cleavage.

[0361] ESI-MS: 486 [M+H] + R t (HPLC): 0.70 min (Method C)

[0362] Intermediate N-10

[0363]

[0364] Prepared from Intermediate N-02 and methyl (2S)-3-allyloxy-2-methylpropanoate in a 3-step sequence similar to that described for the synthesis of N-17.

[0365] ESI-MS: 500.3 [M+H] + R t (HPLC): 2.44 min (Method H)

[0366] Intermediate N-11

[0367]

[0368] It is prepared from intermediate N-02 and methyl (2R)-3-allyloxy-2-methylpropanoate in a 3-step sequence similar to that described for the synthesis of N-17.

[0369] ESI-MS: 500.6 [M+H] + R t (HPLC): 2.44 min (method H)

[0370] Intermediate N-12

[0371]

[0372] It is prepared from intermediate N-02 and 2-(but-3-en-1-yloxy)acetic acid using the following two-step sequence:

[0373] Step 1: According to the general procedure Int-G

[0374] Step 2: According to the general procedure Int-A, the title compound is obtained.

[0375] ESI-MS: 486 [M+H] + R t (HPLC): 0.54 min (method B)

[0376] Intermediate N-13

[0377]

[0378] It is prepared from intermediate N-18 and 2-(but-3-en-1-yloxy)acetic acid using the following two-step sequence:

[0379] N-13 Step 1: According to the general procedure Int-G

[0380] N-13 Step 2: According to the general procedure Int-A, the title compound is obtained.

[0381] ESI-MS: 504 [M+H] + R t (HPLC): 0.45 min (method A)

[0382] Intermediate N-14: 2-(4-{6-[(2S,4S)-2-[(tert-butoxy)carbonyl]-4-hydroxypyrrolidin-1-yl]-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(13),2,4,6,9,11-hexaen-4-yl}-4,4-difluorobutoxy)acetic acid

[0383]

[0384] N-14 Step 1:

[0385] Equip a dry reaction vessel with a magnetic stir bar and charge it with tert-butyl 2-allyloxyacetate (1.00 g; 5.81 mmol), nickel(II) chloride anhydrous (0.038 g; 0.290 mmol), and sodium carbonate (0.61 g; 5.81 mmol). Subsequently, briefly evacuate the reaction vessel and backfill with argon (this sequence is repeated a total of three times). Sequentially add anhydrous DMF (40 mL), ethyl bromodifluoroacetate (1.5 mL, 11.6 mmol), and phenylsilane (2.9 mL; 23.2 mmol) to the reaction vessel via syringe. Heat the vessel in an oil bath at 70 °C and stir until TLC monitoring indicates depletion of the starting materials (here: overnight). Dilute the reaction mixture with 30 mL of EtOAc, and wash the organic layer with 80 mL of saturated aqueous sodium chloride. Thereafter, dry the organic layer over sulfate and concentrate under reduced pressure, and further purify by FC (silica gel; hexane / EtOAc) to afford ethyl 5-(2-tert-butoxy-2-oxo-ethoxy)-2,2-difluoro-pentanoate.

[0386] N-14 Step 2:

[0387] At room temperature, add 1,5,7-triazabicyclo[4.4.0]dec-5-ene (0.79 g; 5.68 mmol) to a solution of 3-aminobenzofuran-2-carboxamide (0.25 g; 1.42 mmol) in 1,4-dioxane (1 mL). Subsequently, add the product of Step 1 (0.84 g; 2.84 mmol), raise the temperature to 120 °C and continue stirring until TLC indicates nearly complete conversion (here: 18 h). Dilute the reaction mixture with water (15 mL) and extract with DCM (3 × 7 mL). Adjust the pH of the resulting aqueous layer to between 4 and 5 and filter off the precipitated solid.

[0388] ESI-MS: 353 [M+H] + R t (HPLC): 1.61 min (Method F)

[0389] N-14 Step 3:

[0390] Subject the product of Step 2 to the General Procedure Int-C reaction.

[0391] ESI-MS: 371 [M+H] + R t (HPLC): 1.82 min (Method F)

[0392] N-14 Step 4:

[0393] Subject the product of Step 3 to the general procedure Int-D reaction to obtain the title compound.

[0394] ESI-MS: 522.6 [M+H] + R t (HPLC): 2.96 min (Method E)

[0395] Intermediate N-15

[0396]

[0397] Prepared using a procedure similar to that described for the synthesis of Intermediate N-14, applying 3-amino-6-fluoro-1-benzofuran-2-carboxamide as the starting material in Step 2.

[0398] The starting material 3-amino-6-fluoro-1-benzofuran-2-carboxamide was prepared as follows:

[0399] Dissolve 4-fluoro-2-hydroxybenzonitrile (2.06 g; 14.3 mmol) in ethanol (80 mL). Add potassium carbonate (3.02 g; 21.8 mmol) and 2-bromoacetamide (2.40 g; 17.4 mmol) and heat the mixture to 78 °C for 1 h. Add potassium hydroxide (powder; 1.91 g; 28.97 mmol), stir and continue heating overnight. Cool the mixture to room temperature, add water, evaporate the ethanol, and filter the precipitate, wash with water and dry in air.

[0400] Intermediate N-16

[0401]

[0402] Prepared using a procedure similar to that described for the synthesis of Intermediate N-14, applying 3-amino-6-chloro-1-benzofuran-2-carboxamide (prepared as described in EP1710233) as the starting material in Step 2.

[0403] Intermediate N-17

[0404]

[0405] Prepared from Intermediate N-02 using the following 3-step sequence: N-17 Step 1:

[0406] General procedure Int-G:

[0407] The intermediate N-02 (300 mg; 0.69 mmol) and methyl 5-hexenoate (298 μL; 2.07 mmol) were dissolved in DMF (10 mL; 123 mmol). Triethylamine (0.39 mL; 2.76 mmol) was added and the mixture was degassed with argon. Palladium(II) acetate (31 mg; 0.14 mmol) and tris(o-tolyl)phosphine (84 mg; 0.28 mmol) were added under argon. The sealed vial was stirred overnight at 95 °C. The mixture was diluted with ACN / water, acidified with TFA, and filtered through a syringe filter. Purification was carried out by preparative RP HPLC (Sunfire C18, ACN / water; modifier: TFA) to give tert-butyl (2S,4S)-4-hydroxy-1-{4-[6-methoxy-6-oxohex-1-en-1-yl]-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(13),2,4,6,9,11-hexaen-6-yl}pyrrolidine-2-carboxylate.

[0408] N-17 Step 2:

[0409] The product of Step 1 was hydrogenated according to General Procedure Int-A to give tert-butyl (2S,4S)-4-hydroxy-1-[4-(6-methoxy-6-oxohexyl)-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(13),2,4,6,9,11-hexaen-6-yl]pyrrolidine-2-carboxylate.

[0410] ESI-MS: 484 [M+H] + R t (HPLC): 0.65 min (Method B)

[0411] N-17 Step 3:

[0412] The product of Step 2 was reacted according to General Procedure Int-E to give the title compound with ester cleavage.

[0413] Intermediate N-18: tert-butyl (2S,4S)-1-{4-bromo-11-fluoro-8-oxa-3,5-diazatricyclo-[7.4.0.0 2,7 trideca-1(13),2,4,6,9,11-hexaen-6-yl}-4-hydroxypyrrolidine-2-carboxylate

[0414]

[0415] Similar to the reaction sequence described for the synthesis of intermediate N-02, from 11-fluoro-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7Prepare 13-1(13),2(7),9,11-tetraene-4,6-dione. Similar to the synthesis of 11-chloro-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 -13-1(13),2(7),9,11-tetraene-4,6-dione described in WO2019059577, prepare the starting material 11-fluoro-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 13-1(13),2(7),9,11-tetraene-4,6-dione.

[0416] Intermediate P-01

[0417] (3S,4R)-4-(5-chloro-2-fluoropyridin-3-yl)-4-fluoro-3-methylpiperidine-1-carboxylic acid tert-butyl ester

[0418]

[0419] Step 1 of P-01: 4-(5-chloro-2-fluoropyridin-3-yl)-4-hydroxy-3-methylpiperidine-1-carboxylic acid tert-butyl ester

[0420] Under an argon atmosphere, cool a solution of 3-bromo-5-chloro-2-fluoropyridine (10.0 g, 48.0 mmol, available from Activate, MFCD04972409) in THF (150 mL; 15.0 V) to -78 °C. Subsequently, add dropwise isopropylmagnesium chloride-lithium chloride complex (1.3 M in THF, 38.4 mL, 50.0 mmol) while maintaining the temperature below -65 °C. Subsequently, stir the reaction mixture at -78 °C for an additional 50 min. Subsequently, add dropwise a premixed solution of tert-butyl 3-methyl-4-oxopiperidine-1-carboxylate (11.6 g; 55.0 mmol, prepared as described in WO2011 / 159852) and lanthanum(III) chloride-lithium chloride complex (0.6 M in THF, 7.92 mL, 5.00 mmol) in THF (20.0 mL; 2.00 V) while maintaining the temperature below -65 °C, and after completion of the addition, allow the reaction mixture to reach room temperature overnight.

[0421] Quench the reaction mixture with 20 mL of saturated NH4Cl solution and remove THF in vacuo. Acidify the mixture with 6 mL of 1N HCl and extract three times with EtOAc. Dry the combined organic phases over sodium sulfate and concentrate in vacuo. Co-evaporate the residue with 15 mL of toluene three times. The crude product is used in the next step without further purification.

[0422] ESI-MS: 289.0 / 291.0 [M+H] + ; R t(HPLC): 0.64 / 0.68 min (Method A)

[0423] 1H NMR: (400 MHz, CDCl3): δ ppm 8.02 - 8.11 (m, 2H) 3.80 - 4.20 (m, 2H) 3.01 - 3.20 (m, 1H) 2.68 - 2.91 (m, 1H) 2.25 - 2.49 (m, 2H) 1.99 (s, 1H) 1.48 (s, 9H) 0.65 (d, J = 6.80 Hz, 3H).

[0424] Step 2 of P-01: tert-Butyl 4-(5-chloro-2-fluoropyridin-3-yl)-4-fluoro-3-methylpiperidine-1-carboxylate

[0425] To a solution of tert-butyl 4-(5-chloro-2-fluoropyridin-3-yl)-4-hydroxy-3-methylpiperidine-1-carboxylate (5.00 g, 15.0 mmol) in anhydrous DCM (70 mL, 14.0 V) was added dropwise DAST (3.83 mL, 29.0 mmol) while cooling the reaction mixture in an ice-MeOH bath. The reactants were kept in the ice-MeOH bath with stirring for 3.5 h, quenched by adding saturated NaHCO3 solution and extracted three times with DCM. The combined organic layers were dried over sodium sulfate, filtered and evaporated, and the crude material was further purified by flash column chromatography (n-heptane / EA = 50 / 1 to 5 / 1).

[0426] ESI-MS: 291.0 / 291.0 [M-isobutene]+, chlorine isotope pattern, R t (HPLC): 0.75 min (racemic-trans) and 0.80 min (racemic-cis) (Method A)

[0427] Step 3 of P-01: (3S,4R)-tert-Butyl 4-(5-chloro-2-fluoropyridin-3-yl)-4-fluoro-3-methylpiperidine-1-carboxylate

[0428] The cis racemate obtained in Step 2 was purified by preparative HPLC (neutral conditions) and then further separated by chiral SFC (column: REGIS (s,s) WHELK-O1 (250 mm * 50 mm, 10 um); mobile phase: [0.1% NH3H2O IPA]; B%: 11% - 11%, 2.3 min).

[0429] Intermediate P-01 (3S,4R) was the first elution peak under these conditions.

[0430] 1H NMR (400 MHz, CDCl3): δ ppm 8.10 - 8.14 (m, 1H) 7.93 (dd, J = 8.19, 2.56 Hz, 1H) 4.08 (br s, 2H) 3.11 (br s, 1H) 2.82 (br s, 1H) 2.18 - 2.49 (m, 2H) 1.83 (br t, J = 12.19 Hz, 1H) 1.50 (s, 9H) 0.71 (d, J = 6.75 Hz, 3H). -19F NMR (400 MHz, CDCl3): δ ppm -69.5, -179.4。

[0431] The absolute configuration of this intermediate was confirmed by single crystal X-ray diffraction.

[0432] Intermediate P-02

[0433] 5-Chloro-2-fluoro-3-[(3S,4R)-4-fluoro-3-methylpiperidin-4-yl]pyridine hydrochloride

[0434]

[0435] (3S,4R)-tert-Butyl 4-(5-chloro-2-fluoropyridin-3-yl)-4-fluoro-3-methylpiperidine-1-carboxylate (8.27 mmol, 3.02 g) was dissolved in 1,4-dioxane (10.3 mL, 41.4 mmol) containing 4N HCl, and the reaction mixture was stirred at room temperature for 1 h. The reaction mixture was diluted with ether, and the formed precipitate was collected by filtration, washed with ether and dried.

[0436] ESI-MS: 247 / 249 (1Cl) [M + H] + ; R t (HPLC): 0.33 min (Method A) Intermediate P-03:

[0437] tert-Butyl 4-(5-chloro-2-fluoropyridin-3-yl)-4-fluoropiperidine-1-carboxylate

[0438]

[0439] This intermediate was prepared from tert-butyl 4-oxopiperidine-1-carboxylate as the starting material according to the synthesis of Intermediate P-01 (Steps 1 - 2).

[0440] Step 1 of P-03: tert-Butyl 4-(5-chloro-2-fluoropyridin-3-yl)-4-hydroxypiperidine-1-carboxylate

[0441] ESI-MS: 275 / 277 (1Cl) [M + H] + ; R t(HPLC): 0.61 min (Method A)

[0442] Step 2 of P-03: tert-Butyl 4-(5-chloro-2-fluoropyridin-3-yl)-4-fluoropiperidine-1-carboxylate

[0443] ESI-MS: 333 / 335 (1Cl) [M+H] + ; R t (HPLC): 0.74 min (Method A) Intermediate P-04:

[0444] (3S,4R)-tert-Butyl 4-(5-bromo-2-fluoropyridin-3-yl)-4-fluoro-3-methylpiperidine-1-carboxylate

[0445]

[0446] This intermediate was prepared starting from 3,5-dibromo-2-fluoropyridine according to the synthesis of Intermediate P-01 (Steps 1-3). Analytical data for Step 2: ESI-MS: 335 / 337 [M-isobutene]+, chlorine isotope pattern, Rt (HPLC): 0.75 min (racemic-cis) and 0.80 min (racemic-trans) (Method A).

[0447] Chiral separation conditions for the cis enantiomer: Separated by chiral SFC (column: DAICEL CHIRALPAK IG (250 mm * 50 mm, 10 μm); mobile phase: MeOH [0.1% NH3] / CO2; 15 / 85).

[0448] Intermediate P-04 (3S,4R) is the second elution peak under these conditions.

[0449] The absolute stereochemistry of these products was finally assigned retrospectively by X-ray based on Example EX-17.

[0450] Intermediates P-05 and P-06:

[0451]

[0452] P-05: (3S,4R)-4-(2-{[(3S,5S)-1-{4-bromo-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-6-yl}-5-[(tert-butoxy)carbonyl]pyrrolidin-3-yl]oxy}-5-chloropyridin-3-yl)-4-fluoro-3-methylpiperidine-1-carboxylate

[0453] This intermediate was prepared according to the general procedure Int-D from intermediates P-01 and N-02.

[0454] ESI-MS:760 / 762 / 764(Cl,Br)[M+H] + ; R t (HPLC):0.99min (Method A)

[0455] P-06: (3S,4R)-4-(2-{[(3S,5S)-5-[(tert-butyloxy)carbonyl]-1-[4-(1,1-difluoro-2-methoxy-2-oxoethyl)-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 ]13-(9),2(7),3,5,10,12-hexaen-6-yl]pyrrolidin-3-yl]oxy}-5-chloropyridin-3-yl)-4-fluoro-3-methylpiperidin-1-carboxylic acid tert-butyl ester

[0456] Under argon, at room temperature, molecular sieves Add to a degassed solution of intermediate P-05 (240 mg, 0.300 mmol) in 4.26 mL DMSO, followed by methyl bromodifluoroacetate (89.7 μL, 0.792 mmol) and bronze powder (101 mg, 1.52 mmol). After stirring at room temperature for 4 days, the reaction mixture was diluted with 10 mL EA and quenched with aqueous KH2PO4 (1.27 M, 5.00 mL). The mixture was filtered through a pad of celite and the solids were washed with EA. The aqueous phase was extracted with EA. The combined organic phases were washed with water, dried over sodium sulfate, filtered and evaporated to give the crude product. The residue was dissolved in DMF / ACN, acidified with 10% TFA and purified by preparative HPLC (XBridge C18; 60-100% ACN / H2O / TFA).

[0457] ESI-MS:790 / 792(1Cl)[M+H] + ; R t (HPLC): 1.00 min (Method A) Intermediate P-07: (3S, 4R)-4-(5-chloro-2-fluoropyridin-3-yl)-4-fluoro-3-methylpiperidine-1-carboxylic acid pent-4-en-1-yl ester

[0458]

[0459] At 80 °C, a mixture of intermediate P-02 (100 mg, 0.279 mmol), DIPEA (144 μL, 0.892 mmol), and 4-nitrophenyl pent-4-ene-1-yl carbonate (94 mg, 0.200 mmol, prepared as described in WO2014 / 11769) in 1.00 mL of THF was stirred for 2 h. Volatiles were removed in vacuo, and the crude mixture was purified by preparative HPLC (ACN, Sunfire, TFA, narrow).

[0460] ESI-MS: 359 [M+H] + ; R t (HPLC): 1.11 min (Method B)

[0461] Intermediate P-08: 3-{2-[(3S,4R)-4-(5-chloro-2-fluoropyridin-3-yl)-4-fluoro-3-methylpiperidin-1-carbonyloxy]ethoxy}propanoic acid

[0462]

[0463] Step 1 of P-08: tert-Butyl 3-(2-{[(4-nitrophenoxy)carbonyl]oxy}ethoxy)propanoate

[0464] tert-Butyl 3-(2-hydroxyethoxy)propanoate (5.10 g, 26.8 mmol, prepared as described in WO2019 / 195609) was dissolved in 40.0 mL of DCM, pyridine (2.16 mL, 26.9 mmol) was added, and the mixture was cooled to 0 °C. At this temperature, a solution of 4-nitrophenyl chloroformate (5.57 g, 26.8 mmol) in 20.0 mL of DCM was added dropwise. Once the addition was complete, the reaction mixture was warmed to room temperature and stirred for 2 h. Water (150 mL) was added and the layers were separated. The organic layer was washed with water and dried over sodium sulfate, filtered, and concentrated in vacuo. The residue was purified by silica gel chromatography (CH / EtOAc = 95 / 5 -> 80 / 20).

[0465] ESI-MS: 378 [M+Na] + , R t (HPLC): 0.93 min (Method B)

[0466] Step 2 of P-08: tert-Butyl 3-{2-[(3S,4R)-4-(5-chloro-2-fluoropyridin-3-yl)-4-fluoro-3-methylpiperidin-1-carbonyloxy]ethoxy}propanoate

[0467] To a mixture of the product obtained in Step 1 of P-08 (3.24 g, 7.31 mmol) and intermediate P-02 (2.30 g, 8.12 mmol) in 46 mL of THF was added DIPEA (4.36 mL), and the mixture was heated to 65 °C for 2 h. The reaction mixture was cooled to room temperature and diluted with EtOAc. The phases were separated and the organic phase was washed with dilute sodium hydroxide solution. The organic layer was dried over sodium sulfate and evaporated to dryness.

[0468] ESI-MS: 463 [M+H] + , R t (HPLC): 1.05 min (Method B)

[0469] Step 3 of P-08: 3-{2-[(3S,4R)-4-(5-chloro-2-fluoropyridin-3-yl)-4-fluoro-3-methylpiperidin-1-carbonyloxy]ethoxy}propanoic acid

[0470] This step was carried out according to General Procedure H.

[0471] ESI-MS: 407 [M+H] + , R t (HPLC): 0.79 min (Method B)

[0472] Intermediate P-09

[0473] 5-Bromo-2-fluoro-3-[(3S,4R)-4-fluoromethylpiperidin-4-yl]pyridine hydrochloride

[0474]

[0475] This intermediate was prepared starting from P-04 according to the synthesis of intermediate P-02.

[0476] ESI-MS: 291 / 293 (1Br) [M+H] + ; R t (HPLC): 0.35 min (Method A)

[0477] Synthesis of the Examples

[0478] The syntheses described below were carried out in part according to the general procedures indicated below.

[0479] General Procedure A: Amidation (TBTU / NMP) (see: Step 1 of EX-01)

[0480] General Procedure B: S N Ar (NaH / DMA) (see: Step 2 of EX-01)

[0481] General Procedure C: Ester Hydrolysis (LiOH / THF) (See: Step 4 of EX-01)

[0482] General Procedure D: Amidation (HATU / DMF) (See: Step 5 of EX-01)

[0483] General Procedure E: Amidation (HATU / DMA) (See: Step 5 of EX-02)

[0484] General Procedure F: Removal of BOC protecting group using PTSA (See: EX-03, Step 2) General Procedure G: Macrolactamization (HATU / DMA) (See: Step 3 of EX-03)

[0485] General Procedure H: Removal of tBu protecting group (TFA / DCM) (See: Step 6 of EX-04) General Procedure I: Cyclization to pyridine (NaH / NMP) (See: Step 2 of EX-05)

[0486] General Procedure J: Amidation using DCC (See: Step 3 of EX-08)

[0487] General Procedure K: Olefin metathesis (See: Step 4 of EX-08)

[0488] General Procedure L: Hydrogenation using Raney-Ni (See: Step 5 of EX-08)

[0489] General Procedure M: Heck coupling (See: Step 1 of EX-22)

[0490] General Procedure N: Hydrogenation using Pd / C (See: Step 3 of EX15)

[0491] General Procedure O: Suzuki coupling (See: Step 1 of EX-20.01)

[0492] Example EX-01

[0493] (1R,9S,11S,34S)-4-chloro-1-fluoro-34-methyl-26,31-dioxo-8,15-dioxa-6,12,23,27,32,37-hexaaza-heptacyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2,4,6,13(37),14(22),16(21),17,19,23-nonaene-11-carboxylic acid

[0494]

[0495] EX-01 Step 1: tert-Butyl N-{4-[(3S,4R)-4-(5-chloro-2-fluoropyridin-3-yl)-4-fluoro-3-methylpiperidin-1-yl]-4-oxobutyl}carbamate

[0496] General Procedure A: Amidation (TBTU / NMP)

[0497] To a solution of 4-{[(tert-butoxy)carbonyl]amino}butanoic acid (BOC-GABA-OH) (59.8 mg, 0.29 mmol) in 1.5 mL of 1-methyl-2-pyrrolidone was added DIPEA (270 μL, 1.20 mmol), followed by TBTU (94.5 mg, 0.290 mmol), and the mixture was stirred at room temperature for 5 min. Subsequently, Intermediate P-02 was added and the reaction mixture was stirred overnight at room temperature. Water and ethyl acetate were added, the phases were separated and the aqueous phase was extracted with ethyl acetate. The combined organic phases were washed successively with water, 5% LiCl solution, saturated NaHCO3 solution and brine, dried over sodium sulfate and evaporated to dryness.

[0498] ESI-MS: 432 / 434 [M+H] + (1Cl); R t (HPLC): 0.70 min (Method A) EX-01 Step 2:

[0499] General Procedure B: S N Ar (NaH / DMA)

[0500] (2S,4S)-4-({3-[(3S,4R)-1-(4-{[(tert-butoxy)carbonyl]amino}butanoyl)-4-fluoro-3-methylpiperidin-4-yl]-5-chloropyridin-2-yl}oxy)-1-[4-(2-ethoxy-2-oxoethyl)-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-6-yl]pyrrolidine-2-carboxylic acid tert-butyl ester

[0501] This reaction was carried out under an argon atmosphere. To a stirred, ice / water bath-cooled solution of hydroxyproline ester N-01 (100 mg, 0.220 mmol) and the product obtained in Step 1 (120 mg, 0.270 mmol) in 2.00 mL of DMA was added sodium hydride (60% dispersion in mineral oil, 25.8 mg, 0.65 mmol). Stirring was continued at 0 °C until the hydroxyproline was exhausted (15 min). Water was added dropwise with ice cooling, followed by EtOAc and water. The mixture was acidified and the phases were separated. The organic phase was washed with brine, dried over sodium sulfate, filtered and evaporated to dryness in vacuo.

[0502] ESI-MS: 853 [M+H] + ; R t (HPLC): 1.18 min (Method B)

[0503] EX-01 Step 3: (2S,4S)-4-({3-[(3S,4R)-1-(4-aminobutanoyl)-4-fluoro-3-methylpiperidin-4-yl]-5-chloropyridin-2-yl}oxy)-1-[4-(2-ethoxy-2-oxoethyl)-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-6-yl]pyrrolidine-2-carboxylic acid tert-butyl ester

[0504] The product obtained in Step 2 (158 mg, 0.185 mmol) was reacted with dioxane containing 4N HCl (0.14 mL, 0.555 mmol) in a mixture of 10 mL of dioxane overnight at room temperature. Another 0.05 mL (0.18 mmol) of dioxane containing 4N HCl was added, and another 0.05 mL (0.18 mmol) was added after 4 h. Water was added to the reaction mixture, and the volatiles were removed under reduced pressure. The residue was co-evaporated with toluene, and the crude product was further purified by preparative HPLC (Sunfire, ACN / TFA, narrow).

[0505] ESI-MS: 753 / 755 [M+H] + (1Cl); R t (HPLC): 0.83 min (Method B) EX-01 Step 4: 2-{6-[(2S,4S)-4-({3-[(3S,4R)-1-(4-aminobutanoyl)-4-fluoro-3-methylpiperidin-4-yl]-5-chloropyridin-2-yl}oxy)-2-[(tert-butoxy)carbonyl]pyrrolidin-1-yl]-8-oxa-3,5-diazatricyclo-[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-4-yl}acetic acid

[0506] General Procedure C: Ester Hydrolysis (LiOH / THF)

[0507] At room temperature, 2M aqueous LiOH solution (0.223 mL, 0.440 mmol) was added to a mixture of the product obtained in Step 3 (59 mg, 70.0 mmol) in 0.59 mL of THF, and the reaction mixture was stirred overnight. The reaction mixture was diluted with EA and water, acidified with acetic acid. The phases were separated, the organic phase was washed with water and brine, dried over sodium sulfate, filtered and evaporated to dryness.

[0508] ESI-MS: 725 / 727 [M+H] + (1Cl); R t (HPLC): 0.75 min (Method B) EX-01 Step 5: (1R,9S,11S,34S)-4-chloro-1-fluoro-34-methyl-26,31-dioxo-8,15-dioxa-6,12,23,27,32,37-hexaazacycloheptacyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2,4,6,13(37),14(22),16(21),17,19,23-nonaene-11-yl tert-butyl carbonate

[0509] General Procedure D: Amidation (HATU / DMF)

[0510] At room temperature, with stirring, the product obtained in Step 4 (39 mg, 0.050 mmol) and a solution of DIPEA (19 μL, 0.11 mmol) in 2.50 mL of DMF were slowly added to a mixture of HATU (25 mg, 0.065 mmol) in 2.50 mL of DMF. The mixture was stirred at room temperature for 1 h, then acidified with acetic acid and diluted with ACN. The crude product was purified using HPLC (Sunfire, ACN, TFA, narrow).

[0511] R t (HPLC): 0.75 min (Method B)

[0512] EX-01 Step 6: (1R,9S,11S,34S)-4-chloro-1-fluoro-34-methyl-26,31-dioxo-8,15-dioxa-6,12,23,27,32,37-hexaaza-heptacyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2,4,6,13(37),14(22),16(21),17,19,23-nonaenoic acid

[0513] React 2.00 mL of DCM containing the product obtained in Step 5 (15 mg, 0.021 mmol) with TFA (16 μL, 0.21 mmol) at room temperature and stir the mixture overnight. Add TFA (32 μL, 0.42 mmol) again and stir the mixture for 4 h. Add TFA (16 μL, 0.21 mmol) again and stir the mixture for 3 days. Dilute the mixture with ACN and water, filter and purify directly by preparative HPLC (Sunfire, ACN, TFA, narrow).

[0514] ESI-MS: 649 / 651 [M+H] + (1Cl); R t (HPLC): 0.85 min (Method B) Example 02

[0515] (1R,9S,11S,34S)-4-chloro-1-fluoro-34-methyl-26,31-dioxo-8,15,30-trioxa-6,12,23,27,32,37-hexaazacycloheptatriaconta 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2,4,6,13(37),14(22),16(21),17,19,23-nonaene-11-carboxylic acid

[0516]

[0517] EX-02 Step 1: 2-{[(tert-Butoxy)carbonyl]amino}ethyl (3S,4R)-4-(5-chloro-2-fluoropyridin-3-yl)-4-fluoro-3-methylpiperidine-1-carboxylate

[0518] Mix 2.70 mL of THF, P-02 (270 mg, 0.906 mmol), 1-{[(2-{[(tert-Butoxy)carbonyl]amino}ethoxy)carbonyl]oxy}-4-nitrobenzene (373 mg, 0.915 mmol, preparation described in US2019 / 192668) and DIPEA (439 μL, 2.72 mmol), and heat at 80 °C for 2 h with stirring. After cooling to room temperature, dilute the reaction mixture with EA and water. Add NaHCO3 solution and separate the phases. Wash the organic phase twice with water and brine, dry over Na2SO4, filter and evaporate. Purify the crude product by preparative HPLC (sunfire, ACN, TFA, narrow).

[0519] ESI-MS: 434 [M+H] + ; R t(HPLC): 0.99 min (Method B)

[0520] EX-02 Step 2: (3S,4R)-4-(2-{[(3S,5S)-5-[(tert-Butoxy)carbonyl]-1-[4-(2-ethoxy-2-oxoethyl)-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-6-yl]pyrrolidin-3-yl]oxy}-5-chloropyridin-3-yl)-4-fluoro-3-methylpiperidine-1-carboxylic acid 2-{[(tert-Butoxy)carbonyl]amino}ethyl ester

[0521] To a mixture of Intermediate N-01 (150 mg, 0.340 mmol) and tert-Butyl N-{4-[(3S,4R)-4-(5-chloro-2-fluoropyridin-3-yl)-4-fluoro-3-methylpiperidin-1-yl]-4-oxobutyl}carbamate (145 mg, 0.334 mmol) in 3.00 mL of DMA was added sodium hydride (60% dispersion in mineral oil, 25.8 mg, 0.65 mmol), and the mixture was stirred at room temperature for 1 h. The reaction mixture was quenched by dropwise addition of slightly acidified water (TFA) and purified directly by preparative HPLC (Sunfire, ACN / TFA, narrow).

[0522] ESI-MS: 855 [M+H] + ; R t (HPLC): 1.17 min (Method B)

[0523] EX-02 Step 3: (3S,4R)-4-(2-{[(3S,5S)-5-[(tert-Butoxy)carbonyl]-1-[4-(2-ethoxy-2-oxoethyl)-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-6-yl]pyrrolidin-3-yl]oxy}-5-chloropyridin-3-yl)-4-fluoro-3-methylpiperidine-1-carboxylic acid 2-aminoethyl ester

[0524] Under an argon atmosphere, 2.0 mL of DCM containing the product obtained in Step 2 was cooled to -15 °C (acetone / ice bath), and TMSI (17 μL, 0.12 mmol) was added slowly. The reaction mixture was kept at -15 °C for 45 min, then at 0 °C for 30 min. TMSI (9 μL, 0.060 mmol) was added again and stirring was continued at 0 °C for 45 min. The reaction was stopped by adding 2.0 mL of methanol. The solvent was removed under reduced pressure, and the residue was purified by preparative HPLC (Sunfire, ACN / TFA, narrow).

[0525] ESI-MS: 755 [M+H] + ; R t (HPLC): 0.82 min (Method B)

[0526] EX-02 Step 4: 2-{6-[(2S,4S)-4-({3-[(3S,4R)-1-[(2-aminoethoxy)carbonyl]-4-fluoro-3-methylpiperidin-4-yl]-5-chloropyridin-2-yl}oxy)-2-[(tert-butoxy)carbonyl]pyrrolidin-1-yl]-8-oxa-3,5-diazatricyclo-[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-4-yl}acetic acid

[0527] To a solution of the product obtained in Step 3 (53 mg, 0.070 mmol) in 0.53 mL of THF was added 2N aqueous LiOH (100 μL, 0.200 mmol) and water (60 μL, 3.33 mmol). The reaction mixture was stirred at room temperature for 4 h, then acidified with acetic acid (13 μL, 0.23 mmol) and diluted with ACN and methanol. The precipitate formed was removed by filtration, and the filtrate was evaporated under reduced pressure to give the title compound.

[0528] ESI-MS: 727 [M+H] + ; R t (HPLC): 0.75 min (Method B)

[0529] EX-02 Step 5: (1R,9S,11S,34S)-4-chloro-1-fluoro-34-methyl-26,31-dioxo-8,15,30-trioxa-6,12,23,27,32,37-hexaazacycloheptacyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21tert-Butyl octatriaconta-2,4,6,13(37),14(22),16(21),17,19,23-nonadeca-11-oate

[0530] General procedure E

[0531] After 30 min, the product obtained in Step 4 (46 mg, 0.063 mmol) and DIPEA (0.030 mL, 0.18 mmol) in a solution of 1.00 mL DMA were slowly added via an infusion pump to a solution of HATU (25 mg, 0.065 mmol) in 3.0 mL DMA, and stirring was continued for 30 min at room temperature. The reaction mixture was acidified with TFA and purified directly by preparative RP-HPLC (Sunfire, ACN / H2O / TFA, narrow).

[0532] ESI-MS: 769 [M+H] + ; R t (HPLC): 1.01 min (Method B)

[0533] EX-02 Step 6: (1R,9S,11S,34S)-4-Chloro-1-fluoro-34-methyl-26,31-dioxo-8,15,30-trioxa-6,12,23,27,32,37-hexaazatricyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 Octatriaconta-2,4,6,13(37),14(22),16(21),17,19,23-nonadecenoic acid

[0534] At room temperature, TFA (100 μL, 1.30 mmol) was added to a solution of the product obtained in Step 5 (5.0 mg, 0.0071 mmol) in 0.25 mL DCM, and the reaction mixture was stirred overnight. TFA (100 μL, 1.30 mmol) was added again and stirring was continued for 6 h. Volatiles were removed under a stream of nitrogen, the residue was dissolved in ACN / water and lyophilized.

[0535] ESI-MS: 653 [M+H] + ; R t (HPLC): 0.91 min (Method B)

[0536] Example 03 (1R,9S,11S,34S)-4-chloro-1,25,25-trifluoro-27,34-dimethyl-26,31-dioxo-8,15-dioxa-6,12,23,27,32,37-hexaazacycloheptacyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2,4,6,13,16,18,20,22,24(37)-nonaene-11-carboxylic acid

[0537]

[0538] EX-03 Step 1: 4-(2-{6-[(2S,4S)-2-[(tert-butoxy)carbonyl]-4-({3-[(3S,4R)-1-[(tert-butoxy)carbonyl]-4-fluoro-3-methylpiperidin-4-yl]-5-chloropyridin-2-yl}oxy)pyrrolidin-1-yl]-8-oxa-3,5-diazatricyclo-[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-4-yl}-2,2-difluoro-N-methylacetamido)butyric acid

[0539] React intermediate N-03 with intermediate P-01 according to the general procedure to obtain the title compound.

[0540] ESI-MS: 875 / 877 (1Cl) [M+H] + ; R t (HPLC): 0.92 min (Method A) EX-03 Step 2: 4-(2-{6-[(2S,4S)-2-[(tert-butoxy)carbonyl]-4-({5-chloro-3-[(3S,4R)-4-fluoro-3-methylpiperidin-4-yl]pyridin-2-yl}oxy)pyrrolidin-1-yl]-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-4-yl}-2,2-difluoro-N-methylacetamido)butyric acid

[0541] General Procedure F: Removal of the BOC protecting group using PTSA

[0542] At 0 °C, a mixture of the BOC-protected amine obtained in the previous step (72 mg, 0.078 mmol) in 2.50 mL of ACN was reacted with PTSA (31.3 mg, 0.156 mmol, 2.00 eq). The cooling bath was removed and the mixture was stirred until complete conversion of the starting material to the free amine was observed (2 - 24 h). The reaction mixture was quenched by the addition of water, diluted with ACN, filtered and purified by semi-preparative HPLC (XBridge C18; ACN / H2O / TFA) to give the title compound.

[0543] ESI-MS: 775 / 777 (1Cl) [M+H] + ; R t (HPLC): 0.65 min (Method A) EX-03 Step 3: (1R,9S,11S,34S)-4-chloro-1,25,25-trifluoro-27,34-dimethyl-26,31-dioxo-8,15-dioxa-6,12,23,27,32,37-hexaazacycloheptatriacont-2,4,6,13,16,18,20,22,24(37)-nonene-11-carboxylic acid tert-butyl ester 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2,4,6,13,16,18,20,22,24(37)-nonene-11-carboxylic acid tert-butyl ester

[0544] General Procedure G: Macrolactamization (HATU / DMA)

[0545] Over 30 min, a mixture of the product obtained in Step 3 (44 mg, 0.054 mmol) and DIPEA (0.030 mL, 0.162 mmol) in 1.10 mL of DMA was slowly added via syringe pump to a solution of HATU (23 mg, 0.060 mmol) in 2.42 mL of DMA, and stirring was continued at room temperature until complete depletion of the starting material was observed. The reaction mixture was acidified with TFA and purified directly by preparative RP-HPLC (Sunfire, ACN / H2O / TFA, narrow).

[0546] ESI-MS: 757 / 759 (1Cl) [M+H] + ; R t (HPLC): 0.86 min (Method A) EX-03 Step 4: (1R,9S,11S,34S)-4-chloro-1,25,25-trifluoro-27,34-dimethyl-26,31-dioxo-8,15-dioxa-6,12,23,27,32,37-hexaazacycloheptatriacont-2,4,6,13,16,18,20,22,24(37)-nonene-11-carboxylic acid tert-butyl ester 9,12 .113,24 .0 2,7 .0 14,22 .0 16,21 38-2,4,6,13,16,18,20,22,24(37)-Nonadeca-11-oic acid

[0547] At room temperature, the product obtained in the previous step was dissolved in 0.50 mL of DCM and treated with TFA (38.7 μL, 0.502 mmol, 20 eq) at 36 °C for 21 h. An additional amount of TFA (19.4 μL; 10.0 eq) was added and stirring was continued at 36 °C for 6 h. TFA (38.7 μL; 20.0 eq) was added again and stirring was continued at 36 °C for 20 h. Volatiles were removed in vacuo, the residue was dissolved in ACN / H2O and purified by semi-preparative HPLC (XBridge C18; ACN / H2O / TFA).

[0548] ESI-MS: 701 / 703 (1Cl) [M+H] + ; R t (HPLC): 0.71 min (Method A)

[0549] The absolute configuration of the final compound was confirmed from the co-crystal of the compound with human cGAS protein.

[0550] Example 04:

[0551] (1R,9S,11S,34S)-4-Chloro-1,25,25-trifluoro-34-methyl-26,31-dioxo-8,15-dioxa-6,12,23,27,32,37-hexaazatricyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 38-2,4,6,13,16,18,20,22,24(37)-Nonadeca-11-oic acid

[0552]

[0553] EX-04 Step 1: 2-{6-[(2S,4S)-2-[(tert-Butoxy)carbonyl]-4-({3-[(3S,4R)-1-[(tert-Butoxy)carbonyl]-4-fluoro-3-methylpiperidin-4-yl]-5-chloropyridin-2-yl}oxy)pyrrolidin-1-yl]-8-oxa-3,5-diazatricyclo-[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-4-yl}-2,2-difluoroacetic acid

[0554] This compound was prepared from P-06 according to General Procedure C.

[0555] ESI-MS: 776 / 778 (1Cl) [M+H] + ; R t (HPLC): 0.94 min (Method A) EX-04 Step 2: (3S,4R)-4-(2-{[(3S,5S)-5-[(tert-butoxy)carbonyl]-1-(4-{difluoro[(4-methoxy-4-oxobutyl)carbamoylmethyl]-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-6-yl)pyrrolidin-3-yl]oxy}-5-chloropyridin-3-yl)-4-fluoro-3-methylpiperidine-1-carboxylic acid tert-butyl ester

[0556] This compound was prepared from the product obtained in the previous step and methyl 4-aminobutyrate hydrochloride according to General Procedure D.

[0557] ESI-MS: 875 / 877 (1Cl) [M+H] + ; R t (HPLC): 0.96 min (Method A) EX-04 Step 3: 4-(2-{6-[(2S,4S)-2-[(tert-butoxy)carbonyl]-4-({3-[(3S,4R)-1-[(tert-butoxy)carbonyl]-4-fluoro-3-methylpiperidin-4-yl]-5-chloropyridin-2-yl}oxy)pyrrolidin-1-yl]-8-oxa-3,5-diazatricyclo-[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-4-yl}-2,2-difluoroacetamido)butyric acid

[0558] This compound was prepared from the product obtained in the previous step according to General Procedure C.

[0559] ESI-MS: 861 / 863 (1Cl) [M+H] + ; R t (HPLC): 0.91 min (Method A) EX-04 Step 4: 4-(2-{6-[(2S,4S)-2-[(tert-butoxy)carbonyl]-4-({5-chloro-3-[(3S,4R)-4-fluoro-3-methylpiperidin-4-yl]pyridin-2-yl}oxy)pyrrolidin-1-yl]-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-4-yl}-2,2-difluoroacetamido)butyric acid

[0560] This compound was prepared from the product obtained in the previous step according to General Procedure D.

[0561] ESI-MS: 761 / 763 (1Cl) [M+H] + ; R t (HPLC): 0.64 min (Method A) EX-04 Step 5: (1R,9S,11S,34S)-4-chloro-1,25,25-trifluoro-34-methyl-26,31-dioxo-8,15-dioxa-6,12,23,27,32,37-hexaazatricyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2,4,6,13,16,18,20,22,24(37)-nonaene-11-yl tert-butyl carbonate

[0562] This compound was prepared from the product obtained in the previous step according to General Procedure G.

[0563] ESI-MS: 743 / 745 (1Cl) [M+H] + ; R t (HPLC): 0.93 min (Method A) EX-04 Step 6: 2-{6-[(2S,4S)-2-[(tert-butoxy)carbonyl]-4-({3-[(3S,4R)-1-[(tert-butoxy)carbonyl]-4-fluoro-3-methylpiperidin-4-yl]-5-chloropyridin-2-yl}oxy)pyrrolidin-1-yl]-8-oxa-3,5-diazatricyclo-[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-4-yl}-2,2-difluoroacetic acid

[0564] General Procedure H: Removal of the tBu protecting group (TFA / DCM)

[0565] At room temperature, the product obtained in the previous step (33 mg, 0.042 mmol) was dissolved in 0.33 mL of DCM and treated with TFA (130 μL, 1.67 mmol, 40 equivalents). The mixture was stirred at room temperature until the starting material was completely consumed (24 h). The volatiles were removed in vacuo, the residue was dissolved in ACN / H2O, and purified by semi-preparative HPLC (XBridge C18; ACN / H2O / TFA).

[0566] ESI-MS: 687 / 689 (1Cl) [M+H] + ; R t(HPLC): 0.73 min (Method A) Example 05:

[0567] (1R,9S,11S,34S)-4-chloro-1,25,25-trifluoro-34-methyl-31-oxo-8,15-dioxa-6,12,23,32,37-pentaazatricyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2,4,6,13,16,18,20,22,24(37)-nonene-11-carboxylic acid

[0568]

[0569] EX-05 Step 1:

[0570] This compound was prepared from Intermediate N-04 and Intermediate P-02 according to General Procedure A.

[0571] ESI-MS: 748 / 750 [M+H] + ; R t (HPLC): 0.79 min (Method A)

[0572] EX-05 Step 2:

[0573] General Procedure I: Pyridine cyclization (NaH / NMP)

[0574] Within 2 min, a mixture of the product (fluoropyridine) (75.0 mg; 0.0095 mmol) obtained in EX-05 Step 1 in 2.00 mL of NMP was added dropwise to a suspension of NaH (16.6 mg; 0.381 mmol) in 1.00 mL of NMP. After the addition was complete, the reaction mixture was stirred at room temperature for 1 h. Another portion of NaH (16.6 mg, 0.381 mmol) was added and stirring was continued at room temperature for 20 min. To avoid the formation of other by-products, the reaction mixture was quenched with water, acidified with TFA, diluted with ACN / H2O, filtered and purified by semi-preparative HPLC (XBridge C18; ACN / H2O / TFA) to give the macrocyclized product.

[0575] ESI-MS: 728 [M+H] + ; R t (HPLC): 0.91 min (Method A)

[0576] EX-05 Step 3:

[0577] React the product obtained in step 2 of EX-05 according to General Procedure H to obtain the final product.

[0578] ESI-MS: 672 / 674 (1Cl) [M+H] + ; R t (HPLC): 0.87 min (Method A)

[0579] Similar to the above Example EX-05, the following compounds were prepared following General Procedures A, I, and H.

[0580]

[0581] Example EX-06: (1R,9S,11S,34S)-4-chloro-1,25,25,30,30-pentafluoro-34-methyl-31-oxo-8,15-dioxa-6,12,23,32,37-pentaazacycloheptatriaconta-2,4,6,13(37),14(22),16(21),17,19,23-nonaene-11-carboxylic acid 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 -octatriaconta-2,4,6,13(37),14(22),16(21),17,19,23-nonaene-11-carboxylic acid

[0582]

[0583] EX-06 Step 1:

[0584] React intermediate N-05 with intermediate P-01 using NMP as the solvent according to General Procedure B to obtain the title compound.

[0585] ESI-MS: 883 / 885 (1Cl) [M+H] + ; R t (HPLC): 0.95 min (Method A) EX-06 Step 2:

[0586] This compound was prepared from the product obtained in step 1 of EX-06 according to General Procedure F.

[0587] ESI-MS: 782 / 784 (1Cl) [M+H] + ; R t (HPLC): 0.69 min (Method A) EX-06 Step 3:

[0588] This compound was prepared from the product obtained in step 2 of EX-06 according to General Procedure G.

[0589] ESI-MS: 765 / 767 (1Cl) [M+H]+ ; R t (HPLC): 0.96 min (Method A) Step 4 of EX-06:

[0590] React the product obtained in Step 3 of EX-06 according to General Procedure H to obtain the final compound.

[0591] ESI-MS: 708 / 710 (1Cl) [M+H] + ; R t (HPLC): 0.86 min (Method A)

[0592] The following compounds can be prepared by applying General Procedures B, F, G, and H similar to Example 06 above.

[0593]

[0594]

[0595]

[0596] Example EX-08: (1R,9S,11S,34S)-4-chloro-1-fluoro-34-methyl-31-oxo-8,15,26-trioxa-6,12,23,32,37-pentaazatricyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2,4,6,13(37),14(22),16(21),17,19,23-nonaene-11-carboxylic acid

[0597]

[0598] Step 1 of EX-08:

[0599] React intermediate N-06 with intermediate P-01 using NMP as the solvent according to General Procedure B to obtain the title compound.

[0600] ESI-MS: 752 / 754 (1Cl) [M+H] + ; R t (HPLC): 0.89 min (Method A) Step 2 of EX-08:

[0601] React the product obtained in Step 1 of EX-08 according to General Procedure F.

[0602] ESI-MS: 652 / 654 (1Cl) [M+H] + ; R t(HPLC): 1.03 min (Method B) Step 3 of EX-08:

[0603] General Procedure J: Amidation Using DCC

[0604] At 0 °C, with stirring, a 1 M solution of DCC (387 μL, 0.387 mmol, 1.30 eq) in DCM was added dropwise to a mixture of piperidine (200 mg, 0.297 mmol), DMAP (4.7 mg, 0.039 mmol), and vinylacetic acid (34.0 μL, 0.387 mmol) in 2.00 mL of DCM obtained in Step 2 of EX-08. The reaction mixture was allowed to reach room temperature and stirred at room temperature until complete conversion of the starting materials was observed (30 h). Subsequently, the reaction mixture was cooled to 0 °C, diluted with DCM, filtered, and the filtrate was concentrated to give a crude product, which was further purified by preparative HPLC (XBridge C18; 30 - 100% ACN / H2O / TFA).

[0605] ESI-MS: 720 / 722 (1Cl) [M + H] + ; R t (HPLC): 0.80 min (Method A) Step 4 of EX-08:

[0606] General Procedure K: Olefin Metathesis

[0607] Under argon, the 2nd generation Grubbs (25.0 mg, 0.028 mmol, 10 mol%) was added to a degassed solution of the product obtained in Step 3 of EX-08 (212 mg, 0.280 mmol) in 8.48 mL of 1,2-dichloroethane. The vial was sealed, the reaction mixture was heated to 60 °C, and stirred at this temperature until the starting materials were exhausted (here: about 45 min). If necessary, an additional amount of catalyst (25.0 mg, 0.028 mmol, 10 mol%) was added and stirring was continued at 60 °C. Once the starting materials were completely converted (here: about 2 h), the volatiles were removed in vacuo, the residue was dissolved in ACN / H2O, filtered, and purified by semi-preparative HPLC (XBridge C18; ACN / H2O / TFA).

[0608] ESI-MS: 692 / 694 (1Cl) [M + H] + ; R t (HPLC): 0.73 min (Method A) Step 5 of EX-08:

[0609] General Procedure L: Hydrogenation Using Raney-Ni

[0610] In a Parr apparatus, Raney-Ni (2 mg) was added to a mixture of the product obtained in Step 5 of EX-08 in 0.10 mL of MeOH and 1.00 mL of EtOAc. The reaction mixture was placed under a hydrogen pressure of 2 bar at room temperature for 2 h. The solid was removed by filtration and the filtrate was concentrated to dryness under vacuum. The residue was dissolved in ACN / H2O and freeze-dried.

[0611] ESI-MS: 694 / 696 (1Cl) [M+H] + ; R t (HPLC): 0.76 min (Method A) Step 6 of EX-08:

[0612] The product obtained in Step 5 of EX-08 was reacted according to General Procedure H to give the final compound.

[0613] ESI-MS: 638 / 640 (1Cl) [M+H] + ; R t (HPLC): 0.63 min (Method A) Example EX-09:

[0614] (1R,9S,11S,30S,34S)-4-chloro-1-fluoro-30,34-dimethyl-31-oxo-8,15,26-trioxa-6,12,23,32,37-pentaazatricyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2,4,6,13,16,18,20,22,24(37)-nonene-11-carboxylic acid

[0615]

[0616] Step 1 of EX-09:

[0617] The product (2S,4S)-4-({5-chloro-3-[(3S,4R)-4-fluoro-3-methylpiperidin-4-yl]pyridin-2-yl}oxy)-1-{4-[(prop-2-en-1-yloxy)methyl]-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-6-yl}pyrrolidine-2-carboxylic acid tert-butyl ester obtained in Step 2 of EX-08 was reacted with 2-methylbut-3-enoic acid according to General Procedure J.

[0618] ESI-MS: 734 / 736 (1Cl) [M+H]+ ; R t (HPLC): 0.83 min (Method A) Step 2 of EX-09:

[0619] React the product obtained in the previous step according to General Procedure K and proceed to the next step as a mixture of diastereomers.

[0620] ESI-MS: 706 / 708 (1Cl) [M+H] + ; R t (HPLC): 0.74 / 0.76 min (Method A)

[0621] Step 3 of EX-09:

[0622] React the product obtained in the previous step according to General Procedure L and proceed to the next step as a mixture of diastereomers.

[0623] ESI-MS: 708 / 710 (1Cl) [M+H] + ; R t (HPLC): 0.77 min (Method A) Step 4 of EX-09:

[0624] The product obtained in the previous step reacts according to General Procedure H to give two diastereomers, which are separated by HPLC (XBridge C18, CAN / H2O / TFA). EX-09 is the first elution peak under the applied conditions. Arbitrarily assign the absolute configuration of the α-methyl substituent.

[0625] ESI-MS: 652 / 654 (1Cl) [M+H] + ; R t (HPLC): 0.61 min (Method A) ds of EX-08

[0626] ESI-MS: 652 / 654 (1Cl) [M+H] + ; R t (HPLC): 0.69 min (Method A) EX-08

[0627] Example EX-10:

[0628] (1R,9S,11S,34S)-4-chloro-1,30,30-trifluoro-34-methyl-31-oxo-8,15,26-trioxa-6,12,23,32,37-pentaazatricyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,2138-2,4,6,13,16,18,20,22,24(37)-Nonadecen-11-oic acid

[0629]

[0630] EX-10 Step 1:

[0631] Under argon at room temperature, ethyl bromodifluoroacetate (41.8 mg, 26.7 μL, 0.200 mmol) and phenylsilane (44.6 mg, 50.9 μL, 0.400 mmol) were added to a degassed suspension of tert-butyl (3S,4R)-4-(2-{[(3S,5S)-5-[(tert-butoxy)carbonyl]-1-{4-[(prop-2-en-1-yloxy)methyl]-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-6-yl}pyrrolidin-3-yl]oxy}-5-chloropyridin-3-yl)-4-fluoro-3-methylpiperidine-1-carboxylate (product obtained from EX-08 Step 1) (80.0 mg, 0.100 mmol), nickel(II) chloride (0.66 mg, 0.0050 mmol), and sodium carbonate (10.6 mg, 0.100 mmol) in 1.00 mL of anhydrous DMF. The vial was sealed and the reaction mixture was stirred at 70 °C for 16 h. The reaction mixture was quenched with 10% aqueous TFA, diluted with ACN / H2O, and purified by semi-preparative HPLC (XBridge C18; ACN / H2O / TFA).

[0632] ESI-MS: 876 / 878 (1Cl) [M+H] + ; R t (HPLC): 0.93 min (Method A)

[0633] The product obtained in EX-10 Step 1 was further reacted using the following reaction sequence:

[0634] EX-10 Step 2: Ester hydrolysis using General Procedure C

[0635] ESI-MS: 848 / 850 (1Cl) [M+H] + ; R t (HPLC): 0.85 min (Method A)

[0636] EX-10 Step 3: Removal of the BOC protecting group using General Procedure F

[0637] ESI-MS: 748 / 750 (1Cl) [M+H] + ; Rt (HPLC): 0.64 min (Method A)

[0638] EX-10 Step 4: Amidation using General Procedure D

[0639] ESI-MS: 730 / 732 (1Cl) [M+H] + ; R t (HPLC): 0.86 min (Method A)

[0640] EX-10 Step 5: Removal of the tert-butyl ester protecting group using General Procedure H.

[0641] ESI-MS: 674 / 676 (1Cl) [M+H] + ; R t (HPLC): 0.68 min (Method A) Example EX-11: (1R,9S,11S,33S)-4-chloro-1-fluoro-33-methyl-27,30-dioxo-8,15,26-trioxa-6,12,23,31,36-pentaazatricyclo[29.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 heptatriaconta-2,4,6,13(36),14(22),16(21),17,19,23-nonaene-11-carboxylic acid

[0642]

[0643] EX-11 Step 1:

[0644] At room temperature, stir a mixture of (3S,4R)-4-(2-{[(3S,5S)-5-[(tert-butoxy)carbonyl]-1-{4-[(prop-2-en-1-yloxy)methyl]-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaene-6-yl}pyrrolidin-3-yl]oxy}-5-chloropyridin-3-yl)-4-fluoro-3-methylpiperidine-1-carboxylic acid tert-butyl ester (see EX-08 Step 1) (31.6 mg, 0.040 mmol), 1,3-dimethylbarbituric acid (12.6 mg, 0.080 mmol), and Pd(PPh3)4 (2.31 mg, 0.0020 mmol) in 0.32 mL of methanol for 5.5 h. Subsequently, heat the reaction mixture to 65 °C for 15 h. After cooling to room temperature, dilute the mixture with ACN / H2O, filter, and purify by semi-preparative HPLC (XBridge C18; ACN / H2O / TFA).

[0645] ESI-MS: 712 / 714 (1Cl) [M+H] + ; R t (HPLC): 0.81 min (Method A) EX-11 Step 2:

[0646] At room temperature, succinic anhydride (25.0 mg, 0.250 mmol) was added to a mixture of the product obtained in the previous step (60.0 mg, 0.0800 mmol) in 0.60 mL of pyridine, and the mixture was stirred at 50 °C for 5 h. The reaction mixture was allowed to reach room temperature and stirred at room temperature for 2 days. Succinic anhydride (25.0 mg; 0.25 mmol) was added again and the mixture was stirred at 50 °C for 6 h. The reaction mixture was acidified by adding TFA, diluted with ACN / H2O and purified by preparative HPLC (ACN / H2O / TFA).

[0647] ESI-MS: 812 [M+H] + ; R t (HPLC): 1.20 min (Method B)

[0648] The product obtained in EX-11 Step 2 was further reacted using the following reaction sequence:

[0649] EX-11 Step 3: Removal of the BOC protecting group using General Procedure F

[0650] ESI-MS: 712 [M+H] + ; R t (HPLC): 0.77 min (Method B)

[0651] EX-11 Step 4: Amidation using General Procedure D

[0652] ESI-MS: 694 [M+H] + ; R t (HPLC): 0.92 min (Method D)

[0653] EX-11 Step 5: Removal of the tert-butyl ester protecting group using General Procedure H

[0654] ESI-MS: 638 [M+H] + ; R t (HPLC): 0.82 min (Method B)

[0655] Example EX-12:

[0656] (2S,4S)-4-({5-chloro-3-[(3S,4R)-4-fluoro-3-methylpiperidin-4-yl]pyridin-2-yl}oxy)-1-{4-[(prop-2-en-1-yloxy)methyl]-8-oxa-3,5-diazatricyclo[7.4.0.0 2,7 trideca-1(9),2(7),3,5,10,12-hexaen-6-yl}pyrrolidine-2-carboxylic acid tert-butyl ester

[0657] Step 1 of EX-12:

[0658] At room temperature, triethylamine (0.084 mL, 0.606 mmol) was added to a mixture of pent-4-enoic acid (30.0 mg, 0.303 mmol) in 2.00 mL of DCM, followed by the addition of EDC hydrochloride (70.0 mg, 0.363 mmol). After stirring for 10 min at room temperature, the product obtained in Step 2 of EX-08 (290 mg, 0.303 mmol) was added, and the mixture was stirred for 4 h at room temperature. The reaction mixture was poured onto ice water and extracted with EtOAc. The organic phases were combined, washed with water, dried over sodium sulfate, and evaporated to dryness.

[0659] ESI-MS: 734 [M+H] + ; R t (HPLC): 1.059 min (Method C)

[0660] The product obtained in Step 2 of EX-11 was further reacted using the following reaction sequence:

[0661] Step 2 of EX-12: Olefin metathesis according to General Procedure K

[0662] ESI-MS: 706 [M+H] + ; R t (HPLC): 0.998 min (Method C)

[0663] Step 3 of EX-12: Hydrogenation using General Procedure L

[0664] ESI-MS: 708 [M+H] + ; R t (HPLC): 1.063 min (Method C)

[0665] Step 4 of EX-12: Removal of the tert-butyl ester protecting group using General Procedure H

[0666] ESI-MS: 652 [M+H] + ; R t (HPLC): 0.951 min (Method C)

[0667] Example EX-13:

[0668] (1R,9S,11S,34S)-4-chloro-1-fluoro-34-methyl-31-oxo-8,15,26,30-tetraoxa-6,12,23,32,37-pentaazatricyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2(7),3,5,13(37),14(22),16(21),17,19,23-nonaene-11-carboxylic acid

[0669]

[0670] Step 1 of EX-13: tert-Butyl 2-(3-{[(4-nitrophenoxy)carbonyl]oxy}propoxy)acetate

[0671] At 0 °C over 30 min, a solution of 4-nitrophenyl chloroformate in DCM (10.0 mL) was added dropwise to a solution of tert-butyl 2-(3-hydroxypropoxy)acetate (1.000 g, 5.10 mmol, prepared as described in WO2019 / 195609) and pyridine (0.441 mL, 5.48 mmol) in 10.0 mL of DCM. The reaction mixture was warmed to room temperature overnight. Water was added and the phases were separated. The organic phase was dried and evaporated to dryness.

[0672] ESI-MS: 300 [M-tBu+H] + ; R t (HPLC): 1.00 min (Method C)

[0673] Step 2 of EX-13: tert-Butyl 2-{3-[(3S,4R)-4-(5-chloro-2-fluoropyridin-3-yl)-4-fluoro-3-methylpiperidin-1-carbonyloxy]propoxy}acetate

[0674] The mixture of intermediate P-02 (250 mg, 0.883 mmol), tert-butyl 2-(3-{[(4-nitrophenoxy)carbonyl]oxy}propoxy)acetate (396 mg, 0.891 mmol) and DIPEA (0.426 mL, 2.65 mmol) in 4.60 mL of THF was heated at reflux for 2 h. After cooling to room temperature, the reaction mixture was diluted with ether and extracted twice with 1 M aqueous NaOH solution. The organic phase was washed with water and saturated aqueous NaCl solution, dried over sodium sulfate and concentrated under reduced pressure. The crude product was used in the next step without further purification.

[0675] ESI-MS: 463 [M+H] + ; R t (HPLC): 0.76 min (Method A)

[0676] EX-13 Step 3 2-{3-[(3S,4R)-4-(5-chloro-2-fluoropyridin-3-yl)-4-fluoro-3-methylpiperidin-1-carbonyloxy]propoxy}acetic acid

[0677] The product obtained from the previous step was reacted according to General Procedure H to give the expected product.

[0678] ESI-MS: 407 / 409 (1Cl) [M+H] + ; R t (HPLC): 0.58 min (Method A) EX-13 Step 4: 3-{[(2-Amino-1-benzofuran-3-yl)carbamoyl]methoxy}propyl (3S,4R)-4-(5-chloro-2-fluoropyridin-3-yl)-4-fluoro-3-methylpiperidine-1-carboxylate

[0679] At room temperature, DIPEA (360 μL, 2.08 mmol) was added to a stirred solution of 2-{3-[(3S,4R)-4-(5-chloro-2-fluoropyridin-3-yl)-4-fluoro-3-methylpiperidin-1-carbonyloxy]propoxy}acetic acid (380 mg, 0.934 mmol) and PFTU (440 mg, 1.03 mmol) in 5.50 mL of DMF, and the mixture was stirred at this temperature for 5 min. Subsequently, 3-amino-1-benzofuran-2-carboxamide (190 mg, 1.035 mmol) was added, and the mixture was stirred at room temperature for 1 h. The reaction mixture was heated to 50 °C and stirred at this temperature for 36 h. It was diluted with ether, and the organic phase was washed with dilute NaOH and brine. The organic layer was dried over sodium sulfate and evaporated to dryness. The residue was dissolved in MeOH acidified with TFA, filtered, and purified by preparative HPLC (XBridge C18; ACN / H2O / TFA).

[0680] ESI-MS: 565 / 567 (1Cl) [M+H] + ; R t(HPLC): 0.68 min (Method A) Step 5 of EX-13: Under reflux, heat a mixture of (3S,4R)-4-(5-chloro-2-fluoropyridin-3-yl)-4-fluoro-3-methylpiperidine-1-carboxylic acid 3-{[(2-amino-1-benzofuran-3-yl)carbamoyl]methoxy}propyl ester (110 mg, 0.195 mmol), chlorotrimethylsilane (0.352 mL, 2.77 mmol), and triethylamine (1.185 mL, 8.44 mmol) in 4.75 mL of dichloroethane for 40 h. After cooling to room temperature, dilute the reaction mixture with DCM. Separate the phases, and wash the organic phase with dilute hydrochloric acid, dry over sodium sulfate, and concentrate in vacuo to obtain the crude product, which is used in the next step without purification.

[0681] ESI-MS: 447 / 449 (1Cl) [M+H] + ; R t (HPLC): 0.68 min (Method A) Step 6 of EX-13: React the product obtained in Step 5 of EX-13 with (2S,4S)-1-[(tert-butoxy)carbonyl]-4-hydroxypyrrolidine-2-carboxylic acid according to General Procedure B.

[0682] ESI-MS: 758 / 760 (1Cl) [M+H] + ; R t (HPLC): 0.81 min (Method A) Step 7 of EX-13: Dissolve the product obtained in Step 5 of EX-13 (140 mg, 0.185 mmol) in methanol, and add thionyl chloride (89 μL, 1.21 mmol). After stirring for 2 h, add another portion of thionyl chloride (55 μL, 0.754 mmol) and continue stirring overnight. Concentrate the reaction mixture in vacuo, and purify the crude product by HPLC (XBridge C18; ACN / H2O / TFA).

[0683] ESI-MS: 672 / 674 (1Cl) [M+H] + ; R t (HPLC): 0.58 min (Method A) Step 8 of EX-13: At room temperature, dissolve the product obtained in Step 7 of EX-13 (21 mg, 0.030 mmol) in 2.00 mL of DMF. Add DBU (22.4 μL, 0.148 mmol) and BOP (35 mg, 0.077 mmol) and stir the mixture at room temperature for 2 h. After adding water, extract the mixture with ether (twice). Wash the combined organic phases with brine, dry over sodium sulfate, and concentrate in vacuo.

[0684] ESI-MS: 654 / 656 (1Cl) [M+H]+ ; R t (HPLC): 0.74 min (Method A) Step 9 of EX-13: To a mixture of the product obtained in Step 8 of EX-13 (25 mg, 0.038 mmol) in 2.00 mL of dioxane was added an aqueous solution of 1 M NaOH (450 μL, 0.450 mmol). The reaction mixture was heated at 60 °C for 30 min. At room temperature, the reaction mixture was acidified, diluted with methanol and purified by HPLC (XBridge C18; ACN / H2O / TFA).

[0685] ESI-MS: 640 / 642 (1Cl) [M+H] + ; R t (HPLC): 0.68 min (Method A) Example EX-15: (1R,9S,11S,34S)-4-chloro-1,25,25-trifluoro-34-methyl-31-oxo-8,15,26-trioxa-6,12,23,32,37-pentaazatetracyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2(7),3,5,13(37),14(22),16(21),17,19,23-nonaene-11-carboxylic acid

[0686] Step 1 of EX-15:

[0687] NaH (130 mg, 3.25 mmol, 60% in mineral oil) was added to a mixture of N-08 (370 mg, 0.802 mmol) and P-01 (285 mg, 0.812 mmol) in 10.0 mL of DMF, and the reaction mixture was stirred at room temperature for 1 h. The reaction mixture was added dropwise to cold water and acidified with an aqueous solution of 1 M HCl. The precipitate was dissolved in ether. The combined organic layers were washed with brine, dried over sodium sulfate and concentrated in vacuo. The residue was further purified by silica gel chromatography using cyclohexane / EA as the eluent.

[0688] ESI-MS: 788 / 790 (1Cl) [M+H] + ; R t (HPLC): 1.01 min (Method A) Step 2 of EX-15:

[0689] Under argon, the 2nd generation Grubbs (4.07 mg, 0.005 mmol, 6 mol%) was added to a degassed solution of the product obtained in Step 1 of EX-15 (63.0 mg, 0.076 mmol) and methyl 3-butenoate (128 μL, 1.139 mmol) in 0.50 mL of DCM. The vial was sealed and the reaction mixture was stirred at room temperature for 2 h. Methyl 3-butenoate (42.7 μL, 5.0 equiv) was added again, and stirring was continued for 2 h. Subsequently, the 2nd generation Grubbs (4.07 mg, 0.005 mmol, 6 mol%) was added, and stirring was continued for 1.5 h. In addition, the catalyst (4.07 mg; 6 mol%) and methyl 3-butenoate (42.7 μL; 3.00 equiv) were added again, and stirring was continued at room temperature for 18 h. Volatiles were removed in vacuo, the residue was dissolved in EtOAc / cyclohexane and purified directly by silica gel chromatography.

[0690] ESI-MS: 860 / 862 (1Cl) [M+H] + ; R t (HPLC): 0.75 min (Method G) EX-15 Step 3:

[0691] General Procedure N: Hydrogenation using Pd / C

[0692] In a Parr apparatus, 10% Pd / C (90.0 mg) was added to a solution of the product obtained in Step 3 of EX-15 (250 mg, 0.276 mmol) in 40 mL of EtOAc, and at room temperature, the reaction mixture was placed under a hydrogen pressure of 3 bar for 18 h. The solid was removed by filtration, and the filtrate was concentrated to dryness in vacuo to give the crude product, which was purified by semi-preparative HPLC (Sunfire C18; ACN / H2O / TFA).

[0693] ESI-MS: 862 / 864 (1Cl) [M+H] + ; R t (HPLC): 1.12 min (Method B)

[0694] The product obtained in Step 3 of EX-15 was further reacted using the following reaction sequence:

[0695] EX-15 Step 4: Ester hydrolysis using General Procedure C

[0696] ESI-MS: 848 / 850 (1Cl) [M+H] + ; R t (HPLC): 0.94 min (Method A)

[0697] EX-15 Step 5: Removal of the BOC protecting group using General Procedure F

[0698] ESI-MS: 748 / 750 (1Cl) [M+H] + ; R t (HPLC): 0.82 min (Method B)

[0699] EX-15 Step 6: Amidation using General Procedure E

[0700] ESI-MS: 730 / 732 (1Cl) [M+H] + ; R t (HPLC): 0.98 min (Method D)

[0701] EX-15 Step 7: Removal of the tert-butyl ester protecting group using General Procedure H

[0702] ESI-MS: 674 / 676 (1Cl) [M+H] + ; R t (HPLC): 0.68 min (Method A) Example EX-18:

[0703] (1R,9S,11S,34S)-4-Ethynyl-1-fluoro-34-methyl-31-oxo-8,15,27-trioxa-6,12,23,32,37-pentaazatricyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2,4,6,13,16,18,20,22,24(37)-nonene-11-carboxylic acid

[0704]

[0705] Under an argon atmosphere, Pd(dppf)Cl2 (9.0 mg, 0.012 mmol) and CuI (3.0 mg, 0.016 mmol) were added to a mixture of Example EX-17 (30 mg, 0.044 mmol), (triisopropylsilyl)acetylene (24 μL, 0.11 mmol) and triethylamine (30 μL, 4.8 equivalents) in 0.75 mL of 2-methyltetrahydrofuran, and the reaction mixture was stirred at 80 °C for 2 h. After cooling to room temperature, it was filtered and the filtrate was concentrated in vacuo. The residue was dissolved in 2.00 mL, TBAF (1 M solution in THF, 100 μL, 0.100 mmol) was added, and the mixture was stirred at room temperature for 30 min. Subsequently, it was added dropwise to cold water, acidified with 1 M aqueous HCl and extracted twice with diethyl ether. The combined organic layers were washed with water and brine and concentrated in vacuo. The product was purified by HPLC (XBridge C18; ACN / H2O / TFA).

[0706] ESI-MS 628 [M+H] + ; R t (HPLC): 0.59 min (Method A)

[0707] Example EX-19:

[0708]

[0709] According to EX-18, the product obtained from the synthesis in Step 3 of EX-17 was subjected to Sonogashira reaction conditions, and then the general procedure EX-D was applied to remove the tert-butyl protecting group to give the title compound.

[0710] ESI-MS 646 [M+H] + ; R t (HPLC): 0.64 min (Method A)

[0711] Example EX-20.01

[0712] (1R,9S,11S,34S)-1-Fluoro-4-(2-methoxypyridin-3-yl)-34-methyl-31-oxo-8,15,27-trioxa-6,12,23,32,37-pentaazatricyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2,4,6,13(37),14(22),16(21),17,19,23-nonaene-11-carboxylic acid

[0713]

[0714] EX-20.01 Step 1:

[0715] General Procedure O: Suzuki Coupling

[0716] At room temperature, under an argon atmosphere, methane sulfonic acid (2-dicyclohexylphosphino-2',4',6'-triisopropyl-1,1'-biphenyl)[2-(2'-amino-1,1'-biphenyl)]palladium(II) (XPhos Pd G3) was added to a degassed mixture of aryl bromide (EX-17 Step 3, 35 mg, 0.047 mmol), (2-methoxypyridin-3-yl)boronic acid (11 mg, 0.071 mmol), and potassium carbonate (2 N aqueous solution, 95 μL, 0.19 mmol). The vial was sealed and heated at 90 °C until complete conversion of the starting material was observed (1 h). Heating was removed and the mixture was allowed to reach room temperature. Water was added and the mixture was extracted twice with DCM. The organic layer was washed with brine, dried over sodium sulfate and evaporated to dryness.

[0717] ESI-MS: 767 [M+H] + ; R t (HPLC): 0.70 min (Method A)

[0718] EX-20.01 Step 2:

[0719] Apply General Procedure H to deprotect the product of the Suzuki coupling reaction to obtain the final compound.

[0720] ESI-MS 711 [M+H] + ; R t (HPLC): 0.61 min (Method A)

[0721] Prepare the following examples according to the above reaction sequence (General Procedures O and H (t-Bu is not applicable if the free acid undergoes Suzuki coupling)).

[0722]

[0723] Example EX-21: (1R,9S,11S,34S)-4-chloro-1,25,25-trifluoro-34-methyl-31-oxo-8,15,27-trioxa-6,12,23,32,37-pentaazatetracyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2,4,6,13(37),14(22),16(21),17,19,23-nonaene-11-carboxylic acid

[0724]

[0725] EX-21 Step 1: Under an argon atmosphere at 0 °C, sodium borohydride (90 mg, 2.38 mmol) was added to a solution of intermediate P-06 (730 mg, 794 mmol) in 8.34 mL of absolute ethanol. The reaction mixture was stirred at 0 °C for 1 h and at room temperature for 2 h. Water was added to the mixture, and the product was extracted three times with EtOAc. The organic phase was then washed with water and brine, dried over MgSO4, filtered, and concentrated under reduced pressure. The crude product was purified by silica column chromatography (10% -> 100% EtOAc / CH).

[0726] ESI-MS 762 / 764 (1Cl) [M+H] + ; R t (HPLC): 0.94 min (Method A)

[0727] EX-21 Step 2: The material obtained in the previous step was dissolved in 2.44 mL of DMA, and sodium hydride (50 mg, 1.15 mmol) was added, followed immediately by allyl bromide (250 μL, 2.86 mmol), and the mixture was stirred at room temperature for 10 min. Ice was added to the reaction mixture, and the product was extracted three times with EtOAc. The organic phase was then washed with water and brine, dried over MgSO4, filtered, and concentrated under reduced pressure. The crude material was purified by semi-preparative HPLC (ACN, Xbridge, TFA), using a narrow gradient of 80 - 100% ACN / water.

[0728] ESI-MS 803 / 805 (1Cl) [M+H] + ; R t (HPLC): 1.03 min (Method A)

[0729] EX-21 Step 3: The product obtained from the previous step (232 mg, 0.289 mmol) was dissolved in 2.00 mL of ethyl acrylate, and the mixture was degassed by bubbling argon for 5 minutes, after which the second-generation Grubbs (24.5 mg, 0.029 mmol) was added to the reactants. The mixture was stirred at room temperature under an argon atmosphere for 1 h, then directly adsorbed onto extrelute adsorbent while concentrating under reduced pressure, and purified by flash chromatography (20% -> 100% EtOAc / CH).

[0730] ESI-MS 875 / 877 (1Cl) [M+H] + ; R t (HPLC): 1.00 min (Method A)

[0731] React the product obtained in step 3 of EX-21 using the following reaction sequence:

[0732] EX-21 Step 4: Hydrogenation using General Procedure L

[0733] EX-21 Step 5: Ester hydrolysis using General Procedure C ESI-MS 848 / 850 (1Cl) [M+H] + ; R t (HPLC): 0.68 min (Method G)

[0734] EX-21 Step 6: Removal of the BOC protecting group using General Procedure F ESI-MS: 748 / 751 (1Cl) [M+H] + ; R t (HPLC): 0.69 min (Method A)

[0735] EX-21 Step 7: Amidation using General Procedure D

[0736] ESI-MS: 731 / 733 (1Cl) [M+H] + ; R t (HPLC): 0.88 min (Method A)

[0737] EX-21 Step 8: Removal of the tert-butyl ester protecting group using General Procedure H

[0738] ESI-MS: 675 / 677 (1Cl) [M+H] + ; R t (HPLC): 0.79 min (Method A) Example EX-22:

[0739] (1R,9S,11S,34S)-4-chloro-1-fluoro-34-methyl-31-oxo-8,15,30-trioxa-6,12,23,32,37-pentaazacyclotetracontane 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2,4,6,13(37),14(22),16(21),17,19,23-nonaene-11-carboxylic acid

[0740]

[0741] EX-22 Step 1: General Procedure M: Heck coupling

[0742] In a sealed tube, under an argon atmosphere, a mixture of intermediate P-07 (85 mg, 0.213 mmol), intermediate N-02 (70 mg, 0.176 mmol), palladium(II) acetate (7.9 mg, 0.035 mmol), tris(ortho-tolyl)phosphine (22 mg, 0.070 mmol), and triethylamine (71 mg, 0.70 mmol) in 2.85 mL of DMF was heated to 95 °C for 6 h. The reaction mixture was cooled to room temperature and stirred overnight at room temperature. Volatiles were removed in vacuo, and the crude mixture was purified by silica gel column chromatography (20 - 50% EE / CH). The product was isolated as a mixture of isomers and used as such in the next reaction step.

[0743] ESI-MS 656 [M+H] + ; R t (HPLC): 0.70 / 0.71 / 0.73 min (Method B)

[0744] Using the following reaction sequence, the product obtained in EX-22 Step 1 was further reacted:

[0745] EX-22 Step 2: Hydrogenation using General Procedure L

[0746] ESI-MS 658 [M+H] + ; R t (HPLC): 0.76 min (Method B)

[0747] EX-22 Step 3: Using General Procedure I, via macrocyclization of S N Ar

[0748] ESI-MS 638 [M+H] + ; R t (HPLC): 0.85 min (Method B)

[0749] Example EX-23: (1R,9S,11S,34S)-4-chloro-1-fluoro-34-methyl-31-oxo-8,15,28-trioxa-6,12,23,32,37-pentaazatricyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2,4,6,13(37),14(22),16(21),17,19,23-nonaene-11-carboxylic acid

[0750]

[0751] The title compound is prepared starting from intermediate P-05 using the following reaction sequence:

[0752] EX-23 Step 1: Apply General Procedure M using Heck coupling of 3-(prop-2-en-1-yloxy)propanoic acid ESI-MS: 810 / 812 (1Cl); R t (HPLC): 0.81 / 0.82 min (Method A)

[0753] EX-23 Step 2: Hydrogenation according to General Procedure N (Pd / C)

[0754] ESI-MS: 812 / 814 (1Cl) [M+H] + ; R t (HPLC): 0.98 min (Method B)

[0755] EX-23 Step 3: Removal of the BOC protecting group using General Procedure F

[0756] ESI-MS 712 / 714 (1Cl) [M+H] + ; R t (HPLC): 0.67 min (Method B)

[0757] EX-23 Step 4: Amidation using General Procedure E

[0758] ESI-MS 694 / 696 (1Cl) [M+H] + ; R t (HPLC): 0.97 min (Method D)

[0759] EX-23 Step 5: Removal of the tert-butyl ester protecting group using General Procedure H

[0760] ESI-MS 638 / 640 (1Cl) [M+H] + ; R t (HPLC): 0.74 min (Method A) Example EX-24:

[0761] (1R,9S,11S,34S)-1-fluoro-34-methyl-31-oxo-8,15,28-trioxa-6,12,23,32,37-pentaazatricyclo-[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2,4,6,13(37),14(22),16(21),17,19,23-nonaene-11-carboxylic acid

[0762]

[0763] This compound was prepared from EX-24 via hydrogenation using general procedure N.

[0764] ESI-MS 604 [M+H] + ; R t (HPLC): 0.65 min (method B)

[0765] Example EX-29:

[0766] (1R,9S,11S,34S)-1-fluoro-34-methyl-31-oxo-4-(prop-1-yn-1-yl)-8,15,29-trioxa-6,12,23,32,37-pentaazatetracyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2(7),3,5,13,16(21),17,19,22,24(37)-nonene-11-carboxylic acid

[0767]

[0768] This compound was prepared from EX-28 via the Sonogashira coupling described for EX-18.

[0769] ESI-MS: 642 [M+H] + ; R t (HPLC): 0.61 min (method A)

[0770] Example EX-34:

[0771] (1R,9S,11S,34S)-4-chloro-1-fluoro-34-methyl-31-oxo-8,15,27,30-tetraoxa-6,12,23,32,37-pentaaza-tetracyclo[30.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 octatriaconta-2,4,6,13(37),14(22),16(21),17,19,23-nonene-11-carboxylic acid

[0772]

[0773] Step 1 of EX-34:

[0774] At room temperature, 1-chloro-N,N,2-trimethylallylamine (50 μL, 0.378 mmol) was added to a solution of intermediate P-08 (115 mg, 0.284 mmol) in 2.00 mL of acetonitrile, and the resulting mixture was stirred for 10 min. Subsequently, pyridine (70 μL, 0.856 mmol) was added to the mixture, followed by 3-aminobenzofuran-2-carboxamide (230 mg, 1.31 mmol), and the mixture was stirred at room temperature for 2 h. The reaction mixture was concentrated to dryness and water and DCM were added. The phases were separated, and the organic phase was dried and concentrated to dryness.

[0775] ESI-MS: 565 / 567 (1Cl) [M+H] + ; R t (HPLC): 0.64 min (Method A) EX-34 Step 2:

[0776] At room temperature, chlorotrimethylsilane (0.512 mL, 4.03 mmol) was added to a solution of the product of Step 1 (160 mg, 0.283 mmol) in 7.00 mL of 1,2-dichloroethane, followed by triethylamine (1.724 mL, 12.3 mmol), and the resulting mixture was stirred under reflux for 40 h. The reaction was cooled to room temperature and diluted with DCM. The organic phase was washed with dilute aqueous HCl, dried over sodium sulfate and concentrated to dryness.

[0777] ESI-MS: 547 / 549 (1Cl) [M+H] + ; R t (HPLC): 0.64 min (Method A) EX-34 Step 3:

[0778] At room temperature, NaH (60% in mineral oil, 50.8 mg, 1.27 mmol) was added to a solution of (2S,4S)-tert-butyl 4-hydroxypyrrolidine-2-carboxylate hydrochloride (151 mg, 0.635 mmol) in 2.17 mL of DMF. After stirring for 5 min, a solution of the product obtained in EX-34 Step 2 (217 mg, 0.32 mmol) in 2.17 mL of DMF was slowly added, and the mixture was stirred at room temperature for 1 h. The reaction mixture was added dropwise to water, acidified with acetic acid and extracted twice with EtOAc. The combined organic phases were washed with brine, dried over sodium sulfate and evaporated to dryness. The crude product was purified by HPLC (Sunfire, ACN / H2O / TFA, narrow).

[0779] ESI-MS: 758 / 760 (1Cl) [M+H] + ; R t(HPLC): 1.06 min (Method B) Step 4 of EX-34:

[0780] Dissolve the product from Step 3 of EX-34 (100 mg, 0.130 mmol) in 2.00 mL of methanol, and slowly add thionyl chloride (0.060 mL, 0.79 mmol) at room temperature. Stir the reaction mixture at room temperature for 48 h, then concentrate under reduced pressure and purify by HPLC (Sunfire, ACN / H2O / TFA, narrow).

[0781] ESI-MS: 672 / 674 (1Cl) [M+H] + ; R t (HPLC): 0.70 min (Method B) Step 5 of EX-34:

[0782] Add DBU (54 μL, 0.353 mmol) to a mixture of the product obtained in Step 4 of EX-34 (50 mg, 0.071 mmol) and BOP (84 mg, 0.184 mmol) in 5.00 mL of acetonitrile and 5.00 mL of DMF. Stir the reaction mixture at room temperature overnight. Remove the volatiles under reduced pressure, and purify the crude product by HPLC (Sunfire, ACN / H2O / TFA).

[0783] ESI-MS: 654 / 656 (1Cl) [M+H] + ; R t (HPLC): 0.84 min (Method B) Step 6 of EX-34:

[0784] Deprotect the product from Step 6 of EX-34 according to General Procedure C to obtain the title compound.

[0785] ESI-MS: 640 / 642 (1Cl) [M+H] + ; R t (HPLC): 0.78 min (Method B) Example EX-35: (1R,9S,11S,33S)-4-chloro-1-fluoro-33-methyl-30-oxo-8,15,29-trioxa-6,12,23,31,36-pentaazatricyclo[29.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 heptatriaconta-2,4,6,13(36),14(22),16(21),17,19,23-nonaene-11-carboxylic acid

[0786]

[0787] EX-35 Step 1:

[0788] Add intermediate P-05 (200 mg, 0.250 mmol) and 3-buten-1-ol (86 μL, 1.00 mmol) to a microwave vial and flush the system with argon. Add 2.31 mL of DMA and triethylamine (696 μL, 4.99 mmol) successively, and degas the tube for 5 min under an argon stream. Then add 1,1'-bis(di-tert-butylphosphino)ferrocene palladium dichloride (24.4 mg, 0.037 mmol), seal the vial and heat the reaction mixture at 115 °C for 4 h. After cooling to room temperature, filter the reaction mixture through a catalyst scavenging cartridge, dilute with additional ACN and acidify with 2.5 M acetic acid. Purify the crude product directly by HPLC (Sunfire, ACN / H2O / TFA).

[0789] ESI-MS: 753 / 755 (1Cl) [M+H] + ; R t (HPLC): 0.79 min (Method A) EX-35 Step 2:

[0790] Hydrogenate the product obtained in EX-35 Step 2 according to General Procedure N.

[0791] ESI-MS: 755 / 757 (1Cl) [M+H] + ; R t (HPLC): 0.77 min (Method A) EX-35 Step 3:

[0792] Dissolve the product obtained in EX-35 Step 2 (21 mg, 0.028 mmol) in 0.36 mL of DCM and add pyridine (2 μL, 0.03 mmol). Cool the mixture to 0 °C and add 4-nitrophenyl chloroformate (6.3 mg, 0.030 mmol). Warm the reactants to room temperature and stir for 3 h. Remove the volatiles under reduced pressure and use the remaining crude material in the next step without further purification.

[0793] ESI-MS: 920 / 922 (1Cl) [M+H] + ; R t (HPLC): 0.85 min (Method A) EX-35 Step 4:

[0794] Deprotect the product obtained in EX-35 Step 3 according to General Procedure F.

[0795] ESI-MS: 820 / 822 (1Cl) [M+H] + ; R t(HPLC): 0.66 min (Method A) Step 5 of EX-35:

[0796] Triethylamine (39 mg, 0.053 mL, 0.384 mmol) was added to a solution of the product obtained in Step 4 of EX-35 (21 mg, 0.026 mmol) in 3.00 mL of THF. The reaction mixture was heated to 70 °C for 3 h, then concentrated under reduced pressure and used in the next step without further purification.

[0797] ESI-MS: 681 / 683 (1Cl) [M+H] + ; R t (HPLC): 0.73 min (Method A) Step 6 of EX-35:

[0798] Deprotect the product obtained in Step 5 of EX-35 according to General Procedure H.

[0799] ESI-MS: 625 [M+H] + ; R t (HPLC): 0.60 min (Method A)

[0800] Example EX-37:

[0801] (1R,9S,11S,33S)-4-chloro-1-fluoro-33-methyl-30-oxo-8,15,28-trioxa-6,12,23,31,36-pentaaza-heptacyclo[29.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 heptatriaconta-2,4,6,13(36),14(22),16(21),17,19,23-nonaene-11-carboxylic acid

[0802]

[0803] The title compound was prepared using the following reaction sequence starting from intermediate P-05:

[0804] Step 1 of EX-37: Heck coupling using 2-(prop-2-en-1-yloxy)acetic acid according to General Procedure M

[0805] ESI-MS: 796 / 798 (1Cl) [M+H] + R t (HPLC): 0.65 / 0.66 min (Method D)

[0806] Step 2 of EX-37: Hydrogenation (Pd / C) according to General Procedure N

[0807] ESI-MS: 798 / 800 (1Cl) [M+H] + ; R t (HPLC): 0.97 min (Method B)

[0808] EX-37 Step 3: Removal of BOC protecting group using General Procedure F

[0809] ESI-MS 698 / 700 (1Cl) [M+H] + ; R t (HPLC): 0.63 min (Method B)

[0810] EX-37 Step 4: Amidation using General Procedure E

[0811] ESI-MS 680 / 682 (1Cl) [M+H] + ; R t (HPLC): 0.93 min (Method D)

[0812] EX-37 Step 5: Removal of tert-butyl ester protecting group using General Procedure H

[0813] ESI-MS 624 / 626 (1Cl) [M+H] + ; R t (HPLC): 0.48 min (Method D) Example EX-38:

[0814] (1R,9S,11S,33S)-4-chloro-1-fluoro-33-methyl-30-oxo-8,15,27-trioxa-6,12,23,31,36-pentaaza-heptacyclo[29.2.2.1 9,12 .1 13,24 .0 2,7 .0 14,22 .0 16,21 heptatriaconta-2,4,6,13(36),14(22),16(21),17,19,23-nonaene-11-carboxylic acid

[0815]

[0816] The title compound is prepared starting from intermediate P-05 using the following reaction sequence:

[0817] EX-38 Step 1: Heck coupling using ethyl 3-(vinyloxy)propionate using General Procedure M

[0818] ESI-MS: 768 / 770 (1Cl) [M-tBu+H] + Rt (HPLC): 0.83 min (Method A)

[0819] Step 2 of EX-38: Hydrogenation according to General Procedure N (Pd / C)

[0820] ESI-MS: 826 / 828 (1Cl) [M+H] + ; R t (HPLC): 1.10 min (Method B)

[0821] Step 3 of EX-38: Ester hydrolysis according to General Procedure C

[0822] ESI-MS: 798 / 800 (1Cl) [M+H] + ; R t (HPLC): 0.99 min (Method B)

[0823] Step 4 of EX-38: Removal of BOC protecting group by applying General Procedure F

[0824] ESI-MS 698 / 700 (1Cl) [M+H] + ; R t (HPLC): 0.65 min (Method B)

[0825] Step 5 of EX-38: Amidation by applying General Procedure E

[0826] ESI-MS 680 / 682 (1Cl) [M+H] + ; R t (HPLC): 0.93 min (Method D)

[0827] Step 6 of EX-37: Removal of tert-butyl ester protecting group by applying General Procedure H

[0828] ESI-MS 624 / 626 (1Cl) [M+H] + ; R t (HPLC): 0.53 min (Method A)

[0829] General Technical Annotations

[0830] The terms "ambient temperature" and "room temperature" are used interchangeably and refer to a temperature of about 20 °C, for example 15 °C to 25 °C.

[0831] Generally, the 1 1H NMR spectra and / or mass spectra of the prepared compounds have been obtained. Unless otherwise stated, all chromatographic operations are carried out at room temperature.

[0832] General Methods

[0833] 1H NMR spectra were recorded on a Bruker Avance 400 MHz. LCMS was performed on a quadrupole mass spectrometer of Shimadzu LCMS2010 (column: sepax ODS 50×2.0 mm, 5 μm) or on an Agilent 1200 HPLC, 1956 MSD operating in ES(+) ionization mode (column: Shim-pack XR-ODS 30×3.0 mm, 2.2 μm). Chromatographic purification was carried out by flash chromatography using silica gel of 100 - 200 mesh. Anhydrous solvents were pretreated with MS columns. Unless otherwise stated, all commercially available reagents were used as received.

[0834] According to the method described by D.J. Patel et al., PNAS 2019, 11946 - 11955

[0835] (doi.org / 10.1073 / pnas.1905013116), the absolute configurations of the selected examples (EX-01, EX-03, EX-11, EX-17, EX-

[0836] 37) were assigned based on the co-crystal structure with human cGAS protein.

[0837] List of abbreviations

[0838] ACN Acetonitrile

[0839] aq. Aqueous solution

[0840] BOP Bis((1H-benzo[d][1,2,3]triazol-1-yl)oxy)tris

[0841] (dimethylamino)-phosphonium(V)

[0842] ℃ Degree Celsius

[0843] CH Cyclohexane

[0844] DAST Diethylaminosulfur trifluoride

[0845] DBU 1,8-Diazabicyclo[5.4.0]undec-7-ene

[0846] DCM Dichloromethane

[0847] DIPEA Diisopropylethylamine

[0848] DMA Dimethylacetamide

[0849] DMAP 4-Dimethylaminopyridine

[0850] DMF N,N-Dimethylformamide

[0851] ds diastereoisomers

[0852] EDC (3-dimethylamino-propyl)-ethyl-carbodiimide

[0853] ESI-MS Electrospray ionization mass spectrometry

[0854] EA or EtOAc Ethyl acetate

[0855] eq Equivalent

[0856] h Hour

[0857] HCl Hydrochloric acid

[0858] HATU [Dimethylamino-(1,2,3-triazolo[4,5-b]pyridin-3-

[0859] yloxy)-methylene]-dimethyl-ammonium hexafluorophosphate

[0860] HPLC High performance liquid chromatography

[0861] 2nd generation Grubbs Dichloro[1,3-bis(2,4,6-trimethylphenyl)-2-imidazolidinylidene

[0862] (imidazolidinyliden)](benzylidene), CAS 246047-72-3

[0863] K2CO3 Potassium carbonate

[0864] L Liter

[0865] MeOH Methanol

[0866] Min Minute

[0867] mL Milliliter

[0868] M Mole

[0869] NMP N-Methyl-2-pyrrolidone

[0870] Pd(dppf)Cl2 (1,1′-Bis-(diphenylphosphino)-ferrocene)-palladium(II) dichloride

[0871] Pd(PPh3)4 Tetrakis(triphenylphosphine)palladium(0)

[0872] PTSA p-Toluenesulfonic acid

[0873] RT Room temperature (ca. 20 °C)

[0874] TBAF Tetrabutylammonium fluoride

[0875] TFA Trifluoroacetic acid

[0876] THF Tetrahydrofuran

[0877] TMS Trimethylsilyl

[0878] TosOH p - Toluenesulfonic acid

[0879] RP - HPLC Reversed - phase HPLC

[0880] R t Retention time in minutes

[0881] Analytical method (HPLC / SFC):

[0882] HPLC method A:

[0883]

[0884] Column: XBridge BEH C18_2.1×30mm_1.7μm (Waters); CT: 60 °C

[0885] HPLC method B:

[0886]

[0887] Column: Sunfire C18_3.0×30mm_2.5μm (Waters); CT: 60 °C

[0888] HPLC method C:

[0889]

[0890]

[0891] Column: Sunfire C18_3.0×30mm_2.5μm (Waters); CT: 60 °C

[0892] HPLC method D:

[0893]

[0894] Column: XBridge BEH C18_2.1×30mm_2.5μm (Waters); CT: 60 °C

[0895] HPLC method E:

[0896]

[0897] Column: Kinetex XB-C18 2.6μm (4.6×50mm), CT: 25°C

[0898] HPLC method F:

[0899]

[0900]

[0901] Column: Acquity UPLC BEH C18 1.7μm (2.1×100mm); CT: 40°C

[0902] HPLC method G:

[0903]

[0904] Column: XBridge BEH C18_2.1×30mm_1.7μm (Waters); CT: 60°C

[0905] HPLC method H:

[0906]

[0907] Column: Kinetex XB-C18 2.6μm (4.6×50mm), CT: 25°C 4. Examples

[0908] 5.1 Example compounds of Formula I or II of the present invention

[0909] The following example compounds of Formula I or II outlined in Table 1 have been synthesized and tested for their pharmacological properties with respect to their efficacy in inhibiting cGAS activity.

[0910] Specifically, the "biochemical (in vitro) IC 50 value" for cGAS inhibition (hcGAS IC 50 ); the "IC 50 value for inhibiting IFN induction in virus-stimulated THP1 cells" (THP (vir) IC 50 ); the "IC 50 value for inhibiting IFN induction in cGAMP-stimulated THP1 cells" (THP (cGAMP) IC 50 ); and the "IC 50 value for inhibiting IFN induction in human whole blood stimulated with dsDNA" (hWB IC50 )。The results are summarized in Table 1.

[0911] The example compounds of Formula I or II summarized in Table 1 simultaneously exhibit the following three properties:

[0912] · A satisfactory "biochemical (in vitro) IC 50 value for cGAS inhibition" (where hcGAS IC 50 ≤ 100 nM, preferably ≤ 50 nM, especially ≤ 10 nM),

[0913] · A satisfactory "cellular IC 50 value for cGAS inhibition" (where THP1 (vir) IC 50 ≤ 1 μM, preferably ≤ 500 nM, more preferably ≤ 100 nM, especially ≤ 50 nM)

[0914] and

[0915] · A satisfactory selectivity for cGAS inhibition

[0916] (where the ratio THP1 (cGAMP) IC 50 / THP1 (vir) IC 50 ≥ 10, more preferably ≥ 50, more preferably ≥ 500, especially ≥ 1000).

[0917] In addition, the example compounds of Formula I or II also exhibit an acceptable IC 50 value for IFN induction in human whole blood stimulated with dsDNA (hWB IC 50 ).

[0918] Table 1: Pharmacological properties of the example compounds of Formula I or II of the present invention

[0919]

[0920]

[0921]

[0922]

[0923]

[0924]

[0925]

[0926]

[0927]

[0928]

[0929] Comparison of the Example Compounds of Formulas I or II of 5.2 with the Compounds of the Background Art

[0930] 5.2.1 Compounds of WO 2020 / 142729

[0931] In WO 2020 / 142729, cGAS inhibitors with partially similar structures have been disclosed.

[0932] On pages 44 and 45 of WO 2020 / 142729, the “Biochemistry (in vitro) IC 50 value” (corresponding to “hcGAS IC 50 ”) regarding cGAS inhibition has been disclosed. Here, compounds with a “Biochemistry (in vitro) IC 50 value” less than 100 nM have been designated in “Group A”, compounds with a “Biochemistry (in vitro) IC 50 value” greater than 100 nM and less than 500 nM have been designated in “Group B”, compounds with a “Biochemistry (in vitro) IC 50 value” greater than 500 nM and less than 1 μM have been designated in “Group C”, compounds with a “Biochemistry (in vitro) IC 50 value” greater than 1 μM and less than 10 μM have been designated in “Group D”, and compounds with a “Biochemistry (in vitro) IC 50 value” greater than 10 μM have been designated in “Group E” (see page 44 of WO 2020 / 142729).

[0933] Page 45 of WO 2020 / 142729 discloses that only Compound No. 25 can be designated in “Group A” with a “Biochemistry (in vitro) IC 50 value” less than 100 nM. All other example compounds of WO 2020 / 142729 exhibit a “Biochemistry (in vitro) IC 50 value” exceeding 100 nM.

[0934] Selected background art compounds of WO 2020 / 142729, including Compound No. 25, have been synthesized and subsequently tested for their pharmacological properties regarding their efficacy in inhibiting the cGAS / STING pathway using the same assays as those used to test the compounds of the present invention. Specifically, the “Biochemistry (in vitro) IC 50Value” (hcGASIC 50 );“Regarding the cell IC of IFN induction in THP1 cells stimulated by virus inhibition 50 Value” (THP1 (vir) IC 50 );“Regarding the cell IC of IFN induction in THP1 cells stimulated by cGAMP inhibition 50 Value” (THP1 (cGAMP) IC 50 );and “Regarding the IC of IFN induction inhibition in human whole blood 50 Value” (hWB) (see Table 2).

[0935] Table 2: Pharmacological properties of a series of example compounds of WO 2020 / 142729

[0936]

[0937]

[0938] The pharmacological properties of the example compounds of the present invention outlined in Table 1 and the corresponding pharmacological properties of the compounds of WO 2020 / 142729 outlined in Table 2 can be compared with each other because the compounds were determined experimentally according to the consistent analysis procedure described in Chapter 6 below.

[0939] It is obvious from the data shown in Table 2 that all the example compounds of WO 2020 / 142729 exhibit a “biochemical (in vitro) IC 50 Value” (= hcGAS IC 50 ) significantly greater than 100 nM, the only exception being Example No. 25 of WO 2020 / 142729 (in WO 2020 / 142729, designated in the “biochemical (in vitro) IC 50 Value” (= hcGAS IC 50 ) less than 100 nM “Group A”). In contrast, the “biochemical (in vitro) IC 50 Value” (hcGASIC 50 ) of the example compounds of the present invention are all less than 100 nM. However, Example No. 25 of WO 2020 / 142729 with a “biochemical (in vitro) IC 50 Value” (hcGAS IC 50 ) of 55 nM simply does not meet the selection criteria of “satisfactory cell inhibition efficacy” exhibited by THP1 (vir) IC 50 below 1 μM, because the THP1 of Example No. 25 of WO 2020 / 142729 (vir) IC50 is 17 μM.

[0940] 5.2.2 Compounds of WO 2022 / 174012

[0941] In WO 2022 / 174012, cGAS inhibitors with partially similar structures have been disclosed.

[0942] “Biochemistry (in vitro) IC 50 value” regarding cGAS inhibition has been disclosed on page 65 of WO 2022 / 174012 and “cellular IC 50 value” (THP-1 stimulation for IFNβ ELISA) has been disclosed on page 67 of WO 2022 / 174012. Compound 5 (BBL0100455) of WO 2022 / 174012 seems to be the only compound in WO 2022 / 174012 that may meet the selection criteria of the present invention, meaning it has

[0943] a) “Biochemistry (or enzyme) (in vitro) IC 50 value” less than 100 nM (in the “enzyme assay of WO 2022 / 174012”, compound 5 was measured to belong to “Group B”, which represents an “enzyme IC 50 value” of 50 nM to 100 nM, see Table 2 on page 65 of WO 2022 / 174012)

[0944] b) and has a “cellular IC 50 value” less than 1 μM (THP-1 stimulation for IFNβ ELISA) (in the “cellular assay of WO 2022 / 174012”, compound 5 was measured to belong to “Group A”, which represents a “cellular IC 50 value” of <1 μM, see Tables 3 on pages 67 and 68 of WO 2022 / 174012).

[0945] However, both the biochemistry / enzyme assay and the cellular assay of WO 2022 / 174012 are different from the corresponding “biochemistry / enzyme assay and cellular assay” of the present invention, and thus the biochemistry / enzyme IC 50 value and the cellular IC 50 value measured in WO 2022 / 174012 cannot be compared with the corresponding IC 50 values measured for the compounds of the present invention. Therefore, compound 5 of WO 2022 / 174012 has been synthesized and then tested for its pharmacological properties regarding the efficacy of inhibiting the cGAS / STING pathway using exactly the same assays as those used for testing the compounds of the present invention and described in Chapter 6 below.

[0946] Table 3: Pharmacological properties of compound 5 of WO 2022 / 174012

[0947]

[0948] As shown in the data of Table 3, compound number 5 (BBL0100455) of WO 2022 / 174012 has an acceptable biochemical / enzyme IC 50 value (hcGAS IC 50 = 55 nM), but the cellular IC 50 value is greater than 10000 nM (THP1 (vir) IC 50 = 10000 nM). Therefore, the compounds of the present invention are comparable to compound number 5 of WO 2022 / 174012 in terms of their biochemical / enzyme IC 50 values, but are significantly superior to compound number 5 of WO 2022 / 174012 in terms of their cellular IC 50 values (wherein, for the compounds of formula I or II of the present invention, all are less than 1000 nM).

[0949] 5.3 Prodrugs

[0950] Esters of active agents known to have a carboxylic acid group can represent viable prodrugs, i.e., they can exhibit improved oral absorption / bioavailability compared to the corresponding active agent. Commonly used prodrugs of active agents having a carboxylic acid group are, for example, methyl esters, ethyl esters, isopropyl esters, etc. (See Beaumont et al., Current Drug Metabolism, 2003, Vol. 4, No. 6, 461-485).

[0951] In addition, Nakamura et al., Bioorganic & Medicinal Chem., Vol. 15, No. 24, pp. 7720-7725 (2007) describe that N-acylsulfonamide derivatives and N-acylsulfonylurea derivatives of specific active agents having a free carboxylic acid group may also be viable prodrugs.

[0952] Furthermore, experiments have suggested that the methyl esters of the exemplified compounds of formula I or II also represent viable prodrugs of the cGAS inhibitors of formula I or II.

[0953] Both PCT / EP2022 / 062480 and PCT / EP2022 / 062496 (both not publicly available yet) disclose structurally similar cGAS inhibitors as the cGAS inhibitors of the present invention, all of which also contain a carboxylic acid group linked to a pyrrolidine moiety. In both PCT / EP2022 / 062480 and PCT / EP2022 / 062496, it has been experimentally shown that the methyl ester derivatives of these cGAS inhibitors carrying a carboxylic acid group linked to a pyrrolidine moiety act as viable prodrugs of cGAS inhibitors with a free carboxylic acid group.

[0954] Compounds P01, P02, P03 and P04 of PCT / EP2022 / 062480 are methyl ester derivatives and putative prodrugs of the corresponding Example Compounds 4.04, 1.10, 1.12 and 3.14 of PCT / EP2022 / 062480 (all of which have a free carboxyl group and are active cGAS inhibitors with a lower biochemical IC 50 value and a lower cellular IC 50 value).

[0955] Compounds P01, P02 and P03 of PCT / EP2022 / 062496 are methyl ester derivatives and putative prodrugs of the respective Example Compounds 2.12, 1.13 and 1.05 of PCT / EP2022 / 062496 (all of which have a free carboxyl group and are active cGAS inhibitors with a lower biochemical IC 50 value and a lower cellular IC 50 value).

[0956] In both PCT / EP2022 / 062480 and PCT / EP2022 / 062496, the "active cGAS inhibitor / Example Compound with its free carboxylic acid" and its "corresponding methyl ester derivative / putative prodrug" have been synthesized and tested for their pharmacological properties regarding their efficacy in inhibiting the cGAS / STING pathway.

[0957] This comparison between the properties of the Example Compounds of PCT / EP2022 / 062480 and PCT / EP2022 / 062496 with their free carboxylic acid on the one hand and the properties of their corresponding methyl ester derivatives / putative prodrugs on the other hand shows that the "biochemical IC 50 value (hcGAS IC 50 value)" of the Example Compounds is always approximately or even less than 10 nM, while the "biochemical IC 50 value (hcGASIC 50 value)" of the corresponding methyl ester derivatives / prodrugs is always extremely high, i.e., generally greater than 7000 nM. On the one hand, the IC 50The large difference between the value and on the other hand the IC of its corresponding methyl ester derivative / prodrug 50 value was never observed in the corresponding cellular IC 50 value (THP1 (vir) IC 50 value). The cellular IC 50 values between the exemplified compounds and their corresponding prodrugs always remained more or less within the same range (see Table 4 below).

[0958] A possible explanation for this observation is that the exemplified compounds all have a free carboxyl group, which seems to be crucial for inhibiting cGAS activity, while in all "methyl ester derivatives / prodrugs", the carboxyl group is masked by a carboxyl-methyl group. Therefore, the methyl ester derivatives / prodrugs lose their inhibitory efficacy in the "in vitro human cGAS enzyme assay" (see Chapter 6.1 below), because in this assay, the intracellular enzymes that cleave the carboxyl-methyl group are absent, and thus the key free carboxylic acid group cannot be restored in the biochemical assay. Therefore, the prodrugs show a very large "biochemical (in vitro) IC 50 value" (= hcGASIC 50 ), while the corresponding exemplified compounds (which already have a free carboxylic acid group from the start) show a smaller "biochemical (in vitro) IC 50 value" (= hcGAS IC 50 ).

[0959] In the cellular assay (= "human cGAS cell and counter-cell assay", see Chapter 6.2 below), there are endogenous cellular enzymes that cleave the carboxyl-methyl group. Therefore, not only do the exemplified compounds of PCT / EP2022 / 062480 and PCT / EP2022 / 062496 themselves (which already carry a free carboxylic acid group) show a smaller THP1 (vir) IC 50 value, but also the corresponding methyl ester derivatives / prodrugs show a relatively smaller "THP1 (vir) IC 50 value", because in this "human cGAS cell assay", the carboxyl-methyl group of the prodrug can be cleaved by endogenous intracellular enzymes, and thereby the "active exemplified compound with a free carboxylic acid group" will be released, which in turn shows cGAS inhibitory efficacy.

[0960] This explanation, together with the measurements shown in Table 4, indicates that the carboxy-methyl ester derivatives of the structurally similar example compounds of PCT / EP2022 / 062480 and PCT / EP2022 / 062496 actually seem to represent viable prodrugs of the respective example compounds with a free carboxylic acid group (which itself has no inhibitory efficacy with respect to in vitro human biochemistry cGAS inhibition). However, after cleavage of the carboxy-methyl ester by the endogenous intracellular enzymes present in the cell assay, the "active example compound" is restored, which again exhibits inhibitory efficacy with respect to the cGAS / STING pathway.

[0961] Since the example compounds of formula I or II of the present invention have a free carboxylic acid linked to the pyrrolidine moiety that is very identical to the example compounds of PCT / EP2022 / 062480 or PCT / EP2022 / 062496, it is expected that the carboxy-methyl ester derivatives of these compounds of formula I or II will also act as prodrugs.

[0962] Table 4: Comparison between selected cGAS inhibitor compounds disclosed in PCT / EP2022 / 062480 and PCT / EP2022 / 062496 and their corresponding methyl ester prodrugs:

[0963]

[0964]

[0965]

[0966]

[0967] 5. Biological experiments

[0968] The activity of the compounds of the present invention can be demonstrated using the following in vitro cGAS enzyme and cell assays:

[0969] 6.1 Method: Human cGAS enzyme assay (hcGAS IC 50 (in vitro))

[0970] The human cGAS enzyme is incubated in the presence of 45 base pair double-stranded DNA to activate the enzyme, and GTP and ATP are used as substrates. The compound activity is determined by measuring the effect of the compound on the formation of the enzyme reaction product cGAMP, which is measured by mass spectrometry.

[0971] Enzyme preparation:

[0972] Human cGAS (amino acids 1 - 522) with an N-terminal 6x-His tag and a SUMO tag was expressed in Escherichia coli (E. coli) BL21(DE3)pLysS (Novagen) cells at 18 °C for 16 h. The cells were lysed in a buffer containing 25 mM Tris (pH 8), 300 mM NaCl, 10 mM imidazole, 10% glycerol, protease inhibitor cocktail (cOmplete TM , EDTA-free, Roche) and DNase (5 μg / mL). The cGAS protein was separated by affinity chromatography on Ni-NTA agarose resin and further purified by size exclusion chromatography using a Superdex 200 column (GE Healthcare) equilibrated in 20 mM Tris (pH 7.5), 500 mM KCl and 1 mM TCEP. The purified protein was concentrated to 1.7 mg / mL and stored at -80 °C.

[0973] Analytical methods

[0974] Compounds were delivered in 10 mM DMSO solutions, serially diluted and transferred to 384-well assay plates (Greiner #781201) using an Echo acoustic dispenser. Typically, 8 concentrations were used, with the highest concentration in the final assay volume being 10 μM, followed by approximately 1:5 dilution steps. The DMSO concentration was set to 1% in the final assay volume. The 384-well assay plates contained 22 test compounds (rows 1 - 22) and DMSO (rows 23 - 24).

[0975] After compound transfer, 15 μL of an enzyme-DNA working solution (12 nM cGAS, 0.32 μM DNA of 45 base pairs in assay buffer, 10 mM Tris pH 7.5 / 10 mM KCl / 5 mM MgCl2 / 1 mM DTT) was added to each well in rows 1 - 23 via a MultiDrop Combi dispenser. In row 24, 15 μl of assay buffer without enzyme / DNA was added as a low control.

[0976] The plates were then pre-incubated at room temperature for 60 min.

[0977] Thereafter, 10 μL of an assay buffer of a GTP (ThermoFisher #R0461)-ATP (Promega #V915B) mixture was added to the assay plates (rows 1 - 24, final concentration of 30 μM each) using a Multidrop Combi.

[0978] The plates were incubated again at room temperature for 90 min.

[0979] After incubation, the reaction was stopped by 80 μL of an analytical buffer containing 5 nM cyclic-di-GMP (Sigma #SML1228) in 0.1% formic acid, used as an internal standard for mass spectrometry. The total volume / well was 105 μL.

[0980] Rapidfire MS detection

[0981] Centrifuge each plate at 4000 rpm at 4 °C for 5 min.

[0982] Couple the RapidFire autosampler to a binary pump (Agilent 1290) and a Triple Quad 6500 (ABSciex, Toronto, Canada). This system is equipped with a 10 μL loop, a C18 [12 μL bed volume] cartridge (Agilent, part number G9210A), which contains 10 mM NH4Ac (aq) in water (pH 7.4) as eluent A (pump 1 at 1.5 mL / min, pump 2 at 1.25 mL / min) and 10 mM NH4Ac in v / v / v 47.5 / 47.5 / 5 ACN / MeOH / H2O (pH 7.4) as eluent B (pump 3 at 1.25 mL / min). Aspiration time: 250 ms; loading time: 3000 ms; elution time: 3000 ms; wash volume: 500 μL.

[0983] Operate the MS in positive ion mode with the HESI ion source, where the source temperature is 550 °C, curtain gas = 35, gas 1 = 65, and gas 2 = 80. Obtain unit mass resolution in the SRM mode. Determine the following transitions and MS parameters (DP: declustering potential and CE: collision energy) for cGAMP and DicGMP:

[0984] Analyte: cGAMP at 675.1 / 524, DP = 130, CE = 30, and

[0985] Internal standard: cyclic-di-GMP at 690.1 / 540, DP = 130, CE = 30.

[0986] Monitor the formation of cGAMP and evaluate it as the ratio to cyclic-di-GMP.

[0987] Data evaluation and calculation:

[0988] For data evaluation and calculation, the measurement results of the low control group were set to 0% control and the measurement results of the high control group were set to 100% control. Calculate the IC using the standard 4-parameter logistic regression formula. 50Value. Calculation: [y = (a - d) / (1 + (x / c)^b) + d], where a = low value, d = high value; x = concentration M; c = IC50 M; b = slope.

[0989] 6.2 Method: Human cGAS cell assay and cGAMP-stimulated reverse cell assay (THP1 (vir) IC 50 and THP1 (cGAMP) IC 50 )

[0990] Use THP1-Dual cells expressing the IRF-dependent Lucia luciferase reporter gene (InvivoGen #thpd-nfis) as the basis for both assays. For the detection of cellular cGAS activity, cells are stimulated by infection with a baculovirus delivering the cGAS enzyme to stimulate double-stranded DNA (pFastbac-1, Invitrogen, no coding insert) (measure THP1 TM IC (vir) IC 50 ).

[0991] For the reverse assay, cells are stimulated with cGAMP (SigmaAldrich #SML1232) to activate the same pathway that is independent of cGAS and directly downstream of cGAS (measure THP1 (cGAMP) IC 50 ).

[0992] Pathway activity is monitored by measuring the Lucia luciferase activity induced by DNA-stimulated cGAS enzyme activity (measure THP1 (vir) IC 50 ) or directly by cGAMP (measure THP1 (cGAMP) IC 50 , reverse assay).

[0993] Analysis method

[0994] Compounds are delivered in a 10 mM DMSO solution, serially diluted and transferred to a 384-well assay plate (Greiner #781201) using an Echo acoustic dispenser. Usually 8 concentrations are used, with the highest concentration in the final assay volume being 10 μM, followed by an approximately 1:5 dilution step. The DMSO concentration is set to 1% in the final assay volume. The 384-well assay plate contains 21 test compounds (rows 1 - 22) and DMSO in rows 23 and 24.

[0995] Cells cultured according to the manufacturer's conditions were collected by centrifugation at 300 g / 10 min, and then resuspended in fresh cell culture medium (RPMI 1640 (Gibco#A10491-01), 10% FCS (Gibco#10500), 1×GlutaMax (Gibco#35050-061), 1×Pen / Strep solution (Gibco#15140-122), 100 μg / ml Normocin (InvivoGen#ant-nr), 100 μg / ml Zeocin (InvivoGen#ant-zn), 10 μg / ml Blasticidin S (LifeTechnologies#A11139-03)) and diluted to 1.66E5 cells / ml. Then the baculovirus solution was added to the cells at a ratio of 1:200 (varies depending on the virus batch) (measuring THP1 (vir) IC 50 ). Alternatively, for the reverse analysis, cGAMP was added to the cells at a final concentration of 10 μM (measuring THP1 (cGAMP) IC 50 ).

[0996] 30 μL of the cell / virus mixture was added to each well (5000 cells / well) in rows 1-23 of the compound plate via a MultiDrop Combi dispenser. In row 24, 30 μl / 5000 cells / well (without virus) was added as a low control group.

[0997] The culture plates were then incubated at 37 °C for 18 h in a humidified incubator.

[0998] Thereafter, 15 μL of QuantiLuc detection reagent (InvivoGen#rep-qlcg5) was added to each well using a MultiDrop Combi. Measurements were taken immediately after addition using an EnVision reader (US - luminescence reading mode).

[0999] Data evaluation and calculation:

[1000] For data evaluation and calculation, the measurement results of the low control group were set to 0% control and the measurement results of the high control group were set to 100% control. The IC 50 value was calculated using the standard 4-parameter logistic regression formula. Calculation: [y = (a - d) / (1 + (x / c)^b) + d], a = low value, d = high value; x = concentration M; c = IC50 M; b = slope.

[1001] 6.3 Method: Human whole blood analysis (Human WB IC 50 )

[1002] For the detection of cellular cGAS activity, human whole blood was stimulated with double-stranded DNA by transfection. Pathway activity was monitored by measuring IFNα2α production.

[1003] Analytical method

[1004] Compounds were delivered as 10 mM DMSO solutions and serially diluted, and transferred using an Echo acoustic dispenser into 96-well cell culture plates (Corning #3595), with each well pre-filled with 20 μl of OptiMEM (Gibco, #11058-021). Usually 8 concentrations were used, with the highest concentration in the final assay volume being 10 μM, followed by approximately 1:5 dilution steps. The DMSO concentration was set at 0.1% in the final assay volume. The 96-well assay plates contained 10 test compounds and DMSO in the control wells.

[1005] Human whole blood was collected simultaneously in the form of sodium citrate blood (e.g., 3.8% in Monovettes from Sarstedt) from 3 or more healthy donors (male or female, off medications for 7 days, except contraceptives and thyroxine). After collection, the whole blood was kept at room temperature for up to 3 hours until used in the analysis.

[1006] 160 μl of whole blood samples were transferred to each well of the 96-well assay plates filled with compound / OptiMEM. All assay plates were prepared in duplicate with blood from different donors. The blood plates were kept at room temperature for 60 minutes and continuously shaken at 450 rpm, covered with a lid but not sealed.

[1007] A DNA-Fugene mixture (Herring DNA, Sigma Aldrich #D6898-1G; Fugene (5×1 mL), Promega #E2312) was prepared in OptiMEM and incubated at room temperature for 10 min (125 ng DNA / 20 μl and Fugene ratio 9.6:1). 20 μl of the DNA Fugene mixture was added to each well, resulting in 125 ng DNA / well / 200 μl and Fugene ratio 9.6:1. 20 μl of OptiMEM and 9.6:1 Fugene were added to all low control wells.

[1008] After covering the assay plates with an aera seal and lid, the blood plates were kept at room temperature for 30 minutes and continuously shaken at 450 rpm, then incubated overnight at 37 °C in an incubator for 22 h without shaking.

[1009] To detect IFNα-2α in human plasma, the biotinylated capture antibody (antibody set IFNA2, Meso Scale Diagnostics #B21VH-3, including coating and capture antibodies) was diluted 1:17.5 in diluent 100 (Meso Scale Diagnostics #R50AA-4) according to the manufacturer's instructions. The U-Plex MSD GOLD 96-well streptavidin SECTOR plate (Meso Scale Diagnostics #L45SA-5) was coated with 25 μl of the diluted capture antibody. The coated plate was incubated for 60 min at room temperature with continuous shaking at 700 rpm. The MSD IFNα-2α plate was washed three times with 150 μl of wash buffer (1×HBSS, 0.05% Tween).

[1010] After each plate was blocked for 60 min at room temperature with 100 μl of blocking solution / well (1×HBSS, containing 0.2% Tween, 2% BSA) with continuous shaking at 700 rpm and then emptied as dry as possible by pouring, human plasma was then used.

[1011] The whole blood analysis plates were centrifuged at 1600 rpm for 10 minutes. 25 μl of the supernatant was transferred from each whole blood plate to the corresponding IFNα-2α plate using a pipetting robot. Each plate was sealed with a microplate seal and incubated again for two hours at room temperature with continuous shaking at 700 rpm.

[1012] Subsequently, the MSD IFNα-2α plate was washed three times with 150 μl of wash buffer (1×HBSS, 0.05% Tween), and then 25 μl of MSD SULFO-TAG IFNα-2α antibody solution (diluted 1:100 in diluent 3 (Meso Scale Diagnostics #R50AP-2)) was added to each well of the plate.

[1013] Subsequently, each plate was sealed with a microplate seal and incubated again for two hours at room temperature with continuous shaking at 700 rpm. Finally, the MSD IFNα-2α plate was washed three times with 150 μl of wash buffer (1×HBSS, 0.05% Tween). 150 μl of 2× read buffer was added to each well, and each plate was immediately measured using an MSD Sector S600 reader with the supplier barcode. Data evaluation and calculation:

[1014] For data evaluation and calculation, the control % for each well was calculated based on the mean of the high (DNA stimulation control) and the mean of the low (unstimulated control) controls using the following formula:

[1015] [(Count(sample) - Count(low)) / (Count(high) - Count(low))]*100

[1016] Calculate the IC using the standard 4-parameter logistic regression formula 50 value. Calculate: [y = (a - d) / (1 + (x / c)^b) + d], where a = low value, d = high value; x = concentration M; c = IC 50 M; b = slope.

[1017] 6. Indications

[1018] As has been found, the compounds of formula I or II are characterized by their scope of application in the field of therapy. Particular mention should be made of those applications of the compounds of formula I or II according to the invention which are preferably used on the basis of their pharmaceutical activity as cGAS inhibitors. Although the cGAS pathway is crucial for host defense against pathogen invasion, such as viral infections and the invasion of some intracellular bacteria, cellular stress and genetic factors can also lead to the production of abnormal cellular dsDNA, for example, through nuclear or mitochondrial leakage, and thereby trigger an autoinflammatory response. Therefore, cGAS inhibitors have strong therapeutic potential for the treatment of different autoinflammatory and autoimmune diseases.

[1019] An et al., Arthritis Rheumatol. April 2017; 69(4):800 - 807 disclosed that the cGAS expression in peripheral blood mononuclear cells (PBMCs) of patients with the autoimmune disease systemic lupus erythematosus (SLE) was significantly higher than that in the normal control group. By targeted measurement of cGAMP by tandem mass spectrometry, cGAMP was detected in 15% of the tested SLE patients, but not in the normal or rheumatoid arthritis control groups. SLE patients with cGAMP had a higher disease activity compared to SLE patients without cGAMP. However, the higher cGAS expression could be the result of exposure to type I interferon (IFN), but the detection of cGAMP in SLE patients with increased disease activity indicates that the cGAS pathway may be involved in disease expression.

[1020] Park et al., Ann Rheum Dis. October 2018; 77(10):1507 - 1515 also disclosed that the cGAS pathway is involved in the development of SLE.

[1021] Thim-Uam et al., iScience September 4, 2020; 23(9), 101530 (doi:10.1016 / j.isci.2020.101530) disclosed that the STING pathway mediates lupus via the activation of conventional dendritic cell maturation and plasmacytoid dendritic cell differentiation.

[1022] Gao et al., Proc. Natl. Acad. Sci. USA. 2015 Oct 20;112(42):E5699-705 describe that cGAS causes certain autoimmune diseases, such as interferonopathy, through activation of self-DNA.

[1023] Tonduti et al., Expert Rev. Clin. Immunol. 2020 Feb; 16(2): 189-198 disclose that cGAS inhibitors have particular therapeutic potential in Icardi-Guterres syndrome and familial lupus chilblains, which are lupus-like severe autoinflammatory immune-mediated disorders.

[1024] Steiner et al., Nat Commun. 2022 Apr 28;13(1):232;doi:10.1038 show that deficiency of coatomer complex I causes aberrant activation of STING signaling and COPA syndrome, and that cGAS is required to drive type I IFN signaling in a cell model of COPA syndrome.

[1025] Li et al. showed that extracellular vesicles containing plasma-derived DNA induced a STING-mediated proinflammatory response in dermatomyositis (Theranostics. 2021; 11(15): 7144-7158). Zhou et al. (J Clin Lab Anal. 2022 Oct; 36(10): e24631) described a correlation between cGAS-STING pathway activation and muscle fiber atrophy / necrosis in dermatomyositis.

[1026] In Yu et al., Cell 2020 Oct 29; 183(3): 636-649, a link between TDP-43-triggered mitochondrial DNA and activation of the cGAS / STING pathway in amyotrophic lateral sclerosis (ALS) is described.

[1027] Ryu et al., Arthritis Rheumatol. 2020 Nov; 72(11): 1905-1915 also showed that bioactive plasma mitochondrial DNA is associated with disease progression in specific fibrotic diseases, such as systemic sclerosis (SSc) or interstitial lung disease (ILD), progressive fibrosing interstitial lung disease (PF-ILD) and idiopathic pulmonary fibrosis (IPF).

[1028] In Schuliga et al., Clin. Sci. (Lond). 2020 Apr 17;134(7):889-905, it is described that self-DNA perpetuates senescence of IPF lung fibroblasts in a cGAS-dependent manner.

[1029] Additional scientific hints linking the etiology of other fibrotic diseases, such as nonalcoholic steatohepatitis (NASH), to the cGAS / STING pathway have been described in Yu et al., J. Clin. Invest. 2019 Feb 1;129(2):546-555 and Cho et al., Hepatology. 2018 Oct;68(4):1331-1346.

[1030] Nascimento et al., Sci. Rep. 2019 Oct 16;9(1):14848 disclose that autologous DNA release and STING-dependent sensing drive cigarette smoke-induced inflammation in mice, suggesting a link between the cGAS-STING pathway and chronic obstructive pulmonary disease (COPD).

[1031] Ma et al., Sci. Adv. 2020 May 20;6(21):eaaz6717 discloses that it can limit ulcerative colitis and inflammatory bowel disease (IBD) by controlling cGAS-mediated inflammation.

[1032] Gratia et al., J. Exp. Med. 2019 May 6; 216(5): 1199-1213 show that Bloom syndrome protein restricts innate immune sensing of micronuclei through cGAS. Therefore, cGAS inhibitors have therapeutic potential for the treatment of Bloom syndrome.

[1033] Kerur et al., Nat. Med. 2018 Jan;24(1):50-61 described that cGAS plays an important role in atypical inflammasome activation in age-related macular degeneration (AMD).

[1034] Visitchanakun et al., Int J Mol Sci. 2021 Oct 23;22(21):11450 showed that GAS-deficient mice had less severe sepsis than wild-type mice in the cecal ligation and puncture (CLP) and lipopolysaccharide (LPS) injection sepsis models.

[1035] Wang et al., Mediators Inflamm. 2015; 2015: 192329, describe cGAS as required for cell proliferation and inflammatory cytokine production in rheumatoid arthritis synovial cells. It has also been reported that cGAS deficiency suppresses interferon response, inflammatory cell infiltration and joint swelling in a mouse model of inflammatory arthritis (Willemsen et al., Cell Rep. 2021 Nov 9; 37(6): 109977).

[1036] Guo et al., Osteoarthritis Cartilage. August 2021; 29(8): 1213-1224 describe damaged DNA as a key pathological factor in osteoarthritis (OA), and this may be mediated by the cGAS / STING pathway, as STING attenuation reduces the instability of medial meniscus-induced OA development in mice.

[1037] Mao et al., Arterioscler Thromb Vasc Biol (2017) 37(5): 920-929 show that in diet-induced obesity, the cGAS / STING pathway mediates endothelial inflammation in response to free fatty acid-induced mitochondrial damage, indicating that cGAS inhibitors also have the potential to treat obesity and diabetes.

[1038] Kerur et al., Nat Med. January 2018; 24(1): 50-61 describe elevated cGAS levels in the retinal pigment epithelium of the eyes of humans with geographic atrophy, and cGAS drives atypical inflammasome activation in age-related macular degeneration.

[1039] cGAS promotes cellular senescence and the senescence-associated secretory phenotype (Yang et al., Proc Natl Acad Sci USA June 6, 2017; 114: E4612-E4620). Cytoplasmic chromatin triggers inflammation during senescence via cGAS / STING, and STING knockout mice have reduced tissue inflammation and senescence (Dou et al., Nature. 2017 550: 402-406). In addition, variants within the human STING gene are associated with healthy aging, likely due to reduced inflammation (Hamann et al., Gerontology 2019; 65: 145-154). Collectively, STING inhibitors will reduce senescence-associated inflammation and senescent cell accumulation, and will improve senescence-related diseases, such as aging, muscle disorders, and fibrosis.

[1040] In addition, the cGAS inhibitors of formula I or II have therapeutic potential in the treatment of cancer (see Hoong et al., Oncotarget. July 28, 2020; 11(30): 2930-2955, and Chen et al., Sci. Adv. October 14, 2020; 6(42): eabb8941).

[1041] Furthermore, the cGAS inhibitors of formula I or II also have therapeutic potential in the treatment of heart failure (Hu et al., Am. J. Physiol. Heart Circ. Physiol. June 1, 2020; 318(6): H1525-H1537).

[1042] There are other scientific indications of a correlation between Parkinson's disease and the cGAS / STING pathway (Sliter et al., Nature. September 2018; 561(7722):258 - 262) and between Sjogren's syndrome and the cGAS / STING pathway (Papinska et al., J. Dent. Res. July 2018; 97(8):893 - 900).

[1043] In addition, the cGAS inhibitors of formula I or II also have therapeutic potential in the treatment of COVID-19 / SARS-CoV-2 infection, as shown in the following literature: Di Domizio et al., Nature. January 19, 2022, doi:10.1038 / s41586-022-04421-w: “The cGAS-STING pathway drives type I IFN immunopathology in COVID-19”, and Neufeldt et al., Commun Biol. January 12, 2022; 5(1):45. doi:10.1038 / s42003-021-02983-5: “SARS-CoV-2 infection induces a pro-inflammatory cytokine response through cGAS-STING and NF-kappaB”.

[1044] Furthermore, the cGAS inhibitors of formula I or II have therapeutic potential in the treatment of kidney inflammation and renal fibrosis, as shown in the following literature: Chung et al., Cell Metab. 2019 30:784 - 799: “Mitochondrial Damage and Activation of the STING Pathway Lead to Renal Inflammation and Fibrosis”, and Maekawa et al., Cell Rep. 2019 29:1261 - 1273: “Mitochondrial Damage Causes Inflammation via cGAS-STING Signaling in Acute Kidney Injury”.

[1045] In addition, cGAS inhibitors of Formula I or II have therapeutic potential in the treatment of cancer, as shown in the following literature: Bakhoum et al., Nature. January 25, 2018; 553(7689):467-472: “Chromosomal instability drives metastasis through a cytosolic DNA response”, and Liu et al., Nature. November 2018; 563(7729):131-136: “Nuclear cGAS suppresses DNA repair and promotes tumorigenesis”.

[1046] In addition, because STING gt Animals show reduced macrophage infiltration in adipose tissue after subchronic high-calorie intake (HFD), and the deficiency of STING gt and IRF3 causes reduced blood glucose and insulin and weight loss, so cGAS inhibitors of Formula I or II have therapeutic potential in the treatment of metabolic disorders (Mao et al., Arterioscler Thromb Vasc Biol, 2017; 37(5):920-929).

[1047] In addition, because mitochondrial DNA release activates the cGAS / STING pathway in the cytosol of endothelial cells and inhibits endothelial cell proliferation, cGAS inhibitors of Formula I or II have therapeutic potential in the treatment of vascular diseases and cause blood vessel repair / regeneration. Additionally, gene knockout of the cGAS gene restores endothelial repair / regeneration in a mouse model of inflammatory lung injury (Huang et al., Immunity, March 2020, 2017; 52(3):475-486.e5.doi:10.1016 / j.immuni.2020,02.002).

[1048] In addition, cGAS inhibitors of Formula I or II have therapeutic potential in the treatment of aging-related and obesity-related cardiovascular diseases (Hamann et al., Immun Ageing, March 14, 2020; 17:7; doi:10.1186 / s12979-020-00176-y.eCollection 2020).

[1049] Therefore, compounds of Formula I or II as cGAS inhibitors are useful for treating autoinflammatory and autoimmune diseases such as systemic lupus erythematosus (SLE), interferonopathy, Acardi-Guterres syndrome (AGS), COPA syndrome, familial lupus pernio, age-related macular degeneration (AMD), amyotrophic lateral sclerosis (ALS), inflammatory bowel disease (IBD), chronic obstructive pulmonary disease (COPD), Bloom's syndrome, Sjögren's syndrome, rheumatoid arthritis and Parkinson's disease.

[1050] In addition, compounds of formula I or II as cGAS inhibitors can be used to treat fibrotic diseases, such as systemic sclerosis (SSc), interferonopathy, non-alcoholic steatohepatitis (NASH), interstitial lung disease (ILD), preferably progressive fibrosing interstitial lung disease (PF-ILD), especially idiopathic pulmonary fibrosis (IPF).

[1051] In addition, compounds of Formula I or II as cGAS inhibitors can be used to treat age-related macular degeneration (AMD), retinopathy, glaucoma, diabetes, obesity, aging, muscle disorders, sepsis, osteoarthritis, heart failure, COVID-19 / SARS-CoV-2 infection, kidney inflammation, renal fibrosis, metabolic disorders, vascular diseases, cardiovascular diseases and cancer.

[1052] 7. Combination

[1053] The compounds of Formula I or II may be administered to a patient alone or in combination with one or more other pharmacologically active agents.

[1054] In a preferred embodiment of the invention, the compounds of formula I or II may be combined with one or more pharmacologically active agents selected from the following groups: anti-inflammatory agents; antifibrotic agents; antiallergic agents / antihistamines; bronchodilators; β2 agonists / β mimetics; adrenergic agonists; anticholinergics; methotrexate; mycophenolate mofetil; leukotriene modulators; JAK inhibitors; anti-interleukin antibodies; non-specific immunotherapeutics such as interferons or other cytokines / chemokines; cytokine / chemokine receptor modulators (i.e., cytokine receptor agonists or antagonists); Toll-like receptor agonists (=TLR agonists); immune checkpoint modulators; anti-TNF antibodies (Humira TM ); and anti-BAFF antibodies (belimumab and etanercept).

[1055] The anti-fibrotic agent is preferably selected from pirfenidone or a tyrosine kinase inhibitor, such as nintedanib, with nintedanib being particularly preferred.

[1056] Preferred embodiments of anti-inflammatory agents are NSAIDs and corticosteroids.

[1057] The NSAID is preferably selected from ibuprofen, naproxen, diclofenac, meloxicam, celecoxib, acetylsalicylic acid (Aspirin TM ), indomethacin, mefenamic acid, and etoricoxib.

[1058] The corticosteroid is preferably selected from flunisolide, beclomethasone, triamcinolone, budesonide, fluticasone, mometasone, ciclesonide, rofleponide, and dexamethasone.

[1059] The anti-allergy agent / antihistamine is preferably selected from epinastine, cetirizine, azelastine, fexofenadine, levocabastine, loratadine, ebastine, desloratidine, and mizolastine.

[1060] The β2 agonist / β-mimetic can be a long-acting β2 agonist (LABA) or a short-acting β agonist (SABA). Particularly preferred β2 agonists / β-mimetics are selected from bambuterol, bitolterol, carbuterol, clenbuterol, fenoterol, formoterol, hexoprenaline, ibuterol, pirbuterol, procaterol, reproterol, salmeterol, sulfonterol, terbutalin, tolubuterol, olodaterol, and salbutamol, especially olodaterol.

[1061] The anticholinergics are preferably selected from ipratropium salts, tiotropium salts, glycopyrronium salts and theophylline, with tiotropium bromide being particularly preferred.

[1062] The leukotriene regulators are preferably selected from Montelukast, Pranlukast, Zafirlukast, Ibudilast and Zileuton.

[1063] The JAK inhibitors are preferably selected from Baricitinib, Cerdulatinib, Fedratinib, Filgotinib, Gandotinib, Lestaurtinib, Momelotinib, Pacritinib, Peficitinib, Ruxolitinib, Tofacitinib and Upadacitinib.

[1064] The anti - interleukin antibodies are preferably selected from anti - IL23 antibodies (such as risankizumab), anti - IL17 antibodies, anti - IL1 antibodies, anti - IL4 antibodies, anti - IL13 antibodies, anti - IL - 5 antibodies, anti - IL - 6 antibodies (such as Actemra TM ), anti - IL - 12 antibodies, anti - IL - 15 antibodies.

[1065] 8. Formulations

[1066] The compounds of the present invention can be administered by any suitable route of administration, including both systemic administration and local administration. Systemic administration includes oral administration, parenteral administration, transdermal administration, rectal administration, and administration by inhalant. Parenteral administration refers to a route of administration other than enteral, transdermal or by inhalant, and is usually by injection or infusion. Parenteral administration includes intravenous, intramuscular, intrasternal and subcutaneous injection or infusion. Inhalation refers to administration to the lungs of a patient, whether inhaled via the oral cavity or via the nasal passages. Local administration includes application to the skin. The compounds of the present invention can be administered via eye drops to treat Sjogren's syndrome.

[1067] The forms suitable for administration are, for example, tablets, capsules, solutions, syrups, emulsions or inhalable powders or aerosols. In each case, the content of the pharmaceutically effective compound should be in the range of 0.1 to 90% by weight, preferably 0.5 to 50% by weight of the total composition, i.e., in an amount sufficient to achieve the dosage range specified hereinafter.

[1068] The preparation can be administered orally in the form of tablets, powders, the powder in capsules (e.g., hard gelatin capsules), solutions or suspensions. When administered by inhalation, the active substance combination can be given in the form of a powder, an aqueous solution or an aqueous ethanol solution or using a propellant gas preparation.

[1069] Therefore, preferably, the pharmaceutical preparation is characterized by the content of one or more compounds of formula I or II according to the above preferred embodiments.

[1070] Oral administration of the compounds of formula I or II is particularly preferred, and their administration once or twice a day is also particularly preferred. Suitable tablets can be obtained, for example, by mixing the active substance with known excipients (such as inert diluents, such as calcium carbonate, calcium phosphate or lactose; disintegrants, such as corn starch or alginic acid; binders, such as starch or gelatin; lubricants, such as magnesium stearate or talc; and / or agents for delayed release, such as carboxymethyl cellulose, cellulose acetate phthalate or polyvinyl acetate)). Tablets can also comprise several layers.

[1071] Therefore, coated tablets can be prepared by coating a core similar to that produced by tablets with substances commonly used for tablet coating (such as kollidone or shellac, gum arabic, talc, titanium dioxide or sugar). To achieve delayed release or prevent incompatibility, the core can also consist of multiple layers. Similarly, the excipients mentioned above with respect to tablets can be used, and the tablet coating can consist of multiple layers to achieve delayed release.

[1072] Syrups containing the active substance or its combination according to the invention can additionally contain sweeteners such as saccharin, cyclamate, glycerol or sugar and flavor enhancers (e.g., flavoring agents such as vanilla essence or orange extract). They can also contain suspending adjuvants or thickeners, such as sodium carboxymethyl cellulose; wetting agents, such as the condensation product of a fatty alcohol and ethylene oxide; or preservatives, such as parabens.

[1073] Capsules containing one or more active substances or active substance combinations can be prepared, for example, by mixing the active substance with an inert carrier such as lactose or sorbitol and filling it into gelatin capsules. Suitable suppositories can be manufactured, for example, by mixing carriers provided for this purpose, such as neutral fats or polyethylene glycols or their derivatives.

[1074] Excipients that can be used include, for example, water; pharmaceutically acceptable organic solvents such as paraffin (e.g., petroleum fractions), vegetable oils (e.g., peanut oil or sesame oil), monofunctional or polyfunctional alcohols (e.g., ethanol or glycerol); carriers such as natural mineral powders (e.g., kaolin, clay, talc, chalk), synthetic mineral powders (e.g., highly dispersed silicic acid and silicates), sugars (e.g., cane sugar, lactose, and glucose), emulsifiers (e.g., lignin, spent sulfite liquor, methyl cellulose, starch, and polyvinylpyrrolidone), and lubricants (e.g., magnesium stearate, talc, stearic acid, and sodium lauryl sulfate).

[1075] Of course, for oral administration, in addition to the above carriers, tablets may also contain additives such as sodium citrate, calcium carbonate, and dibasic calcium phosphate, as well as various additives such as starch (preferably potato starch), gelatin, and the like. In addition, lubricants such as magnesium stearate, sodium lauryl sulfate, and talc can be used simultaneously in the tablet manufacturing process. In the case of aqueous suspensions, the active substance can be combined with various flavor enhancers or colorants in addition to the excipients mentioned above.

Claims

1. A compound of formula I wherein R 1 selected from the group consisting of hydrogen, halogen, methyl, ethyl, -CF3, -CHF2, -CFH2 and methoxy, R 2 selected from the group consisting of hydrogen and methyl; R 3 selected from the group consisting of hydrogen, methyl, halogen, ethynyl, propargyl, -CO-(C 1-3 -alkyl), -CO-NH2, -CO-NHCH3, -CO-N(CH3)2 and a 5- or 6-membered heteroaryl ring having 1 or 2 heteroatoms each independently selected from N, S or O, wherein said heteroaryl ring may optionally be further substituted by one or two additional substituents each independently selected from the group consisting of F, Cl, Br, -O-CH3, methyl, -CF3, -CHF2, CH2F; R 4 selected from the group consisting of hydrogen, -OH and F; and wherein A is selected from the group consisting of: -CH2-, -O-, -CF2- and -CHCH3-; D is selected from the group consisting of: -CH2-, -O-, -CF2- and -CHCH3-; E is selected from the group consisting of: -CH2-, -CO-, -O-, -CF2- and -CHCH3-; G is selected from the group consisting of: -NH-, -NCH3-, -CH2-, -O-, -CF2- and -CHCH3-; J is selected from the group consisting of: -CO-, -CH2-, -O-, -CF2- and -CHCH3-; K is selected from the group consisting of -CH2-, -CF2- and -O- or is absent; L is selected from the group consisting of -CH2-, -CHCH3- and -CF2- or is absent; and prodrugs, deuterated analogs and pharmaceutically acceptable salts thereof.

2. The compound of formula II according to claim 1, wherein R 1 selected from the group consisting of hydrogen and / or halogen, wherein the halogen is selected from the group consisting of F and Cl; R 2 selected from the group consisting of hydrogen and methyl; R 3 selected from the group consisting of hydrogen, Cl, Br, ethynyl, propargyl, -CO-(CH3) and a 5- or 6-membered heteroaryl ring having 1 or 2 heteroatoms each independently selected from N, S or O, wherein said heteroaryl ring may optionally be further substituted with one or two additional substituents each independently selected from the group consisting of F, -O-CH3 and methyl; R 4 is F; and wherein A is selected from the group consisting of: -CH2-, -O-, -CF2- and -CHCH3-; D is selected from the group consisting of -CH2- and -O-; E is selected from the group consisting of: -CH2-, -CO- and -O-; G is selected from the group consisting of: -NH-, -NCH3-, -CH2- and -O-; J is selected from the group consisting of: -CO-, -CH2- and -O-; K is selected from the group consisting of -CH2-, -CF2- and -O- or is absent; L is -CH2- or is absent; and prodrugs, deuterated analogs and pharmaceutically acceptable salts thereof.

3. The compound of formula I according to claim 1 or the compound of formula II according to claim 2, wherein L is absent, and prodrugs, deuterated analogs and pharmaceutically acceptable salts thereof.

4. The compound of formula I according to claim 1 or the compound of formula II according to claim 2, wherein L and K are absent, and prodrugs, deuterated analogs and pharmaceutically acceptable salts thereof.

5. The compound of formula I according to claim 1 or the compound of formula II according to claim 2, wherein L is absent and wherein K is -CF2-, and prodrugs, deuterated analogs and pharmaceutically acceptable salts thereof.

6. The compound of formula I according to claim 1 or the compound of formula II according to claim 2, wherein L is absent and wherein A is selected from the group consisting of -CH2- and -CF2-, and prodrugs, deuterated analogs and pharmaceutically acceptable salts thereof.

7. The compound of formula I according to claim 1 or the compound of formula II according to claim 2, wherein L is absent and wherein A is -O-, and prodrugs, deuterated analogs and pharmaceutically acceptable salts thereof.

8. The compound of formula I according to claim 1 or the compound of formula II according to claim 2, wherein L is absent and wherein D is -O-, and prodrugs, deuterated analogs and pharmaceutically acceptable salts thereof.

9. The compound of formula I according to claim 1 or the compound of formula II according to claim 2, wherein L is absent and wherein R 3 is selected from the group consisting of Cl, Br, ethynyl, propargyl, and a 5- or 6-membered heteroaryl ring selected from the group consisting of pyridyl and pyrazolyl, wherein said heteroaryl ring may optionally be further substituted with one or two additional substituents each independently selected from the group consisting of F, -O-CH3, and methyl; and prodrugs, deuterated analogs and pharmaceutically acceptable salts thereof.

10. The compound of formula I according to claim 1 or the compound of formula II according to claim 2, wherein R 1 is selected from the group consisting of F and Cl, and prodrugs, deuterated analogs and pharmaceutically acceptable salts thereof.

11. The compound of formula I according to claim 1 or the compound of formula II according to claim 2, wherein R 1 is hydrogen, and prodrugs, deuterated analogs and pharmaceutically acceptable salts thereof.

12. The compound of formula I as claimed in claim 1 or the compound of formula II as claimed in claim 2, which is selected from the group consisting of: and its prodrugs, deuterated analogs and pharmaceutically acceptable salts.

13. The compound of formula I as claimed in claim 1 or the compound of formula II as claimed in claim 2, wherein A is selected from the group consisting of -CH2- and -O-; D is selected from the group consisting of -CH2- and -O-; E is selected from the group consisting of -CH2- and -O-; G is selected from the group consisting of -CH2- and -O-; J is selected from the group consisting of -CH2- and -O-; K is selected from the group consisting of -CH2- and -CF2-; L is absent; and its prodrugs, deuterated analogs and pharmaceutically acceptable salts.

14. The compound of formula II according to claim 13, which is selected from the group consisting of: and its prodrugs, deuterated analogs and pharmaceutically acceptable salts.

15. The compound of formula I as claimed in claim 1 or the compound of formula II as claimed in claim 2, wherein A is selected from the group consisting of -CH2- and -O-; D is selected from the group consisting of -CH2- and -O-; E is -CH2-; G is selected from the group consisting of -CH2- and -O-; J is -CH2-; K is selected from the group consisting of -CH2- and -CF2-; L is absent; and its prodrugs, deuterated analogs and pharmaceutically acceptable salts.

16. The compound of formula II according to claim 15, which is selected from the group consisting of: and its prodrugs, deuterated analogs and pharmaceutically acceptable salts.

17. An intermediate compound, which has a) formula (A-I) wherein R 1 , R 2 , R 3 , R 4 , A, D, E, G, J, K and L are as defined above, and wherein R 13 is selected from the group consisting of hydrogen, methyl, ethyl and tert-butyl, b) formula (A-II) wherein R 1 , R 2 , R 3 , R 4 , A, D, E, G, J, K and L are as defined above, wherein R 13 is selected from the group consisting of: hydrogen, methyl, ethyl and tert-butyl, and wherein R is hydrogen or a protecting group selected from the group consisting of tert-butyl, methyl, ethyl and benzyl, c) formula (B-I) wherein R 1 , R 2 , R 3 , R 4 , A, D, E, G, J, K and L are as defined above, and wherein R 13 is selected from the group consisting of hydrogen, methyl, ethyl and tert-butyl, d) formula (C-I) wherein R 1 、R 2 、R 3 、R 4 、A, D, E, G, J, K and L are as defined above, and wherein R 13 is selected from the group consisting of hydrogen, methyl, ethyl and tert-butyl, or has e) formula (C-II) wherein R 1 , R 2 , R 3 , R 4 , A, D, E, G, J, K and L are as defined above, and wherein R 13 is selected from the group consisting of: hydrogen, methyl, ethyl and tert-butyl, wherein PG is selected from the group consisting of: tert-butoxycarbonyl (Boc), allyloxycarbonyl (Alloc), benzyloxycarbonyl (Cbz) and fluorenylmethoxycarbonyl (Fmoc).

18. The compound of formula I as claimed in one or more of claims 1 to 16 or the compound of formula II as claimed in one or more of claims 2 to 16, which is used for treating diseases that can be treated by inhibiting cGAS.

19. A compound of formula I as claimed in one or more of claims 1 to 16 or a compound of formula II as claimed in one or more of claims 2 to 16 for use in the treatment of a disease selected from the group consisting of: systemic lupus erythematosus (SLE); interferonopathy; Aicardi-Goutières syndrome (AGS); COPA syndrome; familial pernio lupus; age-related macular degeneration (AMD); amyotrophic lateral sclerosis (ALS); retinopathy; glaucoma; diabetes; obesity; inflammatory bowel disease (IBD); chronic obstructive pulmonary disease (COPD); Bloom's syndrome; dermatomyositis; Sjogren's syndrome syndrome; Parkinson's disease; heart failure; cancer; aging; muscle disorders; sepsis; rheumatoid arthritis; osteoarthritis; COVID-19; systemic sclerosis (SSc); non-alcoholic steatohepatitis (NASH); interstitial lung disease (ILD), preferably progressive fibrosing interstitial lung disease (PF-ILD), in particular idiopathic pulmonary fibrosis (IPF).

20. A compound of formula I as claimed in one or more of claims 1 to 16 or a compound of formula II as claimed in one or more of claims 2 to 16 for use in the treatment of a disease selected from the group consisting of systemic lupus erythematosus (SLE), interferonopathy, Acardi-Guterres syndrome (AGS), COPA syndrome, familial lupus pernio, dermatomyositis, age-related macular degeneration (AMD), amyotrophic lateral sclerosis (ALS), inflammatory bowel disease (IBD), chronic obstructive pulmonary disease (COPD), Bloom's syndrome, Sjögren's syndrome, rheumatoid arthritis and Parkinson's disease.

21. A compound of formula I as described in one or more of claims 1 to 16 or a compound of formula II as described in one or more of claims 2 to 16 for use in the treatment of a disease selected from the group consisting of: systemic sclerosis (SSc); non-alcoholic steatohepatitis (NASH); interferonopathy; interstitial lung disease (ILD), preferably progressive fibrosing interstitial lung disease (PF-ILD), in particular idiopathic pulmonary fibrosis (IPF).

22. A compound of formula I as described in one or more of claims 1 to 16 or a compound of formula II as described in one or more of claims 2 to 16 for use in the treatment of a disease selected from the group consisting of age-related macular degeneration (AMD), retinopathy, glaucoma, diabetes, obesity, aging, muscle disorders, sepsis, osteoarthritis, heart failure, COVID19 / SARS-CoV-2 infection, kidney inflammation, renal fibrosis, metabolic disorders, vascular diseases, cardiovascular diseases and cancer.

23. A pharmaceutical composition comprising a compound of formula I as claimed in one or more of claims 1 to 16 or a compound of formula II as claimed in one or more of claims 2 to 16 and optionally one or more pharmaceutically acceptable carriers and / or excipients.

24. A pharmaceutical composition comprising a compound of formula I as claimed in one or more of claims 1 to 16 or a compound of formula II as claimed in one or more of claims 2 to 16 and one or more active agents selected from the group consisting of: anti-inflammatory agents; anti-fibrotic agents; anti-allergic agents / anti-histamines; bronchodilators; β2 agonists / β mimetics; adrenergic agonists; anti-cholinergics; methotrexate; mycophenolate mofetil; leukotriene regulators; JAK inhibitors; anti-interleukin antibodies; non-specific immunotherapeutic agents such as interferons or other cytokines / chemokines; cytokine / chemokine receptor regulators; Toll-like receptor agonists; immune checkpoint regulators; anti-TNF antibodies such as Humira TM ; anti-BAFF antibodies such as Belimumab and Etanercept; and optionally one or more pharmaceutically acceptable carriers and / or excipients.

25. A pharmaceutical composition comprising a compound of formula I as claimed in one or more of claims 1 to 16 or a compound of formula II as claimed in one or more of claims 2 to 16 and one or more anti-fibrotic agents selected from the group consisting of pirfenidone and nintedanib, and optionally one or more pharmaceutically acceptable carriers and / or excipients.

26. A pharmaceutical composition comprising a compound of formula I as claimed in one or more of claims 1 to 16 or a compound of formula II as claimed in one or more of claims 2 to 16 and one or more anti-inflammatory agents selected from the group consisting of NSAIDs and corticosteroids, and optionally one or more pharmaceutically acceptable carriers and / or excipients.

27. A pharmaceutical composition comprising a compound of formula I as claimed in one or more of claims 1 to 16 or a compound of formula II as claimed in one or more of claims 2 to 16 and one or more active agents selected from the group consisting of bronchodilators, β2 agonists / β mimetics, adrenergic agonists and anticholinergics; and optionally one or more pharmaceutically acceptable carriers and / or excipients.

28. A pharmaceutical combination comprising a compound of formula I as described in one or more of claims 1 to 16 or a compound of formula II as described in one or more of claims 2 to 16 and one or more anti - interleukin antibodies selected from the group consisting of: anti - IL - 23 antibodies such as Risankizumab; anti - IL - 17 antibodies; anti - IL - 1 antibodies; anti - IL - 4 antibodies; anti - IL - 13 antibodies; anti - IL - 5 antibodies; anti - IL - 6 antibodies such as Actemra TM ; anti - IL - 12 antibodies; and anti - IL - 15 antibodies.

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