N-Cyanopyrrolidines with activity as USP30 inhibitors

Abstract

The present invention relates to a class of N-cyanopyrrolidines (formulas (I)(i) and (I)(ii)) that have activity as inhibitors of the deubiquitinase USP30 and have utility in a variety of therapeutic areas, including conditions involving mitochondrial dysfunction, cancer, and fibrosis. JPEG2023529570000081.jpg2981

Description

[Technical Field]

[0001] FIELD OF THE INVENTION The present invention relates to a class of N-cyanopyrrolidines that have activity as inhibitors of the deubiquitinating enzyme ubiquitin C-terminal hydrolase 30, also known as ubiquitin-specific peptidase 30 (USP30), their uses, methods for their preparation, and compositions containing said inhibitors. These inhibitors are useful in a variety of therapeutic areas, including conditions involving mitochondrial dysfunction, cancer, and fibrosis. All documents cited or relied upon below are expressly incorporated herein by reference. [Background technology]

[0002] Background of the Invention Ubiquitin is a small 76-amino acid protein important for regulating protein function within cells. Ubiquitination and deubiquitination are enzyme-mediated processes in which ubiquitin is covalently attached to or cleaved from target proteins by deubiquitinating enzymes (DUBs). Human cells contain approximately 100 DUBs, which are classified into subfamilies based on sequence homology. The USP family is characterized by a common Cys and His box, which contains Cys and His residues important for DUB activity. The ubiquitination and deubiquitination processes are involved in the regulation of many cellular functions, including cell cycle progression, apoptosis, cell surface receptor modification, DNA transcription, and DNA repair. Therefore, the ubiquitin system is involved in the pathogenesis of numerous disease states, including inflammation, viral infections, metabolic dysfunction, CNS disorders, and carcinogenesis.

[0003] Ubiquitin is a master regulator of mitochondrial dynamics. Mitochondria are dynamic organelles whose biogenesis, fusion, and fission events are regulated by post-translational regulation via ubiquitination of many key factors, including mitofusins. In humans, USP30 is a 517-amino acid protein found in the outer mitochondrial membrane (Nakamura et al., 2008, Mol Biol 19:1903-11). It is the only deubiquitinase with a mitochondrial addressing signal and has been shown to deubiquitinate many mitochondrial proteins. It has been demonstrated that USP30 counteracts Parkin-mediated mitophagy and that reducing USP30 activity can rescue Parkin-mediated mitophagy defects (Bingol et al., 2015, Nature 510:370-5; Gersch et al., 2017, Nat Struct Mol Biol 24(11):920-930; Cunningham et al., 2015, Nat Cell Biol 17(2):160-169). Inactivation of USP30 can also increase mitochondrial protein import, likely through ubiquitination of TOM proteins (Jacoupy et al., 2019, Sci Rep 9(1):11829). A small fraction of USP30 is localized to peroxisomes, which are generated by the fusion of mitochondria and ER vesicles, and USP30 can antagonize the Pex2 / pexophagy pathway (Riccio et al., 2019, J Cell Biol 218(3): 798-807). The E3 Ub ligase March5 and the deubiquitinase USP30 associate with translocase and regulate mitochondrial import; while March5 blocks mitochondrial import and directs substrate degradation, USP30 deubiquitinates substrates and promotes their import (Phu et al., 2020, Molecular Cell 77, 1107-1123).

[0004] Mitochondrial dysfunction can be defined as a decrease in mitochondrial content (mitophagy or mitochondrial biogenesis), as a decrease in mitochondrial activity and oxidative phosphorylation, and as a regulation of reactive oxygen species (ROS) production, hence the role of mitochondrial dysfunction in numerous aging processes and disease states.

[0005] For example, Parkinson's disease, which affects approximately 10 million people worldwide (Parkinson's Disease Foundation), is characterized by the loss of dopaminergic neurons in the substantia nigra. While the exact mechanisms underlying PD are unknown, mitochondrial dysfunction is increasingly recognized as a key determinant of dopaminergic neuronal susceptibility in PD, characterizing both familial and sporadic forms of the disease, as well as toxin-induced parkinsonism. Parkin is one of many proteins implicated in early-onset PD. While most PD cases are associated with defects in α-synuclein, 10% of Parkinson's disease cases are associated with specific genetic defects, one of which is the ubiquitin E3 ligase parkin. Parkin and the protein kinase PTEN-induced putative kinase 1 (PINK1) cooperate to ubiquitinate mitochondrial membrane proteins in damaged mitochondria, triggering mitophagy. Dysregulation of mitophagy leads to increased oxidative stress, which has been described as a hallmark of PD. Therefore, inhibition of USP30 may be a potential strategy for the treatment of PD: for example, PD patients with parkin mutations that result in reduced activity could be therapeutically compensated for by inhibiting USP30.

[0006] Depletion of USP30 has been reported to enhance mitophagy clearance of mitochondria and promote Parkin-induced cell death. USP30 has also been shown to regulate BAX / BAK-dependent apoptosis, independent of Parkin overexpression. USP30 depletion sensitizes cancer cells to BH3 mimetics, such as ABT-737, without the need for Parkin overexpression. Therefore, the demonstrated anti-apoptotic role of USP30 makes it a potential target for anticancer therapy.

[0007] The ubiquitin-proteasome system has attracted interest as a target for cancer therapy following the approval of the proteasome inhibitor bortezomib (Velcade®) for the treatment of multiple myeloma. Long-term treatment with bortezomib is limited by associated toxicity and drug resistance. However, therapeutic strategies targeting specific aspects of the ubiquitin-proteasome pathway upstream of the proteasome, such as DUBs, are predicted to be more tolerable (Bedford et al., 2011, Nature Rev 10:29-46).

[0008] Fibrotic diseases, such as renal fibrosis, liver fibrosis, and pulmonary fibrosis, are major causes of morbidity and mortality and can affect all tissues and organ systems. Fibrosis is thought to be the result of acute or chronic stress on tissues or organs and is characterized by the deposition of extracellular matrix, decreased patency, and dysfunction of blood vessels, tubules, ducts, and airways, ultimately leading to organ failure. While many fibrotic conditions are precipitated by lifestyle or environmental factors, a proportion of fibrotic conditions are initiated by genetic triggers or are idiopathic (i.e., without a known cause) in nature. Certain fibrotic diseases, such as idiopathic pulmonary fibrosis (IPF), can be treated with nonspecific kinase inhibitors (nintedanib) or drugs without a well-characterized mechanism of action (pirfenidone). Other treatments for organ fibrosis, such as renal or liver fibrosis, reduce pressure on the organ itself (e.g., beta-blockers for cirrhosis and angiotensin receptor blockers for chronic kidney disease). Attention to lifestyle factors such as glucose and dietary management can also influence the course and severity of the disease.

[0009] Mitochondrial dysfunction is implicated in many fibrotic diseases, and oxidative stress downstream of dysfunction is a key pathogenic mediator, along with reduced ATP production. In preclinical models, disruption of the mitophagy pathway (by mutation or knockout of Parkin or PINK1) exacerbates pulmonary and renal fibrosis, with evidence of increased oxidative stress.

[0010] Kurita et al., 2017, Respiratory Research 18:114, discloses that the accumulation of profibrotic myofibroblasts is a key process in fibrotic remodeling in IPF. Recent findings have implicated autophagy / mitophagy, part of the lysosomal degradation mechanism, in the pathogenesis of IPF. Mitophagy is said to be involved in myofibroblast differentiation by regulating platelet-derived growth factor receptor (PDGFR) activation via mitochondrial reactive oxygen species (ROS). Kurita's results suggest that pirfenidone induces PARK2-mediated mitophagy and also inhibits the development of pulmonary fibrosis in an environment with insufficient mitophagy, which may at least partially explain the antifibrotic mechanism of IPF treatment.

[0011] Williams et al., 2015, Pharmacol Res. December; 102: 264-269, discussed the role of PINK1-Parkin-mediated autophagy in protecting against alcohol- and acetaminophen-induced liver injury by removing damaged mitochondria via mitophagy. It was suggested that pharmacological stabilization of USP8 or inactivation of USP15 and USP30 could be potential therapeutic targets for upregulating Parkin-induced mitophagy and protecting against drug-induced liver injury. However, it should be noted that DUBs are regulated both transcriptionally and post-translationally, potentially making drug development targeting these specific enzymes challenging. Furthermore, phosphorylated ubiquitin was shown to be resistant to DUBs. The authors concluded that stabilizing PINK1 or increasing its kinase activity may be more effective targets than inhibiting DUBs.

[0012] Williams et al, 2015, Biomolecules 5, 2619-2642 and Williams et al, 2015, Am J Physiol Gastrointest Liver Physiol 309: G324-G340 review the mechanisms involved in regulating mitochondrial homeostasis in the liver and how these mechanisms protect against alcohol-induced liver disease.

[0013] Luciani et al., 2020, Nat. Commun. 11, 970, reported that deregulation of the mitochondrial network in terminally differentiated cells contributes to a wide range of disorders, including methylmalonic acidemia (MMA). MMA is one of the most common inherited metabolic disorders caused by deficiency of mitochondrial methylmalonyl coenzyme A mutase (MMUT). MMUT deficiency induces metabolic and mitochondrial alterations, exacerbated by abnormalities in PINK1 / Parkin-mediated mitophagy, leading to the accumulation of dysfunctional mitochondria, which causes epithelial stress and ultimately cell damage. A link between primary MMUT deficiency, diseased mitochondria, mitophagy dysfunction, and epithelial stress has been suggested, providing insights into potential treatments for MMA.

[0014] Kluge et al., Bioorganic & Medicinal Chemistry Letters, 2018, 28 2655-2659, reported that a selective inhibitor of USP30 accelerates mitophagy.

[0015] A series of derivatives of N-cyano substituted heterocycles are disclosed in PCT applications WO2016 / 046530 (US15 / 513125, US15 / 894025, US16 / 448066), WO2016 / 156816 (US15 / 558632, US16 / 297937, US16 / 419558, US16 / 419747, US16 / 788446), WO2017 / 009650 (US15 / 7 38900), WO2017 / 093718(US15 / 776149), WO2017 / 103614(US15 / 781615), WO2017 / 149313(US16 / 07851 8), WO2017 / 109488(US16 / 060299), WO2017 / 141036(US16 / 070936), WO2017 / 163078(US16 / 087515), WO 2017 / 158381(US16 / 080229), WO2017 / 158388(US16 / 080506), WO2018 / 065768(US16 / 336685), WO2 018 / 060742(US16 / 336202), WO2018 / 060689(US16 / 334836), WO2018 / 060691(US16 / 336363), WO20 and PCT / EP2021 / 064166, each of which is expressly incorporated by reference herein. PCT application WO2019 / 171042 (US16 / 977019), which is expressly incorporated by reference herein, discloses the use of N-cyanopyrrolidines as inhibitors of USP30 for the treatment of fibrotic diseases.

[0016] Falgueyret et al., 2001, J. Med. Chem. 44, 94-104 and PCT application WO 01 / 77073 refer to cyanopyrrolidines as inhibitors of cathepsins K and L, potentially useful for the treatment of osteoporosis and other bone resorption-related conditions. PCT application WO 2015 / 179190 refers to N-acylethanolamine hydrolyzing acid amidase inhibitors, potentially useful for the treatment of ulcerative colitis and Crohn's disease. PCT application WO 2013 / 030218 refers to quinazolin-4-one compounds as inhibitors of ubiquitin-specific proteases (such as USP7), potentially useful for the treatment of cancer, neurodegenerative diseases, inflammatory diseases, and viral infections. PCT applications WO2015 / 017502 and WO2016 / 019237 refer to inhibitors of Bruton's tyrosine kinase, potentially useful in the treatment of diseases such as autoimmune diseases, inflammatory diseases, and cancer. PCT applications WO2009 / 026197, WO2009 / 129365, WO2009 / 129370, and WO2009 / 129371 refer to cyanopyrrolidines as inhibitors of cathepsin C, potentially useful in the treatment of COPD. U.S. patent application US2008 / 0300268 refers to polyaromatic compounds as inhibitors of the tyrosine kinase receptor PDGFR. PCT applications WO2019 / 222468, WO2019 / 071073, WO2020 / 036940, and WO2020 / 072964, Rusilowicz-Jones et al., 2020, bioRxiv 2020.04.16.044206 (20 April 2020), and Tsefou et al., bioRxiv 2021.02.02.429344 (2 February 2021) cite cyanamide-containing compounds as USP30 inhibitors. Yue et al., 2014, Cell Research, 24, 482-496 cite the diterpenoid derivative 15-oxopyramilactone as a USP30 inhibitor that induces mitochondrial fusion.

[0017] PCT application WO2015 / 183987 refers to a pharmaceutical composition comprising a deubiquitinase inhibitor and human serum albumin in a method for treating cancer, fibrosis, an autoimmune disease or condition, an inflammatory disease or condition, a neurodegenerative disease or condition, or an infectious disease. Deubiquitinases, including UCHL5 / UCH37, USP4, USP9X, USP11, and USP15, are said to be involved in regulating the TGF-β signaling pathway, the disruption of which leads to the development of neurodegenerative and fibrotic diseases, autoimmune disorders, and cancer.

[0018] PCT application WO2006 / 067165 refers to a method for treating fibrotic diseases using indolinone kinase inhibitors. PCT application WO2007 / 119214 refers to a method for treating early-stage pulmonary fibrosis using endothelin receptor antagonists. PCT application WO2012 / 170290 refers to a method for treating fibrotic diseases using THC acid. PCT application WO2018 / 213150 refers to sulfonamide USP30 inhibitors, potentially useful for treating conditions involving mitochondrial defects. Larson-Casey et al., 2016, Immunity 44, 582-596, relates to macrophage Akt1 kinase-mediated mitophagy, apoptosis resistance, and pulmonary fibrosis. Tang et al, 2015, Kidney Diseases 1, 71-79, review the potential role of mitophagy in renal pathophysiology.

[0019] There is a need for safe, alternative, and / or improved methods and compositions for the treatment or prevention of conditions involving mitochondrial dysfunction, cancer, and fibrosis, as well as various symptoms and conditions associated therewith. Without wishing to be bound by any particular theory or mechanism, it is believed that the compounds of the present invention act to inhibit the enzyme USP30, upregulating Parkin-induced mitophagy.

[0020] Acute kidney injury (AKI) is defined as a sudden decline in kidney function occurring within 7 days. The severity of injury is graded based on an increase in serum creatine (Scr) and a decrease in urine output, as described in the Kidney Disease Improving Global Outcomes (KDIGO) guidelines. AKI affects approximately 13.3 million people annually, 85% of whom live in developing countries, and is thought to result in approximately 1.7 million deaths annually (Mehta et al., 2015, Lancet 385(9987):2616-2643). AKI is likely to cause permanent kidney damage (i.e., chronic kidney disease; CKD) and may also cause damage to organs other than the kidney. AKI is a significant public health concern, especially considering the sheer number of patients who develop incident CKD, progressive CKD, end-stage renal disease, and cardiovascular events. AKI has been found to be prevalent in patients hospitalized with COVID-19 and strongly associated with in-hospital mortality, and mitochondrial damage and dysfunction have been reported as potential pathophysiological mechanisms and therapeutic targets (Kellum et al, Nephrol Dial Transplant (2020) 35: 1652-1662).

[0021] AKI and CKD are considered to be on the same disease spectrum continuum (Chawla et al, 2017, Nat Rev Nephrol 13(4): 241-257). Patients undergoing coronary artery bypass grafting (CABG) are at increased risk for kidney damage. There is a clear unmet medical need for the development of pharmaceuticals for the treatment and / or prevention of AKI.

[0022] The kidney is a site of high metabolic demand, and high mitophagy rates have been demonstrated in vivo (McWilliams et al., 2018, Cell Metab 27(2): 439-449 e435). Renal proximal tubule epithelial cells (RPTECs), a cell type that requires significant amounts of ATP for solute / ion exchange, are rich in mitochondria, and are key effector cells in acute kidney injury (AKI). Mitochondrial dysfunction has been implicated in AKI / CKD mechanisms through both multiple lines of evidence from preclinical AKI and CKD models and data demonstrating abnormal mitochondrial phenotypes in patient biopsies (Emma et al., 2016, Nat Rev Nephrol 12(5): 267-280; Eirin et al., 2017, Handb Exp Pharmacol 240: 229-250). Furthermore, primary mitochondrial diseases often manifest as renal manifestations, such as focal segmental glomerulosclerosis in patients with MELAS / MIDD (Kawakami et al, 2015, J Am Soc Nephrol 26(5): 1040-1052) and primary tubular lesions in patients with coenzyme Q deficiency. Mutations in mtDNA are maternal It can cause inherited tubulointerstitial disease (Connor et al, 2017, PLoS Genet 13(3): e1006620).

[0023] Regarding mitochondrial quality control in renal injury, a study (Tang et al., 2018, Autophagy 14(5): 880-897) showed that renal injury was exacerbated after ischemic AKI in both PINK1 and PARK2 KO mice, suggesting that PINK1 / Parkin-mediated mitophagy plays a protective role in the kidney following IRI. Furthermore, Parkin / PINK1 mitophagy protects against cisplatin-induced renal injury (Wang et al., 2018, Cell Death Dis 9(11): 1113). Limited CKD models are available for investigating mitophagy, and evidence supporting mitochondrial quality control in fibrosis comes from studies on fibrotic lung conditions such as COPD and IPF. Parkin knockout animals exhibit exacerbated pulmonary fibrosis in response to bleomycin (Kobayashi et al., 2016, J Immunol 197:504-516). Similarly, airway epithelial cells from parkin knockout (KO) animals exhibit exacerbated fibrotic and senescent responses to cigarette smoke (Araya et al., 2019, Autophagy 15(3): 510-526).

[0024] Preclinical models can be utilized to study potential new therapeutic approaches through their ability to model fibrosis pathology (e.g., collagen deposition) consistent with the human condition. Preclinical models can be toxin-mediated (e.g., bleomycin for lung and skin fibrosis), surgical (e.g., ischemia / reperfusion injury and unilateral ureteral obstruction models for acute tubulointerstitial fibrosis), and genetic (e.g., diabetic (db / db) mice for diabetic nephropathy) models. For example, both examples given previously for indicated IPF treatments (nintedanib and pirfenidone) have shown efficacy in the bleomycin pulmonary fibrosis model. Summary of the Invention

[0025] Thus, there is a need for compounds that are inhibitors of USP30 for the treatment or prevention of conditions in which inhibition of USP30 is indicated. In particular, there is a need for USP30 inhibitors with suitable and / or improved properties to maximize efficacy against target diseases.

[0026] Summary of the Invention The present invention relates to compounds of formula (I)(i) and formula (I)(ii):

[0027] [ka]

[0028] (In the above formula, R 1 is selected from (C1-C4) alkyl, (C1-C4) fluoroalkyl, and CH2OCH3; R 2 is selected from (C1-C4) alkyl, CF3, and cyclopropyl, and R 3 , R 4 and R 5 are each independently selected from hydrogen and halogen), a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer.

[0029] The present invention is also directed to the use of compounds of formula (I), particularly in the treatment of conditions involving mitochondrial dysfunction, cancer and fibrosis, as well as to methods for their preparation and pharmaceutical compositions containing said compounds. DETAILED DESCRIPTION OF THE INVENTION

[0030] Detailed Description of the Invention The present invention is directed to USP30 inhibitors with suitable and / or improved properties to maximize efficacy against target diseases, such as potency, selectivity, physicochemical properties, ADME (absorption, distribution, metabolism and excretion) properties including PK (pharmacokinetic) profiles, and safety profiles.

[0031] It is generally desirable to maximize the potency of a drug molecule against a target enzyme in a relevant assay in order to lower the effective / effective dose administered to a patient. Compounds of the present invention can be tested for USP30 affinity using the in vitro biochemical fluorescence polarization (FP) assay described herein.

[0032] USP30 is a transmembrane protein located in the outer membrane of mitochondria, an energy-generating organelle present in cells. Therefore, demonstrating cellular activity in vitro is advantageous, as it is one of many components that may indicate a high ability of USP30 inhibitor compounds to engage their targets in a physiological environment where they can penetrate cells. The USP30 cellular Western blot (WB) assay described herein aims to test the activity of compounds against USP30 in cells using an irreversible activity probe and monitor USP30 activity. Similar to the cellular Western blot assay, target binding assessment (ex vivo) can be performed on either brain or kidney tissue samples from animals administered the compound.

[0033] To extend knowledge of target binding to downstream pharmacodynamics, assessment of TOM20 (mitochondrial outer membrane protein) ubiquitination can be performed.

[0034] In general, it is important that drugs be as selective as possible for the desired target enzyme, as additional activity could result in potential side effects. The precise physiological roles of many DUBs have yet to be fully determined, but regardless of what role these DUBs may or may not play, ensuring that drugs are selective for relevant mechanistic targets of unknown physiological function is a guidepost of sound medicinal chemistry. Representative examples of DUB enzymes that can be screened for compounds of the invention include UCHL1, UCHL3, UCHL5, YOD1, SENP2, SENP6, TRABID, BAP1, Cezanne, MINDY2 / FAM63B, OTU1, OTUD3, OTUD5, OTUD6A, OTUD6B, OTUB1 / UBCH5B, OTUB2, CYLD, VCPIP, AMSH-LP, JOSD1, JOSD2, USP1 / UAF1, USP2, USP4, USP5, USP6, USP7, USP8, USP9x, USP10, USP11, USP12 / UAF1, USP13, USP14, USP15, USP 16, USP 19, USP 20, USP 21, USP 22, USP 24, USP 25, USP 28, USP 32, USP 34, USP 35, USP 36, USP 45, USP 46 / UAF1, USP 47 and USP 48. Preferably, the compounds of the invention have good selectivity for USP 30 over one or more of these DUB enzymes.

[0035] Aside from selectivity for other DUB enzymes, it is important that a drug has low affinity for other targets, and pharmacological profiling can be performed on a panel of targets to assess and minimize potential off-target effects. Examples of targets against which the compounds of the present invention can be screened include GPCR receptors, transporters, ion channels, nuclear receptors, and targets in the industry-standard Eurofins-Cerep SafetyScreen 44 panel, which includes 44 targets as a representative selection of kinase and non-kinase enzymes. Preferably, compounds of the present invention have insignificant affinity for the targets in this screening panel. Further examples of targets against which the compounds of the present invention can be screened include kinases in the Thermo Fisher SelectScreen Kinase Profiling Panel, which includes 39 targets as a representative selection of kinase enzymes. Preferably, compounds of the present invention have insignificant affinity for the targets in this screening panel. Furthermore, examples of specific enzyme classes against which the compounds of the present invention can be screened include cathepsins (e.g., cathepsins A, B, C, H, K, L, L2, S, V, and Z). Preferably, the compounds of the present invention have good selectivity for USP30 over one or more of these enzymes.

[0036] There is also a need for compounds with favorable pharmacokinetic properties to make them suitable for oral administration. Orally administered drugs should have good bioavailability, which is the ability to pass easily through the gastrointestinal (GI) tract and not undergo extensive metabolism as they pass from the GI tract into the systemic circulation. Once a drug enters the systemic circulation, the rate of metabolism is also important in determining the drug's residence time in the body.

[0037] Therefore, it is clearly desirable for a drug molecule to have the properties of easily passing through the GI tract and being metabolized only slowly in the body. The Caco-2 assay is a widely accepted model for predicting the ability of a given molecule to pass through the GI tract. The majority of the metabolism of a drug molecule generally occurs in the liver, and in vitro assays using whole-cell hepatocytes (animal or human) are widely accepted methods for measuring the susceptibility of a given molecule to hepatic metabolism. Such assays aim to predict in vivo clearance values ​​from clearance values ​​calculated in hepatocytes.

[0038] Compounds that have good Caco-2 flux and are stable to hepatocytes are predicted to have good oral bioavailability (good absorption across the GI tract and minimal extraction of the compound as it passes through the liver) and a sufficiently long residence time in the body for the drug to be effective.

[0039] Solubility of a compound is a key factor in achieving the desired concentration of a drug in the systemic circulation for the expected pharmacological response. Poor water solubility is a challenge faced in the formulation development of new chemical entities; for a drug to be absorbed, it must be present in solution at the absorption site. The dynamic solubility of a compound can be measured using a turbidimetric solubility assay, and this data can also be used in combination with Caco-2 permeability data to predict dose-dependent human intestinal absorption.

[0040] Other parameters that can be measured using standard assays that indicate the exposure profile of a compound include, for example, plasma stability (half-life measurements), AUC in blood, Cmax, Cmin and Tmax values.

[0041] Treatment of CNS disorders, including Alzheimer's disease, Parkinson's disease, and other disorders described herein, requires drug molecules to target the brain, which requires sufficient penetration of the blood-brain barrier. Therefore, there is a need for USP30 inhibitors that have effective blood-brain penetration properties and provide adequate residence time in the brain to be effective. The ability of a compound to cross the blood-brain barrier can be measured by an in vitro flux assay utilizing MDR1-MDCK cell monolayers (Madin-Darby Canine Kidney cells transfected with MDR-1, resulting in overexpression of the human efflux transporter P-glycoprotein). Additionally, exposure can also be measured directly in the brain and plasma using in vivo animal models.

[0042] There is also a need for compounds with favorable safety profiles that can be measured by a variety of standard in vitro and in vivo methods. A cytotoxicity counterscreen can be used to assay antiproliferative / cytotoxic effects in specific cell lines (e.g., HCT116) by fluorescent detection of resofurin (alamarBlue™) to resofurin in response to mitochondrial activity.

[0043] Toxicology and safety studies may also be performed to identify potential target organs for adverse reactions and to define a therapeutic index for setting an initial starting dose in clinical trials. Regulatory requirements generally require that studies be conducted in at least two experimental animals: rodent (rat or mouse) and non-rodent (rabbit, dog, non-human primate, or other appropriate species).

[0044] A bacterial reverse mutation assay (Ames test) can be used to assess the mutagenic properties of compounds of the invention, generally using the bacterial strain Salmonella typhimurium, which is mutant in the biosynthesis of the amino acid histidine.

[0045] The micronucleus assay can be used to determine whether a compound is genotoxic by assessing the presence of micronuclei, which can contain chromosome fragments generated by DNA breaks (clastogens) or whole chromosomes generated by disruption of the mitotic apparatus (aneuploidogens).

[0046] The hERG predictive assay provides valuable information regarding the potential of test compounds to bind to potassium channels and their potential to prolong the QT interval on echocardiograms. Inhibition of hERG currents causes QT interval prolongation and potentially fatal ventricular tachyarrhythmias (Torsades de Pointes). Typically, assay data can be generated from an automated patch clamp assay platform.

[0047] Thus, the present invention is directed to USP30 inhibitors that have suitable and / or improved properties to maximize efficacy against target diseases, such as potency, selectivity, physicochemical properties, ADME (absorption, distribution, metabolism and excretion) properties including PK (pharmacokinetic) profiles, and safety profiles.

[0048] The compounds of the present invention have been found to exhibit one or more of the above-identified properties that are important and unexpected. For example, Examples 1-12 of the present invention are highly potent against USP30 as measured by the biochemical assays described herein. All of these Examples (1-12) of the present invention are significantly more selective for USP30 than other DUBs and cathepsins. The important and unexpected properties of the compounds of the present invention make them particularly suitable for use in the treatment and / or prevention of diseases associated with USP30 activity.

[0049] According to a first aspect, the present invention provides a compound of formula (I)(i) and formula (I)(ii):

[0050] [ka]

[0051] (In the above formula, R 1 is selected from (C1-C4) alkyl, (C1-C4) fluoroalkyl, and CH2OCH3; R 2 is selected from (C1-C4) alkyl, CF3, and cyclopropyl, and R3, R4 and R5 are each independently selected from hydrogen and halogen), a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer.

[0052] Compounds of formula (I) exist as single stereoisomers with the absolute stereochemistry shown. Alkyl groups are straight-chain or branched and contain 1 to 4 carbon atoms. Examples of alkyl include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, and sec-butyl.

[0053] Halogen means fluorine, chlorine, bromine or iodine, especially fluorine or chlorine. Fluoroalkyl groups can contain one or more fluorine substituents. Examples are fluoromethyl, difluoromethyl and trifluoromethyl.

[0054] Unless otherwise indicated, the term "substituted" means substituted by one or more specified groups. In cases where groups can be selected from multiple options, the selected groups can be the same or different. The term "independently" means that when multiple substituents are selected from multiple possible substituents, the substituents can be the same or different.

[0055] Preferred embodiments of the compounds of formula (I) are defined below. Preferably, R 1 is selected from methyl, CH2F, CHF2, CF3 and CH2OCH3. More preferably, R 1 is selected from methyl, CH2F and CH2OCH3. Most preferably, R 1 is selected from methyl and CH2OCH3. Preferably, R 2 is selected from methyl, CF3 and cyclopropyl. Most preferably, R 2 is selected from methyl and cyclopropyl. Preferably, R 3 is selected from hydrogen, chlorine and fluorine. More preferably, R 3 is selected from hydrogen and fluorine. Most preferably, R 3 is hydrogen. Preferably, R 4 and R 5 are each independently selected from hydrogen and fluorine. Most preferably, R 4 and R 5 are hydrogen atoms.

[0056] According to one aspect of the present invention, the compound of formula (I) has the formula (I)(i):

[0057] [ka]

[0058] It has. According to another aspect of the present invention, the compound of formula (I) has the formula (I)(ii):

[0059] [ka]

[0060] It has. According to a first preferred embodiment of the present invention, a compound of formula (IA)(i):

[0061] [ka]

[0062] (In the above formula, R 1 is selected from (C1-C4) alkyl, (C1-C4) fluoroalkyl, and CH2OCH3; R 2 is selected from (C1-C4) alkyl, CF3, and cyclopropyl, and R 3 , R 4 and R 5 are each independently selected from hydrogen and halogen), a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer.

[0063] Preferred embodiments of compounds of formula (IA)(i) are defined below. Preferably, R 1 is selected from methyl, CH2F, CHF2, CF3 and CH2OCH3. More preferably, R 1 is selected from methyl, CH2F and CH2OCH3. Most preferably, R 1 is selected from methyl and CH2OCH3. Preferably, R 2 is selected from methyl, CF3 and cyclopropyl. Most preferably, R 2 is selected from methyl and cyclopropyl. Preferably, R 3 is selected from hydrogen, chlorine and fluorine. More preferably, R 3 is selected from hydrogen and fluorine. Most preferably, R 3 is hydrogen. Preferably, R 4 and R 5 are each independently selected from hydrogen and fluorine. Most preferably, R 4 and R 5 are hydrogen atoms.

[0064] Preferred compounds of formula (IA)(i) of the present invention are 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(fluoromethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, and 5-(5-cyano-4-fluoro-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, or a pharmaceutically acceptable salt thereof.

[0065] The most preferred compounds of formula (IA)(i) of the present invention are 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, and 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, or a pharmaceutically acceptable salt thereof.

[0066] According to a second preferred embodiment of the present invention, a compound of formula (IB)(i):

[0067] [ka]

[0068] (In the above formula, R 1 is selected from (C1-C4) alkyl, (C1-C4) fluoroalkyl, and CH2OCH3; R 2 is selected from (C1-C4) alkyl, CF3, and cyclopropyl, and R 3 , R 4 and R 5 are each independently selected from hydrogen and halogen), a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer.

[0069] Preferred embodiments of compounds of formula (IB)(i) are defined below. Preferably, R 1 is selected from methyl, CH2F, CHF2, CF3 and CH2OCH3. More preferably, R 1 is selected from methyl, CH2F and CH2OCH3. Most preferably, R 1 is CH2OCH3. Preferably, R 2 is selected from methyl, CF3 and cyclopropyl. Most preferably, R 2 is methyl. Preferably, R 3 is selected from hydrogen, chlorine and fluorine. More preferably, R 3is selected from hydrogen and fluorine. Most preferably, R 3 is hydrogen. Preferably, R 4 and R 5 are each independently selected from hydrogen and fluorine. Most preferably, R 4 and R 5 are hydrogen atoms.

[0070] Preferred compounds of formula (IB)(i) of the present invention are 4-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, and 4-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, or a pharmaceutically acceptable salt thereof.

[0071] According to a third preferred embodiment of the present invention, a compound of formula (IA)(ii):

[0072] [ka]

[0073] (In the above formula, R 1 is selected from (C1-C4) alkyl, (C1-C4) fluoroalkyl, and CH2OCH3; R 2 is selected from (C1-C4) alkyl, CF3, and cyclopropyl, and R 3 , R 4 and R 5 are each independently selected from hydrogen and halogen), a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer.

[0074] Preferred embodiments of compounds of formula (IA)(ii) are defined below. Preferably, R 1 is selected from methyl, CH2F, CHF2, CF3 and CH2OCH3. More preferably, R 1 is selected from methyl, CH2F and CH2OCH3. Most preferably, R 1 is selected from methyl and CH2OCH3. Preferably, R 2 is selected from methyl, CF3 and cyclopropyl. Most preferably, R 2 is selected from methyl and cyclopropyl. Preferably, R 3 is selected from hydrogen, chlorine and fluorine. More preferably, R 3 is selected from hydrogen and fluorine. Most preferably, R 3 is hydrogen. Preferably, R 4 and R 5 are each independently selected from hydrogen and fluorine. Most preferably, R 4 and R 5 are hydrogen atoms.

[0075] Preferred compounds of formula (IA)(ii) of the present invention are 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5S)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-4-fluoro-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, and 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-(fluoromethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, or a pharmaceutically acceptable salt thereof.

[0076] According to a fourth preferred embodiment of the present invention, a compound of formula (IB)(ii):

[0077] [ka]

[0078] (In the above formula, R 1 is selected from (C1-C4) alkyl, (C1-C4) fluoroalkyl, and CH2OCH3; R 2 is selected from (C1-C4) alkyl, CF3, and cyclopropyl, and R 3 , R 4 and R 5 are each independently selected from hydrogen and halogen), a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer.

[0079] Preferred embodiments of compounds of formula (IB)(ii) are defined below. Preferably, R 1 is selected from methyl, CH2F, CHF2, CF3 and CH2OCH3. More preferably, R 1 is selected from methyl, CH2F and CH2OCH3. Most preferably, R 1 is CH2OCH3. Preferably, R 2 is selected from methyl, CF3 and cyclopropyl. Most preferably, R 2 is methyl. Preferably, R 3 is selected from hydrogen, chlorine and fluorine. More preferably, R 3 is selected from hydrogen and fluorine. Most preferably, R 3 is hydrogen. Preferably, R 4 and R 5 are each independently selected from hydrogen and fluorine. Most preferably, R 4 and R 5 are hydrogen atoms.

[0080] Preferred compounds of formula (IB)(ii) of the present invention are 4-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, and 4-(5-cyano-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, or a pharmaceutically acceptable salt thereof.

[0081] Pharmaceutically acceptable salts of the compounds of formula (I) include the acid addition and base salts (including disalts) thereof. Suitable acid addition salts are formed from acids which form non-toxic salts, and examples include acetate, aspartate, benzoate, besylate, bicarbonate / carbonate, bisulfate, camsylate, citrate, edisylate, esylate, fumarate, gluceptate, gluconate, glucuronate, hybenate, hydrochloride / chloride, hydrobromide / bromide, hydroiodide / iodide, hydrogen phosphate, isethionate, D- and L-lactate, malate, maleate, malonate, mesylate, methylsulfate, 2-napsylate, nicotinate, nitrate, orotate, palmate, phosphate, saccharate, stearate, sulfate succinate, D- and L-tartrate, and tosylate salts. Suitable base salts are formed from bases which form non-toxic salts, examples of which include aluminum, ammonium, arginine, benzathine, calcium, choline, diethylamine, diolamine, glycine, lysine, magnesium, meglumine, olamine, potassium, sodium, tromethamine and zinc salts.

[0082] For a review of suitable salts, see Stahl and Wermuth, Handbook of Pharmaceutical Salts: Properties, Selection, and Use, Wiley-VCH, Weinheim, Germany (2002).

[0083] Pharmaceutically acceptable salts of compounds of formula (I) can be readily prepared by mixing together solutions of a compound of formula (I) and the desired acid or base, as appropriate. The salt may precipitate from solution and be collected by filtration or may be recovered by evaporation of the solvent.

[0084] Pharmaceutically acceptable solvates in accordance with the invention include hydrates and solvates wherein the solvent of crystallization may be isotopically substituted, for example, D2O, acetone-d6, DMSO-d6. Also within the scope of the present invention are clathrates, drug-host inclusion complexes, in which, in contrast to the solvates mentioned above, the drug and host are present in non-stoichiometric amounts. For a review of such complexes, see J. Pharm Sci, 64 (8), 1269-1288 by Haleblian (August, 1975). Hereinafter all references to compounds of formula (I) include references to salts thereof and to solvates and clathrates of compounds of formula (I) and salts thereof. The present invention includes all polymorphic forms of the compounds of formula (I) defined above.

[0085] Also within the scope of the present invention are so-called "prodrugs" of compounds of formula (I). Thus, certain derivatives of compounds of formula (I), which themselves have little or no pharmacological activity, can be metabolized in or upon administration to the body to yield compounds of formula (I) with the desired activity. Such derivatives are referred to as "prodrugs." Prodrugs according to the present invention can be produced, for example, by replacing appropriate functional groups present in compounds of formula (I) with certain moieties known in the art as "promoieties," as described, for example, in "Design of Prodrugs," H Bundgaard (Elsevier, 1985). Finally, certain compounds of formula (I) may themselves act as prodrugs of other compounds of formula (I). Certain derivatives of compounds of formula (I) containing nitrogen atoms can also form the corresponding N-oxides and such compounds are also within the scope of the invention. All tautomers of the compounds of formula (I) are included within the scope of the invention.

[0086] Conventional techniques for the preparation / isolation of individual enantiomers include chiral synthesis from suitable optically pure precursors or resolution of the racemate (or racemate of a salt or derivative) using, for example, chiral high-performance liquid chromatography (HPLC). Alternatively, the racemate (or racemic precursor) can be reacted with a suitable optically active compound, for example, an alcohol, or, if the compound of formula (I) contains an acidic or basic moiety, with a base or acid such as 1-phenylethylamine or tartaric acid. The resulting diastereomeric mixture can be separated by chromatography and / or fractional crystallization, and one or both of the diastereomers can be converted to the corresponding pure enantiomer by means well known to those skilled in the art. The chiral compounds of the present invention (and their chiral precursors) can be obtained in enantiomerically enriched form by chromatography, typically HPLC, on an asymmetric resin using a mobile phase consisting of a hydrocarbon, typically heptane or hexane, containing 0-50% by volume, typically 2-20% by volume, propan-2-ol, and 0-5% by volume of an alkylamine, typically 0.1% by volume, diethylamine. Concentration of the eluate yields the enriched mixture. The present invention encompasses all crystalline forms of the compounds of formula (I), including their racemates and racemic mixtures (conglomerates). Stereoisomeric conglomerates can be separated by conventional techniques known to those skilled in the art. See, for example, "Stereochemistry of Organic Compounds" by EL Eliel and SH Wilen (Wiley, New York, 1994). The compounds of formula (I) are R 1 and amide-substituted pyrrolidine ring carbon atoms, which may exist in either the (R) or (S) configuration. The designation of the absolute configuration (R) and (S) of stereoisomers according to IUPAC nomenclature depends on the nature of the substituents and the application of sequence rules. Thus, compounds of formula (I) can exist in four stereoisomeric forms. The compounds of formula (I) of the present invention exist as single stereoisomers. The pyrrolidine carbon atom of the amide substituent exists as the (R)-stereocenter, but the R 1 The designation of the pyrrolidine carbon atom of the group depends on the nature of the substituent. The compounds of formula (I) can be isolated as single stereoisomers and can exist in stereoisomeric excess of at least 60%, preferably at least 80%, more preferably at least 90%, more preferably at least 95%, for example 96%, 97%, 98%, 99% or 100%. Additional chiral centers can be found in compounds of formula (I) at R 1 may be present within the substituents themselves. All such stereoisomeric forms of the compounds of formula (I) are included within the scope of the present invention.

[0087] The present invention also includes all pharmaceutically acceptable isotopic variations of the compounds of formula (I). An isotopic variation is defined as one in which at least one atom is replaced with an atom having the same atomic number, but an atomic mass different from the atomic mass usually found in nature. Examples of isotopes suitable for inclusion in compounds of the invention include: 2 H and 3 Hydrogen such as H 13 C and 14 Carbon, such as C, 15 Nitrogen such as N 17 O and 18 Oxygen, such as O 32 Phosphorus such as P 35 Sulfur such as S 18 Fluorine such as F and 36 Examples include isotopes of chlorine such as Cl. Substitution of compounds of the invention with isotopes such as deuterium may afford certain therapeutic advantages resulting from greater metabolic stability, for example, increased in vivo half-life or reduced dosage requirements, and therefore may be preferable in some circumstances. Certain isotopic variations of the compounds of formula (I), for example, those incorporating a radioactive isotope, are useful in drug and / or substrate tissue distribution studies. The radioactive isotopes tritium and 14C is particularly useful for this purpose due to its ease of incorporation and ready means of detection. Isotopic variations of compounds of formula (I) may generally be prepared by conventional techniques known to those skilled in the art, or by methods analogous to those described in the accompanying Examples and Preparations, using appropriate isotopic variations of suitable reagents. The compounds of formula (I) are inhibitors of the deubiquitinating enzyme USP30. According to a further aspect, the present invention provides a compound of formula (I) as defined herein, a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer, for use as a medicament.

[0088] According to a further aspect, the present invention provides a method for the treatment or prevention of a disorder or condition in which inhibition of USP30 is known or can be shown to have a beneficial effect in a mammal, comprising administering to said mammal a therapeutically effective amount of a compound of formula (I) as defined herein, a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer. According to a further aspect, the present invention provides the use of a compound of formula (I) as defined herein, a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer, in the manufacture of a medicament for the treatment or prevention of a disorder or condition in which inhibition of USP30 is known or can be shown to have a beneficial effect. The manufacture of a medicament can include, inter alia, the chemical synthesis of a compound of formula (I) or a salt thereof, or the preparation of a composition or formulation comprising the compound or salt, or the packaging of any medicament comprising the compound. According to a further aspect, the present invention provides a method for the inhibition of USP30 in a patient comprising administering to the patient a therapeutically effective amount of a compound of formula (I) as defined herein, or a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer. The disorder or condition that would benefit from USP30 activity is selected from conditions involving mitochondrial dysfunction, cancer and fibrosis. In one preferred embodiment of all aspects of the invention, the disorder or condition that would benefit from USP30 activity is a condition involving mitochondrial dysfunction. Mitochondrial dysfunction is caused by defects in mitochondria, specialized compartments present in all cells of the body except red blood cells. When mitochondria stop functioning, less and less energy is produced within the cell, leading to cell damage or even cell death. When this process is repeated throughout the body, the life of the subject in question is seriously impaired. Mitochondrial diseases most frequently appear in organs with high energy demands, such as the brain, heart, liver, skeletal muscles, kidneys, endocrine system, and respiratory system.

[0089] The condition associated with mitochondrial dysfunction may be selected from a condition associated with a mitophagy defect, a condition associated with a mutation in mitochondrial DNA, a condition associated with mitochondrial oxidative stress, a condition associated with a defect in mitochondrial membrane potential, a condition associated with a defect in mitochondrial biogenesis, mitochondrial shape or morphology, and a condition associated with a lysosomal storage defect.

[0090] In particular, conditions involving mitochondrial dysfunction include neurodegenerative diseases, multiple sclerosis (MS), mitochondrial encephalopathy, lactic acidosis and stroke-like episodes (MELAS) syndrome, maternal Hereditary diabetes and deafness (MIDD), Leber's hereditary optic neuropathy (LHON), cancer (e.g., breast cancer, ovarian cancer, prostate cancer, lung cancer, kidney cancer, stomach cancer, colon cancer, testicular cancer, head and neck cancer, pancreatic cancer, brain cancer, melanoma cancer, bone cancer, other cancers of tissue organs, and cancers of blood cells such as lymphoma and leukemia, multiple myeloma, metastatic carcinoma, osteosarcoma, chondrosarcoma, Ewing's sarcoma, nasopharyngeal cancer, colorectal cancer, and non-small cell lung cancer), neuropathy, ataxia, retinitis pigmentosa, maternalHereditary Leigh syndrome (NARP-MILS), Danon disease, diabetes, diabetic nephropathy, metabolic disorders, heart failure, ischemic heart disease leading to myocardial infarction, psychiatric disorders such as schizophrenia, multiple sulfatase deficiency (MSD), mucolipidosis II (ML II), mucolipidosis III (ML III), mucolipidosis IV (ML IV), GM1-gangliosidosis (GM1), neuronal ceroid-lipofuscinosis (NCL1), Alper's disease, Barth syndrome, beta-oxidation defects, carnitine-acyl-carnitine deficiency, carnitine deficiency, creatine deficiency syndrome, coenzyme Q10 deficiency, complex I deficiency, complex II deficiency, complex III deficiency, complex IV deficiency, complex V deficiency, COX deficiency, chronic progressive external ophthalmoplegia syndrome (CPEO), CPT I deficiency, CPT II Deficiency, glutaric aciduria type II, Kearns-Sayre syndrome, lactic acidosis, long-chain acyl-CoA dehydrogenase deficiency (LCHAD), Leigh's disease or syndrome, Leigh's syndrome French Canadian (LSFC) variant, lethal infantile cardiomyopathy (LIC), Luft's disease, medium-chain acyl-CoA dehydrogenase deficiency (MCAD), myoclonic epilepsy and ragged-red fiber (MERRF) syndrome, mitochondrial cytopathies, mitochondrial recessive ataxia syndrome, mitochondrial DNA depletion syndrome The disorder may be selected from the group consisting of neuromuscular gastrointestinal disorders and encephalopathies, Pearson syndrome, pyruvate dehydrogenase deficiency, pyruvate carboxylase deficiency, POLG mutations, medium-chain / short-chain 3-hydroxyacyl-CoA dehydrogenase (M / SCHAD) deficiency, very long-chain acyl-CoA dehydrogenase (VLCAD) deficiency, peroxisomal disorders, methylmalonic acidemia, mevalonate kinase deficiency, age-related cognitive and muscle decline, cognitive impairment associated with neurodegenerative diseases and neuropsychiatric disorders.

[0091] The condition involving mitochondrial dysfunction can be a CNS disorder, such as a neurodegenerative disease.

[0092] Neurodegenerative diseases include, but are not limited to, Parkinson's disease, Alzheimer's disease, amyotrophic lateral sclerosis (ALS), Huntington's disease, ischemia, stroke, dementia with Lewy bodies, multiple system atrophy (MSA), progressive supranuclear palsy (PSP), corticobasal degeneration (CBD), and frontotemporal dementia. In particular, the compounds of the present invention may be useful for treating or preventing Parkinson's disease, including, but not limited to, PD associated with mutations in alpha-synuclein, parkin, PINK1, GBA, and LRRK2, as well as autosomal recessive juvenile Parkinson's disease (AR-JP) or early-onset Parkinson's disease (EOPD), in which parkin or PINK1 is mutated, truncated, or deleted.

[0093] In particular, the compounds of the invention may be useful in the treatment of cognitive impairment associated with neurodegenerative and neuropsychiatric disorders, including, for example, cognitive impairment associated with Alzheimer's disease and Parkinson's disease, preclinical or prodromal forms of AD and PD, Huntington's disease, dementia with Lewy bodies, cognitive impairment associated with schizophrenia, mood disorders, bipolar and major depressive disorder.

[0094] The compounds of the present invention or pharmaceutical compositions thereof as described herein can be combined with one or more additional drugs when used to treat or prevent conditions associated with mitochondrial dysfunction. The compounds can be combined with one or more additional drugs selected from levodopa, dopamine agonists, monoaminooxygenase (MAO) B inhibitors, catechol O-methyltransferase (COMT) inhibitors, anticholinergics, riluzole, amantadine, cholinesterase inhibitors, memantine, tetrabenazine, antipsychotics, diazepam, clonazepam, antidepressants, and anticonvulsants. The compounds can be combined with drugs that reduce or remove pathogenic protein aggregates in neurodegenerative diseases, such as drugs that reduce or remove alpha-synuclein in Parkinson's disease, multiple system atrophy, or dementia with Lewy bodies, drugs that reduce or remove tau in Alzheimer's disease or progressive supranuclear palsy, and drugs that reduce or remove TDP-43 in ALS or frontotemporal dementia.

[0095] In another preferred embodiment of all aspects of the invention, the disorder or condition that would benefit from USP30 activity is cancer. The cancer can be associated with mitochondrial dysfunction. Preferred cancers include, for example, breast cancer, ovarian cancer, prostate cancer, lung cancer, kidney cancer, stomach cancer, colon cancer, testicular cancer, head and neck cancer, pancreatic cancer, brain cancer, melanoma cancer, bone cancer, or other cancers of tissue organs, and cancers of blood cells, such as lymphoma and leukemia, multiple myeloma, metastatic carcinoma, osteosarcoma, chondrosarcoma, Ewing's sarcoma, nasopharyngeal carcinoma, colorectal cancer, and non-small cell lung cancer.

[0096] In particular, the compounds of the present invention may be useful in the treatment or prevention of cancers in which the apoptotic pathway is dysregulated, more particularly cancers in which proteins of the BCL-2 family are mutated or over- or under-expressed.

[0097] Fibrosis refers to the accumulation of extracellular matrix components that occurs after trauma, inflammation, tissue repair, immune response, cellular hyperplasia, and tumor formation. Fibrotic disorders that can be treated by the compounds and compositions of the present invention include, among others, fibrosis / fibrotic diseases associated with major organ diseases, such as interstitial lung disease (ILD), liver cirrhosis, non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH) (liver fibrosis), kidney disease (renal fibrosis), acute kidney injury (AKI), acute kidney injury (AKD), chronic kidney disease (CKD), delayed kidney graft function, heart or vascular disease (cardiac fibrosis), and eye diseases, fibrosis, and fibrosis. These include proliferative diseases such as systemic and localized scleroderma, keloids and hypertrophic scars, atherosclerosis, restenosis and Dupuytren's contracture, trauma-related scars such as surgical complications, chemotherapy-induced fibrosis (e.g., bleomycin-induced fibrosis), radiation-induced fibrosis, accidental injury and burns, retroperitoneal fibrosis (Ormond's disease), and peritoneal fibrosis / scarring in patients undergoing peritoneal dialysis, commonly occurring after renal transplantation. See, e.g., Wynn et al., 2004, Nat Rev Immunol. August; 4(8): 583-594. Accordingly, the present invention relates to methods for treating or preventing fibrosis / fibrotic disorders of and / or associated with major organs, including, for example, the lung, liver, kidney, heart, skin, eye, gastrointestinal tract, peritoneum, and bone marrow, as well as compounds and compositions for use in such methods.

[0098] The compounds may be combined with drugs used to treat kidney disease, including antidiabetic drugs, cardiovascular disease drugs, and novel drugs that target disease-related pathways such as oxidative stress (including but not limited to the nrf2 / keap-1 pathway) and anti-apoptotic pathways (including but not limited to anti-p53 agents). Interstitial lung diseases (ILDs) include diseases in which pulmonary inflammation and fibrosis are the final common pathway of pathology, such as sarcoidosis, silicosis, drug reactions, infections, and collagen vascular diseases such as rheumatoid arthritis and systemic sclerosis (scleroderma). Fibrotic disorders of the lung include, for example, pulmonary fibrosis, idiopathic pulmonary fibrosis (IPF), common interstitial pneumonia (UIP), interstitial lung disease, idiopathic fibrosing alveolitis (CFA), bronchiolitis obliterans, and bronchiectasis. Idiopathic pulmonary fibrosis (IPF) is the most common type of ILD and its cause is unknown.

[0099] The compounds can be combined with drugs that are treatments for IPF and potentially ILD, including nintedanib and pirfenidone. Cirrhosis of the liver has similar causes to ILD, including cirrhosis associated with viral hepatitis, schistosomiasis, and chronic alcoholism. Kidney disease can be associated with diabetes, which damages the kidneys, causing scarring and leading to progressive loss of function, and also hypertensive disorders. Renal fibrosis can occur at any stage of kidney disease, from acute kidney disease (AKD) and chronic kidney disease (CKD) (such as incident CKD and progressive CKD) following injury to end-stage renal disease (ESRD). Renal fibrosis can also develop as a result of cardiovascular diseases such as hypertension or diabetes, both of which place a significant strain on kidney function, promoting a fibrotic response. However, renal fibrosis can also be idiopathic (without a known cause), and certain inherited mitochondrial diseases also exhibit signs and associated symptoms of renal fibrosis. Heart disease can cause scar tissue that can impair the heart's pumping function.

[0100] Eye diseases include, for example, macular degeneration and retinal and vitreoretinopathy, which can impair vision. In a preferred embodiment, the present invention is directed to the treatment or prevention of idiopathic pulmonary fibrosis (IPF). In another preferred embodiment, the present invention is directed to the treatment or prevention of renal fibrosis.

[0101] In another preferred embodiment, the present invention is directed to the treatment or prevention of acute kidney injury (AKI), particularly in high-risk patients. Examples include postoperative AKI, such as in organ transplantation due to ischemia-reperfusion injury and delayed graft function; oncological conditions, such as chemotherapy-induced AKI; contrast-induced nephropathy, such as direct tubular cytotoxicity, hemodynamic ischemia, and osmotic effects; acute interstitial nephritis, such as from drugs or infections; AKI due to obstruction, such as kidney stones; and COVID-19-induced AKI. Particular high-risk patient subgroups are those undergoing cardiac surgery, e.g., coronary artery bypass graft and / or valve surgery. Patients aged 65 years or older, those with insulin-dependent diabetes, and those with CKD (estimated glomerular filtration rate [eGFR] > 60 ml / min / 1.73 m or greater) are at high risk. 2 There are established static risk factors for AKI, including diabetes (adults under 6 years of age are particularly at risk), heart failure, liver disease, and a history of AKI.

[0102] In another preferred embodiment, the present invention is directed to the treatment or prevention of acute kidney disease (AKD) or chronic kidney disease (CKD) resulting from such AKI, including, for example, tubulointerstitial fibrosis and diabetic nephropathy.

[0103] In another preferred embodiment, the present invention is directed to the treatment or prevention of liver diseases, including, for example, NAFLD, NASH, cirrhosis, portal hypertension, acute liver failure, and hepatocellular carcinoma. Liver diseases such as NAFLD and NASH can be associated with various metabolic conditions, such as metabolic syndrome and type II diabetes, and also increase the risk of various diabetes-related conditions, including diabetic retinopathy and peripheral neuropathy.

[0104] The compounds of the invention or pharmaceutical compositions thereof as described herein may be combined with one or more additional agents when used for the treatment or prevention of conditions involving liver disease and metabolic dysfunction, including metformin, sulfonylureas, DPP-4 inhibitors, GLP-1 agonists, PPAR agonists, SGLT2 inhibitors, angiotensin-converting enzyme (ACE) inhibitors and angiotensin II receptor blockers (ARBs).

[0105] Leigh syndrome is a rare inherited neurometabolic disorder affecting the central nervous system. This progressive disorder begins in infants between the ages of 3 months and 2 years. Rarely, it occurs in teenagers and adults. Leigh syndrome can be caused by mutations in nuclear DNA that encodes mitochondrial proteins, mutations in mitochondrial DNA (maternally inherited Leigh syndrome - MILS), or a deficiency of an enzyme called pyruvate dehydrogenase, located on the short arm of the X chromosome (X-linked Leigh syndrome). Symptoms of Leigh syndrome usually progress rapidly. Early signs may be a decrease in sucking ability, loss of head control, and motor skills. These symptoms may be accompanied by loss of appetite, vomiting, irritability, constant crying, and seizures. As the disorder progresses, symptoms may include generalized weakness, lack of muscle tone, and episodes of lactic acidosis, which can lead to impaired respiratory and renal function.

[0106] In maternally inherited Leigh syndrome (MILS), genetic mutations in mitochondrial DNA (a high prevalence, exceeding 90%) disrupt the energy source that powers cells in the brain region responsible for movement. These mutations cause chronic energy deficiency in these cells, affecting the central nervous system and resulting in progressive degeneration of motor function. When the mitochondrial DNA mutations responsible for MILS are rare (less than 90%), the condition is known as neuropathic ataxia and retinitis pigmentosa (NARP). There is also Leigh disease (also known as X-linked Leigh disease), which is the result of mutations in genes that produce another group of substances important for cellular metabolism. Another variant of Leigh syndrome, called the French Canadian variant, exists, characterized by mutations in a gene called LRPPRC. Similar neurological symptoms are present as in Leigh syndrome, but fatty liver is commonly observed in the French Canadian variant.

[0107] In preferred embodiments, the present invention is directed to the treatment or prevention of Leigh syndrome or disease, including, for example, X-linked Leigh disease, Leigh syndrome French Canadian variant, and / or symptoms associated with Leigh disease.

[0108] The compounds may be combined with novel agents that can be used as treatments for mitochondrial diseases, including but not limited to nicotinamide riboside.

[0109] Reference to "treatment" includes measures that temporarily or permanently improve, alleviate, or eliminate the cause of the symptoms. The compounds of the present invention are useful in treating the diseases disclosed herein in humans and other mammals.

[0110] In another embodiment, the present invention encompasses prophylactic therapy of the diseases disclosed herein, including means for preventing or delaying the onset of symptoms of the named disorder or condition. The compounds of the present invention are useful for the prevention of the diseases disclosed herein in humans and other mammals.

[0111] A patient in need of treatment or prevention can be, for example, a human or other mammal suffering from or at risk of suffering from the condition.

[0112] According to a further aspect, the present invention provides a pharmaceutical composition comprising a compound of formula (I) as defined herein, or a pharmaceutically acceptable salt of said compound or tautomer, together with a pharmaceutically acceptable diluent or carrier.

[0113] The pharmaceutical compositions of the present invention include any compound of the present invention combined with any pharmaceutically acceptable carrier, adjuvant, or vehicle. Examples of pharmaceutically acceptable carriers are known to those skilled in the art and include, but are not limited to, preservatives, fillers, disintegrants, wetting agents, emulsifiers, suspending agents, sweeteners, flavorings, fragrances, antibacterial agents, antifungal agents, lubricants, and dispersing agents, depending on the mode of administration and the nature of the dosage form. The compositions can be in the form of, for example, tablets, capsules, powders, granules, elixirs, lozenges, suppositories, syrups, and liquid preparations, including suspensions and solutions. The term "pharmaceutical composition" in the context of the present invention means a composition comprising an active agent and further comprising one or more pharmaceutically acceptable carriers. The composition may further comprise components selected from, for example, diluents, adjuvants, excipients, vehicles, preservatives, fillers, disintegrating agents, wetting agents, emulsifying agents, suspending agents, sweetening agents, flavoring agents, perfuming agents, antibacterial agents, antifungal agents, lubricants and dispersing agents, depending on the mode of administration and the nature of the dosage form.

[0114] The compounds of the present invention or pharmaceutical compositions thereof as described herein can be used alone or in combination with one or more additional pharmaceutical agents. The compounds can be combined with additional anti-tumor therapeutic agents, such as chemotherapeutic agents or inhibitors of other regulatory proteins. In one embodiment, the additional anti-tumor therapeutic agent is a BH-3 mimetic. In a further embodiment, the BH-3 mimetic can be selected from, but is not limited to, one or more of ABT-737, ABT-199, ABT-263, and Obatoclax. In a further embodiment, the additional anti-tumor agent is a chemotherapeutic agent. The chemotherapeutic agent can be selected from, but is not limited to, olaparib, mitomycin C, cisplatin, carboplatin, oxaliplatin, ionizing radiation (IR), camptothecin, irinotecan, topotecan, temozolomide, taxanes, 5-fluoropyrimidines, gemcitabine, and doxorubicin.

[0115] For the treatment or prevention of fibrotic disorders, for example, the compounds of the present invention or pharmaceutical compositions thereof as described herein may be used alone or in combination with one or more additional pharmaceutical agents selected from the group consisting of anticholinergics, beta-2 mimetics, steroids, PDE-IV inhibitors, p38 MAP kinase inhibitors, NK1 antagonists, LTD4 antagonists, EGFR inhibitors, and endothelin antagonists.

[0116] In particular, the compounds of the present invention or pharmaceutical compositions thereof as described herein can be used alone or in combination with one or more additional pharmaceutical agents selected from the group consisting of general immunosuppressants such as corticosteroids, immunosuppressants or cytotoxic agents, or antifibrotic agents such as pirfenidone or nonspecific kinase inhibitors (e.g., nintedanib).

[0117] The pharmaceutical compositions of the present invention can be administered in any suitably effective manner, including orally, parenterally, topically, by inhalation, intranasally, rectally, intravaginally, ophthalmically, and otically. Pharmaceutical compositions suitable for delivery of the compounds of the present invention and methods for their preparation will be readily apparent to those skilled in the art. Such compositions and methods for their preparation are described, for example, in "Remington's Pharmaceutical Sciences," 1999. th Edition (Mack Publishing Company, 1995).

[0118] Oral administration The compounds of the present invention can be administered orally, which may involve swallowing, so that the compound enters the gastrointestinal tract, or buccal or sublingual administration may be employed by which the compound enters the blood stream directly from the mouth.

[0119] Formulations suitable for oral administration include solid formulations such as tablets, microparticles, liquid or powder containing capsules, lozenges (including liquid fills), chews, multiparticulates and nanoparticles, gels, films (including mucoadhesives), ovules, sprays and liquid formulations.

[0120] Liquid preparations include suspension, solution, syrup and elixir.Such preparations can be used as filler in soft or hard capsule, and typically comprise carrier such as water, ethanol, propylene glycol, methylcellulose or suitable oil, and one or more emulsifiers and / or suspending agents.Liquid preparations can also be prepared by reconstituting solid from, for example, sachet.

[0121] The compounds of the invention may also be used in fast dissolving, fast disintegrating dosage forms such as those described in Expert Opinion in Therapeutic Patents, 11 (6), 981-986 by Liang and Chen (2001).

[0122] A typical tablet can be prepared using standard methods known to formulation chemists, for example, by direct compression, granulation (dry, wet, or melt), melt congealing, or extrusion. Tablet formulations can comprise one or more layers and can be coated or uncoated.

[0123] Examples of excipients suitable for oral administration include carriers such as cellulose, calcium carbonate, dibasic calcium phosphate, mannitol, and sodium citrate, granulating binders such as polyvinylpyrrolidone, hydroxypropyl cellulose, hydroxypropylmethylcellulose, and gelatin, disintegrating agents such as starch sodium glycolate and silicate, lubricants such as magnesium stearate and stearic acid, wetting agents such as sodium lauryl sulfate, preservatives, antioxidants, flavorings, and coloring agents.

[0124] Solid dosage forms for oral administration can be formulated for immediate and / or modified release. Modified release formulations include delayed-release, sustained-release, pulsed-release, controlled dual-release, targeted-release, and programmed-release. Details of suitable modified release technologies, such as high-energy dispersions, osmotic, and coated particles, can be found in Verma et al., Pharmaceutical Technology Online, 25 (2), 1-14 (2001). Other modified release formulations are described in U.S. Pat. No. 6,106,864.

[0125] Parenteral administration The compound of the present invention can also be administered directly into the bloodstream, muscle or internal organs.Suitable means for parenteral administration include intravenous, intraarterial, intraperitoneal, intrathecal, intraventricular, intraurethral, ​​intrasternal, intracranial, intramuscular and subcutaneous.Suitable devices for parenteral administration include needle (including microneedle) injectors, needle-free injectors and infusion techniques.

[0126] Parenteral formulations are typically aqueous solutions (preferably pH 3-9) which may contain excipients such as salts, carbohydrates, buffers, etc., although for some applications they may be more suitably formulated as sterile non-aqueous solutions or as a dry form to be used in combination with a suitable vehicle such as sterile, pyrogen-free water.

[0127] The preparation of parenteral formulations under sterile conditions, for example, by lyophilization, may readily be accomplished using standard pharmaceutical techniques well-known to those skilled in the art.

[0128] The solubility of compounds of formula (I) used in the preparation of parenteral solutions may be increased by the use of appropriate formulation techniques, such as appropriate processing, e.g. the use of high-energy spray-dried dispersions and / or the use of solubility enhancers.

[0129] Formulations for parenteral administration may be formulated to be immediate and / or modified release, including delayed-, sustained-, pulsed-, controlled dual-, targeted-, and programmed-release formulations.

[0130] The pharmaceutical compositions of the present invention also include compositions and methods known in the art for bypassing the blood-brain barrier, or can be injected directly into the brain. Suitable areas for injection include the cerebral cortex, cerebellum, midbrain, brainstem, hypothalamus, spinal cord, and ventricular tissue, as well as areas of the PNS, including the carotid bodies and adrenal medulla.

[0131] dose Of course, the magnitude of the effective dose of a compound will vary depending on the nature of the severity of the condition being treated and the route of administration. The selection of an appropriate dose is within the competence of a physician. The daily dose range for humans and non-human animals is about 10 μg to about 100 mg per kg of body weight, and generally about 10 μg to 30 mg per kg of body weight per dose. The above doses can be administered one to three times a day.

[0132] For example, oral administration may require a total daily dose of 5 mg to 1000 mg, such as 5 mg to 500 mg, while an intravenous dose may require only 0.01 to 30 mg / kg body weight, such as 0.1 to 10 mg / kg, more preferably 0.1 to 1 mg / kg body weight. The total daily dose may be administered in single or divided doses. Those skilled in the art will also appreciate that in treating or preventing certain conditions, the compounds of the invention can be taken as a single dose "on demand" (i.e., as needed or desired).

[0133] Synthesis method Compounds of formula (I) may be prepared using the methods as described below in the general reaction schemes and representative examples. If desired, the individual transformations within the schemes may be completed in a different order. The invention is illustrated by the following non-limiting examples in which the following abbreviations and definitions are used. Compounds may be analyzed by liquid chromatography mass spectrometry (LCMS) or 1 Characterized by 1 H NMR or both.

[0134] In a further aspect, the present invention provides a method for preparing a compound of formula (I)(i), comprising reacting a compound of formula (IV) (wherein Y is OH) with an amine of formula (V)(i) (wherein PG is a protecting group such as BOC or CBZ) to provide an amide of formula (III)(i) (Scheme 1). The amide coupling reaction can be carried out using standard methods, for example, by reaction using coupling reagents such as DCC, HATU, HBTU, EDC, or via a mixed anhydride. Alternatively, acid (IV) (wherein Y is OH) can be converted to acid chloride (IV) (wherein Y is Cl) using SOCl, PCl, or PCl, which can then be reacted with amine (V)(i), preferably in a suitable solvent in the presence of a suitable base. Alternatively, compound (IV) (wherein Y forms an ester) can be reacted directly with amine (V)(i), preferably in a suitable solvent. Compounds of formula (III)(i) can be deprotected using standard methods to give amines (II)(i), which can then be reacted with cyanogen bromide to provide the corresponding compounds of formula (I)(i).

[0135] [ka]

[0136] In a further aspect, the present invention provides a compound selected from formula (II)(i) and (III)(i).

[0137] [ka]

[0138] where PG is a protecting group, preferably BOC or CBZ, and R 1 , R 2 , R 3 , R 4 and R 5 is as defined herein for compounds of formula (I) and preferred embodiments thereof, tautomers thereof, or salts of said compounds or tautomers).

[0139] When the compound of formula (I)(i) has formula (IA)(i), formulas (II)(i), (III)(i) and (IV) are (IIA)(i), (IIIA)(i) and (IVA), respectively. When the compound of formula (I)(i) has formula (IB)(i), formulas (II)(i), (III)(i) and (IV) are (IIB)(i), (IIIB)(i) and (IVB)(i), respectively.

[0140] In a further aspect, the present invention provides a method for preparing a compound of formula (I)(ii), comprising reacting a compound of formula (IV) (wherein Y is OH) with an amine of formula (V)(ii) (wherein PG is a protecting group such as BOC or CBZ) to provide an amide of formula (III)(ii) (Scheme 2). The amide coupling reaction can be carried out using standard methods, for example, by reaction using coupling reagents such as DCC, HATU, HBTU, EDC, or via a mixed anhydride. Alternatively, acid (IV) (wherein Y is OH) can be converted to acid chloride (IV) (wherein Y is Cl) using SOCl, PCl, or PCl, which can then be reacted with amine (V)(ii), preferably in a suitable solvent in the presence of a suitable base. Alternatively, compound (IV) (wherein Y forms an ester) can be reacted directly with amine (V)(ii), preferably in a suitable solvent. The compound of formula (III)(ii) can be deprotected using standard methods to provide the amine (II)(ii), which can then be reacted with cyanogen bromide to provide the corresponding compound of formula (I)(ii).

[0141] [ka]

[0142] In a further aspect, the present invention provides a compound selected from formula (II)(ii) and (III)(ii).

[0143] [ka]

[0144] where PG is a protecting group, preferably BOC or CBZ, and R 1 , R 2 , R 3 , R 4 and R 5 is as defined herein for compounds of formula (I) and preferred embodiments thereof, tautomers thereof, or salts of said compounds or tautomers).

[0145] When the compound of formula (I)(ii) has formula (IA)(ii), formulas (II)(ii), (III)(ii) and (IV) are (IIA)(ii), (IIIA)(ii) and (IVA), respectively. When the compound of formula (I)(ii) has formula (IB)(ii), formulas (II)(ii), (III)(ii) and (IV) are (IIB)(ii), (IIIB)(ii) and (IVB)(ii), respectively.

[0146] The protecting groups are preferably tert-butyloxycarbonyl (BOC), benzyloxycarbonyl (Cbz), p-methoxybenzylcarbonyl (MeOZ), 9-fluorenylmethyloxycarbonyl (Fmoc), acetyl (Ac), benzoyl (Bz), benzyl (Bn), carbamate, p-methoxybenzyl (PMB), 3,4-dimethoxybenzyl (DMPM), p-methoxyphenyl (PMP), tosyl (Ts), trichloroethoxycarbonyl (Troc), 4-nitrobenzenesulfonyl (Nosyl), and 2-nitrophenylsulfenyl (Nps). BOC and Cbz are most preferred.

[0147] JPEG0007796047000013.jpg99164

[0148] [Table 1]

[0149] Table 2

[0150] Table 3

[0151] Table 4

[0152] Table 5

[0153] Table 6

[0154] Table 7

[0155] Table 8

[0156] Table 9

[0157] Table 10

[0158] Table 11

[0159] Table 12

[0160] [Table 13]

[0161] [Table 14]

[0162] [Table 15]

[0163] [Table 16]

[0164] [Table 17]

[0165] Intermediate A Ethyl 5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxylate

[0166] [ka]

[0167] (i) NaH, THF, 0 °C, 2 h; (ii) phenyltrimethylammonium tribromide, THF, 0 °C to rt, 16 h; (iii) NaN(CHO)2, MeCN, 70 °C, 1 h, then concentrated HCl, 70 °C, 16 h; (iv) K2CO3, DCM, 0 °C to rt, 2 h; (v) POCl3, 100 °C, 7 h

[0168] Method (i) 3-Acetyl-4-cyclopropoxybenzonitrile This reaction was carried out three times. To a stirred solution of cyclopropanol (CAS 16545-68-9, from Synthonix, 0.71 g, 12.27 mmol) in THF (6 mL) was added NaH (60% in oil, 0.49 g, 12.27 mmol) portionwise at 0 °C and stirred for 30 min. A solution of 3-acetyl-4-fluorobenzonitrile (CAS 267875-54-7, from Combi-blocks, 1.0 g, 6.13 mmol) in THF (4 mL) was added dropwise at 0 °C, and the mixture was stirred at 0 °C for 2 h. The reaction mixture was combined with two more identical batches, then poured into ice-cold water (500 mL), and extracted with EtOAc (2 × 500 mL). The combined organic phase was dried over Na SO and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 12% EtOAc in n-hexane) to give 3-acetyl-4-cyclopropoxybenzonitrile (3.18 g, 15.82 mmol, 85% yield). LCMS: m / z not supported; 1 H NMR (400 MHz, DMSO-d6) δ ppm: 8.05 (dd, J = 8.8, 2.4 Hz, 1H), 7.96 (d, J = 2.0 Hz, 1H), 7.64 (d, J = 8.8 Hz, 1H), 4.13 - 4.17 (m, 1H), 2.50 (s, 3H), 0.81 - 0.93 (m, 4H).

[0169] Project (ii) 3-(2-Bromoacetyl)-4-cyclopropoxybenzonitrile To a stirred solution of 3-acetyl-4-cyclopropoxybenzonitrile (3.18 g, 15.82 mmol) in THF (40 mL) was added phenyltrimethylammonium tribromide (5.94 g, 15.82 mmol) at 0 °C. The mixture was slowly warmed to room temperature and stirred for 16 h, then poured into water (200 mL) and extracted with EtOAc (2 × 200 mL). The combined organic phases were dried over NaSO and concentrated under reduced pressure to provide 3-(2-bromoacetyl)-4-cyclopropoxybenzonitrile (4.3 g, 15.41 mmol, 97% yield). This crude material was used directly in the next step. LCMS: Method F, 6.61 min; MS: ES+: 297.0, 299.0 (M+18).

[0170] Project (iii) 4-Cyclopropoxy-3-glycylbenzonitrile hydrochloride To a stirred solution of 3-(2-bromoacetyl)-4-cyclopropoxybenzonitrile (4.30 g, 15.41 mmol) in acetonitrile (43 mL) was added sodium diformylamide (1.75 g, 18.49 mmol) and heated at 70° C. for 3 hours. The reaction mixture was cooled to room temperature and concentrated under reduced pressure. The residue was diluted with MeOH (43 mL) and concentrated HCl (4.3 mL). The mixture was further heated at 70° C. for 16 hours and then allowed to cool to room temperature. The mixture was concentrated under reduced pressure, and the residue was stirred with isopropyl alcohol (25 mL) to form a precipitate. The solid was collected by filtration under reduced pressure to provide 4-cyclopropoxy-3-glycylbenzonitrile hydrochloride (5.0 g, quantitative yield). LCMS: Method C, 1.30 min; MS: ES+: 217.4.

[0171] Engineering (iv) Ethyl 2-((2-(5-cyano-2-cyclopropoxyphenyl)-2-oxoethyl)amino)-2-oxoacetate To a stirred solution of 4-cyclopropoxy-3-glycylbenzonitrile HCl salt (5.0 g, 19.80 mmol) in DCM (50 mL) was added KCO (10.92 g, 79.2 mmol) at 0 °C. Ethyl oxalyl chloride (5.4 g, 4.43 mL, 39.60 mmol) was added dropwise at 0 °C. The mixture was allowed to cool to room temperature and stirred for 2 h, then poured into water (500 mL) and extracted with DCM (2 × 300 mL). The combined organic phases were dried over NaSO and concentrated under reduced pressure to provide ethyl 2-((2-(5-cyano-2-cyclopropoxyphenyl)-2-oxoethyl)amino)-2-oxoacetate (5.0 g, quantitative yield). LCMS: Method C, 1.52 min; MS: ES+ 316.9.

[0172] Engineering (v) Ethyl 5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxylate A stirred solution of ethyl 2-((2-(5-cyano-2-cyclopropoxyphenyl)-2-oxoethyl)amino)-2-oxoacetate (5.0 g, 15.81 mmol) in POCl (50 mL, 10 vol) was heated at 100° C. for 7 h. The mixture was cooled to room temperature, poured into ice-cold water (500 mL), and extracted with EtOAc (2×200 mL). The combined organic phases were dried over NaSO and concentrated under reduced pressure. The residue was suspended in MeOH (40 mL) and stirred at −78° C. for 15 min. The solid was collected by filtration under reduced pressure to provide ethyl 5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxylate (1.0 g, 3.35 mmol, 21% yield over three steps). LCMS: Method C, 1.77 min; MS: ES+: 299.5.

[0173] Intermediate B Ethyl 5-(5-cyano-2-methoxyphenyl)oxazole-2-carboxylate [ka]

[0174] (i) Zn(CN)2, Zn dust, PdCl2(dppf).DCM complex, DMA, 120°C, 16 h; (ii) pyridinium tribromide, THF, 0°C to rt, 16 h; (iii) NaN(CHO)2, MeCN, 80°C, 5 h, followed by concentrated HCl, 80°C, 16 h; (iv) K2CO3, DCM, 0°C to rt, 4 h; (v) POCl3, 100°C, 16 h.

[0175] Project (i) 3-Acetyl-4-methoxybenzonitrile To a stirred solution of 1-(5-bromo-2-methoxyphenyl)ethan-1-one (CAS 16740-73-1, from Combi-blocks, 60.0 g, 261.89 mmol) in DMA (600 mL) was added zinc cyanide (92.25 g, 785.67 mmol) and zinc dust (17.22 g, 261.89 mmol) at room temperature. The mixture was degassed with N gas for 30 minutes and then provided [1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium(II) (10.69 g, 13.09 mmol) complexed with DCM. The mixture was heated at 120°C for 16 hours. The mixture was allowed to cool to room temperature, filtered through Celite Hyflow™, and the filtrate was poured into ice-cold water (800 mL) and extracted with EtOAc (2 x 1000 mL). The combined organic phase was washed with ice-cold water (4x1000 mL), dried over Na2SO4, and concentrated under reduced pressure. The residue was purified by column chromatography (60-120# silica gel, 15% EtOAc in n-hexane) to give 3-acetyl-4-methoxybenzonitrile (40.0 g, 228.57 mmol, 87% yield). LCMS: Method C, 1.52 min; MS: ES+: 176.08.

[0176] Project (ii) 3-(2-Bromoacetyl)-4-methoxybenzonitrile To a stirred solution of 3-acetyl-4-methoxybenzonitrile (7.20 g, 41.14 mmol) in THF (72 mL) was added pyridinium tribromide (14.47 g, 45.25 mmol) at 0 °C. The mixture was slowly warmed to room temperature and stirred for 16 h, then poured into water (400 mL) and extracted with EtOAc (2 × 400 mL). The combined organic phases were dried over NaSO and concentrated under reduced pressure to provide 3-(2-bromoacetyl)-4-methoxybenzonitrile (13.0 g, quantitative yield). This crude material was used directly in the next step.

[0177] Project (iii) 3-Glycyl-4-methoxybenzonitrile hydrochloride To a stirred solution of 3-(2-bromoacetyl)-4-methoxybenzonitrile (10.5 g, 41.51 mmol) in acetonitrile (105 mL) was added sodium diformylamide (5.91 g, 62.26 mmol), and the mixture was heated at 80° C. for 8 hours. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, and the residue was diluted with MeOH (105 mL) and concentrated HCl (10.5 mL). The mixture was further heated at 80° C. for 16 hours and then allowed to cool to room temperature. The mixture was concentrated under reduced pressure, and the residue was stirred with isopropyl alcohol (40 mL), forming a precipitate which was collected by filtration under reduced pressure to provide 3-glycyl-4-methoxybenzonitrile hydrochloride (9.0 g, 39.73 mmol, 95% yield). LCMS: Method C2, 0.40 min; MS: ES+: 191.0.

[0178] Engineering (iv) Ethyl 2-((2-(5-cyano-2-methoxyphenyl)-2-oxoethyl)amino)-2-oxoacetate To a stirred solution of 3-glycyl-4-methoxybenzonitrile hydrochloride HCl salt (9.0 g, 39.73 mmol) in DCM (90 mL) was added KCO (21.93 g, 158.92 mmol) at 0 °C. Ethyl oxalyl chloride (10.84 g, 8.89 mL, 79.46 mmol) was added dropwise at 0 °C. The mixture was warmed to room temperature and stirred for 4 h. The mixture was added to water (900 mL) and extracted with DCM (2 × 500 mL). The combined organic phases were dried over NaSO and concentrated under reduced pressure to provide ethyl 2-((2-(5-cyano-2-methoxyphenyl)-2-oxoethyl)amino)-2-oxoacetate (9.0 g, 31.03 mmol, 78% yield). LCMS: Method C, 1.34 min; MS: ES+ 291.1.

[0179] Engineering (v) Ethyl 5-(5-cyano-2-methoxyphenyl)oxazole-2-carboxylate A stirred solution of ethyl 2-((2-(5-cyano-2-methoxyphenyl)-2-oxoethyl)amino)-2-oxoacetate (1.2 g, 4.14 mmol) in POCl (12 mL, 10 vol) was heated at 100 °C for 2 h. The mixture was cooled to room temperature, poured into ice-cold water (200 mL), and extracted with EtOAc (3 × 100 mL). The combined organic phases were washed with saturated NaHCO solution (3 × 100 mL), dried over NaSO, and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 32% EtOAc in n-hexane) to provide ethyl 5-(5-cyano-2-methoxyphenyl)oxazole-2-carboxylate (0.60 g, 2.20 mmol, 53% yield). LCMS: Method C, 1.53 min; MS: ES+: 272.6.

[0180] Intermediate C Ethyl 5-(5-bromo-2-(trifluoromethoxy)phenyl)oxazole-2-carboxylate

[0181] [ka]

[0182] (i) HATU, DIPEA, THF, 0 °C to rt, 3 h; (ii) methylmagnesium bromide, THF, -10 °C to 0 °C, 4 h; (iii) phenyltrimethylammonium tribromide, MeCN, 0 °C to rt, 9 h; (iv) NaN(CHO)2, MeCN, 80 °C, 1 h, followed by concentrated HCl, 80 °C, 3 h; (v) K2CO3, DCM, 0 °C to rt, 3 h; (vi) POCl3, 110 °C, 16 h.

[0183] Project (i) 5-Bromo-N-methoxy-N-methyl-2-(trifluoromethoxy)benzamide To a stirred solution of 5-bromo-2-(trifluoromethoxy)benzoic acid (CAS 403646-47-9, from Combi-blocks, 10.0 g, 35.08 mmol) in THF (100 mL) was added DIPEA (13.57 g, 17.9 mL, 105.25 mmol) and HATU (20.0 g, 52.62 mmol) in small portions at 0° C. After 30 min, N,O-dimethylhydroxylamine HCl (4.45 g, 45.61 mmol) was added at 0° C. The mixture was slowly warmed to room temperature and stirred for 3 h, then poured into ice-cold water (300 mL) and extracted with EtOAc (3×150 mL). The combined organic phases were dried over NaSO and concentrated under reduced pressure to give 5-bromo-N-methoxy-N-methyl-2-(trifluoromethoxy)benzamide (12.2 g, quantitative yield), which crude material was used directly in the next step. LCMS: Method C, 1.69 min; MS: ES+: 327.8, 329.8.

[0184] Project (ii) 1-(5-bromo-2-(trifluoromethoxy)phenyl)ethan-1-one To a stirred solution of 5-bromo-N-methoxy-N-methyl-2-(trifluoromethoxy)benzamide (12.2 g, 37.31 mmol) in THF (130 mL) was added dropwise methylmagnesium bromide (3 M in diethyl ether, 24.9 mL, 74.62 mmol) at −10° C. The mixture was slowly warmed to 0° C. and stirred for 4 h, then poured into saturated NH4Cl solution (300 mL) and extracted with EtOAc (3×200 mL). The combined organic phases were dried over Na2SO4 and concentrated under reduced pressure to provide 1-(5-bromo-2-(trifluoromethoxy)phenyl)ethan-1-one (10.0 g, quantitative yield). This crude material was used directly in the next step. LCMS: Method C, m / z not supported; 1 H NMR (400 MHz, CDCl3) δ ppm: 7.92 (s, 1H), 7.67 - 7.71 (m, 1H), 7.24 (d, J = 8.0 Hz, 1H), 2.62 (s, 3H).

[0185] Project (iii) 2-Bromo-1-(5-bromo-2-(trifluoromethoxy)phenyl)ethan-1-one To a stirred solution of 1-(5-bromo-2-(trifluoromethoxy)phenyl)ethan-1-one (10.0 g, 35.46 mmol) in acetonitrile (100 mL) was added phenyltrimethylammonium tribromide (13.33 g, 35.46 mmol) in small portions at 0 °C. The mixture was slowly warmed to room temperature and stirred for 9 h, then poured into water (200 mL) and extracted with EtOAc (3 × 100 mL). The combined organic phases were dried over NaSO and concentrated under reduced pressure. The residue was purified by column chromatography (silica gel, 5% EtOAc in n-hexane) to give 2-bromo-1-(5-bromo-2-(trifluoromethoxy)phenyl)ethan-1-one (11.5 g, 31.95 mmol, 91% yield for three steps). LCMS: m / z not supported; 1 H NMR (400 MHz, CDCl3) δ ppm: 7.93 (s, 1H), 7.73 (dd, J = 8.8, 2.4 Hz, 1H), 7.25 (d, J = 8.0 Hz, 1H), 4.45 (s, 2H).

[0186] Engineering (iv) 2-Amino-1-(5-bromo-2-(trifluoromethoxy)phenyl)ethan-1-one hydrochloride To a stirred solution of 2-bromo-1-(5-bromo-2-(trifluoromethoxy)phenyl)ethan-1-one (10.0 g, 27.78 mmol) in acetonitrile (100 mL) was added sodium diformylamide (5.28 g, 55.58 mmol), and the mixture was heated at 80° C. for 1 h. The mixture was cooled to room temperature, concentrated under reduced pressure, and the residue was diluted with MeOH (100 mL) and concentrated HCl (10.0 mL). The mixture was heated at 80° C. for 3 h and then allowed to cool to room temperature. The mixture was concentrated under reduced pressure, and the residue was stirred with diethyl ether (4×30 mL), forming a precipitate which was collected by filtration under reduced pressure to provide 2-amino-1-(5-bromo-2-(trifluoromethoxy)phenyl)ethan-1-one hydrochloride (9.41 g, quantitative yield). LCMS: Method C, 1.65 min; MS: ES+: 299.9, 301.9.

[0187] Engineering (v) Ethyl 2-((2-(5-bromo-2-(trifluoromethoxy)phenyl)-2-oxoethyl)amino)-2-oxoacetate To a solution of 2-amino-1-(5-bromo-2-(trifluoromethoxy)phenyl)ethan-1-one HCl salt (9.4 g, 28.19 mmol) in DCM (94 mL) was added KCO (11.67 g, 84.56 mmol) at 0 °C. Ethyl oxalyl chloride (5.77 g, 4.73 mL, 42.28 mmol) was added dropwise at 0 °C. The mixture was warmed to room temperature and stirred for 3 h, then poured into water (150 mL) and extracted with DCM (3 × 120 mL). The combined organic phases were dried over NaSO and concentrated under reduced pressure to provide ethyl 2-((2-(5-bromo-2-(trifluoromethoxy)phenyl)-2-oxoethyl)amino)-2-oxoacetate (3.4 g, 8.56 mmol, 31% yield in two steps). LCMS: Method C, 2.09 min; MS: ES- 396.0, 398.0.

[0188] Engineering (vi) Ethyl 5-(5-bromo-2-(trifluoromethoxy)phenyl)oxazole-2-carboxylate A stirred solution of ethyl 2-((2-(5-bromo-2-(trifluoromethoxy)phenyl)-2-oxoethyl)amino)-2-oxoacetate (3.4 g, 8.56 mmol) in POCl (17 mL, 5 vol) was heated at 110 °C for 16 h. The mixture was cooled to room temperature, poured into ice-cold water (250 mL), and extracted with EtOAc (3 × 100 mL). The combined organic phases were washed with saturated NaHCO solution (3 × 100 mL), dried over NaSO, and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 18% EtOAc in n-hexane) to provide ethyl 5-(5-bromo-2-(trifluoromethoxy)phenyl)oxazole-2-carboxylate (1.3 g, 3.43 mmol, 40% yield). LCMS: Method C, 2.39 min; MS: ES+: 379.8, 381.8.

[0189] Intermediate D Ethyl 5-(5-cyano-4-fluoro-2-methoxyphenyl)oxazole-2-carboxylate

[0190] [ka]

[0191] (i) triflic acid, NBS, MeCN, -30 °C to rt, 18 h; (ii) K2CO3, MeI, DMF, 0 °C to rt, 3 h; (iii) tributyl(1-ethoxyvinyl)tin, PdCl2(PPh3)2, 1,4-dioxane, 100 °C, 3 h, then NBS, THF:water, 0 °C, 10 min; (iv) NaN(CHO)2, MeCN, 80 °C, 2 h, then concentrated HCl, 80 °C, 16 h; (v) ethyl chlorooxoacetate, K2CO3, DCM, 0 °C to rt, 2 h; (vi) POCl3, 100 °C, 5 h.

[0192] Project (i) 5-Bromo-2-fluoro-4-hydroxybenzonitrile To a stirred solution of 2-fluoro-4-hydroxybenzonitrile (CAS 82380-18-5, from Combi-blocks, 8.0 g, 58.39 mmol) in acetonitrile (80 mL) was added triflic acid (10.51 g, 6.18 mL, 70.07 mmol) dropwise at −30° C. After 10 min, N-bromosuccinimide (10.39 g, 58.39 mmol) was added portionwise at −30° C. The mixture was allowed to warm slowly to room temperature and stirred for 18 h, then poured into saturated NaHCO solution (150 mL) and extracted with EtOAc (2×150 mL). The combined organic phases were dried over NaSO and concentrated under reduced pressure to give 5-bromo-2-fluoro-4-hydroxybenzonitrile (5.08 g, 23.63 mmol, 40% yield). This crude material was used directly in the next step. LCMS: Method C, 1.58 min; MS: ES-: 214.0, 216.0.

[0193] Project (ii) 5-Bromo-2-fluoro-4-methoxybenzonitrile To a stirred solution of 5-bromo-2-fluoro-4-hydroxybenzonitrile (5.08 g, 23.63 mmol) in DMF (30 mL) was added dropwise K2CO3 (6.52 g, 47.26 mmol) at 0 °C. After 10 min, methyl iodide (5.03 g, 2.21 mL, 35.45 mmol) was added dropwise at 0 °C. The mixture was slowly warmed to room temperature and stirred for 3 h, then poured into water (100 mL) and extracted with EtOAc (2 × 100 mL). The combined organic phases were dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by column chromatography (silica gel, 2% EtOAc in n-hexane) to give 5-bromo-2-fluoro-4-methoxybenzonitrile (5.70 g, quantitative yield). LCMS: m / z not supported; 1 H NMR (400 MHz, DMSO-d6) δ ppm: 8.30 (d, J = 7.2 Hz, 1H), 7.46 (d, J = 12.0 Hz, 1H), 4.01 (s, 3H).

[0194] Project (iii) 5-(2-Bromoacetyl)-2-fluoro-4-methoxybenzonitrile A stirred solution of 5-bromo-2-fluoro-4-methoxybenzonitrile (5.70 g, 24.89 mmol) and tributyl(1-ethoxyvinyl)tin (CAS 97674-02-7, from Combi-blocks, 13.46 g, 37.34 mmol) in 1,4-dioxane (70 mL) was degassed with N for 15 minutes, and then bis(triphenylphosphine)palladium(II) dichloride (0.87 g, 1.24 mmol) was added at room temperature. The mixture was heated at 100° C. for 3 hours. The mixture was cooled to 0° C., and THF:water (2:1, 120 mL) was added. N-bromosuccinimide (8.85 g, 49.78 mmol) was added at 0° C. The mixture was stirred at 0° C. for 10 minutes, then poured into water (100 mL) and extracted with EtOAc (2×100 mL). The combined organic phases were dried over NaSO and concentrated under reduced pressure. The residue was purified by column chromatography (silica gel, 2% EtOAc in n-hexane) to give 5-(2-bromoacetyl)-2-fluoro-4-methoxybenzonitrile (6.3 g, 23.25 mmol, 98% yield in two steps). LCMS: m / z not supported; 1 H NMR (400 MHz, DMSO-d6) δ ppm: 8.27 (dd, J = 7.6, 2.4 Hz, 1H), 7.54 (dd, J = 12.0, 2.4 Hz, 1H), 4.84 (s, 2H), 4.05 (s, 3H).

[0195] Engineering (iv) 2-Fluoro-5-glycyl-4-methoxybenzonitrile hydrochloride To a stirred solution of 5-(2-bromoacetyl)-2-fluoro-4-methoxybenzonitrile (6.30 g, 23.25 mmol) in acetonitrile (65 mL) was added sodium diformylamide (25.85 g, 27.9 mmol), and the mixture was heated at 80° C. for 2 hours. The mixture was cooled to room temperature, concentrated under reduced pressure, and the residue was diluted with MeOH (65 mL) and concentrated HCl (6.5 mL). The mixture was heated at 80° C. for 16 hours and then allowed to cool to room temperature. The mixture was concentrated under reduced pressure, and the residue was stirred with isopropyl alcohol (100 mL), forming a precipitate that was collected by filtration under reduced pressure to provide 2-fluoro-5-glycyl-4-methoxybenzonitrile hydrochloride (5.3 g, quantitative yield). LCMS: Method H, 1.96 min; MS: ES+: 209.1.

[0196] Engineering (v) Ethyl 2-((2-(5-cyano-4-fluoro-2-methoxyphenyl)-2-oxoethyl)amino)-2-oxoacetate To a stirred solution of 2-fluoro-5-glycyl-4-methoxybenzonitrile HCl salt (5.30 g, 21.67 mmol) in DCM (60 mL) was added KCO (11.96 g, 86.68 mmol) at 0 °C. Ethyl oxalyl chloride (5.89 g, 4.83 mL, 43.34 mmol) was added at 0 °C. The mixture was warmed to room temperature and stirred for 2 h, then poured into water (150 mL) and extracted with EtOAc (2 × 150 mL). The combined organic phases were dried over NaSO and concentrated under reduced pressure to provide ethyl 2-((2-(5-cyano-4-fluoro-2-methoxyphenyl)-2-oxoethyl)amino)-2-oxoacetate (4.20 g, 13.63 mmol, 58% yield in two steps). LCMS: Method C, 1.50 min; MS: ES+ 309.4.

[0197] Engineering (vi) Ethyl 5-(5-cyano-4-fluoro-2-methoxyphenyl)oxazole-2-carboxylate A stirred solution of ethyl 2-((2-(5-cyano-4-fluoro-2-methoxyphenyl)-2-oxoethyl)amino)-2-oxoacetate (4.2 g, 13.63 mmol) in POCl (42 mL, 10 vol) was heated at 100 °C for 5 h. The mixture was cooled to room temperature, poured into ice-cold water (100 mL), and extracted with EtOAc (2 x 150 mL). The combined organic phases were washed with saturated NaHCO solution (2 x 100 mL), dried over NaSO, and concentrated under reduced pressure. The residue was suspended in MeOH (30 mL) and stirred at -78 °C for 15 min. The solid was collected by filtration under reduced pressure to provide ethyl 5-(5-cyano-4-fluoro-2-methoxyphenyl)oxazole-2-carboxylate (1.60 g, 5.51 mmol, 40% yield). LCMS: Method C, 1.63 min; MS: ES+: 291.2.

[0198] Example 1 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide

[0199] [ka]

[0200] (i) TBD, THF, 0 °C to rt; (ii) TFA, DCM, 0 °C to rt; (iii) K2CO3, CNBr, THF, 0 °C to rt.

[0201] Project (i) tert-Butyl (2R,4R)-4-(5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamido)-2-methylpyrrolidine-1-carboxylate To a stirred solution of ethyl 5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxylate (0.60 g, 2.01 mmol) and tert-butyl (2R,4R)-4-amino-2-methylpyrrolidine-1-carboxylate (CAS 348165-63-9, 0.60 g, 3.02 mmol) in THF (10 mL) was added TBD (0.42 g, 3.02 mmol) in small portions at 0 °C. The mixture was warmed to room temperature and stirred for 1 h, then poured into water (70 mL) and extracted with EtOAc (2 × 70 mL). The combined organic phase was dried over anhydrous NaSO and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 30% EtOAc in n-hexane) to give tert-butyl (2R,4R)-4-(5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamido)-2-methylpyrrolidine-1-carboxylate (0.32 g, 0.71 mmol, 35% yield). LCMS: Method C, 1.86 min; MS: ES- 451.4.

[0202] Project (ii) 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of tert-butyl (2R,4R)-4-(5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamide)-2-methylpyrrolidine-1-carboxylate (0.31 g, 0.68 mmol) in DCM (7 mL) was added TFA (0.93 mL, 3 vol) dropwise at 0° C. The mixture was warmed to room temperature and stirred for 45 minutes, then concentrated under reduced pressure to provide 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.3 g, 0.64 mmol, 94% yield). LCMS: Method C, 1.38 min; MS: ES+ 353.3.

[0203] Project (iii) 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.3 g, 0.64 mmol) in THF (8 mL) was added K2CO3 (0.27 g, 1.93 mmol) at room temperature, and the mixture was stirred for 10 min. Cyanogen bromide (0.07 g, 0.64 mmol) was added at 0 °C, and the mixture was stirred at room temperature for 45 min, then poured into water (70 mL) and extracted with EtOAc (2 x 70 mL). The combined organic phases were dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 40% EtOAc in n-hexane) to give 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide (0.1 g, 0.27 mmol, 42% yield). LCMS: Method H, 2.96 min; MS: ES+ 378.0; 1 H NMR (400 MHz, DMSO-d6) δ ppm: 9.36 (d, J = 6.4 Hz, 1H), 8.27 (s, 1H), 7.98 (d, J = 8.4 Hz, 1H), 7.68 - 7.70 (m, 2H), 4.51 - 4.53 (m, 1H), 4.21 - 4.23 (m, 1H), 3.89 - 3.95 (m, 1H), 3.77 - 3.81 (m, 1H), 3.43 - 3.46 (m, 1H), 2.16 - 2.18 (m, 1H), 1.76 - 1.83 (m, 1H), 1.26 (d, J = 6.4 Hz, 3H), 0.92 (s, 4H). Chiral SFC: Method Y5, 5.18 min.

[0204] Example 2 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide

[0205] [ka]

[0206] (i) TBD, THF, 0 °C to rt; (ii) TFA, DCM, 0 °C to rt; (iii) K2CO3, CNBr, THF, 0 °C to rt.

[0207] Project (i) tert-Butyl (2S,4R)-4-(5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamido)-2-(methoxymethyl)pyrrolidine-1-carboxylate To a stirred solution of ethyl 5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxylate (0.50 g, 1.67 mmol) and tert-butyl-(2S,4R)-4-amino-2-(methoxymethyl)pyrrolidine-1-carboxylate (CAS 1207853-53-9, 0.46 g, 2.01 mmol) in THF (8 mL) was added TBD (0.35 g, 2.51 mmol) in small portions at 0 °C. The mixture was warmed to room temperature and stirred for 3 h, then poured into water (100 mL) and extracted with EtOAc (2 × 80 mL). The combined organic phase was dried over anhydrous NaSO and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 55% EtOAc in n-hexane) to give tert-butyl (2S,4R)-4-(5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamido)-2-(methoxymethyl)pyrrolidine-1-carboxylate (0.32 g, 0.66 mmol, 39% yield). LCMS: Method C1, 1.35 min; MS: ES+ 483.4.

[0208] Project (ii) 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of tert-butyl (2S,4R)-4-(5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamide)-2-(methoxymethyl)pyrrolidine-1-carboxylate (0.32 g, 0.66 mmol) in DCM (8 mL) was added TFA (0.96 mL, 3 vol) dropwise at 0° C. The mixture was allowed to warm to room temperature and stirred for 1 hour, then concentrated under reduced pressure to provide 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.32 g, 0.64 mmol, 97% yield). LCMS: Method C1, 1.06 min; MS: ES+ 383.4.

[0209] Project (iii) 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.32 g, 0.64 mmol) in THF (8 mL) was added K2CO3 (0.27 g, 1.93 mmol) at room temperature and stirred for 10 min. Cyanogen bromide (0.07 g, 0.64 mmol) was added at 0 °C. The mixture was stirred at room temperature for 1 h, then poured into water (80 mL) and extracted with EtOAc (2 x 80 mL). The combined organic phases were dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 70% EtOAc in n-hexane) to give 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide (0.10 g, 0.24 mmol, 38% yield). LCMS: Method H, 2.94 min; MS: ES+ 408.2; 1 H NMR (400 MHz, DMSO-d6) δ ppm: 9.32 (d, J = 6.4 Hz, 1H), 8.26 (d, J = 1.6 Hz, 1H), 7.98 (d, J = 8.8 Hz, 1H), 7.69 -7.70 (m, 2H), 4.47 - 4.59 (m, 1H), 4.19 - 4.27 (m, 1H), 4.00 - 4.10 (m, 1H), 3.70 - 3.74 (m, 1H), 3.39 - 3.51 (m, 3H), 3.36 (s, 3H), 2.11 - 2.18 (m, 1H), 1.98 - 2.03 (m, 1H), 0.92 (s, 4H). Chiral SFC: Method Y3, 3.56 min.

[0210] Engineering (iii) Replacement synthesis 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (600.0 g, 1209.67 mmol) in THF (6 L) under N2 atmosphere, K2CO3 (500.80 g, 3629.01 mmol) was added at room temperature under N2 atmosphere and stirred for 15-20 min. A solution of cyanogen bromide (153.87 g, 1451.60 mmol) in THF (1.2 L) was added dropwise at 0 °C. The mixture was allowed to warm to room temperature and stirred at room temperature for 1 h. The progress of the reaction was monitored by TLC (mobile phase: 50% EtOAc in n-hexane, product R f 0.05; 10% MeOH / DCM product R f The reaction mixture was monitored by HPLC (0.4). The mixture was quenched with water (3 L) and extracted with EtOAc (3 x 6 L). The combined organic phases were washed with brine solution (1.5 L), dried over anhydrous NaSO, and concentrated under reduced pressure to give crude 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide (505 g). The crude material (505 g) was suspended in IPA (5 L, 10 vol) and heated at 80 °C for 1 h to give a clear solution. The mixture was allowed to cool slowly to room temperature and then cooled at 0° C., forming a crystalline solid which was collected by filtration under reduced pressure, quenched with cold IPA (505 mL), and dried under vacuum at 50° C. to give 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)-pyrrolidin-3-yl)oxazole-2-carboxamide as a white crystalline solid (400.0 g, 982.80 mmol).

[0211] The material from the IPA recrystallization (400 g) was suspended in IPA (10 L, 25 vol), charcoal (100 g) was added, and the mixture was heated at 80° C. for 1 h. The hot mixture was filtered through Celite Hyflow™ and washed with hot IPA (800 mL, 2 vol). The filtrate was allowed to cool slowly to room temperature and then chilled at 0° C., forming a crystalline solid which was collected by suction filtration, washed with cold IPA (400 L, 1 vol), and dried under vacuum at 50° C. to provide 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide as a white crystalline solid (320.0 g, 786.24 mmol, 63% yield). LCMS: Method H1, 2.81 min, MS: ES+ 408.2; HPLC: Method P3, 24.22 min; Chiral HPLC: Method Y, 28.97 min; 1H NMR (400 MHz, DMSO-d6) δ ppm: 9.29 (d, J = 6.8 Hz, 1H), 8.22 (d, J = 2.0 Hz, 1H), 7.95 (dd, J = 8.8, 2.4 Hz, 1H), 7.67 (d, J = 8.8 Hz, 1H), 7.65 (s, 1H), 4.48 - 4.58 (m, 1H), 4.18 - 4.22 (m, 1H), 4.01 - 4.07 (m, 1H), 3.70 - 3.74 (m, 1H), 3.40 - 3.51 (m, 3H), 3.35 (s, 3H), 2.12 - 2.18 (m, 1H), 1.97 - 2.03 (m, 1H), 0.88 - 0.96 (m, 4H); Chiral HPLC: Method Y15, 15.11 min; 99.8% ee; HPLC: Method Y26, 27.87 min; DSC peak temperature (melting) = 132.7 °C; High-resolution MS: ES- 406.1521 (calculated exact mass 407.1594). XRPD data in the table below was prepared on a Bruker AXS D8 Advance: Cu, kα: Kα1 (Å): 1.540598; Kα2 (Å): 1.544426; Kα2 / Kα1 = 0.50.

[0212] [Table 18]

[0213] Instead of recrystallization 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide Crude 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide (134.4 g, 98.9% purity by LC) was suspended in ethanol (1340 mL, 10 vol) and heated to 55-65°C to form a brown solution. The mixture was allowed to cool to 40-45°C and seed material from a similar smaller scale batch was added (135 mg) and the mixture was stirred at 40-45°C for 30 minutes. The mixture was cooled to room temperature and stirred overnight. The beige suspension was filtered and the solid collected, washed with ethanol (2x260 mL), and the filter cake was sucked dry and then dried at 60°C overnight to give 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide as an off-white solid (116.2 g, 86% yield from crude). MS: ES+ 408.2; 1 H NMR (400 MHz, CDCl3) δ ppm: 8.24 (d, J = 2.0 Hz, 1H), 7.66 (dd, J = 8.8, 2.0 Hz, 1H), 7.56 (s, 1H), 7.47 (d, J = 8.4 Hz, 1H), 4.68 - 4.75 (m, 1H), 3.97 - 4.00 (m, 2H), 3.83 - 3.85 (m, 1H), 3.60 - 3.66 (m, 1H), 3.45 - 3.53 (m, 2H), 3.42 (s, 3H), 2.24 - 2.32 (m, 1H), 2.09 - 2.15 (m, 1H), 0.89 - 1.02 (m, 4H); residual solvent by NMR = EtOH (2260 ppm); HPLC purity 99.6%; DSC peak temperature (melting) = 86.96 °C.

[0214] Example 2 was prepared as an aqueous suspension containing 0.1% (w / v) Tween-80 and 0.5% (w / v) hydroxypropyl-methylcellulose.

[0215] Example 3 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(fluoromethyl)pyrrolidin-3-yl)oxazole-2-carboxamide

[0216] [ka]

[0217] (i) TBD, THF, 0 °C to rt; (ii) TFA, DCM, 0 °C to rt; (iii) K2CO3, CNBr, THF, 0 °C to rt.

[0218] Project (i) tert-Butyl (2S,4R)-4-(5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamido)-2-(fluoromethyl)pyrrolidine-1-carboxylate To a stirred solution of ethyl 5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxylate (0.15 g, 0.50 mmol) and tert-butyl (2S,4R)-4-amino-2-(fluoromethyl)pyrrolidine-1-carboxylate (CAS 1207853-03-9, from Angene, 0.11 g, 0.50 mmol) in THF (4 mL) was added TBD (0.07 g, 0.50 mmol) in small portions at 0 °C. The mixture was warmed to room temperature and stirred for 8 h, then poured into water (40 mL) and extracted with EtOAc (2 × 50 mL). The combined organic phase was dried over anhydrous NaSO and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 51% EtOAc in n-hexane) to give tert-butyl (2S,4R)-4-(5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamido)-2-(fluoromethyl)pyrrolidine-1-carboxylate (0.09 g, 0.19 mmol, 37% yield). LCMS: Method C, 1.76 min; MS: ES- 469.1.

[0219] Project (ii) 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-5-(fluoromethyl)pyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of tert-butyl (2S,4R)-4-(5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamide)-2-(fluoromethyl)pyrrolidine-1-carboxylate (0.09 g, 0.19 mmol) in DCM (5 mL) was added TFA (0.9 mL, 10 vol) dropwise at 0° C. The mixture was allowed to warm to room temperature and stirred for 1 hour, then concentrated under reduced pressure to give 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-5-(fluoromethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.16 g, quantitative yield). LCMS: Method C, 1.35 min; MS: ES- 368.9.

[0220] Project (iii) 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(fluoromethyl)pyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-5-(fluoromethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.15 g, 0.32 mmol) in THF (5 mL) was added K2CO3 (0.13 g, 0.96 mmol) at room temperature, and the mixture was stirred for 10 min. Cyanogen bromide (0.03 g, 0.32 mmol) was added at 0 °C. The mixture was stirred at room temperature for 1 h, then poured into water (40 mL) and extracted with EtOAc (2 x 40 mL). The combined organic phases were dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 69% EtOAc in n-hexane) to give 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(fluoromethyl)-pyrrolidin-3-yl)oxazole-2-carboxamide (0.01 g, 0.03 mmol, 16% yield in two steps). LCMS: Method H, 2.96 min; MS: ES+ 396.1; 1 H NMR (400 MHz, DMSO-d6) δ ppm: 9.39 (d, J = 6.8 Hz, 1H), 8.25 (s, 1H), 7.98 (d, J = 7.2 Hz, 1H), 7.68 - 7.70 (m, 2H), 4.41 - 4.69 (m, 3H), 4.10 - 4.26 (m, 2H), 3.61 - 3.80 (m, 1H), 3.43 - 3.53 (m, 1H), 2.12 - 2.25 (m, 1H), 1.95 - 2.07 (m, 1H), 0.92 (s, 4H). Chiral SFC: Method Y4, 4.96 min.

[0221] Example 4 5-(5-cyano-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide

[0222] [ka]

[0223] (i) TBD, THF, 0 °C to rt; (ii) TFA, DCM, 0 °C to rt; (iii) K2CO3, CNBr, THF, 0 °C to rt.

[0224] Project (i) tert-Butyl (2R,4R)-4-(5-(5-cyano-2-methoxyphenyl)oxazole-2-carboxamido)-2-methylpyrrolidine-1-carboxylate To a stirred solution of ethyl 5-(5-cyano-2-methoxyphenyl)oxazole-2-carboxylate (0.5 g, 1.84 mmol) and tert-butyl (2R,4R)-4-amino-2-methylpyrrolidine-1-carboxylate (CAS 348165-63-9, 0.37 g, 1.84 mmol) in THF (10 mL) was added TBD (0.38 g, 2.76 mmol) in small portions at 0 °C. The mixture was warmed to room temperature and stirred for 2 h, then poured into water (100 mL) and extracted with EtOAc (2 × 70 mL). The combined organic phase was dried over anhydrous NaSO and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 30% EtOAc in n-hexane) to give tert-butyl (2R,4R)-4-(5-(5-cyano-2-methoxyphenyl)oxazole-2-carboxamido)-2-methylpyrrolidine-1-carboxylate (0.30 g, 0.70 mmol, 38% yield). LCMS: Method C, 1.61 min; MS: ES- 425.1.

[0225] Project (ii) 5-(5-cyano-2-methoxyphenyl)-N-((3R,5R)-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of tert-butyl (2R,4R)-4-(5-(5-cyano-2-methoxyphenyl)oxazole-2-carboxamide)-2-methylpyrrolidine-1-carboxylate (0.30 g, 0.70 mmol) in DCM (6 mL) was added TFA (0.9 mL, 3 vol) dropwise at 0° C. The mixture was allowed to warm to room temperature and stirred for 1 h, then concentrated under reduced pressure to give 5-(5-cyano-2-methoxyphenyl)-N-((3R,5R)-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.30 g, 0.68 mmol, 97% yield). LCMS: Method C, 1.22 min; MS: ES+ 327.1.

[0226] Project (iii) 5-(5-cyano-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of 5-(5-cyano-2-methoxyphenyl)-N-((3R,5R)-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.30 g, 0.68 mmol) in THF (7 mL) was added K2CO3 (0.28 g, 2.04 mmol) at room temperature and stirred for 10 min. Cyanogen bromide (0.07 g, 0.68 mmol) was added at 0 °C. The mixture was stirred at room temperature for 1 h, then poured into water (50 mL) and extracted with EtOAc (2 x 50 mL). The combined organic phase was dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 1% MeOH in DCM) to give 5-(5-cyano-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide (0.10 g, 0.28 mmol, 42% yield). LCMS: Method H, 2.73 min; MS: ES+ 352.2; 1H NMR (400 MHz, DMSO-d6) δ ppm: 9.35 (d, J = 6.8 Hz, 1H), 8.25 (s, 1H), 7.95 (d, J = 7.2 Hz, 1H), 7.84 (s, 1H), 7.41 (d, J = 8.4 Hz, 1H), 4.47 - 4.57 (m, 1H), 4.08 (s, 3H), 3.89 - 3.94 (m, 1H), 3.76 - 3.80 (m, 1H), 3.42 - 3.49 (m, 1H), 2.16 - 2.20 (m, 1H), 1.76 - 1.83 (m, 1H), 1.26 (d, J = 6.4 Hz, 3H). Chiral SFC: Method Y6, 3.76 min.

[0227] Example 5 5-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide

[0228] [ka]

[0229] (i) TBD, THF, 0 °C to rt; (ii) TFA, DCM, 0 °C to rt; (iii) K2CO3, CNBr, THF, 0 °C to rt.

[0230] Project (i) tert-Butyl (2S,4R)-4-(5-(5-cyano-2-methoxyphenyl)oxazole-2-carboxamido)-2-(methoxymethyl)pyrrolidine-1-carboxylate To a stirred solution of ethyl 5-(5-cyano-2-methoxyphenyl)oxazole-2-carboxylate (0.80 g, 2.94 mmol) and tert-butyl (2S,4R)-4-amino-2-(methoxymethyl)pyrrolidine-1-carboxylate (CAS 1207853-53-9, 0.67 g, 2.94 mmol) in THF (20 mL) was added TBD (0.61 g, 4.41 mmol) in small portions at 0 °C. The mixture was stirred at 0 °C for 6 h, then poured into water (50 mL) and extracted with EtOAc (2 × 70 mL). The combined organic phase was dried over anhydrous NaSO and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 50% EtOAc in n-hexane) to give tert-butyl (2S,4R)-4-(5-(5-cyano-2-methoxyphenyl)oxazole-2-carboxamido)-2-(methoxymethyl)pyrrolidine-1-carboxylate (0.25 g, 0.55 mmol, 18% yield). LCMS: Method C1, 1.29 min; MS: ES+ 457.2.

[0231] Project (ii) 5-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of tert-butyl (2S,4R)-4-(5-(5-cyano-2-methoxyphenyl)oxazole-2-carboxamide)-2-(methoxymethyl)pyrrolidine-1-carboxylate (0.25 g, 0.55 mmol) in DCM (5 mL) was added TFA (2.5 mL, 10 vol) dropwise at 0° C. The mixture was allowed to warm to room temperature and stirred for 2 hours, then concentrated under reduced pressure to give 5-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.32 g, quantitative yield). LCMS: Method C1, 1.00 min; MS: ES+ 357.3.

[0232] Project (iii) 5-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of 5-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.31 g, 0.66 mmol) in THF (15 mL) was added KCO (0.27 g, 1.98 mmol) at room temperature, and after stirring for 10 min, cyanogen bromide (0.07 g, 0.66 mmol) was added to the mixture at 0 °C. The mixture was stirred at 0 °C for 0.5 h, then poured into water (50 mL) and extracted with EtOAc (2 × 50 mL). The combined organic phase was dried over NaSO and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 60% EtOAc in n-hexane) to give 5-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide (0.11 g, 0.29 mmol, 52% yield in two steps). LCMS: Method H, 2.66 min; MS: ES+ 382.2; 1 H NMR (400 MHz, DMSO-d6) δ ppm: 9.34 (d, J = 6.8 Hz, 1H), 8.25 (s, 1H), 7.96 (d, J = 8.0 Hz, 1H), 7.85 (s, 1H), 7.43 (d, J = 8.8 Hz, 1H), 4.47 - 4.58 (m, 1H), 4.09 (s, 3H), 4.00 - 4.10 (m, 1H), 3.70 - 3.74 (m, 1H), 3.41 - 3.53 (m, 3H), 3.36 (s, 3H), 2.08 - 2.20 (m, 1H), 1.95 - 2.06 (m, 1H). Chiral SFC: Method Y7, 5.42 min.

[0233] Example 6 4-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide

[0234] [ka]

[0235] (i) AgOTf, EtOAc, 80 °C; (ii) LiOH.H2O, H2O, THF, 0 °C to rt; (iii) POCl3, pyridine, 0 °C to rt; (iv) TFA, DCM, 0 °C to rt; (v) K2CO3, CNBr, THF, 0 °C to rt.

[0236] Project (i) Ethyl 4-(5-cyano-2-methoxyphenyl)oxazole-2-carboxylate To a stirred solution of ethyl 5-(5-cyano-2-methoxyphenyl)oxazole-2-carboxylate (3.5 g, 13.83 mmol) and methyl oxamate (CAS 617-36-7, from Sigma-Aldrich, 4.86 g, 41.51 mmol) in EtOAc (35 mL) was added silver triflate (CAS 2923-28-6, from Combi-blocks, 10.66 g, 41.51 mmol) at room temperature. The mixture was heated at 80 °C for 24 h, then poured into water (120 mL), filtered through Celite Hyflow™, and the filtrate was extracted with EtOAc (3 × 70 mL). The combined organic phase was dried over anhydrous NaSO and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 16% EtOAc in n-hexane) to give ethyl 4-(5-cyano-2-methoxyphenyl) oxazole-2-carboxylate (0.53 g, 1.95 mmol, 14% yield). LCMS: Method C, 1.65 min; MS: ES+ 273.1.

[0237] Project (ii) 4-(5-cyano-2-methoxyphenyl)oxazole-2-carboxylic acid To a stirred solution of ethyl 4-(5-cyano-2-methoxyphenyl)oxazole-2-carboxylate (0.50 g, 1.84 mmol) in THF:water (1:1, 10 mL) was added lithium hydroxide monohydrate (0.23 g, 5.51 mmol) in small portions at 0 °C. The mixture was stirred at 0 °C for 2 h, then poured into water (70 mL), acidified with 1 N HCl, and extracted with DCM (3 × 70 mL). The combined organic phases were dried over anhydrous NaSO and concentrated under reduced pressure to give 4-(5-cyano-2-methoxyphenyl)oxazole-2-carboxylic acid (0.15 g, 0.61 mmol, 33% yield). LCMS: Method C2, 0.81 min; MS: ES+ 245.1.

[0238] Project (iii) tert-Butyl (2S,4R)-4-(4-(5-cyano-2-methoxyphenyl)oxazole-2-carboxamido)-2-(methoxymethyl)pyrrolidine-1-carboxylate To a stirred solution of 4-(5-cyano-2-methoxyphenyl)oxazole-2-carboxylic acid (0.16 g, 0.65 mmol) and tert-butyl (2S,4R)-4-amino-2-(methoxymethyl)pyrrolidine-1-carboxylate (CAS 1207853-53-9, 0.15 g, 0.65 mmol) in pyridine (3 mL) was added POCl (0.3 g, 0.18 mL, 1.97 mmol) dropwise at 0 °C. The mixture was stirred at 0 °C for 10 min, then poured into water (50 mL) and extracted with EtOAc (2 × 30 mL). The combined organic phase was dried over anhydrous NaSO and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 22% EtOAc in n-hexane) to give tert-butyl (2S,4R)-4-(4-(5-cyano-2-methoxyphenyl)oxazole-2-carboxamido)-2-(methoxymethyl)pyrrolidine-1-carboxylate (0.18 g, 0.39 mmol, 60% yield). LCMS: Method C, 1.70 min; MS: ES- 455.1.

[0239] Engineering (iv) 4-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of tert-butyl (2S,4R)-4-(4-(5-cyano-2-methoxyphenyl)oxazole-2-carboxamide)-2-(methoxymethyl)pyrrolidine-1-carboxylate (0.17 g, 0.37 mmol) in DCM (5 mL) was added TFA (0.85 mL, 5 vol) dropwise at 0° C. The mixture was allowed to warm to room temperature and stirred for 1 hour, then concentrated under reduced pressure to give 4-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.25 g, quantitative yield). LCMS: Method C, 1.32 min; MS: ES+ 357.1.

[0240] Engineering (v) 4-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of 4-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.24 g, 0.51 mmol) in THF (5 mL) was added K2CO3 (0.21 g, 1.53 mmol) at room temperature and stirred for 10 min. Cyanogen bromide (0.04 g, 0.41 mmol) was added at 0 °C. The mixture was stirred at room temperature for 1 h, then poured into water (50 mL) and extracted with EtOAc (3 × 50 mL). The combined organic phase was dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 71% EtOAc in n-hexane) to provide 4-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide (0.05 g, 0.14 mmol, 36% yield in two steps). LCMS: Method H, 2.80 min; MS: ES- 380.1; 1 H NMR (400 MHz, DMSO-d6) δ ppm: 9.25 (d, J = 6.8 Hz, 1H), 8.72 (s, 1H), 8.44 (d, J = 1.6 Hz, 1H), 7.89 (dd, J = 8.8, 2.0 Hz, 1H), 7.37 (d, J = 8.8 Hz, 1H), 4.47 - 4.58 (m, 1H), 4.07 (s, 3H), 4.00 - 4.18 (m, 1H), 3.69 - 3.73 (m, 1H), 3.43 - 3.50 (m, 3H), 3.34 (s, 3H), 2.09 - 2.19 (m, 1H), 1.96 - 2.02 (m, 1H). Chiral SFC: Method Y3, 3.49 min.

[0241] Example 7 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide

[0242] [ka]

[0243] (i) TBD, THF, 0 °C to rt; (ii) Zn(CN)2, Zn, Pd2(dba)3, phosphine ligand, DMA, 140 °C; (iii) TFA, DCM, 0 °C to rt; (iv) K2CO3, CNBr, THF, 0 °C to rt.

[0244] Project (i) tert-Butyl (2S,4R)-4-(5-(5-bromo-2-(trifluoromethoxy)phenyl)oxazole-2-carboxamido)-2-(methoxymethyl)pyrrolidine-1-carboxylate To a stirred solution of ethyl 5-(5-bromo-2-(trifluoromethoxy)phenyl)oxazole-2-carboxylate (0.70 g, 1.84 mmol) and tert-butyl (2S,4R)-4-amino-2-(methoxymethyl)pyrrolidine-1-carboxylate (CAS 1207853-53-9, 0.42 g, 1.84 mmol) in THF (8 mL) was added TBD (0.38 g, 2.77 mmol) in small portions at 0 °C. The mixture was stirred at 0 °C for 2 h, then poured into water (100 mL) and extracted with EtOAc (3 × 70 mL). The combined organic phase was dried over anhydrous NaSO and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 25% EtOAc in n-hexane) to give tert-butyl (2S,4R)-4-(5-(5-bromo-2-(trifluoromethoxy)phenyl)oxazole-2-carboxamido)-2-(methoxymethyl)pyrrolidine-1-carboxylate (0.37 g, 0.66 mmol, 35% yield). LCMS: Method C, 2.00 min; MS: ES+ 564.4, 566.4.

[0245] Project (ii) tert-Butyl (2S,4R)-4-(5-(5-cyano-2-(trifluoromethoxy)phenyl)oxazole-2-carboxamido)-2-(methoxymethyl)pyrrolidine-1-carboxylate To a stirred solution of tert-butyl (2S,4R)-4-(5-(5-bromo-2-(trifluoromethoxy)phenyl)oxazole-2-carboxamido)-2-(methoxymethyl)pyrrolidine-1-carboxylate (0.35 g, 0.62 mmol) in DMA (3 mL) was added zinc cyanide (0.18 g, 1.55 mmol), zinc dust (0.02 g, 0.31 mmol), and 1,1′-ferrocenediyl-bis(diphenylphosphine) (0.07 g, 0.12 mmol) at room temperature. The mixture was degassed with N for 15 minutes, and then tris(dibenzylideneacetone)dipalladium(0) (0.11 g, 0.12 mmol) was added. The mixture was heated at 140° C. in a microwave oven for 1 h, then allowed to cool to room temperature, poured into ice-cold water (70 mL), and extracted with EtOAc (3×70 mL). The combined organic phases were dried over NaSO and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 22% EtOAc in n-hexane) to give tert-butyl (2S,4R)-4-(5-(5-cyano-2-(trifluoromethoxy)-phenyl)oxazole-2-carboxamido)-2-(methoxymethyl)pyrrolidine-1-carboxylate (0.25 g, 0.5 mmol, 80% yield). LCMS: Method C, 1.83 min; MS: ES+: 511.4.

[0246] Project (iii) 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5S)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of tert-butyl (2S,4R)-4-(5-(5-cyano-2-(trifluoromethoxy)phenyl)oxazole-2-carboxamide)-2-(methoxymethyl)pyrrolidine-1-carboxylate (0.25 g, 0.49 mmol) in DCM (3 mL) was added TFA (1.25 mL, 5 vol) dropwise at 0° C. The mixture was allowed to warm to room temperature and stirred for 2 hours, then concentrated under reduced pressure to provide 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5S)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.39 g, quantitative yield). LCMS: Method C, 1.38 min; MS: ES+ 411.1.

[0247] Engineering (iv) 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5S)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.39 g, 0.74 mmol) in THF (5 mL) was added K2CO3 (0.31 g, 2.23 mmol) at room temperature and stirred for 10 min. Cyanogen bromide (0.06 g, 0.59 mmol) was added at 0 °C. The mixture was stirred at room temperature for 1 h, then poured into water (70 mL) and extracted with EtOAc (3 x 70 mL). The combined organic phases were dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 40% EtOAc in n-hexane) to give 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide (0.14 g, 0.33 mmol, 66% yield in two steps). LCMS: Method H, 3.03 min; MS: ES- 434.1;1 H NMR (400 MHz, DMSO-d6) δ ppm: 9.40 (d, J = 6.8 Hz, 1H), 8.51 (d, J = 1.6 Hz, 1H), 8.12 (dd, J = 8.8, 2.0 Hz, 1H), 7.86 (s, 1H), 7.82 (d, J = 8.4 Hz, 1H), 4.51 - 4.55 (m, 1H), 3.99 - 4.08 (m, 1H), 3.69 - 3.74 (m, 1H), 3.42 - 3.51 (m, 3H), 3.34 (s, 3H), 2.11 - 2.17 (m, 1H), 1.97 - 2.03 (m, 1H). Chiral SFC: Method Y4, 3.57 min.

[0248] Example 8 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide

[0249] [ka]

[0250] (i) TBD, THF, 0 °C to rt; (ii) Zn(CN)2, Zn, Pd2(dba)3, phosphine ligand, DMA, 140 °C; (iii) TFA, DCM, 0 °C to rt; (iv) K2CO3, CNBr, THF, 0 °C to rt.

[0251] Project (i) tert-Butyl (2R,4R)-4-(5-(5-bromo-2-(trifluoromethoxy)phenyl)oxazole-2-carboxamido)-2-methylpyrrolidine-1-carboxylate To a stirred solution of ethyl 5-(5-bromo-2-(trifluoromethoxy)phenyl)oxazole-2-carboxylate (0.40 g, 1.05 mmol) and tert-butyl (2R,4R)-4-amino-2-methylpyrrolidine-1-carboxylate (CAS 348165-63-9, 0.23 g, 1.16 mmol) in THF (8 mL) was added TBD (0.22 g, 1.57 mmol) in small portions at 0 °C. The mixture was warmed to room temperature and stirred for 3 h, then poured into water (50 mL) and extracted with EtOAc (3 × 60 mL). The combined organic phase was dried over anhydrous NaSO and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 32% EtOAc in n-hexane) to give tert-butyl (2R,4R)-4-(5-(5-bromo-2-(trifluoromethoxy)phenyl)oxazole-2-carboxamido)-2-methylpyrrolidine-1-carboxylate (0.14 g, 0.26 mmol, 24% yield). LCMS: Method C, 2.01 min; MS: ES+ 534.2, 536.2.

[0252] Project (ii) tert-Butyl (2R,4R)-4-(5-(5-cyano-2-(trifluoromethoxy)phenyl)oxazole-2-carboxamido)-2-methylpyrrolidine-1-carboxylate To a stirred solution of tert-butyl (2R,4R)-4-(5-(5-bromo-2-(trifluoromethoxy)phenyl)oxazole-2-carboxamido)-2-methylpyrrolidine-1-carboxylate (0.13 g, 0.24 mmol) in DMA (1.3 mL) was added zinc cyanide (0.07 g, 0.61 mmol), zinc dust (0.01 g, 0.12 mmol), and 1,1′-ferrocenediyl-bis(diphenylphosphine) (0.03 g, 0.05 mmol) at room temperature. The mixture was degassed with N for 10 minutes, and then tris(dibenzylideneacetone)dipalladium(0) (0.04 g, 0.05 mmol) was added. The mixture was heated at 140° C. in a microwave oven for 1 h, then cooled to room temperature, poured into water (20 mL), and extracted with EtOAc (3×30 mL). The combined organic phases were dried over NaSO and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 38% EtOAc in n-hexane) to give tert-butyl (2R,4R)-4-(5-(5-cyano-2-(trifluoromethoxy)phenyl)-oxazole-2-carboxamido)-2-methylpyrrolidine-1-carboxylate (0.05 g, 0.10 mmol, 42% yield). LCMS: Method C, 1.88 min; MS: ES+: 425.6 (M-56).

[0253] Project (iii) 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5R)-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of tert-butyl (2R,4R)-4-(5-(5-cyano-2-(trifluoromethoxy)phenyl)oxazole-2-carboxamide)-2-methylpyrrolidine-1-carboxylate (0.05 g, 0.1 mmol) in DCM (2 mL) was added TFA (0.15 mL, 3 vol) dropwise at 0° C. The mixture was allowed to warm to room temperature and stirred for 1 hour, then concentrated under reduced pressure to give 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5R)-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.08 g, quantitative yield). LCMS: Method C, 1.38 min; MS: ES+ 381.2.

[0254] Engineering (iv) 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5R)-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.08 g, 0.16 mmol) in THF (4 mL) was added K2CO3 (0.07 g, 0.48 mmol) at room temperature and stirred for 10 min. Cyanogen bromide (0.02 g, 0.16 mmol) was added at 0 °C. The mixture was stirred at room temperature for 1 h, then poured into water (20 mL) and extracted with EtOAc (3 × 25 mL). The combined organic phases were dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 47% EtOAc in n-hexane) to give 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide (0.02 g, 0.05 mmol, 47% yield in two steps). LCMS: Method H, 3.02 min; MS: ES- 404.0; 1H NMR (400 MHz, DMSO-d6) δ ppm: 9.43 (d, J = 6.8 Hz, 1H), 8.51 (s, 1H), 8.11 (d, J = 8.4 Hz, 1H), 7.84 (s, 1H), 7.82 (d, J = 8.4 Hz, 1H), 4.48 - 4.57 (m, 1H), 3.89 - 3.94 (m, 1H), 3.76 - 3.80 (m, 1H), 3.42 - 3.46 (m, 1H), 2.15 - 2.18 (m, 1H), 1.76 - 1.83 (m, 1H), 1.25 (d, J = 6.4 Hz, 3H). Chiral SFC: Method Y8, 5.02 min.

[0255] Example 9 5-(5-cyano-4-fluoro-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide

[0256] [ka]

[0257] (i) TBD, THF, 0 °C to rt; (ii) TFA, DCM, 0 °C to rt; (iii) K2CO3, CNBr, THF, 0 °C to rt.

[0258] Project (i) tert-Butyl (2R,4R)-4-(5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamido)-2-methylpyrrolidine-1-carboxylate To a stirred solution of ethyl 5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxylate (0.50 g, 1.72 mmol) and tert-butyl (2R,4R)-4-amino-2-methylpyrrolidine-1-carboxylate (CAS 348165-63-9, 0.27 g, 1.38 mmol) in THF (10 mL) was added TBD (0.29 g, 2.07 mmol) in small portions at 0 °C. The mixture was warmed to room temperature and stirred for 2 h, then poured into water (50 mL) and extracted with EtOAc (2 × 50 mL). The combined organic phase was dried over anhydrous NaSO and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 40% EtOAc in n-hexane) to give tert-butyl (2R,4R)-4-(5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamido)-2-methylpyrrolidine-1-carboxylate (0.3 g, 0.67 mmol, 38% yield). LCMS: Method C, 1.78 min; MS: ES+ 389.3 (M-56).

[0259] Project (ii) 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of tert-butyl (2R,4R)-4-(5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamide)-2-methylpyrrolidine-1-carboxylate (0.3 g, 0.67 mmol) in DCM (7 mL) was added TFA (0.9 mL, 3 vol) dropwise at 0° C. The mixture was allowed to warm to room temperature and stirred for 1 hour, then concentrated under reduced pressure to give 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.29 g, quantitative yield). LCMS: Method C, 1.39 min; MS: ES+ 345.3.

[0260] Project (iii) 5-(5-cyano-4-fluoro-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.29 g, 0.63 mmol) in THF (7 mL) was added KCO (0.26 g, 1.89 mmol) at room temperature and stirred for 10 min. Cyanogen bromide (0.07 g, 0.63 mmol) was added at 0 °C. The mixture was stirred at room temperature for 1 hour, then poured into water (70 mL), a precipitate formed, and the solid was collected by filtration under reduced pressure to give 5-(5-cyano-4-fluoro-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide (0.13 g, 0.35 mmol, 52% yield in two steps). LCMS: Method H, 2.75 min; MS: ES+ 370.1; 1 H NMR (400 MHz, DMSO-d6) δ ppm: 9.31 (d, J = 6.4 Hz, 1H), 8.28 - 8.31 (m, 1H), 7.79 - 7.82 (m, 1H), 7.49 - 7.52 (m, 1H), 4.47 - 4.59 (m, 1H), 4.08 (s, 3H), 3.85 - 3.99 (m, 1H), 3.76 - 3.80 (m, 1H), 3.42 - 3.45 (m, 1H), 2.12 - 2.23 (m, 1H), 1.73 - 1.87 (m, 1H), 1.24 -1.29 (m, 3H). Chiral SFC: Method Y5, 4.98 min.

[0261] Example 10 4-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide

[0262] [ka]

[0263] Project (i) Ethyl 4-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxylate To a stirred solution of 3-(2-bromoacetyl)-4-cyclopropoxybenzonitrile (0.70 g, 2.50 mmol) and methyl oxamate (CAS 617-36-7, from Sigma-Aldrich, 0.66 g, 3.75 mmol) in EtOAc (10 mL) was added silver triflate (CAS 2923-28-6, from Combi-blocks, 0.96 g, 3.75 mmol) at room temperature. The mixture was heated at 80 °C for 16 h. Another identical batch was combined, then poured into water (100 mL), filtered through Celite Hyflow™, and the filtrate was extracted with EtOAc (2 × 100 mL). The combined organic phase was dried over anhydrous NaSO and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 10% EtOAc in n-hexane) to give ethyl 4-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxylate (0.35 g, 1.17 mmol, 23% yield). LCMS: Method H1, 3.40 min; MS: ES+ 299.0.

[0264] Project (ii) tert-Butyl (2S,4R)-4-(4-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamido)-2-(methoxymethyl)pyrrolidine-1-carboxylate To a stirred solution of ethyl 4-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxylate (0.32 g, 1.07 mmol) and tert-butyl (2S,4R)-4-amino-2-(methoxymethyl)pyrrolidine-1-carboxylate (CAS 1207853-53-9, 0.25 g, 1.07 mmol) in DMF (7 mL) was added TBD (0.18 g, 1.28 mmol) in small portions at 0 °C. The mixture was warmed to room temperature and stirred for 2 h, then poured into water (50 mL) and extracted with EtOAc (2 × 50 mL). The combined organic phase was dried over anhydrous NaSO, filtered, and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 30% EtOAc in n-hexane) to give tert-butyl (2S,4R)-4-(4-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamido)-2-(methoxymethyl)pyrrolidine-1-carboxylate (0.10 g, 0.22 mmol, 20% yield). LCMS: Method H1, 3.51 min; MS: ES- 481.3.

[0265] Project (iii) 4-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt To a stirred solution of tert-butyl (2S,4R)-4-(4-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamide)-2-(methoxymethyl)pyrrolidine-1-carboxylate (0.1 g, 0.22 mmol) in DCM (5 mL) was added TFA (1 mL, 10 vol) dropwise at 0° C. The mixture was allowed to warm to room temperature and stirred for 1 hour, then concentrated under reduced pressure to give 4-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.14 g, quantitative yield). LCMS: Method J, 3.61 min; MS: ES+ 383.0.

[0266] Engineering (iv) 4-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of 4-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.14 g, 0.28 mmol) in THF (5 mL) was added K2CO3 (0.12 g, 0.84 mmol) at room temperature and stirred for 5 min. Cyanogen bromide (0.03 g, 0.28 mmol) was added at 0 °C. The mixture was stirred at room temperature for 1 h, then poured into water (30 mL) and extracted with EtOAc (2 × 30 mL). The combined organic phases were dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 45% EtOAc in n-hexane) to give 4-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide (0.04 g, 0.09 mmol, 47% yield in two steps). LCMS: Method H1, 2.97 min; MS: ES- 406.2; 1 H NMR (400 MHz, DMSO-d6) δ ppm: 9.24 (d, J = 6.8 Hz, 1H), 8.52 (s, 1H), 8.43 (d, J = 2.0 Hz, 1H), 7.92 (dd, J = 8.8, 2.0 Hz, 1H), 7.64 (d, J = 8.4 Hz, 1H), 4.48 - 4.56 (m, 1H), 4.15 - 4.23 (m, 1H), 4.0 - 4.08 (m, 1H), 3.68 - 3.72 (m, 1H), 3.42 - 3.50 (m, 3H), 3.32 (s, 3H), 2.10 - 2.15 (m, 1H), 1.96 - 2.01 (m, 1H), 0.86 - 0.97 (m, 4H). Chiral HPLC: Method Y15, 10.0 min.

[0267] Example 11 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide

[0268] [ka]

[0269] Project (i) tert-Butyl (2R,4R)-4-(5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamido)-2-(methoxymethyl)pyrrolidine-1-carboxylate To a stirred solution of ethyl 5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxylate (0.25 g, 0.83 mmol) and tert-butyl (2R,4R)-4-amino-2-(methoxymethyl)pyrrolidine-1-carboxylate (CAS 1123305-98-5, 0.19 g, 0.83 mmol) in toluene (5 mL) was added TBD (0.12 g, 0.83 mmol) in small portions at 0 °C. The mixture was warmed to room temperature and stirred for 1 h, then poured into water (50 mL) and extracted with EtOAc (2 × 50 mL). The combined organic phase was dried over anhydrous NaSO, filtered, and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 35% EtOAc in n-hexane) to give tert-butyl (2R,4R)-4-(5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamido)-2-(methoxymethyl)pyrrolidine-1-carboxylate (0.08 g, 0.16 mmol, 19% yield). LCMS: Method C1, 1.41 min; MS: ES+ 483.4.

[0270] Project (ii) 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt To a stirred solution of tert-butyl (2R,4R)-4-(5-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamide)-2-(methoxymethyl)pyrrolidine-1-carboxylate (0.07 g, 0.15 mmol) in DCM (1 mL) was added TFA (0.22 mL, 3 vol) dropwise at 0° C. The mixture was allowed to warm to room temperature and stirred for 1 hour, then concentrated under reduced pressure to give 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.10 g, quantitative yield). LCMS: Method C1, 1.06 min; MS: ES+ 383.2.

[0271] Project (iii) 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.10 g, 0.20 mmol) in THF (5 mL) was added K2CO3 (0.08 g, 0.60 mmol) at room temperature and stirred for 5 min. Cyanogen bromide (0.02 g, 0.20 mmol) was added at 0 °C. The mixture was stirred at room temperature for 1 h, then poured into water (50 mL) and extracted with EtOAc (2 × 50 mL). The combined organic phase was dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 45% EtOAc in n-hexane) to give 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide (0.02 g, 0.04 mmol, 29% yield in two steps). LCMS: Method H1, 2.94 min; MS: ES+ 408.2; 1 H NMR (400 MHz, DMSO-d6) δ ppm: 9.18 (d, J = 6.8 Hz, 1H), 8.24 (s, 1H), 7.96 (d, J = 8.4 Hz, 1H), 7.67 - 7.69 (m, 2H), 4.50 - 4.61 (m, 1H), 4.15 - 4.25 (m, 1H), 3.88 - 3.97 (m, 1H), 3.63 - 3.72 (m, 1H), 3.46 - 3.60 (m, 2H), 3.37 - 3.41 (m, 4H), 2.27 - 2.38 (m, 1H), 1.80 - 1.90 (m, 1H), 0.83 - 0.98 (m, 4H). Chiral SFC: Method Y12, 3.53 min.

[0272] Example 12 4-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide

[0273] [ka]

[0274] Project (i) tert-Butyl (2R,4R)-4-(4-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamido)-2-(methoxymethyl)pyrrolidine-1-carboxylate To a stirred solution of ethyl 4-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxylate (0.40 g, 1.34 mmol) [Example 10; Step (i)] and tert-butyl (2R,4R)-4-amino-2-(methoxymethyl)pyrrolidine-1-carboxylate (CAS 1123305-98-5, 0.31 g, 1.34 mmol) in toluene (5 mL) was added TBD (0.19 g, 1.34 mmol) in small portions at 0 °C. The mixture was warmed to room temperature and stirred for 4 h, then poured into water (50 mL) and extracted with EtOAc (2 × 50 mL). The combined organic phase was dried over anhydrous NaSO, filtered, and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 40% EtOAc in n-hexane) to give tert-butyl (2R,4R)-4-(4-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamide)-2-(methoxymethyl)pyrrolidine-1-carboxylate (0.22 g, 0.46 mmol, 34% yield). LCMS: Method C1, 1.42 min, MS: ES+ 483.3.

[0275] Project (ii) 4-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt To a stirred solution of tert-butyl (2R,4R)-4-(4-(5-cyano-2-cyclopropoxyphenyl)oxazole-2-carboxamide)-2-(methoxymethyl)pyrrolidine-1-carboxylate (0.22 g, 0.46 mmol) in DCM (5 mL) was added TFA (0.66 mL, 3 vol) dropwise at 0° C. The mixture was allowed to warm to room temperature and stirred for 1 hour, then concentrated under reduced pressure to give 4-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.28 g, quantitative yield). LCMS: Method C1, 1.09 min. MS: ES+ 383.2.

[0276] Project (iii) 4-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide To a stirred solution of 4-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide TFA salt (0.28 g, 0.56 mmol) in THF (5 mL) was added K2CO3 (0.23 g, 1.69 mmol) at room temperature and stirred for 5 min. Cyanogen bromide (0.06 g, 0.56 mmol) was added at 0 °C. The mixture was stirred at room temperature for 1 h, then poured into water (50 mL) and extracted with EtOAc (2 x 50 mL). The combined organic phases were dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 65% EtOAc in n-hexane) to give 4-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide (0.06 g, 0.14 mmol, 31% yield in two steps). LCMS: Method H1, 3.01 min; MS: ES+ 408.2; 1H NMR (400 MHz, DMSO-d6) δ ppm: 9.10 (d, J = 7.6 Hz, 1H), 8.52 (s, 1H), 8.38 (s, 1H), 7.91 (d, J = 8.4, 2.0 Hz, 1H), 7.64 (d, J = 8.8 Hz, 1H), 4.51 - 4.56 (m, 1H), 4.16 - 4.23 (m, 1H), 3.88 - 3.97 (m, 1H), 3.63 - 3.67 (m, 1H), 3.46 - 3.56 (m, 2H), 3.38 - 3.41 (m, 4H), 2.29 - 2.36 (m, 1H), 1.83 - 1.90 (m, 1H), 0.92 (br s, 4H). Chiral SFC: Method Y16, 5.11 min.

[0277] Example 13 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide

[0278] [ka]

[0279] The title compound was prepared in a manner analogous to Example 1, using tert-butyl (2S,4R)-4-amino-2-methylpyrrolidine-1-carboxylate (CAS 708274-46-8) in step (i).

[0280] Example 14 5-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide

[0281] [ka]

[0282] The title compound was prepared in a manner analogous to Example 4, using tert-butyl (2S,4R)-4-amino-2-methylpyrrolidine-1-carboxylate (CAS 708274-46-8) in step (i).

[0283] Example 15 5-(5-cyano-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide

[0284] [ka]

[0285] The title compound was prepared in a manner analogous to Example 5, using tert-butyl (2R,4R)-4-amino-2-(methoxymethyl)pyrrolidine-1-carboxylate (CAS 1123305-98-5) in step (i).

[0286] Example 16 4-(5-cyano-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide

[0287] [ka]

[0288] The title compound was prepared in a manner similar to Example 6, using tert-butyl (2R,4R)-4-amino-2-(methoxymethyl)pyrrolidine-1-carboxylate (CAS 1123305-98-5) in step (iii).

[0289] Example 17 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide

[0290] [ka]

[0291] The title compound was prepared in a manner analogous to Example 7, using tert-butyl (2R,4R)-4-amino-2-(methoxymethyl)pyrrolidine-1-carboxylate (CAS 1123305-98-5) in step (i).

[0292] Example 18 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5S)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide

[0293] [ka]

[0294] The title compound was prepared in a manner analogous to Example 8, using tert-butyl (2S,4R)-4-amino-2-methylpyrrolidine-1-carboxylate (CAS 708274-46-8) in step (i).

[0295] Example 19 5-(5-cyano-4-fluoro-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide

[0296] [ka]

[0297] The title compound was prepared in a manner analogous to Example 9, using tert-butyl (2S,4R)-4-amino-2-methylpyrrolidine-1-carboxylate (CAS 708274-46-8) in step (i).

[0298] Biological activity of this compound Abbreviation: TAMRA carboxytetramethylrhodamine PCR polymerase chain reaction PBS Phosphate-buffered saline EDTA Ethylenediaminetetraacetic acid Tris 2-amino-2-(hydroxymethyl)-1,3-propanediol NP-40 Nonidet P-40, Octylphenoxypolyethoxyethanol BSA Bovine serum albumin PNS Peripheral Nervous System BH3 Bcl-2 homology domain 3 PTEN phosphatase and tensin homolog SDS-PAGE sodium dodecyl sulfate polyacrylamide gel electrophoresis DMSO dimethyl sulfoxide YFP Yellow Fluorescent Protein VME Vinyl Methyl Ester HA hemagglutinin Ahx aminohexanoic acid

[0299] USP30 Biochemistry IC 50 Assay Dilution plates were prepared at 21x the final concentration (2100 μM for a final concentration of 100 μM) in 50% DMSO in 96-well polypropylene V-bottom plates (Greiner #651201). A typical 8-point dilution series resulted in final concentrations of 100, 30, 10, 3, 1, 0.3, 0.1, and 0.03 μM. Reactions were performed in duplicate in a black 384-well plate (small volume, Greiner 784076) with a final reaction volume of 21 μl. 1 μl of either 50% DMSO or diluted compound was added to the plate. USP30 (Boston Biochem #E582) was diluted in reaction buffer (40 mM Tris, pH 7.5, 0.005% Tween 20, 0.5 mg / ml BSA, 5 mM β-mercaptoethanol) to give a final assay concentration of 4 nM, and 10 μl of diluted USP30 was added to the compound. The enzyme and compound were incubated at room temperature for 30 minutes. The reaction was initiated by the addition of 50 nM of TAMRA-labeled peptide linked to ubiquitin via an isopeptide bond as a fluorescence polarization substrate. The reaction was read immediately after the addition of the substrate and after a 2-hour incubation at room temperature. Readings were performed on a Pherastar Plus (BMG Labtech). λ excitation 540 nm; λ emission 590 nm.

[0300] [Table 19]

[0301] Reference Example [Table 20]

[0302] Off-target pharmacology Example 2 underwent pharmacological profiling in the Eurofins CEREP Safety Screen44 panel. At a single concentration of 10 μM, less than 50% inhibition of binding or enzymatic activity was observed for all targets in the panel. Example 1 has low affinity for its targets in this assay, making it less likely to have off-target interactions.

[0303] Safety Pharmacology Example 2 was evaluated for its effect on the hERG potassium channel in stably expressed CHO cells at concentrations ranging from 0.01 to 30 μM. Example 2 shows that 30 μM produces a maximal inhibition of 35% of the hERG current amplitude and has little tendency to affect the QT interval.

[0304] Genetic Toxicology Example 2 was evaluated in bacterial reverse mutation assays (Ames) and in vitro micronucleus assays. All in vivo tests were performed with or without exogenous metabolic activation, using concentrations up to those limited by cytotoxicity or insolubility. Example 2 did not induce mutations when tested up to 5000 μg / plate in reverse mutation assays in Salmonella typhimurium strains TA98, TA100, TA1535, and TA97a and E. coli strain WP2 uvrA pKM101, with or without metabolic activation. Induction of chromosomal damage was assessed using an in vitro micronucleus assay in TK6 cells. Example 2 was negative for induction of micronuclei when incubated for 3 hours in the presence of exogenous metabolic activation followed by 27 hours of recovery, and also when incubated for 27 hours in the absence of exogenous metabolic activation followed by 27 hours of recovery.

[0305] USP30 endogenous cellular target engagement assay HeLa cells stably overexpressing YFP-Parkin were seeded in six-well dishes. Once attached, cells were treated with the appropriate concentration of test compound or vehicle control for 1 h at 37°C and 5% CO2. Whole cell lysates were prepared by scraping cells into cold PBS, centrifuging, and lysing them in lysis buffer (50 mM Tris-base, pH 7.5, 50 mM NaCl, 1% NP-40 / Igepal CA-630, 2 mM MgCl2, 10% glycerol, 5 mM β-mercaptoethanol, Complete Mini Tablets EDTA-free (Roche), PhosStop tablets (Roche)) for 10 min. 20 μg of protein equivalent from the clarified cell lysate was incubated with the HA-Ahx-Ahx-Ub-VME probe at a final concentration of 2.5 μM at room temperature. The reaction was stopped by adding 5x SDS sample loading buffer, and proteins were separated by SDS-PAGE and Western blotting. USP30 was detected using anti-USP30 sheep S746D antibody (MRC PPU Reagents and Services) and rabbit anti-sheep secondary IgG (H+L) horseradish peroxidase conjugate (Thermo #31480), and visualized on a GE LAS4000 imager using ECL reagents (GE #RPN2109). Target engagement was measured by quantification of bands corresponding to USP30 and USP30 bound to the Ub-VME probe and the ratio of expression to that of vehicle-treated controls.

[0306] TOM20 ubiquitination assay Human cell lines can be challenged with mitochondrial depolarizing agents (ionophores (e.g., CCCP, valinomycin) or mitochondrial complex inhibitors (oligomycin, antimycin A)) to induce TOM20 ubiquitination, which is then further enhanced in the presence of USP30 inhibitors. TOM20 ubiquitination is then assessed by Western blotting of cell lysates. Detection of TOM20 ubiquitin adducts is enabled by an 8 kDa molecular weight increase per molecule of ubiquitin added, resulting in a ladder of TOM20 immunoreactive bands. TOM20 ubiquitination levels can be quantified using chemiluminescence densitometry of the ladder immunoreactive bands.

[0307] In vitro cytotoxicity (Cell Tox): The assay endpoint was measured in HCT116 human colorectal cancer cells using alamarBlue™. Compound cytotoxicity was measured over a 96 hour continuous exposure period to the compounds.

[0308] Further research log P: Partition coefficient; lipophilicity measure log D: partition coefficient; lipophilicity measurement TPSA: Topological Polar Surface Area Turbidimetric solubility: Test compound solutions prepared in DMSO diluted in aqueous buffer. Turbidity measurements are used as endpoints by measuring absorbance at 620 nm. FaSSIF: Fasted simulated intestinal fluid measured at pH 6.5 Hep Cl Mouse: In vitro hepatocyte clearance in mouse cells Hep Cl Human: In vitro hepatocyte clearance in human cells Plasma fu,p: Free fraction of compounds in plasma preparations determined by in vitro equilibrium dialysis Brain fu,br: Free fraction of compounds in brain homogenate preparations determined by in vitro equilibrium dialysis Clu: In vitro clearance. Clu, as defined herein, is the scaled clearance and is calculated from the intrinsic clearance. The intrinsic clearance is the predicted clearance due to hepatic metabolic reactions, determined from incubation of the compound in hepatocyte preparations. The lower the mL / min / kg value, the more stable the compound. Cl in vivo clearance: A pharmacokinetic measure of the volume of plasma (or any matrix) from which a substance is completely cleared per unit time. The lower the value in mL / min / kg, the more stable the compound. Oral F: Oral bioavailability MDR1-MDCK (Madin-Darby canine kidney cell monolayer) in vitro flux assay WT-MDCK (wild type) in vitro flux Kpuu is the ratio of unbound drug in the brain to unbound drug in the plasma and can be an indicator of the potential to treat peripheral and / or CNS indications.

[0309] [Table 21]

[0310] [Table 22]

[0311] The Examples possess beneficial properties that indicate their potential superiority over other compounds. For example, the observed IV plasma clearances of 41 and 20 mL / min / kg, respectively, as measured in mice for Examples 1 and 2, are low, indicating valuable plasma stability, and the compounds have very good oral bioavailability of 37 and 47%, respectively.

[0312] [Table 23]

[0313] [Table 24]

[0314] [Table 25]

[0315] [Table 26]

[0316] [Table 27]

[0317] Comparison Data Reference Examples A, B, C, D, and E are known DUB inhibitors identified as active as inhibitors of USP30 and share some structural similarity with the compounds of the present invention, which have cyanamide structural features. Reference Examples B, C, D, and E are disclosed in WO2016 / 046530 as having UCHL1 inhibitory activity.

[0318] USP30 efficacy Examples 1-11 of the present invention are significantly more potent against USP 30 than Reference Examples A, B, C, D, and E, as measured by biochemical assays. Example 12 is significantly more potent against USP 30 than Reference Examples B, C, D, and E. For example, Examples 1-11 are 6.8-34 times more potent than Reference Example A, 16-155 times more potent than Reference Examples B, C, and D, and at least 440 times more potent than Reference Example E.

[0319] Selectivity of USP30 over other DUBs The data provided demonstrate that Examples 1, 2, 4, 6, 7, 11, and 12 are significantly more selective for USP 30 than for nine DUBs (USP 2, USP 6, USP 10, USP 15, USP 16, USP 21, USP 25, USP 28, and USP 46) compared to Reference Example A. These Examples are at least 110-7820 times more potent against USP 30 than against each of the nine DUBs (Example 7 was not tested against USP 15). This is a significant selectivity advantage over Reference Example A, being 2.2 times more potent.

[0320] Selectivity of USP30 for UCHL1 The data provided show that Examples 1, 2, 4, 6, 7, 11 and 12 are significantly more selective for USP30 over UCHL1 compared to Reference Examples B, C, D and E. Examples 1, 2, 4, 6, 7, 11 are over 30,000 times more potent for USP30 than UCHL1, and Example 12 is over 4,412 times more potent, whereas Reference Examples B, C, D and E are only 3.0, 1.3, 0.8 and 1.5 times more potent, respectively, with C being more selective for UCHL1.

[0321] Selectivity of USP30 for cathepsins B, K, L, S, and V The data provided show that Examples 1, 2, 4, 6, 7, 11, and 12 are significantly more selective for USP30 over cathepsins (B, K, L, S, and V) compared to Reference Example A. These Examples are over 1540-fold more potent against USP30 than against each cathepsin, a significant selectivity advantage over Reference Example A. Reference Example A is only 11.6-fold more potent against cathepsin K. The above-identified advantages of the compounds of the present invention over the prior art references are both significant and unexpected, and alone, and especially in combination, these advantages make the compounds of the present invention particularly suitable for use in the treatment or prevention of diseases associated with USP30 activity.

[0322] Preclinical in vivo models (a) A modified human renal proximal tubule cell cisplatin injury model for testing renal protective agents. Primary human PTCs are isolated from rejected kidney transplants. Dissociated cells were plated onto 96-well Transwell inserts (surface area = 0.143 cm). 2 ) were seeded at a density of 20,000 cells / insert. The medium was refreshed 24 hours after initial seeding and on days 3 and 5 of culture. The monolayers were measured to a resistance of 80-120 Ω.cm before use in experiments. 2 The results showed a TEER in the range of . Human PTC monolayers were pre-exposed to Example 2 (0.01, 0.03, 0.1, 0.3, 1.0, and 3.0 μM) for 1 hour on both the apical and basolateral sides of the cell layer before exposing the cells to cisplatin (20 μM). Subsequently, the cells were exposed to both Example 2 (0.01, 0.03, 0.1, 0.3, 1.0, and 3.0 μM) and cisplatin (20 μM) for 24 and 48 hours on both the apical and basolateral sides. Additionally, negative controls (Example 2 only, 0.5% DMSO in Example 2, 0.2% dimethylformamide (DMF) for cisplatin vehicle control), positive controls (cisplatin 20 μM), and cisplatin plus a cisplatin uptake inhibitor (dolutegravir 100 μM) were also included in parallel. Experiments were performed in triplicate on human cell monolayers (n=3) and three different donors (N=3). Cell viability assays based on ATP production and LDH release were performed on monolayers treated with test compounds performed in 96-well Transwell inserts. Example 2 protected against cisplatin-induced ATP loss in a concentration-dependent manner. Example 2 also reduced the cisplatin-induced increase in LDH. These data indicate that Example 2 can protect against cisplatin-induced toxicity in human proximal tubule epithelial cells.

[0323] Preclinical in vivo models The compounds of the invention may be tested for efficacy in representative in vivo disease models using standard research procedures from the published literature, such as, for example: (a) The bleomycin-induced pulmonary fibrosis model, which is the main preclinical in vivo model of idiopathic pulmonary fibrosis [Kobayashi et al, 2016, J Immunol, 197(2):504-516]. (b) Diet-induced model of NAFLD and glucose homeostasis [Nishida et al, 2013, Lab Invest; Feb;93(2):230-41], (c) The MPTP model of Parkinson's disease, a commonly used paradigm to investigate neurodegeneration of the brain's dopaminergic system caused by chemically induced mitochondrial dysfunction [Karuppagouner et al, 2014, Sci Rep. 2014 May 2;4:4874]. (d) Ndufs4KO Leigh syndrome model [Kruse et al, 2008, Cell Metab. Apr;7(4):312-20], (e) Aged rodent models: effects on hippocampus, cognition, and motor function [Kobilo et al, 2014, Learn Mem. Jan 17;21(2):119-26; Creed et al, 2019, Neuroscience. Jun 15;409:169-179; Van Skike et al, 2020, Aging Cell. 19; e13057] (f) Unilateral ureteral obstruction (UUO) model [Chevalier et al., 2009, Kidney Int 75(11): 1145-1152] Includes:

[0324] UUO causes renal injury characterized by tubular cell damage, interstitial inflammation, and fibrosis. It serves as a model for irreversible postrenal acute kidney injury (AKI). Experimental UUO illustrates the molecular mechanisms of apoptosis, inflammation, and fibrosis, all of which are important processes in renal injury, regardless of the primary insult. Consequently, the UUO model provides researchers with information beyond injury (Chevalier et al., 2009, Kidney Int 75(11): 1145-1152). Example 2 was evaluated in a UUO model to determine the compound's ability to reduce progressive tubulointerstitial fibrosis and chronic kidney disease (CKD). On study day 1, adult C57BL / 6 mice were orally gavaged according to one of the following dosing regimens: vehicle, 5, or 1.5 mg / kg twice daily. Two hours after dosing on study day 1, mice underwent surgery to ligate the left ureter at two points. Successful UUO surgery was later confirmed by the observation of dilated renal pelvis due to hydronephrosis. Animals were administered the prescribed regimen for 10 days, at which point kidneys were harvested or for histopathological and protein / RNA evaluation. Picrosirius red staining was performed to assess the degree of collagen deposition, and IHC was used to evaluate relative α-smooth muscle actin (αSMA) expression. Results showed that administration of 5 and 1.5 mg / kg of Example 2 (po) BID statistically reduced collagen deposition as evidenced by decreased picrosirius red staining in ligated kidneys. Evaluation of α-SMA staining revealed that oral administration of 5 and 1.5 mg / kg of Example 2 BID resulted in a statistically reduced α-SMA levels in UUO-injured kidneys when compared to vehicle-treated controls. (g) AKI can be induced by bilateral renal pedicle clamping, which results in ischemia-reperfusion injury (IRI), causing tubular damage and inflammation that can lead to severe loss of renal function [Lu et al. 2012. J Nephrol. 25 (5): 738-45].

[0325] Prophylaxis: Example 2 was administered to C57BL / 6 mice at 5 and 1.5 mg / kg BID (po) and compared to vehicle treatment from day -1 to day +21. On day 0, mice were anesthetized and the left renal pedicle was clamped for 45 minutes and then released to induce IRI. On day +21, kidneys were harvested. Morphology and fibrosis were evaluated. Body weight was similar between groups and remained constant throughout the observation period. Masson's trichome staining showed significantly less tubular atrophy and collagen content in the outer medulla from Example 2-treated animals (both dose levels) on day +21. Fibronectin expression in the cortex and outer medulla was significantly reduced in Example 2-treated animals (both dose levels) on day +21. Example 2 demonstrated efficacy in this model of IR-induced CKD, with daily treatment showing significant benefits in attenuating tubular atrophy and reducing fibrosis.

[0326] Post-injury administration: On day 0 (zero), C57BL / 6 mice were anesthetized, and the left renal pedicle was clamped for 45 minutes and released to induce IRI. Mice were then administered either vehicle or Example 2: 5 mg / kg (po) BID for 21 days. The first treatment began 5 hours after IRI surgery (i.e., administration of the therapeutic drug). Mice were monitored, and kidneys were harvested on day +21. Kidney sections were quantitatively evaluated for relative cellular morphology and fibrosis using a blinded histological scoring method. Body weight was similar between groups and remained constant throughout the observation period. Fibronectin staining in the cortex was significantly reduced in Ex. 2 treated mice on day +21. Example 2, when administered therapeutically, showed partial efficacy in this model of IR-induced CKD. Initiating treatment after the establishment of ischemia-reperfusion injury showed a significant benefit toward reducing cortical fibrosis.

[0327] Inventive paragraph The present invention is directed to the following: 1. Formula (I)(i) and Formula (I)(ii):

[0328] [ka]

[0329] (In the above formula, R 1is selected from (C1-C4) alkyl, (C1-C4) fluoroalkyl, and CH2OCH3; R 2 is selected from (C1-C4) alkyl, CF3, and cyclopropyl, and R 3 , R 4 and R 5 are each independently selected from hydrogen and halogen), a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer. 2.R 1 2. The compound of paragraph 1, wherein is selected from methyl, CH2F, CHF2, CF3, and CH2OCH3. 3.R 1 3. The compound of paragraph 2, wherein is selected from methyl and CH2OCH3. 4.R 2 4. The compound of any one of paragraphs 1 to 3, wherein is selected from methyl, CF3, and cyclopropyl. 5.R 2 5. The compound according to paragraph 4, wherein is selected from methyl and cyclopropyl. 6.R 3 6. The compound of any one of paragraphs 1 to 5, wherein is selected from hydrogen and fluorine. 7.R 3 7. The compound of paragraph 6, wherein is hydrogen. 8.R 4 and R 5 Paragraph 8. The compound of any one of paragraphs 1 to 7, wherein each is hydrogen.

[0330] 9. Formula (I)(i):

[0331] [ka] 9. The compound according to any one of paragraphs 1 to 8, or a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer, wherein 10. Formula (IA)(i):

[0332] [ka]

[0333] 10. The compound of paragraph 9, or a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer. 11. 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(fluoromethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, and 11. The compound according to paragraph 10 selected from 5-(5-cyano-4-fluoro-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, or a pharmaceutically acceptable salt thereof. 12. Formula (IB)(i):

[0334] [ka]

[0335] 10. The compound of paragraph 9, or a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer. 13. 4-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, and 4-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 13. The compound according to paragraph 12, selected from: or a pharmaceutically acceptable salt thereof. 14. Formula (I)(ii):

[0336] [ka]

[0337] 9. The compound according to any one of paragraphs 1 to 8, or a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer, wherein 15. Formula (IA)(ii):

[0338] [ka]

[0339] 15. The compound of paragraph 14, or a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer. 16. 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5S)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-4-fluoro-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, and 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-(fluoromethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 16. The compound according to paragraph 15, selected from: or a pharmaceutically acceptable salt thereof. 17. Formula (IB)(II):

[0340] [ka] 15. The compound of paragraph 14, or a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer. 18. 4-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, and 4-(5-cyano-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 18. The compound according to paragraph 17, selected from: or a pharmaceutically acceptable salt thereof. 19. A compound according to any one of paragraphs 1 to 18, a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer, for use as a pharmaceutical.

[0341] 20. A compound according to any one of paragraphs 1 to 18, a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer, for use in the treatment or prevention of a condition involving mitochondrial dysfunction, cancer, or fibrosis. 21. Use of a compound according to any one of paragraphs 1 to 18, or a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer, in the manufacture of a medicament for use in the treatment or prevention of a condition involving mitochondrial dysfunction, cancer, or fibrosis. 22. A method for the treatment or prevention of a condition involving mitochondrial dysfunction, cancer or fibrosis, comprising administering to a patient in need thereof an effective amount of a compound according to any one of paragraphs 1 to 18, a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer. 23. The conditions associated with mitochondrial dysfunction include CNS disorders, neurodegenerative diseases, Parkinson's disease, Alzheimer's disease, amyotrophic lateral sclerosis, Huntington's disease, ischemia, stroke, dementia with Lewy bodies, frontotemporal dementia, multiple sclerosis, mitochondrial encephalopathy, lactic acidosis, and stroke-like episode syndromes; maternal Hereditary diabetes and hearing loss, Leber's hereditary optic neuropathy, neuropathy, ataxia, retinitis pigmentosa maternalGenetic Leigh syndrome, Danon disease, diabetes, diabetic nephropathy, metabolic disorders, heart failure, ischemic heart disease leading to myocardial infarction, psychiatric disorders, schizophrenia, multiple sulfatase deficiency, mucolipidosis II, mucolipidosis III, mucolipidosis IV, GM1-gangliosidosis, neuronal ceroid-lipofuscinosis, Alper's disease, Barth syndrome, beta-oxidation defects, carnitine-acyl-carnitine deficiency, carnitine deficiency, creatine deficiency syndrome, coenzyme Q10 deficiency, complex I deficiency, complex II deficiency, complex III deficiency, complex IV deficiency, complex V deficiency, COX deficiency, chronic progressive external ophthalmoplegia syndrome, CPT I deficiency, CPT II 23. The compound, use or method of any one of paragraphs 20 to 22, wherein the compound is selected from the group consisting of: deficiency, glutaric aciduria type II, Kearns-Sayre syndrome, lactic acidosis, long-chain acyl-CoA dehydrogenase deficiency, Leigh's disease or syndrome, Leigh's syndrome French-Canadian variant, fatal infantile cardiomyopathy, Luft's disease, medium-chain acyl-CoA dehydrogenase deficiency, myoclonic epilepsy and ragged-red fiber syndrome, mitochondrial cytopathies, mitochondrial recessive ataxia syndromes, mitochondrial DNA depletion syndromes, neuromuscular gastrointestinal disorders and encephalopathies, Pearson's syndrome, pyruvate dehydrogenase deficiency, pyruvate carboxylase deficiency, POLG mutations, medium / short-chain 3-hydroacyl-CoA dehydrogenase deficiency, very long-chain acyl-CoA dehydrogenase deficiency, peroxisomal disorders, methylmalonic acidemia, mevalonate kinase deficiency, age-related decline in cognitive function and muscle strength, cognitive impairment associated with all neurodegenerative and neuropsychiatric disorders.

[0342] 24. The compound, use or method according to paragraph 23, wherein the neurodegenerative disease is selected from Parkinson's disease, Alzheimer's disease, amyotrophic lateral sclerosis, Huntington's disease, ischemia, stroke, dementia with Lewy bodies, multiple system atrophy, progressive supranuclear palsy, corticobasal degeneration, frontotemporal dementia, Parkinson's disease associated with mutations in alpha-synuclein, parkin, PINK1, GBA and LRRK2, and autosomal recessive juvenile-onset Parkinson's disease or early-onset parkinsonism (EOPD) in which parkin or PINK1 is mutated, truncated or deficient. 25. The compound, use or method according to paragraph 23, wherein said neurodegenerative disorder is Leigh syndrome or Leigh disease, X-linked Leigh disease, Leigh syndrome French Canadian variant and / or a condition associated with Leigh disease. 26. The compound, use or method according to paragraphs 20 to 22, wherein the cancer is selected from breast cancer, ovarian cancer, prostate cancer, lung cancer, kidney cancer, stomach cancer, colon cancer, testicular cancer, head and neck cancer, pancreatic cancer, brain cancer, melanoma, bone cancer, liver cancer, soft tissue cancer, cancer of tissue organs, cancer of blood cells, CML, AML, mantle cell lymphoma, neuroblastoma, melanoma, soft tissue sarcoma, liposarcoma, fibroblastic sarcoma, leiomyosarcoma, hepatocellular carcinoma, osteosarcoma, esophageal cancer, leukemia, lymphoma, multiple myeloma, metastatic carcinoma, osteosarcoma, chondrosarcoma, Ewing's sarcoma, nasopharyngeal carcinoma, colorectal cancer, non-small cell lung cancer, cancers in which the apoptotic pathway is dysregulated, and cancers in which proteins of the BCL2 family are mutated or over- or under-expressed. 27. The compound, use or method according to any one of paragraphs 20 to 22, wherein the fibrosis is selected from fibrosis or fibrotic disorders associated with the accumulation of extracellular matrix components following trauma, inflammation, tissue repair, immune response, cellular hyperplasia and neoplasia.

[0343] 28. The compound, use or method according to paragraph 27, wherein said fibrosis is selected from fibrotic disorders associated with fibrosis or major organ disease, fibroproliferative disorders, and scarring associated with trauma. 29. The compound, use or method according to paragraph 28, wherein the fibrosis is selected from fibrosis or fibrotic disorders associated with interstitial lung disease, cirrhosis of the liver, non-alcoholic fatty liver disease, non-alcoholic fatty liver disease and non-alcoholic steatohepatitis, kidney disease, acute kidney disease, acute kidney injury, chronic kidney disease, delayed renal graft function, heart or vascular disease, eye disease, systemic and localized scleroderma, keloids, hypertrophic scars, atherosclerosis, restenosis, Dupuytren's contracture, surgical complications, chemotherapy-induced fibrosis, radiation-induced fibrosis, accidental injury and burns, retroperitoneal fibrosis and peritoneal fibrosis / peritoneal scarring. 30. The compound, use or method according to paragraph 29, wherein the fibrosis associated with interstitial lung disease is selected from sarcoidosis, silicosis, drug reaction, infection, collagen vascular disease, rheumatoid arthritis, systemic sclerosis, scleroderma, pulmonary fibrosis, idiopathic pulmonary fibrosis, common interstitial pneumonia, interstitial lung disease, idiopathic fibrosing alveolitis, bronchiolitis obliterans and bronchiectasis. 31. The compound, use or method according to paragraph 29, wherein said renal disease is acute kidney disease, acute kidney injury or chronic kidney disease. 32. A pharmaceutical composition comprising a compound of formula (I) as defined in any one of paragraphs 1 to 18, a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer, together with one or more pharmaceutically acceptable excipients. 33. Formulas (II)(i), (III)(i), (II)(II) and (III)(ii):

[0344] [ka]

[0345] (In the above formula, R 1 , R 2 , R 3 , R 4 and R 5 is as defined for a compound of formula (I) according to any one of paragraphs 1 to 18, and PG is a protecting group, preferably tert-butyloxycarbonyl, benzyloxycarbonyl, p-methoxybenzylcarbonyl, 9-fluorenylmethyloxycarbonyl, acetyl, benzoyl, benzyl, carbamate, p-methoxybenzyl, 3,4-dimethoxybenzyl, p-methoxyphenyl, tosyl, trichloroethoxycarbonyl, 4-nitrobenzenesulfonyl, and 2-nitrophenylsulfenyl), a tautomer thereof, or a salt of said compound or tautomer.

Claims

1. Formula (I)(i) and Formula (I)(ii): 【Chemistry 1】 (In the above formula, R 1 (C 1 -C 4 ) alkyl, (C 1 -C 4 ) Fluoroalkyl and CH 2 OCH 3 Selected from R 2 (C 1 -C 4 ) alkyl, CF 3 and cyclopropyl, and R 3 , R 4 and R 5 are each independently selected from hydrogen and halogen), a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer.

2. R 1 is methyl, CH 2 F, CHF 2 , C.F. 3 and CH 2 OCH 3 The compound of claim 1 selected from:

3. R 1 is methyl and CH 2 OCH 3 The compound of claim 2 selected from:

4. R 2 is methyl, CF 3 The compound according to any one of claims 1 to 3, wherein the compound is selected from the group consisting of cyclopropyl, ...

5. R 2 5. The compound of claim 4, wherein is selected from methyl and cyclopropyl.

6. R 3 The compound according to any one of claims 1 to 5, wherein is selected from hydrogen and fluorine.

7. R 3 The compound of claim 6, wherein is hydrogen.

8. R 4 and R 5 The compound of any one of claims 1 to 7, wherein each is hydrogen.

9. Formula (I)(i): 【Chemistry 2】 9. The compound according to any one of claims 1 to 8, or a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer.

10. Formula (IA)(i): 【Transformation 3】 10. The compound of claim 9, having the formula:

11. 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(fluoromethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, and 11. The compound according to claim 10, or a pharmaceutically acceptable salt thereof, selected from 5-(5-cyano-4-fluoro-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide.

12. Formula (IB)(i): 【Chemistry 4】 10. The compound of claim 9, having the formula:

13. 4-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, and 4-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 13. The compound according to claim 12, selected from: or a pharmaceutically acceptable salt thereof.

14. Formula (I)(ii): 【Transformation 5】 9. The compound according to any one of claims 1 to 8, or a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer.

15. Formula (IA)(ii): 【Transformation 6】 15. The compound of claim 14, having the formula:

16. 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5S)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-2-(trifluoromethoxy)phenyl)-N-((3R,5S)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, 5-(5-cyano-4-fluoro-2-methoxyphenyl)-N-((3R,5S)-1-cyano-5-methylpyrrolidin-3-yl)oxazole-2-carboxamide, and 5-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-(fluoromethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 16. The compound of claim 15, selected from: or a pharmaceutically acceptable salt thereof.

17. Formula (IB) (II): 【Transformation 7】 15. The compound of claim 14, having the formula:

18. 4-(5-cyano-2-cyclopropoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, and 4-(5-cyano-2-methoxyphenyl)-N-((3R,5R)-1-cyano-5-(methoxymethyl)pyrrolidin-3-yl)oxazole-2-carboxamide, 18. The compound of claim 17, selected from: or a pharmaceutically acceptable salt thereof.

19. 19. A compound according to any one of claims 1 to 18, a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer for use as a pharmaceutical.

20. CNS disorders, neurodegenerative diseases, Parkinson's disease, Alzheimer's disease, amyotrophic lateral sclerosis, Huntington's disease, ischemia, stroke, Lewy body dementia, frontotemporal dementia, multiple sclerosis, mitochondrial encephalopathy, lactic acidosis and stroke-like episode syndrome, maternally inherited diabetes and hearing loss, Leber's hereditary optic neuropathy, neuropathy, ataxia, retinitis pigmentosa - maternally inherited Leigh syndrome, Danon disease, diabetes, diabetic nephropathy, metabolic disorders, heart failure, ischemic heart disease leading to myocardial infarction, psychiatric disorders, Schizophrenia, Multiple Sulfatase Deficiency, Mucolipidosis II, Mucolipidosis III, Mucolipidosis IV, GM1-Gangliosidosis, Neuronal Ceroid-Lipofuscinosis, Alper's Disease, Barth Syndrome, Beta-Oxidation Defect, Carnitine-Acyl-Carnitine Deficiency, Carnitine Deficiency, Creatine Deficiency Syndrome, Coenzyme Q10 Deficiency, Complex I Deficiency, Complex II Deficiency, Complex III Deficiency, Complex IV Deficiency, Complex V Deficiency, COX Deficiency, Chronic Progressive External Ophthalmoplegia Syndrome, CPT I Deficiency, CPT II Deficiency, glutaric aciduria type II, Kearns-Sayre syndrome, lactic acidosis, long-chain acyl-CoA dehydrogenase deficiency, Leigh's disease or syndrome, Leigh's syndrome French-Canadian variant, fatal infantile cardiomyopathy, Luft's disease, medium-chain acyl-CoA dehydrogenase deficiency, myoclonic epilepsy and ragged-red fiber syndrome, mitochondrial cytopathies, mitochondrial recessive ataxia syndrome, mitochondrial DNA depletion syndrome, neuromuscular gastrointestinal disorders and encephalopathies, Pearson's syndrome, pyruvate dehydrogenase deficiency, pyruvate carboxylase deficiency 20. A pharmaceutical composition comprising the compound of any one of claims 1 to 18, a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer, for use in the treatment or prevention of a condition associated with mitochondrial dysfunction selected from the group consisting of deficiency, POLG mutation, medium / short-chain 3-hydroacyl-CoA dehydrogenase deficiency, very long-chain acyl-CoA dehydrogenase deficiency, peroxisomal disorders, methylmalonic acidemia, mevalonate kinase deficiency, age-related decline in cognitive function and muscle strength, and cognitive impairment associated with all neurodegenerative and neuropsychiatric disorders.

21. 21. The pharmaceutical composition of claim 20, wherein the neurodegenerative disease is selected from Parkinson's disease, Alzheimer's disease, amyotrophic lateral sclerosis, Huntington's disease, ischemia, stroke, dementia with Lewy bodies, multiple system atrophy, progressive supranuclear palsy, corticobasal degeneration, frontotemporal dementia, Parkinson's disease associated with mutations in alpha-synuclein, parkin, PINK1, GBA and LRRK2, and autosomal recessive juvenile-onset Parkinson's disease or early-onset Parkinson's disease (EOPD), in which parkin or PINK1 is mutated, truncated or deficient.

22. 21. The pharmaceutical composition of claim 20, wherein the neurodegenerative disorder is Leigh syndrome or Leigh disease, X-linked Leigh disease, Leigh syndrome French Canadian variant and / or a condition associated with Leigh disease.

23. 20. A pharmaceutical composition comprising the compound of any one of claims 1 to 18, a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer, for use in the treatment or prevention of a cancer selected from breast cancer, ovarian cancer, prostate cancer, lung cancer, kidney cancer, stomach cancer, colon cancer, testicular cancer, head and neck cancer, pancreatic cancer, brain cancer, melanoma, bone cancer, liver cancer, soft tissue cancer, tissue organ cancer, cancer of blood cells, CML, AML, mantle cell lymphoma, neuroblastoma, soft tissue sarcoma, liposarcoma, fibroblastic sarcoma, leiomyosarcoma, hepatocellular carcinoma, osteosarcoma, esophageal cancer, leukemia, lymphoma, multiple myeloma, metastatic carcinoma, chondrosarcoma, Ewing's sarcoma, nasopharyngeal carcinoma, colorectal cancer, non-small cell lung cancer, cancer in which the apoptotic pathway is dysregulated, and cancer in which the BCL2 family of proteins is mutated or over- or under-expressed.

24. 20. A pharmaceutical composition comprising a compound according to any one of claims 1 to 18, a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer, for use in the treatment or prevention of a fibrosis selected from fibrosis or fibrotic disorders associated with the accumulation of extracellular matrix components following trauma, inflammation, tissue repair, immune response, cellular hyperplasia, and neoplasia.

25. 25. The pharmaceutical composition of claim 24, wherein the fibrosis is selected from fibrosis or fibrotic disorders associated with major organ disease, fibroproliferative disorders, and scarring associated with trauma.

26. 26. The pharmaceutical composition of claim 25, wherein the fibrosis is selected from fibrosis or fibrotic disorders associated with interstitial lung disease, cirrhosis, non-alcoholic fatty liver disease, non-alcoholic steatohepatitis, kidney disease, acute kidney disease, acute kidney injury, chronic kidney disease, delayed renal graft function, heart or vascular disease, eye disease, systemic and localized scleroderma, keloids, hypertrophic scars, atherosclerosis, restenosis, Dupuytren's contracture, surgical complications, chemotherapy-induced fibrosis, radiation-induced fibrosis, accidental injury and burns, retroperitoneal fibrosis and peritoneal fibrosis / scarring.

27. 27. The pharmaceutical composition of claim 26, wherein the fibrosis associated with interstitial lung disease is selected from sarcoidosis, silicosis, drug reactions, infectious diseases, collagen vascular diseases, rheumatoid arthritis, systemic sclerosis, scleroderma, pulmonary fibrosis, idiopathic pulmonary fibrosis, common interstitial pneumonia, interstitial lung disease, idiopathic fibrosing alveolitis, bronchiolitis obliterans, and bronchiectasis.

28. 27. The pharmaceutical composition of claim 26, wherein the renal disease is acute renal disease, acute renal injury, or chronic renal disease.

29. 19. A pharmaceutical composition comprising a compound of formula (I) as defined in any one of claims 1 to 18, a tautomer thereof, or a pharmaceutically acceptable salt of said compound or tautomer, together with one or more pharmaceutically acceptable excipients.

30. Formulas (II)(i), (III)(i), (II)(II) and (III)(ii): 【Transformation 8】 (In the above formula, R 1 , R 2 , R 3 , R 4 and R 5 is as defined for compounds of formula (I) according to any one of claims 1 to 18, and PG is a protecting group selected from tert-butyloxycarbonyl or benzyloxycarbonyl, a compound, a tautomer thereof, or a salt of said compound or tautomer.

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